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Literature Search No. 721 Toxicity and Pyrolysis of Guaiacol

Date: 14 Dec 1987
Length: 363 pages
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December 14., 1987 LITERATURE SEARCH NO. 721 TOXICITY AND FYROLYSIS OF GUAIACOL Requested by
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LITERATURE SEARCH NO. 721 T(1`1ICT:".X PT1?l PY'^n7.Y^~Ic Cn. M' n:7 nc[1L L~_
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LITERATURE SEARCH NO. 721 SEARCH TERMSe GUAIACOL r r]-HYDROX YAN I~ SOLE -JETHOXYPHENOL PYROGUAIAC.ACID ~ 2-ETHOXYPNENOL 2-HYDROXYANISOLE O-METHYLCATECHOL ANASTIL SUAIASTIL PYROCATECHOL MONOMETHYLETHER RN=90-05-1 PRINT SOURCES SEARCHEDe Ar-ctander, S. 1969. Perfume and Flavor Chemicals: Aroma Chemicals. By the Author, Elizabeth JN, Chemical Sources Association. 1985. Flavor and Frangrance Materials. Allured Publishing Corp., Wtneaton;IL, . Alimentarius Commission.198,;..Codex Alimentarius. Vol. 14. CAC/Vol. 14 - Ed. 1. FAO, Rome,
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Commerce Clearing House. 1980. Food Drug Comestic Law Reporter. CommerceCleariing House, Chicago, IL, Council of Europe. 19.81. F1:avouring Substances and Natural Sources of Flavourings. 3dd ed. Maisonneuve, Strasbourg. Department of Health and Sociall Security. 1979. CHunter] List of Permitted Additives to Tobacco Products. InoSecond Report of thei, I.ndEpendent Scientific Committee of Smoking and.Health:. Develomentss in Tobacco Products and the Possibility of "Lower- Ri.sk"Cioarette.s. Her Majesty.'s°$tationery Office, London, Fend.roli's Handbook of Flavor~Inoredients. 1975. (Edited, tra'fislated, and:revised by T.£.:'..f~uria and N.. Bellanca7, 2nd ed. ChE;mical Rubber Co., Cleveland, OH, ~j + Fla~or and Extract Marn.ifacturers' Association. 1985. Scientific Li ~rature Review of Phenolsin Flavor Usage. Report no. P086- 15 68. NT3S., Springfield, VA. Foorti Chemicical Codex. 1981. 3d>ed. National Academy of Sciences, Washington, D.C.,. . A Gos°selin,, R'. E.; Hodge, H. Smith,R.. F'.; and Gleason, M. N. 19~6. Clinical Toxicology`'-~o~f Commercial Products. 4th ed'~. WiYaiams and Wilkins, Baltimore, MD, doi',nt FA0/WHO Expert Committee on Food Additives. 1964. FAO-WHO Fodd Additi:ves Datay-System. FAOFood and Nutrition Paper no. 30. FAO, Rome, Mer.;'ck. Index.. 1983. 1S?thed.Merck & Co.,.Rahway,, N.1. Opc'fyk:e, D.L.J'., ed. 1979. Monora hs on Fra rance Raw Materials.Pergamon Press, Okford, .. Sax, N. I. 1984.. Dangerous Properties of Industriali Materials. -,6th ed. Van Nostrand Reinhold, New York,
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ONLINE SOURCES; NAME . 1975-87/SEPT 19h3-87/NOV d196?-87/DEC. A 1987/DEC ~~ 19Q;4/AUG 19~~~8-87 / AUG- 6-87/DEC 19&4-87/ISS24 t. ~. 19~--57/AUG' 19G7/JUL ~ $ 197€()-07/AUG 1966-87/DEC' ABRIS INTERNATIONAL CANCERLIT CHEMICAL ABSTRACTS CHEMICAL CARCINOGENESSS RESEARCH CHEMICAL•EXPOSURE CRIS/USDA FEDERAL RESEARCH IN PROGRESS FOODSADLIbRAFSTA HAZARDOUS.SUBSTANCES DATABASE LIFE SCIENCES.COLLECTYON MEDLINE NTIS` OCCUPATIONAL SAFETY AND HEALTH (NIOSH) REGISTRY OF TOXIC EFFECTS OF CHEMICAL SUBSTANCES SMOKING AND HEALTH TOXLINE . , . ..
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(Jl 0 Cl , RN 90-05-1 INPhenoi, 2-methoxy- (9CI) 8Y Guaiacol . SY o-Hydroxyanisole SY ',o-Methoxyphenol SY ~,;Pyroguaiac acid SY,'Phenol „o-methoxy- (8CI) SY :2-Methoxyphenol SY <2-Hydroxyanisole SY 1,1-Hydroay-2-methoxyhenzene SY ,,3ro-Methyl.` catechol SY P~.Anastil' " !Guaiastil SY~ ~ o-Guaiacol ~~ - SY; Pyrocatechol monomethyl ether MF' C7 H8 `02 CI COM LCi`CASREACT, TSCA C. .OMe C: . C, Cc' C, ;~ :C. OH REFERENCES IN FILE CAOLD (PRfORTO 1967) 221"1 REFERENCES IN FILE CA (1967 TO DATE)
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Molecular weight Melting point Boiling point Specificgravily Refractive index Solubility o-Hydroxyanisole o-Methoxyphenol MethylcatecholPyrocatechol monomethyl ether CiH.Or OH ~ OCHr ,'Yellowish, low-mclling crystals or liquid; tends to darken on exposure to light J 124.14 J 27,9°C J 205`C; 107'C at 24 mm,Hg ~ 1.1395 (J t I 12 liquid) . - 1.5341 at 34.8°C . - Siightlysoluble in waler; soluble inalco- hol, ether, and chloroform , Characteristic sweet odor; slightly pheno-lit Obtaine& from hardwood tar or syn- thetically from o-nitrophenol via uanisidine° Discovered in the distillation product from guaiacresin: Guaiacol is found in . castoreum oil,7 in the essential oil from flowers of Pandaaas odorarissimas L,° in the distillation waters of orange.' leavcs,' in the essential oil of Rura nnurtatm L.,'" in'the essential oil from celery seeds,!` and in the oil from tobacco leaves" Non-alcoholic beverages 0.95 ppm Ice cream,, ites,.etc. 0.52 ppm Candy 0.96 ppm Baked goods 0.75 ppm FDA 121.1164; FEMA No. 2532 REFERENCES For References 1-5, see end of Part III. 6. Kalle, German Patent 95,339, 1896. .:. 7. Ledcrer,Prog. Clrin Subst.Org. Mat, 6, 113, 1950. 8. lanistyn,Cbem. Zentra(bL, 1, 2263, 1938. . 9. Igolen, C7tem. %emmllil., 2, 750, 1939. . - , 10. St. Ptau,Hefe Grsm. Acta, 22, 382, 1939. . 1 t. Small, @em. Absts, 42, 6149, 1948. . Oni.thi et al., Clrem. Abstr., 50. 15028, 1956. Synthetic.Flavors 225 Guaiacyl acetate Acetyl guaiacol o-Methoxyphcnyl acetate CyH1oOs 0 O-C-CH, O-CH, Colorless liquid 166.18 i 235-240°C; 123-124'C at 13 mm Hg 1-156 af-0'C Almost insoluble in water;; miscible with alcohol and ether Odor similar to that of guaiacol From guaiacol and excess acetic anhydride in the presence of trace amounts of HiSO. FDA 121.1164
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rtn.A +'OA CaE rvaT GB SLR GA> roa: RIeM EUA 1HiA Gua..c gum .traca (gusiacum epp.f 2531 172.510 220 1 2 9000-29-7 XSOURCE. Chart Gualaop1532 172.515 J173 2 5 C12. 90-05-1 X SOURCE: GUAR.NCC Guaiaeoll phenylacetate SEE Guaiacyl phenylacetate Guaiaowood acetate ' 61789-17-1 Gualac wood extract. (gualacum app.) 2533 172.510 220. 11 2 SOURCE: Nk:kstadt-Moeller Gualae wood oil (qualaeum spp.) 2534 172.510 220 1. 28o16-23-7 X X SOURCE Amigo & ArditCFDL.Ungerer,Zimmermam,Manhaimer Polarome,RoureGualacyl acetate 3687 172.515 552 2 5 C12 613-70-7 _ SOURCE CTC;NCC,FDO,Ungerer;Caro , Guaiacyl phanylacatate 2535 172.515 238 3 5C12 4112-89-4 SOURCE NCC.Givaudan.Oxford Guaic acetateSEE. Guaiol acetate Guaicwood acetate SEEGuaioi acetate Gu'alena 172.515 2 C2 688-84-6 X SOURCEFDO Guai-l-en-it-o1 acetate SEE Guaiol acetateGualol aeatata 172.515 3 n5 61789-17-1 X SOURCE EIan.IFF.Ungerer,Polarome,CMA,Raure Guaiyl acetate SEE Guaiol acetateGuarana gum (paullinla cupana hbk) 2536 172.510 323 1 2 SOURCE Meer Guarana saed extract SOURCE Meer Guaranine SEE Caffeine Guar flour SEE Guar gum (cyamopsis tatragoralobus 0.11 Guar gum (cyamopsis tatragonolobus (IJ) 2537 184,1339 1661. 3 9000-30-0 SOURCE Meer.Madis.Tic Guava juice .. SOURCE Pembroek Guava. (psldulm sppd 182.20 1 Gum benjamin SEE Benzoin, resin Istyrax spp.) Gum camphor SEE d-Camphor Gum cyamopsis SEE Guar gum (cyamopsis tetragonolbbus fq) Gurjun balsam SOURCE Polarome Gyrane SEE 2-8utyl-4,6-dimethyldihydropyran Hagan phosphate SEE Sodum hexametaphosphateHamameils extract SOURCE CAL Haw bark, black, extract (vibumumprunlfol{um I.T 2538 172-510 48o 1 3 SOURCE: Meen,Pembroek Hawthanol OFF I SEE artho,meza,para-Methyl benzene ethanol ~ Mero is now CMA INTERNATIONAL 'NKRwYFS 8030-55-5 X HAY absolute 104 Flavor &Fragranoe Materials - 1985 - Allured Publishing
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1E I Introduction This is the fourth compilation of this sedes,'• 2-' and the only comprehensive directory of materials used In the creation of flavors and fragrances. Dr. Bruce K.. Bernard, Director of Scientific Research Associates, Inc.', and Consulting Director of Scientific Affairs for the Flavor and Extract Manufac- turers' Association (FEMA)'served as Consulting Editor. The original document containing GRAS flavor substances with much additional international reg- ulatory and scientific information was compiled by Dr. Richard A. Ford, who was then directing the scientific work of the FEMA. The flavor materials In this publication include the current materials listed in the FEMA GRAS lists, including GRAS 13. Additional flavor materials have been included from fnfonnation supplied by the Chemical Sources Association and from many users and suppliers of flavor materials. The fragrance materials were compiled from the list of materials which RIFM has Investigated for their safety in fragrance use, plus the list of substances for which there are published EOA/FMA standards, plus additional materials which have been submitted by fragrance companies and suppliers of fragrance materials. Anyone having knowledge of, or Interest in, flavor and fragrance materials is aware of at least two major problems. First, a substance may be referred to or identified by a multitude of different terms or names. Secondly, a single name or term may be employed for a variety of separate and distinct substances. While we have tried to avoid either error, instances of both will more than likely be found~ in-tFiis volume. When found,, we appreciate you bringing these to the attention of the Editor. References are given after each principal name, providing the following information on each ap- plicable material:
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at~ sfuJii3dsa3{i;aludCm:~S:.l;,,:. SLR SLR Code Title *85A Primary alcohols, aldehydes, acids and related esters with thiol or sulfide functions "B58 Selected oxygenated derivatives of inercaptansand sulfides *B5C Aliphatic thiol esters *66 Substituted pyrazines C1 Piperonal and relatedd substances C2 Aromatic hydrocarbons *C3 Salicvlates and ~_salicylaldehyde *C4 Aryl substituted tertiary alcohols and esters *C5 Aromatic thiols and sulfides C6 Phenethyl alcohol and related'compounds C7 Fused ring aromatic lactones C8 Eugenol and related substances "C9 Benzyl alcohol, benzaldehyde, benzoic acid and related compounds C10 3-Phenyl-l-propanol and related substances = C11 Cinnamyl alcohol and ~. related substances ~' •C12 Phenols C13 Acetophenones and related substances tll . ~..+odfi'sl~i:ii~'•ii;11ti1il{i~A&9llFNiifeBFG~tl S Number of Substances Covered Date Com- lp eted. rder No. rice SLR Cod C1i 17 2/84 P88 5141141 A05 C1_ 14 10/83 P88 5141158 A04 C1( 5 2/84 PB8 5141265 A05 C17 34 1/84 P88 5141174 A09 Clf 5 11/77 PB 283-499/AS A04 Clf 7 11/78 P8 291-117/AS A09 C2( 10 1/84 PB8 5141182 A07 C21 10 10/83 PB8 5141190 A04 11 12/83 PB8 5141208 A06 C2Z C2: D1 41 9/78 PB 291-114/AS A14 *D2 5 2/78 PB 283-502/AS A04 D3 12 5/78 PB 283-501/AS A07 D4 31 2/84 P88 5141216 A14 D5 m %I D6 11 5/78 PR 283-504/AS A05 la tQ 07 31 7/78 PB 284-961/AS A10 O ~ D8 46 1/84 P88 5141224 A22 14 10/78 PB 291-105/AS A06 *D9
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LIST I LIST OF FLAVOURING SUBSTANCES WHICH MAY BE ADDED TO FOODSTUFFS WITHOUT HAZARD TO PUBLIC HEALTH LISTE I LISTE DES MATII:RES AROMATISANTES POUVANT BTRE AJOUTEES A L'ALIMENTATION SANS RISQUE POUR LA SANTE PUBLIQUE Note: When calculating the usable dose, it should be understood that - if the limits indicate only the ADI - other possible dietary intakes of the same substance must be taken into aceount' in arriving at.a technologically useful limih For the definition of group limits see comments on page 117. En ce qui concerne k calcul de Ia doseadmissibk,.il est bien entendu que, si les limites mentionnenY seulement Ia DJA, & faut tenir compte des autres apports didt8tiques possibles de Ia m8me substance pour en arriver & la dose technologique pratique. Pour Ia dEfinition des limites de groupe voir commentaires page 121. 123 I
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fi (~,v~- L i s -7- Ct . List 2 Additives, that may beapplied either at a stage in the manufacture of, or directly to, the.Gnal tobacco blend suchthat.the total level in the GnaUtobacco blend does not exceed 0,15 per cent in cigarette, cigar or hand-rollingtobacco and 0.5 per cent in pipe tobacco, calculated on a dry weight basis. The maximum permissible level at which.any individual additive may be.added shall be as indicated in the list under the appropriate heading. Cigarette, cigar and hand- rolling rolling Pipe Additive. Biological origin Synonym tobacco tobacco (p8/g) (pg/g) ginger oil. (Zingiber ojfrcinale) glycerol 1,2-diacetate 'tk' guaiacol guaiac wood oil (Bulnesia sarmienti, Guaiacum oJfrcinale, Guaiacum sanctum) guaiyl acetate q A t (I-methyl-1[1,2,3;4,5,6,7, 8-octahydro-3,8- dimethylazulen-5-yl] ethyl acetate) heptan-4-olide ( 7-heptalactone) hexanoic acidhex-2tny1 acetate hexyl acetate 3-hydroxybutanone (acetoin) 7-hydroxy-3,7-dimethyloctanal (hydroxycitronellal) 2-hydroxy-3-methytcyclopent-2-en 1-one (methylcyclopentenolone) 50 100 isoeugenol (4-prop-I-enylguaiacol) isopentyl alcohol (3-methylbutan-l-ol). 4-isopropylbenzaldehyde isovaleraldehyde isovaleric acid 3-hydroxy-2-methyl-4-pyrone (maltol) 4-(4-hydroxypheny)butanone indole a -ionone (Lionone a -irone isobutyl phenylacetate . jasmine oil (Jasminum o/'feeinale grand{Jlorum)) juniper berry oil (Juniperus communis)'labdanum (Cisrns spp ) labdanum oil (Ci.rtus.spp)lavender oil (Lavandula offieinali.r, Lavandula vera) 1 2 140 140 I
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"AJag 66i1ptlela, 'sOilewJo;uI .m!t Itl,, II EXPLANATORY NOTES Column I - Council of Europe serial number Column 2 - Formula Column 3 - Systematianame (Brussels nomenclature) Column 4 - Synonym(s) Column 5 - ADI in mg/kg bodyweight, where allocated Columns 6, 7 and 8- Specific limits in mg/kg for various food items, namely: Beverages, Food and Exceptions Abbreviations: OFAU - Other food additive uses JECFA - ADI in mg/kg bodyweight allocated by the Joint FAO/WHO Expert Committee on Food Additives (JECFA), with the meeting number given in brackets CE - ADI in mg/kg bodyweight allocated by the Council of Europe CEp - Provisional ADI in mg/kg bodyweight allocated by the Council of Europe * - According.to information currently available to the Committee of Experts the substances marked with an asterisk in the list either have no intlrinsic flavouring properties or are used primarily for other purposes a) candy,.confectionery, b) condiments, seasonings c) alcoholic beverages d) soups, savouries 125
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I GENERAL TOX.i_.CITY STUDIES ROUTE: sut+cut_aneous SPECSESct pigeon STUDY TYPEa LDLo DOSE: 400 mg/kg EFf'ECT:MO TOXIC EFFECT NOTED ftEFERENCE:Pood Chem Tmx.ical, vol 20 (SUpP ') , pg. 69'7, 1,.8: (FCTOD7) REPRODUCTIVE STUDIES TOXICOLOGY REVIEW CANCER REVIEW THi;ESHULDLIMIT VALUE NIOSH RECOMMENDED LIMITS STANDARDS AND F'iEGULATIONS. FEDERAL PFnOGRAM STATUS ND ND ND ND ND ND EPA GENETOXPfiOGRAM 1986, ncnmclusive: FEDERAL PROGRAM STA'IUS Histidine reversion-Ames tes.t. EPATSCA.CHEMICAL.INVENTOFY, 1996 FEDEFAL PROGRAM STATUS EPA TBCA.TEST SUBMISSION (TSCATS) DATA BASE, JUNE 19e7 FEDERAL PROGRAM STATUS MEETS r.F7.ITE[xIAFOfi PROPOSED OSHA MEDICAL RECORDS RULE; FEREAC 47,30420,82: Fed Regist 0
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RTECS - JULY 1987 CI RTECS RECORD NUMBER NAME OF SUBSTANCE CAS REGISTRY NUMBER L.AST REVISION DATE RECORDLENGTH RTECS ACCESSION NUMBER NAME OF SUBSTANCE CAS REGISTRY NUMBER SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS. SYNONYMS SYNONYMS SYNONYMS MOLECULAR FORMULA MOLECULAR WEIGHT CLASS OF COMPOUND~ CL-ASSOF COM6'O! !ND CHEMICAL DEFINITION WISWESSER LINE NOTATION DATA TYPE DATA TYPE DATA TYPE 5::3p21 PHENOL, o-METHOXY- 9i i-C15-1 8707 1947 NIOSH/SL7525600 PHENOL, o--MEfHOXY- 90--05-1 GUAIACOL GUAICOL o-H}7DROXYANISOLE ?-HYDROXYANISOLE 1-HYDROXY-2-METHOXYBENZENE o-METHOXYPHF_NOL- 2--METHOXYPHENOL PYROGUAIAC ACID METHYLCATECHOL C7-H8-02 124.15 Mutation d .ta Sk:in/EyeIrritant ND ND Muta.geni.c~ity Skin/Eye~Iriritatiun. General 'Tox~.ici~.ty MUTAGENTCITYSTUDIESTEST SYSTEM: sister chromatid excEi«ngeSFECIESLROUTE/ CELL TYPE:a human:lymphocyte DOSE: 500 umol/L REFERENCEo. Mutat Res,., vol 169, pg 1^c9, 1986 (MUREAVX, CARCINOGENIC.ITY STUDIES ND 24
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( !l `( i Introduction The first number following the principal name is that assigned the substance in one of the eleven FEMA GRAS lists "" Numbers 2001-3124, GRAS 3; 3125-3249, GRAS 4; 3250-3325, GRAS 5; 3326-3390, GRAS 6; 3391 3423, GRAS 7; 3424-3444,.GRAS 8; 3445-3475, GRAS 9; 3476-3525, GRAS 10; 3526-3596,.GRAS 11; 3597-3650, GRAS 12; and 3651-3739, GRAS 13. A separate list of all of these FEMA substances and their synonyms in numerical order begins on page 229. The second number following each principal name refers to the section of Tltle21 of the Code of Federal Regulations where the substance is listed. The third number refers to the number by which the substance is listed in the publication "Flavour- ing Substances and Naturaf Sources of Flavourings" of the Council of Europe (CoE).10' The fourth number indicates the class of. flavouring substances. The division of flavouring sub- stances into three categories was proposed by the International Organization of the Flavor Industry (IOFI) and has been adopted by the FAOMIHO Food Standard Programme, the Codex Alimen- tarius Commission.10• " The official definitions for the three classes of flavouring substances, indicated by the numbers 1, 2 or 3 read as follows: 1. Natural Flavours and Natural Flavouring Substanaes. For the purpose of the Codex Allmen- tarius, "natural flavours" and "natural flavouring substances" are preparations and single sub- stances, respectively, acceptable for human consumption, obtained exclusively by physical pro- cesses from vegetable, sometimes animal raw materials either in their natural state or proces- sed, for human consumption. 2. Nature-Identical. Flavouring Substances. For the purpose of the Codex Alimentarius, "nature- identical flavouring substances" are substances chemically isolated from aromatic raw matedais or obtained synthetically; they are chemically identical to substances present in natural products intended for human consumption, either processed or not. 3. Artificial Flavoudng Substances. For the purpose of the Codex Alimentarius, "artificial fiavour- ing substances" are those substances which have not yet been identified in natural products in- tended for human-consumption, either processed or not. The fifth number refers to one of the six appendices, numbers 1 through 6, of the "Report on Re- view of Flavouring in Food; "" published by the Ministry of Agriculture, Fisheries and Food of the United Kingdom (UK list). Not all substances on the UK list are included here. The sixth column is an alphanumeric code to the Scientific Literature Revlew (SLR) in which the substance is reviewed. This code refers primarily to SLRs produced by FEMA, originally under contract to the FDA. The titfes of these SLRs together with their order number are listed. These SLRs are available either paper bound or on microfiche from the Customer Service Natlonal Technical Information Services (NTIS), 5285 Port Royal Road, Springfield,. VA 22161, 703/487- 4660. The exact price can be obtained from NTIS by giving the order (PB) number. The FEMA SLRs cover groups of substances with related chemical structures and, therefore, presumably, related metabolic and toxicologic properties. For example, SLR Al covers 65 sub- stances from related chemical classes. Also included is, the date the SLR was completed and subsequently delivered to FDA. vi 3
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Introduction In those instances where the substance is covered only in a non-FEMA report, "PB" is indicated in the SLR reference column.ln these cases, the order numbers and price codes can be obtained from FDA. In those instances wliere FEMA SLRs and non-FEMA reports are both available, only the former is referenced as the laher will be noted therein. The seventh column gives tlte Chemical Abstracts Service (CAS) number for the substance. A number is assigned by CAS to each material which appears in the literature, and this number is be- cwming widely used as prime identification. An "X" in the eighth column indicates if the substance appears among the monographs published in the Third Editlon of the Food Chemicals Codex.'• CP An "X" in the ninth column indicates that the substance has a monograph in the series published by the Research Institute forfragrance Matenals." An "X" in the tenth column indicates'd there is a standard for the material published by the Essential Oil Assoclation of the U.S. (now known as the Fragrance Materials Association)." A letter in the 11th column indicates that material has a reference in the Guidelines of the Interna- tional Fragrance Association (IFRA)."These guidelines give the recommendations of IFRA for the safe use of fragrance materials, based on the latest research information available from published and unpublished sources, worldwide, of adverse reactions to the use of fragrance materials. The recommendations are grouped into three categories as follows: R-The use of the material should be limited quantitatively. S-The material should only be used if it meets certain purity criteria or'rf it is used in conjunction with other materials. P-The materials should not be used as a fragrance ingredient. Finally, the suppliers of bottt fiavor and fragrance materials have been listed. The information under "sources" In this volume has been taken from CAS" publications and files, as well as from information sent to us specifically for this Edition. Our aim Is to provide a full and complete source listing of all these materials woddwide. All the information that our readers may be able to contribute will be greatly appreciated. (I This puWication is intended to aid in the search for a particular substance in one of the categories mentioned above-Although a reasonable effort was made to find all listings, it is probable that some were missed. Accurate cross-referencing was difficult. This is especially true of the botanicals. In this case, it was assumed that where the species were the same on two or more referenced lists, the substance was the same unless it was deady differentiated by reference to a particular part of the plant, i.e., roots on one list and leaves on another. It is important, therefore, to emphasizedhat if accuracy if desired, the original reference should be consulted for verification. If you find any errors in this edition, it would be greatly appreciated if you could let us know so corrections can be made in the next edition. viB I I
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SKIN AND EYE IRRITATION ROUTE: skin SPECIES: rabbit DOSE; 5qr?mg../24H' EFFEL:T:SEVERE REFERENCE:. Food Chem Tor::i col , vol 20(Suppl ), pg198.^_' (FCTOD7) SKIN AND EYE 778, 1964 GENERAL TOXICITY STUDIES' ROUTE: oral SPEyIES: human STUDY TYPE: LDLoDOSE: 43 mg/kg EFFECT: BEHAVIORAL (Tremor) E:FFECT:. GASTROINTESTINAL (Other changes) REFERENCE: Toxicol Drugs Chem 1969,, pg 29.°,y 1969 (T4ZIAG) GENERAL TOXICITY STUDIiESRUUTE: oral SPECIES: rat STUDY TYPE: LD5r} DO SE: 725 mg/kg EFFECT: TOXIC EFFECTS NOT YET REVIEWED. REFERENCE: Toxicol Appl Pharmacol, vol 6, QTXAPA9) pg GENERAL TOXICITY STUDIES ROUTE: Subcu.tan_ous SPECIES: rat. STUDY TYPE.:LDLo~ DOSE: 900mg/kg EFFECT: NO TOXICEFFECT NOTED REFERENCE:. Food Chem Toxicol, vol 20(Suppl), 1'982 (FCTOD7) GENERAL. TOXICITY STUDIES ROUTE: oral SPECIES: mouse FTUDYTYPEe LD50DOSE: 621 mg/kg EFFECT: NO TOXIC EFFECT NOTED REFERENCE:. Drugs.Future, vol 5, pg :,-`_T.9, 1920 IRRITATION ROUTE: eye SPECIES: rabbit DOSE: 5 mg EFFECT: MILD REFERENCE: Food Chern Toxicol, vol 20(Suppl), pg 697, 1962 (FCTOD7) pg 697, . 697, (DRFUD4)
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GENERAL 'COX.I:CITY S'7UD:(ES ROUTE: inhalatzon I SPECIES; mous Je STUDY TYPE: LC50 DOSE: 7570 mq:/m3_. F_FFECT: NO TOXIC EFFECT NOTED REFER:ENCE`a Food Chem Too-;,icol, vol :°<'i(Supp.I), 1902 (FCT007)' 2c0( SI_lp p l) 9. 697, GENERAL TOXICITY STUDIES ROUTE: intravenou.rs. J SPECIES: mouse STUDY TYPE: LD5r) DOSE: 170 mg/Ic.g. EFFECT: NO TOXIC EFFECT NOTED REFERENCE: Food Chem Tox,icol, vol 20(Suppl),, pg 697, 1982 (.FCTOD7) . GENERAL TOXICI'TY STUDIES / ROUTE: ora J1 SPECIESe cat STUDY TYPE: LDLo DOSE: 1500 mglk9 EFFECT: NO TOXICEFFF_CT NOTED REFERENCE: Food Chem Tox,icoi, vol 2(J(Suppl), 19a2 (FCTOD7) OENER(iL TOXICITY STUDIES ROUTE: skin SPECIES: rabbit STUDY TYPE: LD5r7 DOSE; 4600 mg/kg EFFECT: NO TOXIC EFFECT NOTED REFERENCE: Food Chem Tox.icol, vol 1902 (FCTOD7) CENERAL T!OX.ICIITYSTUDIES ROUTE: subr_ut.aneousSPECIES: rabbit STUDY TYF'E: LDLo DOSE: 125<J mg/k:g EFFECT: NO TOXIC EFFECT NOTED REFERENCE: Food Chem Toxicol, vol 1962: (FCTOD7) GENERAL TOX.ICITY'STUDIES ROUTE: subcutaneous . SPECIES: guinea.pig. STUDY TYPE: LDLo DOSEc 901) mg/kg. EFFECT: NO TOXIC EFFECT NOTED REFERENCE: Food Chem Toxicol, vol 1982 (FCTOD7) 26 20 (Supp 1 ) ,. p9 pg 697, p9 697, J Pg 697, ~ ~ N 20(Suppl.),. pg 697, ~ N ~ 1
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Serfel Fo ula ti S t m a S D1 k Specific limits mg kg Food numbtr rm ys e a c n me ynonym A g bw me: , Bes'erages Fa~M Fscep- _ linns Llmites spEcifiques mg kg Numfro Formule . Nomenclature Synonyme DJA mg'kg pc do denri•es atimentuirec de sfrie systEmatique Bnissons :\liments Excep- hnns 169 cx=i:=e=o ~ I I 3-melhyl•4-(2,6,6- ¢-isomethylionon j 5 x3 x3 trimethyl-cyclo- hex-2-enylj-but- 3-en-2-oae , 170 cxycxrax:excxs 5-propenyl 2-efhoxy3• CE I P euaethol propenyl phenol ox 171 xo-=-~-cxzcx-oxr euHenol 2.5 T JECFA (23). CE: Tota . Y eugenol from 171, 172. 183, 18 and their esters CH30 172 Np-~ ( 7)-Cx~eNCx3 lSn°eagen0l 2-methozY-4- cf 171 ,.°. propenyl phenol . 4 Cx ~V x0 p l73 guaiaml 1 2 6:3 Cx3e \/ Specific limits mg'kg rood Serial Formula Systematic name Synonym AD[ mg,'kg bn ExccP- number Bevemges Food tiunc Limiles p' "iqims in>; kg "c imenlairrs de denr Numbro Nans~ ture - - - k DJh Formule , m Syaonyme mg F Pr dc sdrie .Kiiqnc srslc
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Introduction (I Cosmetic Materials A publication similar in style and design to Flavor and Fragrance Materials covering cosmetic raw materials called Cosmetic Bench Reference. This volumeJs published annually as the August issue of Cosmetics & Toiletries magazine. For a description and order form for the Cosmetic Bench Reference, write to Allured Publishing Corporation, P.O. Box 318, Wheaton, IL 60189- 0318, USA. References 1. Ford, R. A., and Cramer, G. M., Reference list ot flavoring substances in use in the United States. Perf. Flavodst 2(1),1, 1977 2. Ford, R. A., Flavor Materials 1979, Allured Publishing Corporation, Wheaton, IL 60189, USA 3. Flavor and Fragrance Materlafs-1981, Allured Publishing Corporation, Wheaton, IL 60189, USA 4. Scientific Research Associates, Inc., 200 St. Lawrence Drive, Silver Spring, Maryland 20901 USA, (202) 223-3283 5. Flavor & Extract Manufacturers Association, 900 17th Street, N.W., Suite 650, Washington, D.C. 20006, (202) 293-5800 6. Hall, R. L., and Oser, B. L., Recent progress in the consideration of flavoring ingredients under the Food Additives Amendment. FEMA GRAS Substances 3. Food Technol. 19(2), Part 2, 151, 1965; 4. Food Technol. 24(5), 25, 1970 7. Oser, B. L., and Hall, R. L., Recent progress in the consideration of flavoring ingredients under the Food Additives Amendment FEMA GRAS Substances 5. Food Technol. 26(5), 35, 1972 8. Oser, B. L., and Ford, R. A., Recent progress in the consideration of flavoring ingredients under the Food Additives Amendment. FEMA GRAS Substances 6. Food Technol.17(1), 64, 1973; FEMA GRAS Substances 7. Food Technol. 27(11), 56, 1973; FEMA GRAS Substances 8. Food Technol. 28(9), 76, 1974; FEMA GRAS Substances 9. Food Technol. 29(9), 70,1975; 1 FEMA GRAS Substances 10. Food Technol. 31(1), 65, 1977; FEMA GRAS Substances 11. Food Technol. 32(2),.60,1978; FEMA GRAS Substances 12. Food Technol.33(7), 65, 1979;. and FEMA GRAS Substances 13. Food Technol. 38(10), 65,.1984 9. Oser, B. L.; Ford, R. A.; and Bernard, B. K., Recent progress in the consideration of flavoring ingredients under the Food Additives Amendment. FEMA GRAS Substances 13. Food Technol. 38(10), 65, 1984 10. Council of Europe, Flavouring Substances and Natural Sources of Flavourings, Stras- bourg, 1981, p. 376 11. Grundschober, F.; Hall, R. L; Stofberg, J.; and Vodoz, C. A., Survey of Worldwide Use Levels of Artificial Flavouring Substances. Flavours, 1975, pp. 223-230 12. Guide to the Safe Use of Food Addhlves, FAO, CAC/FAL 5-1979, p. 6 13. Ministry of Agriculture, Fisheries and Food; Food Additives and Contaminants Committee, Report on the Review of Flavourings in Food, Her Majesty's Stationary Office, London, 1976, p. 190 m 3- 14. Flavor and Extract Manufacturers Association, 900 - 17th Street NW, Washington, DC 20006 15. Food Chemicals Codex, National Academy Press, 2101 Constitution Avenue NW, Washing- ton, DC 20418, 1981 a N N Cp 16. Research Institute for Fragrance Materials, 375 Sylvan Avenue, Englewoof Cliffs, NJ 07632, USA 17. Fragrance Materials Association of the United States, 900 - 17th Street NW, Washington, DC 20006 O tD ib 18. International Fragrance Association, 8 rue Charles-Hubert, CH-1205 Geneva. Switzerland 19. Chemical Sources Association, 900- 17th Street NW, Washington, DC 20006 fx i,
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OH f7h•tt'riprion mu! plp:vir+d.t/ruprrrirs: dlrrrA Utdr.v (1976C Oi'vurrence: Discovered in tlte dislilbuion prmhleP from guaiac resin, aud reported tit (wxur in calor- cum,in thcessenlial uil fronrllowers of Piaalaml.s rsfnrnrissinurs L., in the distillation watersof urung9 . leaves, in the essential oil of Rwa mnnruna L, in the essential oil from celery sceds and in the oil from t, nba.cc~av~(Finarufi s Nnndbook of FBunr Intpxvlirnts, 1975). Also rcported,to have been found in asparagus: celery, roasted onion, tomato, cinnamon, crisphrcad, milk and milk products, beur, rum, whisky, wine,.cocoa,, coffee, tea, ro•asted frlberl. roasted pe:mul, pupcorn, soya bean, arctic bramble, cloudbcrry, passion fruit undlroasted ses:rmeseed(CIVO-TNO. 1977kPrepninrirw: From o-nitrophenol rra o-anisidine and the diazo compound (Arctander, 1969):. Uses: In public use since the 1930s, Use in fragrances in the USA amounts lo.lessth:m IIq01b/yr. Concentralion in final product ("„1: _ Usual Maximum StwP 0-01 0~06 Detergent 0001 1 &006. Creanrs, furions G005 . (Y02 Perlinne 0ilg D2 FR1Af: F&GT Uo1. 20. (SUp, ) Nw. 1982 Fragrance raw materials munagmplu 697 GUAIACOI. L , . I Synonyms: o-Methaxyphenol; n_hydroxyanisole; 2-methuxyphenul: methylcmnhoL Strur'rure. I Analctiraf dara: Gas chromatogram, RIFM no. 77-199; infra-redcun•e. RIFM no. 77 199. ~ y y pp s a s ven , was g Guaiaco 172515)and ~ . ~ ~ -- - Status the FDA for food uso(21 CFR rovlid b b FBMA (19651 is a GRAS t m l i 'AS~.RegiAry Biological data' $ftgjU(9xicuy,jThe lowest.dosage resulting.in death following ingestion by humans was estimated . u.50mg/kg;tn mosl cases of clinical poisoning; guaiacol has been ingested as wood-tar creosote (Gosselin, Hodge, Smith & Gleason, 1976):..Guaiacol ingestion in humans has been associated with irritation, burning pain with vomiting,, and possibly bloody diarrhoea. Repeated small doses of guaiacol mayhave.produced tolerance. Cardiovascular collapse has been reported: to accampanythc ingestion of unstated doses of guaiacol used as an expectorant. The deathh of a 9-yr-old girl was attributed to the ingestion of 5 mhguaiacol.(Hake & Rowe,.1963): In rats, the LDsu of guaiacoL administered as a 20";, solution in corn oil by intubation, was 1-0-2-Og/kg, while the lethal oraldose was 4 g for rabbits and 60 drops.for cats (Hake & Rowe. 1963)., An oralLDSO of 725 mp/kg has been reported for young rats fasted for Ig hr; marked depressiomand a comalose condition were noted. withh death. occurring from I hr to 44 days a0cr trcmment. (Taylor, Jenncr & Jones, 1964). Liver lesions were noted inihcx rats at autopsy. - The following sc do:e.s of gtmiacol were lethal: 0-2 g in the pigeon. ].5 g.in the rabbit and 09 grRg in guinea-pigs and rats (Hake & Rowe. 1963). The ivLDsn in white mice was 170 mg,3,g (Siemoneit, Zipf & Dillmann, 1966). A mixture of guaiacol, sage und',cucalyplol injected im into it rabbit producerl no skin reactionor lesions of muscle tissue (Cioc &von Schilling, 19651. Thedermnl Ll),,, , valure in rabbits hasbeemreported as 4-6 gll:g (3'g-5-4.g/kgl (Moreno, 197g). vlnhalyjiqp. In mice, the maximumm tolerated concentration of guaiucol vapour was 203 mg/litre, Ihe LCsv was 7-57 mg/litre, and the LCiou was 1719 mg/litre (Ostrovskii, 1964). +ylrritn(ig,((, Gosselin er n7. (1976) reported that percutanr.~ousubsurplion of guaiacol is dangerous. Apparenrly, guaiacul may be absorbed byy human skin, with >2g reported as haz:adous:.. Serious irritation has been produced by exposure of rabbit skin to undiluted guaiacol for several 24-hr periods (Hake & Rowe, 1963). Guaiacol did not produce depigmentatiun when applied to guinea-pig ears as.a.20f;;; cream in lanolin once daily for up to 8 wk (Rilcy, 1969). The eyes of rabbits ttcre seriuuslyy injured byy adminiaralion uf I drop of Imdilincdpoaioculd corneal necrosis and inluryy to the conjunctival nremhranes heingrcpunedg hut, one dropuf a Ify';, solution impropylencEly~col was'.onlys milJly'irritallnullJake&Rowu. 196}I. • I.llcr:Unrc .e:nrchea Irom I'/b-' tlrrouph I'179.
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n3790?iAGR.IS No: 487677 Comparison of two.anestesic mixtures to dispersed toxicity in horse.: glyceryl-guaiacol-ether-penthothal~, chloral hydrate-magnesium sulfate-pentobarbital (Comparacion de dos me-<c:las.anestesicas a toxicidad di.sper-sa.en el caballo: gliceri.l-guayacol--et•er-pentotal,, hidrato de r_loral-sulfato d'ee magnesi~o--pentobarb.ital) Domdnguez Boxer, A.R. Undversidad Naci..mnal Autonoma d'e Mexica,. Mexico, DF. Facultad deM>_di.cini,Veterinaria y Zootecnia. Tesis (MVZ) Mexico, DF., 1977, 42 p. Notes: 11 ref Language: Spanish Availabilit.y: MexicoCent:.er Document Typee Monograph, Dissertation, Nonconventi.onal, . Literature Journal Announcement: 0603 Record input by IICA(Inter-Ameri:.can Inst of Agr Sciences) (l 19
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I DOT EMERGENCY GUIDELINES FIRE POTENTIAL NFPA HAZARDCLASS'IFICATION FLAMMAt?LE.L.IMITS FLASH POINT AUTOIGNITION TEMPERATURE FIRE FIGHTINGPROCEDURES INTENSITY OF HEAT TOXIC COMBUSTION PRODUCTS OTHER.F'IRE FIGHTING HAZARDS EXPLOSIVE LIMITS AND POTENTIAL REACTIVITIES.& INCOMPATIBSLITIES DECOMPOSITION POLYMERIZATION OTHER HAZARDOUS REACTIONS ODOR THRESHOLD y SKIN, EYE AND RESPIRATORY C~ IRRITATIONS faROTECT I VE EQUIPMENT & CLOTHING OTHER PREVENTIVE MEASURES STABILITY/SHELF LIFE (1, SHIPMENT METHODSAND.. REGUti.ATIONS SHIPMENT METHODS AND. REGULATIONS STORAGE CONDITIONS CLEANUP METHODS DISPOSAL METHODS RADIATION LIMITS AND POTENT IiAL TOXICITY SUMMARY ND ND ND ND iBO DEG F OC [Hawley, G.G. The Condensed Chemical'L`ici-ionary. 9th ed.. New Yor4t: Van NostrandRei~nholdCo., 1977.: , p. 4243 **ACCEPTED** ND ND ND ND ND ND ND ND ND ND ND ND ND ND DARK.ENS ON EXPOSURE 70 AIR'.E+. LIGHT [The MerckIndex. 9th ed. . Rahway,, New Jersey; Merck & Co.,. Inc., 1976. , p. 5913 *#ACCEPTED** CONTAINERS: (CRYSTALS) DRUMS & TINSg (LIQUID) DRUMS & CARBOYS. CHawley,. G.G. The Condensed Chemical Dictionary. 9th ed. New. York: Va.m.Nostrand Reinhold Co. , 1977. , p. 4243 **ACCEPTED** PROTECT FROM LIGHT. [The Merck: Index. 9th ed. Rahway, New Jersey: Merck &Co.,. Inc., 1976. i, p. 5917'**ACCEPTED*+r ND ND ND ND ND 16 ...... .__ .__ _ Jid
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MAJOR USES STIMULANTEX'.Pk=.CTOi"iANT, CHEM INT' FOR OTHER EXPECTORANTS, Eti, GUAIACOI. CAft6ON4:rTE; FLAVORINGAGENT FOR NON'--ALCOHOLICSEVERAGES AND FOODS., CHEM INL!- FOR PAPAVERkNE & VANILLIN C£;ftI ]**ACCEPTEDxw MAJOR USES METHOXYPHENOL. IS USED IN MFF: OF MEDICINES, GUAIACOL COMPD, & F'ERFUMES. CNational. Research Council. Drinking Water & Health Volume 1. Washington, DC: National Academy Press, 1977. ,, p. 7403 **ACCEPTED** CONSUMPTION PATTERNS ND CSRr_, I**ACCEPTED** CI TASTE WHITE OR SLIGHTL_Y YELLOW CRYSTALLINE MASS OR COLORL.ESS TO, YELLOWISH L.The Merck Index. 9th ed. Rahway, New.Jersey: Merck & Co., Inc., 1976. ,, p. 5917. **ACCEPTED** HEXAGONAL PRISMS [Weast, R.C. (ed.>. Handbook of Chemistry and Physics. 60th ed. Boca Raton, Florida: CRC Press Inc., 1979. C-4T17**ACCEPTED** CRYSTALS ORLIL?UI'DCFenaroli's Handbook ofFlavor Ingredients. Volume 2. Edited, translated, and revised by T.E. Furia:and N1.. 6ellanca.. 2nd ed. Cleveland: The Chemical F.ubber Co., 1975. , p. 2251 ->F*ACCEPTED** CHARACTERISTIC SWEET ODOR, SLIGHTLY PHENOLIC CFenaroli's. Handbook of Flavor Ingredients. Volume 2. E...dited,, translated, and revised by T.E. Furia and N. Bellanca. 2nd ed. Cleveland: The Chemical Rubber Co., 1975, , p. 2251 **ACCEPTED*# ND BOILING POINT 204-206 DES C CThe. Merck Index. 9th ed. Rahway, New Jersey: Merck & Ca.,. Inc., 1976. , p. 59i7 **ACCEPTED** MELTING POINT 32 DF_OCfWerist, R.C. (ed.). Handbook of Chemistry and Physics. 60th ed. Boca Ratnn,, Florida. CRC:Press Inc., 1979. C-4317. #*ACr_.EPTED** MOLF_CUU-AR'WEIGHT 1'24..13 CThe Merck Index. 9th ed. Rahway, New Jerseyc Merck: R. Co., Inc., 1976. ,. p. 5917 *MACCEPTEm*:# /F.. CORROSIVITY ND l{ CRITICAL TEMPERATURE &. ND PRESSURE 14 c
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GRISOFULVIN CAS RN: 126078 IOSH #: WG 9800000 SYNS: '1/ GUAICOL O-HYDROXYANISOLE GUAIAC WOOD OIL 1487 2-METHOIrYPHENOL PYROGUAIAC ACID mECvH1aC1O6;. mw; 352.79 SYNS: 2-HYDROXYANISOLE 1-HYDRO%Y•2-METNOEYBEN- ZENE 2-METHOXY-4-PROPENYLPHENOL PROPENYLGUAIACOL 4-PROPENYLGUAIACOL GRISCOFULVIN ORISEDFULVINUM GRISEOFULVIN NSC 34533 (+)-GRISEUPULVIN SPIROFl/LVIN GRISEOEULVIN-mRTEURAPSC-2 TOXICITY DATA: 3 CODEN: cyt-asn 400 mg/L CIMIAZ 18,683,72 cyl-hmn:lyn140 mg/L/3D MUREAV 25,123,74 spm-mus-ipr 3 gm/kg/5D PNASA6 72.4425.75 orl-mt TDLO:2500 mg/k8/(6-I5D SCIEAS 175,1483,72 preg) TFX:TER orl~ TDLa:2000 mg/kg/(1-22D TJADAB 11,79,75 preg) TFX:TER orl-nt TDLo:462 gm/kg/2Y-t BICAAI 38,237,78 TFX:NEO orl-mus TDLO:440 gm/kg/52W-C CNREAe 26,721,66 ' TFX:NEO orl-mus TD:730gmCkg/52WC CNREAB. 26,721,66TFX:NEO ivn-rat LD50:400 mg/tcg NATUAS 182,1320,58 ipr-mus LD50:200 mg/kg NTIS•• AD277b89 ivn-mus LD50:280 mg/hg 85ERAY 3,1766,78 cyt-nt-ivn 200 mg/kg NATUAS182,1320,58 ar1-nc TDLo:I250 mg/kg (6-lSD SQEAS 175,1483,72pre8) orl-rat.TDLo:2gm/ag(11-14D preg) ANTBAL 14,44,69 orl-ret TDLo: 12500 mg/kg (6-15D SCIEAS' 175,1483,72 preg) orl-rat TDLo:500 mg/kg (9D preg) AN78AL 14,44,69 acu-mus TDLo: 120 mg/kg/49w.t CNRFAS 27,1900,67 TFX:NEO Carcinogenic Determination: Animal Positive IARC•• 10,153,76. Toxicology Review: LLOYA2 38(1),21,75;. 32XPAD -,49,75; ARMIAZ 26,279,72. THR: MUT data. An exper TER, NEO, CARC. HIGH ivn, ipr. GRUBILIN CAS RN: 62851574 NIOSH #: ME 4685000 An antibiotic produced by an unidentified strain of STREPTOMYCES sp (85ERAY 2,1053,78) TOXICITY DATA: 3 CODEN: ipr-mus LDS0:30 mg/kg 85 ERAY 2,1053,78 ivn-mus LD50:15 mg/kg 85IiRAY 2,1053,78 THR: HIGH ipr and ivn. Disaater Hazard: When heated to decomp it~t emits tox fumes: OMETHOXYPHENOL TOXICITY DATA: 2 CODEN: mmo-rat 400 uL/plate. BECTA6 24,590,80 Grl-rat LD50:725 mg/Iig TXAPA9 6,378,64. Toxicology Review: 27ZTAP 3,74,69. Reported in EPA TSCA Inventory, 1980. EPA TSCA 8(a) Preliminary Assessment Information Proposed Rule FERREAC 45,13646,80. THR: MUT data.. Protect from light. MOD via oral route. Ingestion produces burning ih mouth and throat,. GI distress, tremorsands collapse. See also phenol. Fire Hazard: Mod, when exposed to heat or flame; can react with oxidizing materials. To Fight Fire.• Foam, COP, dry chemical. Dlsaster Hazard: When heated to decomp it emits acrid smoke and fumes. UAIACOL GLYCERYL ETHER-CARBAMATE' CAS RN: 532036 mL CrrHrsNOs; mw: 241.27 SYNS: GLYCERYLGUAIACOLATE CARBA- MATE OLVCERYLGUAIACOL{ARBAMAT GUTACOL-0LICERILETERE MONO CAR9AMMATO 2-NYDROXY-3-(o-METHOXY- PHENOXY)PROPYL-I-CARBA- MATE TOXICITY DATA: orl-rat LD50:1320 mg/kg ipr-rat LD50:820 mg/kg or1-mua LD50:1530 mg/kg ipr-mus LD50:955 mg/kg acu-mus LD50: 780 mg/kg orl-0og LD50:2000 mg/kg irn-rbl LD50:680 mg/kg orl-hem LD50:1410 mg/kg ipr-ham LD50:1050 mg/kg NIOSH #:TY 8750000 3-(2-METHoxYTHENoxY)-I- GLYCERYL CARBAAtATE 3{O-MEn WXYPNENOxY-2-HY- DROXYPROPYL CARBAMATE 3{D-MerxoxYPHPxoxY)-1 i2- PROPANEDIOL-I-CARBAMATE 2 CODEN: JPETAB 129,75,60 3PETAB 129,75,60 JPBTAB 129,75,60 AB2NAD 17,242,67 APTOA6 19,247,62 27ZQAG -,398,72 UNEAQ 5,305,66 JPETAB 129,75,60 JPETAB 129,73,60 Reported in EPA TSCA Inventory, 1980. THR.• MOD orl, ipr, scu, ivn. See also ethers and carba- mates. Disaster Hazard: When heated to decomp it emits tox fumes of NOr. GUAIACOL CAS RN: 90051 NIOSH #: SL 7525000 mF. CrHLOP; mw: 124.15 Clear, pale yellow liquid or solid. Characteristic odor,, darkens on exposure to air and light; d (crystals): 1.129;. d(liq): about 1.112. Misc. with alc,, chloroform, ether,: oils, glacial acetic acid; Slightly sol in petr. ether, sol in NaOH soln. mp: 28°; bp:Q02°-209°, flash p: 180°F (OC), d: 1.097 ® 25°/25°: GUAIAC WOOD OIL CAS RN: 8016237 NIOSH #: ME 6270000 From steam distillation of BULNESIA SARMIENTI LOR. wood chips or sawdust (FCTXAV 12,807,74) TOXICITY DATA: 2 CODEN: skn-rbt 500 mg/24H MOD FCI7CAV12,807,74 Reported in EPA TSCA Inventory, 1980. THR: Strong irr to rbts..
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FORMJLATIONSIP'REPARATIONS GUAI'ACOL PLUS FROM MYZON (VETERINARY EXf='ECTORANT):EACH LP: IODINE QAS POTASSIUM I.1]DIDE AND ETI-1YLENEDIAMINE DPHYDROIODSDE, 10.9 G,GUAIACOL GLYCERYL ETHER, 22.7 G; AMMONIUI'1 CHLORIDE, 227.0 ? G; SODIUM CITRATE, , ?.0 6;, SALT;. FLA'JORINrr SWL=ETENING U.S. THIS PRODUCT DISCONTINUEDAS OF JUNE 1978. tRumack:, IB.H. ('.ed..) Poisindex. Micro4iche Edition. Deriver, Colora.do: Microm-n-edex, Inc., in associationn with the National Center for Poison InFormation, with update=, 197,°,-present. I **ACCEPTED** I f•'ORMULATIONS/PREPARATIONSGUAIACOL COMPDINJF_CTION FROM GENEVA: EACH 2 CC: NAIOUL.I OIL, 0.10 G;EUCALYPTOL, 0.08 6; Gr_iATACOL, 0.10 G; IODOFORM, 0.02 G;, CAMPHOR, c_r,h5G,. [Rt.rmacF::, Er:.H'. (ed.) Poisindex. Microfiche Edition. Denver, Colorado: Micromedex, Inc.,, in associat:ionwiththe National Center for Poison Information, with updates, 1975-present. 3 **ACCEPTED** OTHER MANUFACTURING FEMA, NO. 2532. REF`ORTED~ USES~: NONALCOHOLIC INFORMATION BEVERAGES,. 0.95 PPM;ICE CREAM, ICES, ETC, 0.52 PPM,. CANDY, 0.96 PPMp: BAKED GOODS, R.75 WPM. /FROM TABLE/ CFenarol.i's Handbook: of Flavor Ingredients. Volume 2'. Edited, translated, and revised by T.E. Furia and N. BelLanca. 2nd ed. Cleve3and: The Chemical Rubber Co., 1975. , p. C<5] **ACCEPTED** OTHER MANUFACTURING INFORMATION OF DIRECT FOOD ADDITIVES, LIMITATIONS: SYNTHETIC FLAVOR... GFuria, T.E. (ed.). CRC Handbook of Food Additives. 2nd ed.. Cieveland: The Chemical Rubber Co.,, 1972. , MAJOR USES p. 56"7 **ACCEPTED** LOCAL ANESTHETIC AGENT COsol, A. and J.E. Hoover„ et a:. (.ed's..). Remington's. AJOR USES Pharmaceutical Sciences. 15th ed. Easton, Pennsylvania: Mack Publishing. Co.,, 1975., P, Eti?] **ACCEPTED** ANTIOYIDANT; ANT.I-SUMMING AGENT IN HYDROCARBON SOLVENTS; ANTI-SKINNING AGENT IN SURFACE Cl7ATINGS;. CHEM INT CPatty,, F. (:ed.),. Industrial Hygiene and To:.icology: Volume ~ II: Toxicology. 2nd ed. New York: ~ Interscience Publishers, 1963. , p. 1683] IPA **ACCE.PTED+* ~ MAJOR USES (VET): ANTITUSSIVE; ANALGESIC; ANTISEPTIC Fb 1.i EP.ossoff, I.S.. Handbook.k of Veterinary Drugs. New York:: Springer Publishing Company, 1974. ,, p. 2497 *+,ACCEPTED** . 13
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( I METHODS OF MANUFACTURING OBTAINED FROM HARDWOOD: TAR OR SYNTHETICALLY FROM O-NITROF'HENOL.. VIA~ ORTHO--ANISIDINF_. CFenar-o].i'sz Handbook of Flavor Ingredients. Volume2. Edited, ti^anslated,, and revised by T.E. Furia and N. Dellarrca.. 2ndad. , Cleveland: The.Chemi..al! Rubber Co.., 1975. , p 2257 **ACCEF'TED#*' METHODS OF MANUFACTURI'.NG ...BY MERCURIC OXIDE OXIDN OF LIGNIN: LEWIS, PEARL, USF'ATENT 2,433,227 (1947 TO SULPHITE PROD); E+Y OXIDN OF ANISOLE WITH TIRIFLUOROPEROXYACETIC ACID: MCCLURE, WIL.LIAMS, J OftG CHEM27, 627 (1962); FROM ACETOVANILLONE & ZNCL: READ, US PATENT 3,057,927 (1962 TO ONTARIO RES FOUND)... [The Merck Ind'ex. 9th ed. Rahway, New ,7erseye Mer°ck & Co.,I'nc., 1976. , p. 5917 **ACCEPTED** METHODS OF MANUFACTURING ...FFFOM DIAZONIUM SALT! OF O-ANIS'IDSNE: HERBST, GERMAN PATENT 1,14E,2_6. (196:3 TO HOECHST). CTheMercR I!ndex.. 9th ed. Rahway, New.dersey: Merck & Co., Inc.., 1976. , p. 5917 **ACCEPTED** METHODS OF MANUFACTURING BY EXTRACTING DEECHWOOD CREOSOTE WITH. ALCOHOLICfiOTASH, WASHING .WITI-iIETHER,. CRYSTALLIZING FOTASH!.COMPD FROM ALCOHOL 'a DECOMP W.ITH.DILUTE SULFURIC ACID; ALSO FROM ORTHO•-ANISIDINE BY DIAZOTIZATION AND. SUBSEQUENT ACTION OF DILUTE SULFURIC ACID. /GUAIACOL/ IHawley, G.G. The Condensed Chemical'. Dir_tionary.. 9th ed. New York: VanNostrand'Reinhold'Co. , 1977. , p. 4247 **ACCEPTED** IMF'URI'rIiES ND FOiiMULATIONS/FREPARATIONS. GUAIACOL FROM JENKINS:.. EACH 2 CC CONTAINS: GOMENOL, 0.1 G; GUAIACOL,, 0.1 G; EUCALYPTOL, 0.08 G; I ODOFORM, 0.02 G; CAMPHOR, 0.05 G; SESAME OIL Q.S. GUAIACOL COMFD1U'/. FROM MED TEC: CREOSOTE, 0.527+; GUAIACOL, 9.70%; CRESYLIC ACID, 4.3p'/., OIL EUCALYPTUS, 0.50%; CAMPHOR, 6.y5%;EMULSIFYING BASE Q.S. LRumack, G.H. (ed.) Poisindex. Microfiche Edition. Denver, Color-ado: Micromedex, Inc., imassociation with the National Center for F'oison.Information, with updates,. 1975-present. 7 **ACrEFTED*+ 12
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Poilnorr Information, with updates, I975-presenh. MGMNT 1] **ACCF_PTED# :* ( J ANTIDOTE AND EMERGENCY SEE PHEhIOL. S). ALC IS CONTRAINDICATED IN TREA! -MENT THIS INGESTION, 6) CONVULSIONS... TREATED WITH IiV DIAZEPAM 0.1-0.3 MG/f`:G/DOSE IN CHILD •°.a UP TO lU MG IN'ADULT. 7) ACID-BASE BALANCE....INVESTIGATED & TREATED W.ITH. APPRUPRIATE.MEANS MOS7: UStUALLY'BICARHONATE. /PHENOL./ CRumack, E+.H. (ed'.) Poisindex. Microfiche Edition. Denver, CoSoradosMi.cromedex, Inc., in association with the National Center for- Poison Iroformati~on, withh updates, 1975-present. MGMNT ].l **ACCEPTED** ANTIDOTE AND EMERGENCY SEE PHENOL.. B) BASE LINE LIVER & RENAL TREATMENT MEASUREMENTS SHOULDFE OBTAINED & FOLLOWED IF ANYSYMPTOMATOLOGY IS DEMONSTRATED. 9) FORCED DIURESIS, PERITONEAL DIA.LYSIS'.& HEMODIALYSIS.ARE OF NO VALUE. /PHENOL/ CRumack, D.H. (ed.) Poisindex. Microfiche Edition. Denver, Colorado: Mic:romedex., Inc., in association with the National Center for Poison Information, with updates, . 1975-present. MGMNT 17l **ACCEPTED** MEDICAL SURVEILLANCE ND HUMANTCXIC.ITY EXCERPTS MEDICAL EXPERIENCE INDICATESTHAT TOXIC. RUANTITIES /OF GUAIACOL/ CAN.BE AErSOREEDTHROUGH SKIN QUITE READILY. MATERIAL IS NOT ESP IRRITATING TO SKIN HUT PROLONGED. CONTACT MAY CAUSE INJURY, PARTICULARLY IF SKI!N IS ABRADED. SEVERELY INJURIOUS TO EYES. CF•atty, F. (ed.). Industri.al Fiygiene and Toxicology; Volume II: Toxicology. 2nd ed. New York.: Intersciemce Publishers, 198". ,, p.. 16^043 **ACCEPTED#*: HUMAN TOxICITYEXCERPTS ...APPEARS TO BE ABOUT ONE THIRD AS TOXIC AS PHENOL & TO'HAVE PHARMACOLOGICAL PROPERTIES G!UITE SIMILAR TO PHENOL. CAUSES MUSCULARWEA};NESS, CARDIOVASCULAR COLLAPSE, PARALYSIS OF VASOMQTOR CENTERS. CPntty„ F. (ed.). Industrial Hygiene and Toiic.ology: Volume bI:To:cicology. 2hd ed.: New York:: Interscience. Publishers, 1963. ,, p. 16441 **ACCEPTED** HUMAN'..TOX.ICITY EXCERPTS A.9-YR OLD GIRL DIED AFTERSWALLOWING 5 ML & m DOSES.OF MORE THAN 2 G ARE HAZARDOUS WHEN ~ \ ~ APPLIED TO SKIN. MEDICAL USE AS EXPECTORANT IN LARGE DOSES CAN CAUSE CARDIOVASCULAR N N t0 COLLAPSE & CLINICAL DOSES...CAUSC: GI IRRITATION. [Patty, F. (ed.). Industrial Fr N ~ li8
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File 20z:ASRIS INTERNATIONAL 74-87/SEPT © 0 06E2492 2 AGRIS Nor 942A5T: Reduction of the inductive period in equine anaesthesia by using mixtures of glyceril-guaiacol-ether-sodium thiopental and chloral hydrate-magnesiumsulphate-sodium.pentobarbital combined with a injection of sodium thiopental (Disminucion del pcariodo de induccion; en la anestesia deequinns con las mezclas g:licer il--g±rayacol-eter-tiopental sodico e hidrato de r-loral-sulfato de magnesi.o pentoharbitaL socitco, combinandolas con imyecci.on unica de tiopental sodico) Mal'donado Montiel, C. Universidad Nacional Autonoma de Mexico, Mexico, DP-. Facu.ltad de MedicinaVeterinaria y l,ootecniaTesis (MVZ)~ Mexico, DF, 1982, 24. p. Notes: Illus. Numerical data; 12 ref'_ C.angu.age: Spanish Summary Language:: Spanish Availabil.ity: Bi.blioteca, Facultad Medicina Veteri~,narria y Zootecnia,, Univ. Nacional Automoma, Mexico (Mexico) Document. Type: MonographyNumerical Data, Dissertation, Summary, Noncanventional L.iter•atureJomrnal Announcement: 0911 Record input by .T.I•_^,A(Inter-American Inst o{Agr Sciences.) ti63p852 AGF;IS Nb: B.:r913.5 Toxicity and biodegradability of pulp mill waste constituents to anaerobic bacteria anaerobic degradation, ATA, BMP, furfural, guaiacol, acclimation, toxicity, methane, biogas Benjamin, M.M.;: Woods, S.L.g, Ferguson, J.F. (Washington Univ,,, Seattle (USA). Environmental Engineering~.andScienceF'rog:ram) Anaerobic Treatment of W'astewater in FixedFi.lm.Reactors, Copenhagen (Denimark.) ,. 1b--P.B Jun 1982IAWPF:speci~alic-ed seminarJune16-1@,, 1.982.Copenhagen Denmark: Anaerobic treatment of wastew.ater- in fixed fil.m reactors; Preprints of papers presenteclat the seminar Danmark:s Tekniske H'oejsk:ole, Copenhagen. (Denmark).. Lab. for Teknisk Hygiejne Copenhagen (DenmarF;):: DTH, 1982,, p.. 163-173 Notes: 10 ref. L-anguage: English Summary Language: English Place of Publications Denmark Availability: Denmark Center Document Type: Analytic, Monograph, Conference, Summary, Nonconventional Literature Journal Announcement: 0902 iCecord'i.nput by CEC (Commission ofEuropean Commt.rniti~es)~ N N M+ N 18
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44,0 AN C,A106 (:15):: 118"'49f TI The toxicological implications of the interaction of butylated hydroxytoluene with other antioxidants and phenolic chemicals AU Thompson, D. C. i Trush, M. A. CS Sch. Hyg. Public HLalth, Johns Hopkins !!nsva I:.C1 Baltimore, MD 21^05,,USA SO Food Chem. Ta>::i cnl ., 24 (.1'.0--11 ), 1.1't39-95, . SC 17-6 (Food' and Feed Chemistry) DT J Ci7 FCT0177 IS n^7:3-691.5 PY 1986 LA Eng, IT 5C>--28•-2, Estradiol, biologicall studies 56-53-1, Diethylstil.bestrol **#9n-p.5-14*~ r , Guai.acol99-76-3,Methylparaben 103-90-2, Acetaminophen, 19F3-9s-^<.,. Phenol,, biological studies 121--::3-5, 'Janillin 128:-37-C?, Butylated hydroxytoluene, biological studies 1135-24-6, Ferulic acid 25U13--16-5, DUtylated hydroxyanisole (RHT metab.. by micr-os.omea response to, toxicity in relation to) AN CA106(4):22957b TI Effect of individual phenolic compounds and of their mixtures on luminous bacteria. Part 3. Combined action of phenolic compounds on. aquatic organisms AIJ Stom, D. I.; G'i I,, T. 01. a, Bal ayan,. A. E. CS Sci. Fes.Inst. Diol., A. A. I.danov State Univ. LQ Ir-kutsl<,, SU^664003, USSR SCi Acta Hydroe:him. Hydrobi~ol:., 14(U), 539-49 SC 61-2 (Water) DT J CO AHCEtAU RS 0323"4720 PY 1986 LA Eng IT xrat9c)-i~5~ 7:~~* 1p6-44-5P',, p-.Cresol, pr-epa.ra.t'ron 106-51--4P, preparation 168-46-v, Fesorcinol, biological stu.idies 1'.08-95-2, Phenol, biological studies 120-EiU-9P„ Pyrocatechol,, preparation 123-31-9, Hydroquinone, biological studies (toxicity of, to aqpatic organisms, water pollution in relation to)
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DE_NSI''ITY/GPECIFIC: Gf.AVITY DISSOC'IATION C4NSTANTS HEAT OF COMBUSTION HEAT OF VAPORIZATION OCTANOL/WATER.PARTITTONCCEFFICIENT PH SOLUBILITIES SOLUL;ILITIES SPECTRAL PROI'ERT I ES SURFACE TENSION VAF'ORDENSITY VAPOR PRESSURE RELATIVE EVAPORATION RATE VISCOSITY OTHER CHEMICAL/PHYSICALPROI-'EFiTIF_S 1. 1:'87 @21 DEG ~~~/4DEG C L Weast , R. C. (edl. ). Handbook: of Chemistry and Physics. SCrth ed.Eoca Ral:on,, Florida: CRC Press Inc..,. 1979. C-4'1) **ACCEPTED** ND ND ND ND ND 1 G. SOL IN 60-76ML WATER, IN 1 ML GLYCEROL; MISCI6LE WITH AL.COI-1OL,, CHLOROFORM, ETHER, OILS,GL.ACIAL ACETIC ACID; SLIGHTLY SOL IN.. PETROL.EUM'ETHER; SOL INI SODIUM HYDROXIDE SOLN [The Merck Inde>:.. 9th ed. Rahway, New Jersey: Merck & Co., Inc,,, 1976. , p.591I **ACCEPTED'* -~ SOL IN ALI<.ALIES; MISCIBLE WITH ESTERS, BENZENE, TOLUENE [Patty, F. (ed.). Industrial Hygi~ene and To>oicology: Volume II: Toxicology. 2nd ed'. New York: Interscience Publishers, 1963. , p. 16041 ##ACCEPTED** INDEX OF REFRACTION: 1.5429 @ 20 DEG C/Da MAJ(: ABSORPTION (ALCOfiOL): 274 NM (LOG E_ '..41)CWeastq R.C. (ed.). Handbook of Chemistry and Physics. 60th ed. Lioca Raton, Florida: CRC Press Inc., 1979. C-4z1] *+ACCEPTED*?t ND ND 0.103 MM HG @ 25'DEG'C [Patty, F. (ed.). Industrial Hygiene and To:.icology: Volume II: To::i.colagy. 2md ed. New York: Interscience Publishers, 1963. , p. 16841 **ACCEPTED>+# ND ND VERY REFRACTIVE LIQUID; FORMS SPARINGLY SOL COMPDWITH MODERATELY CONCN KOHCThe Merck Inde::. 9th ed. Rahwa,v,. New Jersey: Merck & Co., Inc., 1976. , p. 591] **ACCEPTED**' m ND C}. HAZARDS SUMMARY 15 -2 N (Q N W iA
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CI1 NATURAL OCCIJRRING SOURC_.F_S' ARTIFICIAL SOURCES RROBABLE ROUTES OF HUMAN EXF'OSURE AVERAGE DAILY INl"A1;E.. PROBABLE EXPOSIJRES BODY BURDENS IMMEDIATELY DANGEROUS TO LIFE OR',HF_ALTH ACCEPTABLE DAILY INTAKES ALLOWABLETOLEft"ANCESOSHA STANDARDS NIOSH.RF_COMMENDATIONS THRESHOLDLIMIT VALUESOTHER.OCCUP.>TIONAL PERMISSIBLE LEVELS WATER. STANDARDS. ATMOSPHERIC STANDARDS SOIL STANDARDS CERCLA REPORTABLE OUANTITIES TSCA RERUIREMENTSRCRA REQUIREMENTS FIFRA REQUIREMENTS FDA REQUIREMENTS SPECIAL REPORTS ON-LINE DATABASES TEST STATUS. PRIOR HISTORY OF ACCIDENTS DISCOVERED IN DISTIL_LATION PRODUCT FROM. GUAIAC RESIN... FOUND IN CASTOREUM'OIL, IN ESSENT:IAL. OIL FROM FLOWERS OF PANDANUS ODORATISSIMUS L, IN.DISTILLATION WATERS OF~ ORANGE LEAVES, IN ESSENTIAL OIL FROM CELERY SEEDS, & IN OIL FROM TOBACCO LEAVES. CFenarol.i's Handbook ofl Flavor Ingredient_.. Volume 2. Edited,, translated',, and.revised by T.E. Furia and'. N. Dellanca. 2nd: ed. Cleveland: The Chemical Rubber Co. , 1975. , p. .^•_25] **ACCEF•TED** ND ND ND DIRECT CONTACT WITH EYES & SKIN WOULDBE. MOST LIk:ELY'INDUST EXPOSURE. SINCE VAPOR PRESSURE IS'LOW & ORDINARY USE DOES NOT • INVOLVE ELEVATED TEMP, VAPOR.EXPOSURE OF SIGNIFICANCE WOULD NOT SEEM LIKELY. .../UNLIKELYI THAT INHALATION OF VAPORS.....SERIOUS INDUST HAZARD UNLESS' MATERIAL HANDLED HOT OR FUMES WERE GENERATED. EPatty„ F. (ed.). Industrial Hygiene and To:;icoloqy: Volume II. Toxicology. 2nd:ed. New York: InterscienceF`ublishers, 1943. , p. 16£34] **ACCEPTED** ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND 22
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0 Ct HAZARDOUS SUBSTANCES DATAi+ANK NUMBER NAME OF SUBSTANCE CA-SREGISTRY NUMBER LAST REVISION DATE RECORD LENGTH RELATED HSD$,RECORDS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMSSYNONYMS SYNONYMS SYNONYMSSYNONYMS SYNONYMS SYNONYMS SYNONYMS SYNONYMS MOLECULARFORMUILAWISWESSER LINE NOTATION RTECS NUMBER OhIM.-TADS NUMEERSH liPP I NG t!dAME/N3JMBER. DOT/UN/NA/IMCO STCC NUMBER EPA HAZARDOUS WASTE NUMBER ASSOCIATEDCHEMICAL-S HSDB - AUG 1984 424'1 O-METHOXYPHENOL 9p--0S-4 040024 15127 7 ND 1-{-EYDROXY--2'-METHOXYBEN'ZENE.*#ACCEF'TED**'2r-HYDROXYANISOLG= **ACCEPTED** 2-METHOXY-4--PROPENYLPHENOL +t*,ACCEPTED*-Y-2--METHOXYPHE.NOL **ACCEPTED** 4--F'ROPENYLGUAIACOL **ACCEPTED** ANAST I L **ACCEPTED** 6UAIACOL **Ar.CEPTED** GUAIASTIL **ACCEPTED** GUAICOL **ACCE'PTED**METHYLCATECHOL **ACCEPTED** 0-GUAIACOL **ACCEPTED** ' O-HYDROXYAN!ISOLE **ACCEPTED** O-METHYL CATECHUL **ACCEPTED** PHENOL, 2-METHOXY-- **ACCEPTED** PHENOL, O-METHOXY-- **ACCEPTED** PROPENYLGUAIACOI-- **ACCEPTED** PYROCATEr_FIOL METHYL ESTER **ACCEPTED** PYROCr'aTECHOL MONOMETHYL ETHER**ACCEPTED** . PYROGUAIAC ACID **ACCEPTED** C7-H£3-Q2 •**ACCE!*'TED#?E QRBn.I **ACi;F_F'TED** NIOSH/SL7525000 ND ND ND ND ND METFfODSOF MANUFACTURING DISTILLATION OF WOOD TAR.OR COAL TAR; ( REACTION OF O--DTr_HLO1;:OE?ENZENE WITH ALF:ALI TO C_.r FORM CATECHOL, FOLLOWED BY METHYLATION CSRIi I. **ACCEPTED** 11 ®:,
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AN L'.A66 (.9 ): ,7557a. TI Pyrolysis of methyl ethers of pyrocatechol and pyrogallol AU SO Shaposhnikovy, Yu.. F:.g hMOSyuk:ova, L. V. khim. F"e.rerabotka Drcvesiny, Rei.Inform., No. 3, 6-9 SC Dr ^5 (Noncondenser.i Aromatic Compounds) r l: PY , 1965 e: LA IT R ieF*9t7-.05:_1'**i . 9.1-1b-7 9-.5--51-6 1irt1-66--3 29799-69-4 (pyrolysis of). 20
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AN CA107 (2):9101u. TI Chemical and biological characteristics of chlorinated organic compounds in spent kraft bLeaching liquors AU CS LO SO SC DI CO IS Shime.da, kinji Ri.ngyo Shik:a:njo Japan Ringyo Shikenjo KenF,yu' Hokoku,. 740, , 1--11r:rb. 4::~-6 (Cellcelose, Lignin, Paper, and Other Wood Products) J RSHKA6 n08^-472ta PY 1986 LA IT .7_a_y__.,an 5C7-84-r>, 2,4-Dichlorobenzoic acid 58-90-2, .'_,.T-.,4,6-Tetrachloroprhenol 87-86-5, Pentachlorophenol 87-88-7, , Chlorranilic acid 88-f-r6-t^., ^,4,6-Trichl.orophenol *+*91?-i55-1*ae* G'.aiacoL 91-10-1, S"yringol 9Ei-57-S, o-Chlorophenol 106-48--9, p-Chlorophenol 1Q8-43--Cr, m-Ch.lorophenol 108-95-2, Pfoenol, biological studies113-80--1, Oleic acid,, biological studies. 12~(D-83--2, 2',4--Di.chlorophenol 5i4--10-3, Abietic acid. SW-57-4, Syr'ingic acid 617-43--6 62,2-°B-6y Tetrachlorophthalic acid. 94i4-61'--6, Tetrachloroveratrole 1135-24--6, Ferulic acid11198-55-6, Tetrachlorocatechol. 1740-19-8 1825-21-4,, F•entachloroanisole 24:-35-5=•-2, Tetrachloro-o-quinome 253:-26-6, Tr-ichlorosyringel 54?8-4U-4 56961.-21-8, Tri~.chlaropyrogall.ol (toxi~city of,, spent kraft pulp: bleaching liquor in' relation to) AtV C:A106(21): 17r1412kTI Effect of individual phenolic compounds and of their mixtures on luminous bacteria. Part 4; causes of the weakening of p- and o-diphenal toxicity by other phenols AU Stom',, D. I...g Geel, T. A.y, Ealaya.n, A. E. CS Sci. P4es.. Inst. Biol., A. A. Idanov State Univ.. t-C Irka.rtsk:, USSR SQ Acta Hydrochim. Hydrobiol.,, 14(6).,, 65~~-9 _+(_ 4._-._ (Tnx i col ogy)' nT :7 CO AHU;EAU IS 03'2 ;--.43,2t> PY 1986 LA Eng IT **49 108-4.6-3, bi'ological *s*Gua»r-r,r 1f-r6--4~4--5, p--Cresol, L-iological'l studies Resorcinol, biological studies 108-95-2, Phenol, studies 1iU-8O-9, Pyrocatec:hol.,:, biological studiesI-iydroquinone, biological studies (Wenzoquinone toxicity to Eencck:ea harveyi in relation to?' _.;<
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AN CAStr5.(9):.7u.962m TI Effect of individuaU phenolic compounds and their mixtures on luminous bacteria. Part 1: use of bacterial luminescence extinguishing for biotesting of phenals AU StomyD, J'.; reel, T. A.; $alayan, A. E. CS 3ri~.. Res. Inst. Bi.ol„ Irkutsk State Univ. LO Ir4:utr-k 6640ri :, U6SR BO Ac_t.a Hydrochi!m. HydrobioL.,: 14(3),. 283-92 SC 4-_F (lars.icology). DT J CD AHCI+AU IS n323--43;?(:) PY 1986 LA Eng IT ***90-Otw-1**#, 106-51-4, biological studies LO8-46-3,, biologicall studies108--95--2, biologi.calstuodies 108--95-2D., derivs. 12O.--BQy--9, biological stt.rdies 1233-31-9, biological studies (toxicity of, bacterial Puminescence assay for) AN CA1p5(9').72664d TI Effect of guaiacol on cultured cells AU Chou, Ming Yung CS Dep. Pharmacol., Nippon Dent. Univ. LO Tokyo 102, Japan SO Shi gaku,. 73. ( 1),.-T•f3-51 SC 1-12 (Pharmacology) DT J CO SHIIGAZIS p372-n068 PY 1985 L.A Tanart .. IT ***90--05-1*** (toxicity er', to lung) AN CA1.04(19):163'551k f TI In vitro studies of biological, effects of cigaret smoke condensate. II. Induction of sister-chromatid exchanges in human lymphocytes by weakly acidic, semivolatile constituents AU Janssoni„ Tommy; Curvall, Margareta; Hedin, Annica, Enzell,, Curt R. CS Med. Nobel. Inst.,. f6arod'inskaInst. Ltl Stock:halm 8-1r14 01, Swed'. m SO Mutat. Res., 169(3), 129-39 ~ SC 4--8 (Toxicology) ~ CO MUREAV (D IS 0027-5107 l'b PY 1986 C12 LA Eng I.T 80-71-7 ***9O-t15-1*** 97-54-1 118-71-8 1'2U-8p-9,. biological studies ' 121-33''-5 6._T.SO-2i-S 16493-98--8D, alkyl derivs. (genotoxicity of,, in human lymphocytes, tobacco smoke in relation to) E; F it 5
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0264714 HFI:NJ1320p58135MUTAGENICITY OF COMPOUNDS IDENTIFIED IN PULP MILL. EFFLUENTS IN SACCHAROMYCES CEREVISIAE (MEETING ABSTRACT) Nest.mann E;;: Lee G6 Mutagenesis Section.,. Environmental and Oc=Apait?.ona]. ]"C4(icology Divi~sier, Department of National H,ealth and blelfar-e,. (.7ttaw.,~; Ontario K1A, CrL_2, Ca.nada 12th Annual. Meeting of the Envi~ronmental Mutager Society, March 5-S, 1901, San Diego, California. The Soci.ety„ 176 pp., 19a1. L anguragese ENGLISH, Document Type: Meeting Abstract Journal Announcement. 82C}2Of the compounds reported to be presentt in ef-fluents of pulpp and paper' :nills,, 42 were tested for ab.il':ity in Sar_cfiaromycescerevisiae to induce gene conversion in strain D7 and reverse mr.rtation in strain XV185-14C. Three of +these 42: compounds, nedabietic acid, tetrar_hloropropene and pentachloropropene werefnund to be mutag2ni.c in the standard '~al:nonell.a!mammalian-micro=sorne plate incorporation assay in the absence of met.abolic activation. The present experiments were performed with growing cultures of yeast. The cells were exposed to test agents for 24 hr in a shaking water bath at 30C. None of the 42 compounds increasedttre spontaneous frequency of Trp+ gene convertants in strain D7. However, the 7 compounds mutag.eni'.c in Salmonellaand 5 others (dichlorocatechol, d.ichloroguaiacol,l chloro-ci.s--muconic acid, 7-oxodehydroabietic acid and acc-tovanillone). were found to induce increased frequsncies oi Trp+ revers.ians,, but not reversions of the hi..yt:id.ine or homoserine markers, in str-airn XV1E35-14C. (4'. Re-Fs) 0223208 I CDB/f3nq5E317r? MUTAGENICITY PRODUCED BY AQUEOUS CHLORINATION OF OR6ANIC COMPOUNDS Rapson WH,. Nazar MA; Butsky VV Dept. Chemical Eng.ineering' and Appllied'; Chemistry, Uhi.v. Tor-onto., Toronto, Canada F.ull'Environ Contam Toxicol; <41.(4.).54n--546 1,'UU Coden: BECTA ISSN. 00r}7-4r361 Languag!es: ENGLISH,. Document Type: Journal Ar-t i c Le 7ournal: . Announcement: 8006 Pure compoundss typical o-f structures found ini the i.ignin moleculee were treated with irncreasingamounts of chloriine and chlorine dioxide, and thee ms.rtageni.c.ity o=r the result.ant chlorinated compound-s was tested using Salmonella typhimurium! tce_ster- strain TA10Cr. Qf the 43 compounds so farchlor-i;nated, 7._ were mutageni.c and toxic, 7 were.toxi;_ but not m!rt.agenic, and ~' were neither toxic nor mutage•nic, (5 Refs)
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AN CA98 (.25.) : ^.109o^u TI Measuring the lipophiliic properties using reversed-phase HPLC and HPTLC AL1 Butte, Werner; Foo4::cen, C.; Ka ussmanm., R.; Schull.er, D.. C5 Fachber. Chem., Univ.. Oldenburg LO SO SC DT O1denburg ^9u0, Fed.. Rep. Ger. k:omtaka:e (Darmstadt) 25-'1 4-1 (Toxicology) 7 . CO IS F::ONTDT (7172-e717 PY 1982 LA IT eir ->f*.*9n-Ci5-1->E** 95-48-7r properties 1+?v--02-7,properties C4 n*#9C1-n5--1**# 91-1v--1 95-95-4 97-53-0 97-54-1 iixj--4'1'-4,. 77241-44-2 77LSC7-B6"5 85514-32-5. (mutagenicity of, in Saccharomyces cerevisiae, paper and pulp mill effluent in relation to)~ 9 106-41-2 106--44-5,, properties 11f3-79--6 150-19-6 (lipophilicity of,, chromatog. detn. of,, toxicity to Daphnia magna in relation to) AN CA98(23.):191.340s TI A study on acute toxicity of Formalin-guaiacol AU A(uroki,. Ka.yokao; Murakami, Yu.j'.i; Osumi,TomoF:o; R:i-m!,.ra, Tamaki CS Dep. Dent. Pharm., t;yushu Dent. Coll!. L-O t:i tak:yushu, Japan Kyushu Shi4?a Gakkai Z'assh~i, 36(6), 932-9 1-S (Pharmacoloqy) J KSIgZA': IS 4~68-6833 PY 1982' LA .~ ;~y.An IT *>E*9()--C5-.1*** 1ZS19--77-3 372C.T••_87-5 (toxi:cityof',y formalin-guaiacol in relation to) SO ^C DT CO AN CA98(21):174537u. TI Mutagenici~ty of constituents of pulp and paper mill effluent in growing cells of Saccharomyces cerevisiae ALI CS LO Nestmann~, Earle F.; L.ee,. Ernie G'. H. Environ. Occup. Toxical.. Div.,. Dep. Ottawa, ON kaA 0L-2', Can. Natl . Health and Wel+are Q9 %I SO SC DT Mu±at. Re_., 119(3-4), 273-80 4--6 (Toxicology) J ~ Mb ~Q W CO MUREAV W IS 0027-5107 F'Y 1957. LA IT Eng 79--;.4-.i 79-54-9 83-w2'-987-65-0 87-88-7 89-98-5
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AN CA93(2f):198768t TI Study of concentration dependences of the toxic action of monophenols, bisphenols, and their mixtures on Saccharomyces cerevi~siaestrai.n 15V-p4 AU Pavle.nk:-ny V. V.; Trubacheeva, l.-, Ya. . (I (I LO SO SC DT PY LA IT AN effect on) CA9"`(1) :167r?a TI Effect of phenols on the adaptation of Daphnia magna and Daphnia l pu ex ~ AU Grrechanyi, G. V.; K)asPoirtsev a, I. N.; Naumova, I. A. ~ t I L.C USSF2 SO 6i~drobi~ol. Ikhtiol. Issl'.ed.. Vost. Sib., 2,. 198-20 1 ~ ~ SC 4-3 (Toxicology) DT J GIISDH ~ CO PY 1978 LA IT Russ ***9fr'--05--1*-x-*108-95--2, biologic.ali studies. 123-31-9, biological studies USSFi Issled. Biel. Deistviya Antropogen. Faktornv, 7agryaznyayushch. Vo-Joemy, Ir kutsl.: 69-911 Froms: Fef. 7-h., Khim. 1980, Abstr. N o. 1.SI.-97 4-3 (Toxi.cology) J 1979 Russ 95-48--7, biological studies 108--46-3, biological studies 1.08-95--2, biological studies. 120--80-9, biologi.call studies 1.L.1-T3--5 123-31-9, biological studies (toxicity of, to yeast) AN CA93(6)~:53877t TI Toxicities of formocresol and formalin-guaiacol and the changes of constituents in process of time AU Tsutsumi, Shoji; hawaguchi,, Mitsuru; Futaki, Shozui CS Dep. F'harmacol.,. Tokyo. Dent. ColL. LU Tokyo, Japan SO 5h i ka Gakuho, 79(12), 24::17-41 SC 6--5 (Pharmaceuticals): DT J CO SWGh.A3 IS 0371-0041 h?Y 1979 LA ,7ap p IT **+r9n-r>5--1+r#* 1319--77-3. (toxicity and stability of, in prepns. with for-mabin,, storage (toxicity of,, to Daphnia) 12
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# CI AN CA10:2(:i6)c137411~d TI Removal of quinones from an aqueous environment by phenols and the effect of their mixing on the luminescence of the bacteria Beneckea harveyi AU. Sil„ T. A..;, Nechaeva,. V.. I.; Gtalayan, A. E.;.. Shal:.hova, G. V.g. st,om, D.. Ii.; l.:ioryak:ovtsev, A. A. CS. Irk. Gos. Univ. L6 I.rkutsk, USm3 S0. Eai ol . Nauki (Moscow) , (1) ,. 5f3--6^SC 61.-2 (WLat.err) CObINfCUT IS Cr47r?-46CJ6 PY 19Ei5 LA Rus3 IT ***90-05-1*** 1Ci6-44--5, biological studies. 1C:8-46-3, biological studies. 108-95-2, biological studies 12<:r-54.-.9,, biological s+-ud'ies('.toxi~city of water coritg.. benzcq!kinone and) AN CA101(r').:9946? TI Anaerobic toxicity and biodegradability of pulp mill waste constituents AU Benjamin, Mark: M.p Woods, Sandra L.; Ferguson, John F. CS Dep. Civ. Eng., Univ.. Washington. L0Seattle, WA 98195, USASQ Water Res., 18(.5), 601•-7 SC 52-1 (Electrochemi.cal, Radiational,, and Thermal EnergyTer-hnology)CO WATRAG IS0043-1:554 PY 1984 LA Eng IT #**40-Q6--1~## 97-53-ii 98-ir1-L, properties 99-67-613Ei-86-3' 4437-2n•-.1 (anaerobic toxicity and biodegradability of puQ.p mill waste contg.) AN CA100( 14) : 108574n TI Removal of organic and toxic substances from debarking and kraft pulp bleaching effluents by activated sludge treatment AL' Junna, Juhani; ILa.mmi, Reino; Miettinen, Veijo CS Kokkola Water-Dist., Natl. Board of Waters 1-0 Finland SO Vesientiatki.muslaito4::sen Julk.,. 49, 17-29 SC 60-1 (Waste Treatment and Disposal)m r-.n VSMJA4 ~ IS ri355-0922 ~ F'Y 19B2 LA Eng ~ IT 57-DeT-3., uses and miscellaneous 57-11--A, uses and miscellaneous 6U-33,-T,, uses and miscellaneous (33-45-4 83--46-5 84-74-2 BS'-C76-2 ***90-05-1*** 111-02-4 112--50-1 , uses and3'978-67-4'. 547).-51 't. 5835- 26-756961 2ir-7 57r?57`£i 3-7 (removal of,.from pulping wastewater, by activated-sludge process, fish, toxicity redn.. by) 7
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Jo55ct_~~ 11-18f1SECTION II. INGREDIENTS INDEX Simple.monohyddc phenols and their esters 488 488 Amyl Phenol p-tert-Amylphenol,p-tert.Pentylphenol , Toxicity Rating:3(?). An intermediate in industrial chemicai'syntheses. This and various other alkyi derivatives of phenol (and of creaul) are alsn used as germicides, usually on, non-biological surfaces. They are less.corrosive than phenol and produce ~-less skin irritation, but they may, sliare.phenol's ability to penetrate intactlskin:.Relevant.oral tox- See also: Phenol, Reference ConRener tnSection III. Ref.: von Oettingen,.1949. icity data were not located, but among mono- and di:elkyllsubstituted phenols, water sulubilityy and systemic toxicity decrease with increasing size of the alkyl constituents. The differences are panic- ularly promihent in comparing n-amyl with n-hexyl phenol and in comparing 6-n:amyl-3-methvlphenol with 6-n-hexyl-3-methylphenol. symptoms and treatmenG as phenol. 490 490 Creosol faopropyl-o-cresol, 2-Hydroxy-p-cymene, Isothymol Toxicity Rating: 4. An isomer of thymol. Somewhat more toxic than thymol in rats and rabbits. Same See afso:Phenol, Reference Congener in Section III. Ref.: vom0ettingen, 1949. . 2-Metho;y-4-methyl phenol . ToricitgRating:4:OneoStheactiveingredienffiof index..Slightly less.toxic and less.corrosive than wqod creosote, not.to be confused with cresol; see phenol. See alsa: Phenol,.Reference Congener in. Section 111. Ref.: von Oettingen, 1949. 491 491 m-Cresyl Acetate Cresatin Toxicity Rating: 4. Presumably benign until hydro- latter in the index. The.latter is less corrosive and lyaed in the bowel to acetic acid and m-cresol; see less toxic than phenol. See also: Phenol,:Reference Congener in Section III. Ref.: von Oettingen, 1949: 492 492 Guaiacol ` Methylcatechol, o-Hydroxyanisole, o-Methoxyphenol Toxicity Rating: 4. Slightly less corrosive and less guaiacol wass taken as wood-tar creosote. percuto- toxic than phenol. In most: . clinical poisonings, neousabsorption is dangerous... See also: Phenol, Reftrence Congener in Section IR. Ref.: von Oettingen, 1949. 93 493 ,6-Naphthol Beta-naphthol, R-Hydroxynaphthalene Toxicity Rating 4. Produces crampy abdominal anemia. Lensopacities.and retinal changes have pain,nausea,vomiting,andsometimesconvulsions, been described. Alpha isomer is said to be even Intestinal or percutaneous absorption may lead to more toxic. a severe nephritia, liver injury, and acute hemulytic See afaoc Phenol, Reference Congener in Section III.. Ref.: Anonymous, 1922. 494 494 Phenol Carbolic acid;.Phenylic acid, Hydroxybensene,..Oxybenxene Toxicity Rating: 4:. See also: Phenol, Reference Congener in Section III:.. / Go OrthophenviF Toxicitv. Ilatio Howicide A. P through skin. on human ski dium salt arr, 0.5%. Irritatin - injury (necros are also irrita See afso:. Phet Ref.: I)ow Chr Maclntosh, 19 Hydroxymeth, Toxicity Ratii toxicity used'u is 509 mg. rep may be releeu ingested;4 but. I S-Meth,vl-2-isa Toxicity flathn mintic and mil its systemic ac is less soluble. between toxici See also: Pher Ref:von qetti Dimethylphen Toxicity Ratin See also: Pher Ref.: von Oetti Polyhydric p 9,10-Anthracei A bird repellen.t an important s of vat dyet TI a low acute or:awuth showed Ref.: Spentxr, e.g., Edwal C.1 Toxicity Ratir toxicity data v Toxicity Ratin ortho isomer 1
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AN GA7:=,.c^5 ). : 1.,`1u 1 12a . TI Absorption of phosgene by aqueous solutions and its relation.to toxicity AU Nash, T.; Pattle, FL E.. CS Micr-ohi.m'1. Res. Esi_a(,1. ~. LO Porton Dr_~wn/Salisbur,yfWiltshire, Engl. - SO Ann. Occup. Hyg., 227-33 SC 14. (Toxic.ology) DT •7 CU AUI-1YA3 PY 1971 LA Eng IT 59-52-9 75-12-7 tj~ -x104--94-91p9-12:-6 5Cr4-29`U -l 784--46-c (reaction of,, with phosgene, tox'icity in relation to) AN CA70(<5):113591g TI Hydroxyanisole depigmentation; in vitro studies ~ AU Riley, Patrick A. " CS Univ. Call. Med.. Sch. LO London, Engl. SO ,7. Pathol. , 97(2), 19^°-2t76 SC 15'(Pharmacodynami.cs)T DT ,] CO JPTLAS PY 1969 LA Eng IT ***9Cr-tJ5-1**"* 15r?-19-.6 150-76-5 (to:;ir_i.ty of, skin depigmentation in relation to)' (I AN CA94(17):133$L?9q..j TI Effects of tobacco smoke compounds on the noradrenaline induced oxidative metabolism in isolated brown fat cells AU Petterrsson,: Bertilg C!rrvall, Margaretag Enzell, G-ut. R. CS Wenner-Gren Inst., Univ. Stockholm LU Stockholm S-113 45, Swed. SO Toxi!calogy, 15(1), t--15 SC 4-13 (Toxicology) DT .7, CO TXCYAC IS n30n-4fl-.T.•x PY 1990 LA Eng IT 50-32-8, biological studies 51-17-2 54-11-5 56-55-3 57--1U-Z-, 9c7-p2-S, biological studies 9i;,-12-n 9(i-1~1--3 (of tobacco smoke,, noradrenaline-induced oxidative meta.b, in brown fat cells response to) 1J. 0 I .
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he same . id it has and no orm Di- i 140' C. :oods or 100 to clyused scs, the eetener: te sweet of acid. :g color which it wbacks, tener in s about troduct. -d from (mainly a 6 to subject ......-,.....-......__ . alcohol, soluble in hot alcohol. Insoluble in oils. Very faihl, sweet caramcllic-rootlikc odor. Intensely swucl taste. Im hi3hcr concentra- tions producing a slightly burning sensation in the backoP the mouth and in thcthroat,e shortly aftcr the sweet taste. In exlreme dilu- tiun there is only a sweet taste. Glycyrrhizin (immoniatcd) as such is not widely usedin Ilavors,bul lh4impure Glycyrr- ortho-M clltox,vphenol.. mvdn. Hydtoxy'anisolc. (I'.rohatechol monomcthylcthcr. / Ox) 2-metitoxybenzenc. Mcthylcata:hol. The normal conccntmtionused is about 5 to 60 ppm in the finished consumer product. Prod::. Glycyrrhizic acid is extracted from.m the rhizomesof Glycyrrhiza glabra (mainly from the Spanish variety: typica) in a 6 to l4';;, yield. The Anintonium salt is thcsubject article. ! G.R.A.S. f•.E.M.A. No.2528. 10(t-492; 124-1( 0; scc also: 104-366; 1472: GUAICOLO-CH, 6O fl CU Colorless prismatic crystals. Darkens inair and daylight. M.P. 28° C:-melts to a color• less oily liquid. B.P. 205° C. Sp.Gr. 1.13. 1.T°;, soluble in water, 50% soluble in Glycerin, soluble in Propylene glycol, mis- cible with alcohol and oils. Powerfulsmoke-likc, somewhat medicinal odor, sweeter than the prototype phenolic odor. 'Warm,.medicinal and somewhat sweettaste; but accompanied by a burning sensation, even in high dilution. This Phenolether is used in perfume com- positions in very modest amounts for certain floml types, such as Cananga and YlangYlang, minute traces in Lilac, Narcissus, Lily, etc. It is also useful in Carnation, while somewhat higher proportions can be used in Spice com- plexes. It is accompanied bythc usual draw- backs of simple Phenols - sensitivity to alkali, air, daylight, iron, etc. CiHslJe = 124:14 Guaiacol finds some application in flavor compositions, suchas.imitation Coffee, Van- illa,. Whisky. (smokey effect), Tobacco,. Rum, etc. and in many fruit and Spice complexes:- "Ihe concentration used is normally very low, about I ppm in the finished producf... The toxicity of Guaiacol is estimated to be_ somewhat lower than that of Phenol. G.R.A.S. F.E:M.A.. No.2532. Prod:: 1) from ordm-Nitrophcnolvianrlho-Anis- idine and the-Diazo-compound. . 2) from Pyrocatechol (CatcchoB by. Mono- methylation with Potassium mcthyl sulfite, or with Dimethylsulfate in aqueous alkali. 4-57; 31-146:.26-596; 68-467; 85-67; 86-56; 90-376; 100-499;.Ifi0-1022; B-VI-768;. , 1473: GUAIACOL ALLYLETHER Catechol mcthylallylethcr. NOTE: There is.considcmblc risk of confusingg this name with the names of the following Aroma-chcmicals: para-Allylguaiacol - EuSenoL 5-Allyl guaiacol = Chavibctol. O -CH, ! J-O-CH_-CH=CH_ ~r CmHtaOa == IG-1.21 Pale ycllowislt or almost colorless viscouss oil.. Insolublb in watcr,, soluble in alcuhol and', oils. r~-,._....,.,.~,... ....,r,.....t...t.~.....,., r........, s..r~. This Phenolether which was developed in searchh of perfume and'lfavor materials other than theconvcntionaDmcmbcrsor the Eugcnol family, has never achieved much popularity, even when it was freely available sonte 10 years ago. It could find somcapplieation in hc:n~ier floral fragrances, perhapsIraces amounts in Carnation and. Wa110owcr bascs, variations of Rose fragrnnces, etc. To Ihc author's kno+dcalgc, (his material is nut used in.llavor contpaisilions. Prod.: I)) from -Pyrocatcchol by treatment with . Allvlhromide in Acetone inn the presence of Potassiwn carbonatc undcr..cooling, ~ follo)vcd by nlahylatiiin with Dimelhyl-: . ... salfatc in nqucou.alkali. +c R, r:, ..,i i., ,r,i.... ,.irh Allvl- . NorE: Ther ~ this natiie ' ~~ Aroma-chC- V. ~. pnrn-Allylg' ~ ~ S-Allyl gUir ~. .. ~~O--CH, ,: ~t ~-° :
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AN CA94(21);16L-«6'yTI Biological action of volatile phenols from pulp and paper industry wastewater under acute, subacute, and chroni,c test conditions. AU KUurova„ i3. A. LO USSR S"O Okhrana Ok:ru<h. 8r E=d,)r ot.. Zagr-yazneniya Prom. Vybrosarni vTsellyuloa.-hum. From-sti, (Leningrad), (7), 104--7 Fromr Ref. Zh. , K:him. 1981, Abstr. No. 4I'-16Cr8C 4-3 (Toxi.cology): DT J PY 1979 LA Russ IT ~ *~r 9CJ-O5-1*#* (toxicity of, of paper and pulp industrial wastewater) I AN cA94(19):151207q TI Mutagenic activity in pulp mill effluents AU Dougl.as,GeorgeR.; Nestmann, Earle h.{: C-retts, John L.; Mueller, Joseph C.; Lee, Ernie:8. H..;, Stich, Hans F.; San, Richard H. C.;.. CS Bror.azes.,. Raymond J. F'.; Chmelar-+st::as,, Alben L. Environ.. Occup. Toxicol. Div., Dep. Nat1. Health Wa?lf'ar-e LO 60 Otfi.awa, ON, Can. Water Chlorination: Environ. Impact Health Eff., 3,, 865-Sr> SC DT C:O FY 4--"i (Toxicology) J WCEEDL 198u LA Eng IT 67-.66-3, biological studies 71-55-6 75-09-2, biological: studies 78--93--3, biol.ogice.l..l stud'ies. 79-34:-8 79-54-9 83--32-9 87-65-iJ:. 87-88--7 89-98-5 91-10-195-48-7, biological studies 95-95-4 97-53-0 97-54-1 100-41-4, bi!ologir-al studies 104-46-1 106--4~4-5, bio7.ogical :;tudies 107-Cr6--2, biological studies iON-39--4, biological studies inB--Cag--:r, biological studies. 120--14-9 121-' 34-'6.. 1 2-2--3<^-7' 127--a7--53P6--C>8-1 471-74-9 471-77-2 ' 498-02-2 514-IQ--3rM.-_24-9555-4i-1 11=1-62--G 1L98-55-6 1518.-.54-316v0-3'7-9 1740-19-8 1835-14-9 2435-53-2 2478-'35-8 2539--17-5 3428-? 4-8 583 5-26-7 1t)4T$-39-7- 139Bir-ir7-9' 167ea6---T-•1.._7 ].8684-55-4.. ?ii665-95-6 24560-98-3 311._5-63-4 56961-2n-.7 57055-=&-6. 57055--39-7 61966-36:7 77.^2v-5(r-4 77241--44i--i 7728rJ-86--5 (mutagenicity o•f, pulp milll effluent in relation to) II
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AN CA1(Jp ( 1'3) : 1i5C1f3fJp TI Topical irritation by formalin-guaiacol AU Osumi , Tomoko.g K:.irrok:i , k;ayoko;, k:imur-a, TamaR:i ; CS k:yushu DFnt.. Co1L... LO k':i.t.ak:yushu„ Japan SO }'yushl-f Shi 6:a Gakka:i Zasshii,. 37(6), 1p.zT-'i' SC 4-3 (.To:;icology) L)T i~ CO k:;SGZrl? IS 6:36E3-6®M' PY 1903 LA Japan IT +*91(-05--11319--77-:1. (s-k:in irritation from formaldehyde and) Murak~~ami, Yu.ii~~ APd. CA91(21) :169139g. 11 TI Tri- and tetrachloroguaiacol: results of a three and six-month feeding study in rats AU! Chu, I.; Villeneuve, D. C.; Yagminas, A. P.; Valli, V. E. C:il Environ. Occup. Toxicol. Div.,, Environ. Ffealth Di..r. l-O Ottawa, ON,. Can. SO Arch. Environ. Contam.Tox:icol.,8(S), 589-97 SC4-3 (Toxicology) DT J CO AECTCV I5. 0090-434 i. ; PY 1979 C ¢ LA Eng IT (chlorination of) AhS. CA97(25)2195103n. TI Regeneration of rat liver in the presence of essential oils and their components AU Gershhein,. L... L. CS. Northwest Inst. Med. 1'.e_. LO Chicago, Ill., USA SO Food Cosmet. Toxicol.:, 15(-~), 173-81 8C4--6 (Toxicology) DT ,? CO FCTXAV PY 1977 LA Eng IT' *#*90-05:-I*** 94-5&'-6 94-59--7 94-62-2 97-53-0 97:-54?--i 100-66-3, biological studies 1'.C-i3-65-1104-45-Cr 1nn,--q..:6-1 1 0S--13-5. 120-57-0 120-58-1 122-43-'2 1 4p--67--0 3 00 ` 5 '7- 2 489--?4-.9 495-76-1 637-50-' 672-13-9 67 3- 2 ?..._:174!5-01-9 1746-..13--U54n6-1:8-[3 _.i2T9-54-9 ~ (liver regenerationn res:Fponse to) ~ N N ~ ~ i8
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( 8 Chem. 33(6): 1C129-1032. MILLERfl.. W., CLARDY, J., KOZLOWSKI, J., MIt;OLA.7CZAK, K. L.., PLATTNER, R: D., POWELL, R. G.,SMITH,C. R., WEISLEDER, D. and GI-TAI, ZHENG. 1985. Phytoecdysteroids 0+ Diploclisia ylur_escens seed. Planta Medica 51'.. ( 1 )a 40--42. PGWELL,, R. G., WEISLEDER, D., and SMITH, C. h. 1985. Daphnane diterpenes from Diarthron vesiculosum:: Vesiculosin„ isovesiculosin. J. Nat. Prod. 48(1):110:°1i17. 21
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TOXIC HAZARD RATING 4. 4= VERY TOXIC:PROBABLE ORAL. LETHAL DOSE Sir--SO0 MG/R:G OR IJETWEEN. 1', . TEASPUON& 1 OZ FOR 7C+,K:G PERSON (150 LB). SLIGHTLY LESSCORh:OSIVE & LESS TOX.IC.THAN PHENOL. IN MOST CLINICAL POISONINGS,GUAIACOL WASTAK:EN AS, WOn_D--fARC'REOSOTE.. CGosseSin, R.E., H.C. Hodge,. R.P. Smith., an&M.N. Gleason. Clinical Toxicology of Commercial Products. 4th ed. Baltimore: Williams and Wilkins, 1976. II-126] *>fACCEPTED*N. CI POI.SON'ING POTENTIAL MEDICAL EXPERIENCE INDICATES THAT /GUAIACOL/ MAY BE MORE HAZARDOUS 'T~O HUMANSTHAP3 TO LOWER ANIMALS.. CPatty, F. (ed.). Industrial Hygienc, andi Tox.icology: Volume II: Tes:icology. 2nd ed. New York: Inter-science. Publishers, 1963. , p. 16851 **ACCEPTF_D*# ANTIDOTE AND EMERGENCY SEE PHENOL. 1) ESTABLISHMENT OF RESPIRATIONS TREATMENT & CREATION OF ARTIFICIAL AIRWAY IF NECESSARY. CHECK TIDAL VOL.(NORMAL 166-15. CC/N:G). 2) DSLUTION'.WITH WATER OR MILK SS IMPERATIVE AS SOON AS POSSIBLE FOLLOWING INGESTION TO'PREVENT CORROSION OF UPPER GITRACT. /PHENOL/ GRumac4:, B:.H. (ed.) Poisindex. Microfiche Edition. Denver, Colorado: Micromedex, Inc., in association with the National Center for Poison Informati.on., with updates, 1I975-present. MGMNT17#*ACCEPTED** ANTIDOTE AND EMERGENCY SEE PHENOL.. 2) EYE EXPOSURES....IRRIGATED 15 TREATMENT MIN'...TEPID WATER'ORSALINE &... OPHTHALMOLOGIST /CONSULTED/... 3) ACTIVATED CHARCOAL IS VERY EFFECTIVE & SHOULD BE ADMIN. AS SOON AS POSSIBLE...DOSE .:;-iGTIMES EST WT OF PHENOL INGESTED OR MIN 30 G DISSOLVED IN WATEF°. /PHENOL/ CRumack, B'.H. (ed'.) Poisindex. Microfiche F_dition. Denver, Colorado:. Micromedex, Inr_., in association with the National. Center for Poison Inrfor-mat.ion,,, with updates, 1975-present. MGMNT 17 #*ACCEPTED>Ex ANTIDOTE. AND~EMERGENCY SEE PHENOL.. 4) CASTOR OIL MAY BE TREATMENT ADMIN... (30--60 ML)...ITABSORBS. PHENOL v:IS CAPABLE OF /RETARDING/...ITS ABSORPTION. ...AVOIDASPIRATION. HYDROCARBON CATHARTIiS...NOT GENERALLY ADVISED BUT IN ~ THIS RARF_.. INSTANCE SEEM...OFVAL7JE. DO NOT io.1 IJSE IN OVERDOSE WITH CAMPHOR AS. OILS WILL N INCR ABSORPTION OF CAMPHOR. /PHENOL/ ~ IFiumack, B.H. (ed.) Poisindex. Microfiche ~ Edition. Denver, Coloradoa Micromedex, Ihc. in assoc.iatiorn,with.the National Center for 17
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AN i:A1ri4(13) : 1f):,.8ir41. TI Effect of aromatic compounds, humic acids and lignins on growth of the earthworm Eisenia foetida AU Hartenstnim, Roy Caitl.. Environ. Sci:. For., State Ureiv. New York. Syracuer.=, NY 13210, USA Soil £io1. Hiochem., 14(F);: 595--9 -T-T (To::icology) . J :iEs 1 UAH b6T8-c971.7 1982 Eng IT 5U`7S-2 65--85-Cr, biological studies 69-72-7, biological studies 71--4T-2,, biologi,cal studies 87-66-1 9r:--C11-7 90-02-8, biologicalstud'ies +o-*#'9U-05-1*** 91-16-7 95-65-8 99'-iiO-S~ 99=50-3 99-96-7,., biological studies 1UCa-CZb-1 1irir-n9-4, 100-5'2-7, biological studies 19V--66-3., W:.ologi¢al studies 1US-46-3y biological studies 108-,58--9 148-73:-6 1Ci8--88-3, biological studies 108--95-+',biological studies 12Qr--St}-9, biologicall stu.dies CS LO SO SC DT CO I.S PY LA 121-Z3-51 121-34-6 123-31-9, biological studies 149-91-7, biological studies 150-19-6 15U-76--5 150-78-7 161-1ii-0 49t_t•-79_.9495-78°-.3 498-n2.-2. .:,n4-15-4 4Cr61•-53--8 8p62-15-5D, derivs. S068•-trJ-1 t di 1135-24-6 248p-86-6 9005-53-2, biological s u es (toxicity of, to earthworm, growth in relation.to.) AN CA1rl4 ( 1-'_+) :10375._T•t TI Mechanism for decreasing hydroquinone toxicity in the presence of AU other phenols Stom„ D.. I.; Gil, T. A.; $alayan, A.. E. CS L4 NII Biol.,, Irkutsk. 6os. Univ. Irkutsk, USSR Sa', SC DT CO IS Gidrobiol. Zh.,. 21.(6)., 88--9n 4-3 (Toxd col ogy) L GBZUAM 0375-8990 PY 1985 LA Russ I'T 05-11~,O6-44--5, biologica.ll studies 108-46-3, biologica.l'l studies 1CrL'a-95-2, biological studies12U-8r?-9, biol'ogicall studies (hydi--oquinone toxicity lowering by,, mechanism uf) AN CA193(C));18258@ BIBLIOGRAPHIC DATA NOT AVAILABLE AT THIS TIME IT 5n-32."8.,, biological studies 51--17-2 54-11-5 56-'5J-3 57-55-6, 00 89-'81i-6 90-00-6 9D-r92-8 bi l i ll studies ~ , o og: ca 1+ 90 --12--r:r 9Q--15-3 91-10-1. 91-20--3,, biol.ogical' studies91--22-5, N (to>;icity of, of tobacco smoke, cytotoxicitypredictionfrom.in tD vitro tests in relation to) Ir W N 6,
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Hygiene and Tmxicology: Volume II: 'fio:;icalogy. 2nd ed. New York: Intersciar.ce Puhlisherrs, 1963. , p. 16E.°11 **ACCEPTED#* CE~ HUMAN.TOXICITY EXCERPTS WHEN GIVEN TO HLIMANS...CAUSES IRRITATION, , BURNING PAIN W.ITHVOMITINr., & DIARRHEA THAT MAYE+E BLOODY. A TOLERANCE DEVELOPS WHEN REPEATEDSMAL.L DOSESAR.E Ci_VEN. ... /ABSORBED/ DOSES LARGER THAN 2r/MAN MAY FiESULT IN CHILLS, SUDDEN FALL IN TEMP, WEAKNESS, EVEN COLLAPSE AND DEATH DUE TO RESPIRATORY FAILURE. LF'atty, F. (ed.)'. Industrial Hygiene and Torr,icology: Volume II: Toxicology. 2nded.. New York-: Interscience. Publishers, 1963. , p, 16857 **ACCEPTED** NONrHUMAN TOXICITY SEVERAL REPEATED 24-HR EXF'OSWRES OF INTACTExCERPTS RABBIT SKIN TO UNDILUTED GUAIACOL WERE REQUIREDTO CAUSE SERIOUS IRRITATION. INJURY TO ABRADED Sk:IN OCCURRED MUCH MORE READILY. ....CAUSES BURNING AND PARTIAL LOSS.OF SENSATION SI'MILAR.TO THAT CAUSED BY PHENOL. [Patty, F. (ed.).. Industrial Hygiene and Toxicology: Volume I1: Toxicology. 2nd ed. New. York: Interscience Pi.L1:_shers, 1963. „ p. 168°.,] **ACCEPTED** IN RATS. i.National Research Council. Drinking. Water & Health Volume 1. Washington, DC: National Academy Press, 1977, i. , p. 7413 **ACCEETED~* ' NON-HUMAN TOXICITY METHOXYPHENOL. HAS BEEN'FOUND TO CONTRIBUTE EXCERPTS. TO THE CARCIiNOGEN.IC EFFECT OF TOBACCO SMOKE NON-HUMAN TOXICITY LETHAL ORAL DOSE OF O-METHOXYPHr-NOL IN MICE EXCERPTS. G.4.GIF;G. SOLN CAUSED PARALYSIS OF HEART MUSCLE & 0.6'l.SOLN, PARALYSIS OF IiNTESTINAL SMOOTH MUSCLE. IN TOXICOLOGIC. STUDIES....EXERTED HEMOLYTIC ACTION ON'.CATTLE F+LOOD...INTERffERED WITH RNA SYNTH, PROTEIN SYNTH,. 8<:...MITOCHONDRI'AL RESP IN RATS. [National Research Council. Dri.nking~.Water &, Health Volume 1. Washington, DC: National Academy Fress, 1977. ,:. p. 7403 **ACCEF'TED+.# NON--HUMAhI TOXICITY LD50 SHOWN TO BE T.74 MG/i.r IN'RABBITS. EXCERPTS MINIMAL LETHAL INJECTED DOSE. IN RATS.SHOWN 'fO BE 50 MG/k:G. ...WHEN O-METHOXYPHENOL INJECTED~INTO PREGNANT RATS... FATAL TO~ FETUS; WHEN SIMILAR DOSESINJECTEDIINTOMAL_E ANIMALS, SERIOUS DISORDERS OF TESTES & DESTRUCTION OF GERMINAL EPITHELIUM WERE OBSERVED. CNational Research Council. Drinking Water & Health Volume 1. 19 i
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AN CAi.~.05.(.25):238435d TI Some molecular aspects of the chemoprevention of cancer AU Voiculetz, N.p N&r_i.,lescu--Duvaz, I.g Muresan,. Zoiag; Safirman, Ci11yq Stoica, G. q Trestioreani..i, Al.exandri-r CS Chem.. C!arcinogenes:is vep,,, Inst. Oneilogy LO Bucharest, Rom. SO Rev. Chir., Oncoli., Radiol.,. ORL, Oitalmol.,, Stomatc.S.s, oncol.,, ;'-:;(2) , +31--943 SC 1-6 (Pharmacology) DT CO •2 ONCODU !'Y LA 1986 Eng IT 60-24-2 70-18-8, biological stu dies ***90-05-1*** 91-53--2 94--36-0, biological studies 95- 55-6 99-Cr7-(] 121'.-Ox?-6. 123-3r=r--8 1.23-31-9, biological studies15 6-75-4. 150-76-53-02-79-4 0 619'-6ii--:?:7t]1-56-4 1953-02-2 2099-84-5 3i1p9-_::4._5 4.053-5t?-,z-, 415?-..89-3 6051-87-2 13406-56-9 250 13-16--5 28OBg-64-4 70786-92:-4 1 G5553-v 4-^< (chemi. carcinogenesis prevention, by) AN CA105(10.):60956r TI Biologically active substances in pulping waste liquors. III. Determination of simple monomers in pine and birch kraft pulp waste liquors and their toxicity to fish AU Sameshima, K:aruhik:o; Shigematsu, Akihib:o; Takamura, Nori'a CS Fac. Agric.., k:ochi. Univ. LO Kochi 763, Japan SO Mokuzai Gakk:aishi, 32(5), 344•-50SC 43-6(Cellulose,, Lignin,.f'aper, and Other Wood k-r-oduc.ts-)DT J CO MK:ZGA7 IS PY LA IT 51.4--10-32478--38-8 (in black sulfite l.iquor,, tor:icity of,, to Oryz.ias latipes) 0021--4795 1986 Eng 57--1u-T, uses and miscel.l'aneocos 57-11-4, uses and miscellaneous 64--19-7,, uses and miscellaneous 91-10--1 112-86-1, uses and miscellaneous121-33-5'114':-96-3 498-02--2
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FUN C7A7'., (^T•~ c .I..if17S6e. TI Production of guaiacol glycerine ether labeled with carbon-14 and deuterium and'its metabolism in animal bodies AU L-uther, D.; Piossbvrg, F.9, Koch, Hartwi.g, CS In st. Angew,. Radioakl:iv. , K.ar-1-Marx-Univ. LO Leip::i:y, Ger. SO Isotopenprcar,i:,, 6(C3)., 25q--3' SC 15 (Pharmacodynamics) DT ,3 CO IPFXA9PY 1970 LA 6er IT TTVSO-05-i*#~ 120-80-9, biological studies (guai'acol glycc_rol ether metabolite) (d C_/ AN C071 (23) : 1.1.1084r TI Effect of essential oils on drug metabolism AU Jori, A.;, Bianchetti, A.g. Prestini, P. E. CS Ist.. Ric. Farmacol. "Mario Negri" LO Mila.n,. I.tally ~ 60 Ei ochem. Pharmacol.., IS (9) , 2081-5 SC 15(Pharmacodynamics Y ) DT J CO DCPCA6 PY 1969 LA Eng 1 IT B0-°;6--8&., biological studies ***9n-f)5-1*** 1r17-q+._"470-02-6 (phar-macc•ruticals rnetabolism response to, in liver-) AN CA10.:i(2.:):19i:92c3q TI Additive-containing paints AU Raffaellini, F'e)api.no; Benatti, Gas.toneg, Vernizzi, Gior-gio; Trevisan, Lucio CS OECE-Industrin. Chimi.r_he S.p.A. LO ItalyF'I Eur. Pat. Appl. EF' 190449AS; 13 Aug 1986, 7pp, D;=signated States: AT,, BE, CH, DE, FR., r_1-3, IT,, LI, LU, NL, SE A1 Appl. 85/1.16U46, 16. Dec 1985; IT Appl. 85/19(77a]8, 3 7'an 1995 .CL Cn9D7P12A ;C 4.^-5 (Coatings, I'nl<; s, and Related Products) DT P CD EPXXDW PY 1906 LA Eng IT 78-69--3 7i3-70._;u1+t+r+t 91-64-'_5 : 94-59-7 q7"5a-fi 97..-54--1. 90-55-5 99-48'-9 104--46-1 lf%6-.-22-9 106--24-1i 1Vr..._25-212Qr-11-612r7-57-0 . 12t-511-0 49i.~:-71.-5. 5n7-70-0 L64L9-39-C! 4F,02--84--0 5D53-66-5 681.2-7&-8 8000-41-7 16150--4~5-i: 3n418--38--3 3458n--1.s-7 53tz82...12--5 6Q667-34-3 68844-77-9 (paintss..cantg., toredLiceirritatingo effects of vol.at.iUes) 17
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I AN CAf_7(.1.-=:) r 97077n TI Mutagenicity of benzidine and related compounds employed in the detection of hemoglobin AU Ferr-etti, Joseph. J.; L-i,, Wuan; Li..u,. Mong--Bimg CC Dep. Micr-ob.iol. Immurnal,, Umiv. Oklahoma Vde~lth, Sci. Cent. L.0 Okl ahoma City,. L74:la.,,, USIA GQfim, Jw Clin. f'a:thol., 67(6), 526-7 SC 4--7 Cfoaicology) DT .7, CO AJCPAI PY 1977 L.A Eng IT 9<}-..04-0 **-x•9n-05-1*+~~ 9'2-87_..`J. 95-5_;-4, biological studies 1.I9-9U--4 119-97-7 12:?-'9-4i, biological stud'.ies (mutagenici.ty of, carcinogenicit:v in relation t.o)~ AN, CA84.(7) : T9555u z"s J TI and tobacco smoke constituents an cell Effects of tobac AU multiplication in vitro Filotti, Ake; Ancker, Klasg; Arrhenius, Eri~kg Enzell, Curt CS LO SOi SC Wallenberg Lab.., Stockholm Univ. Stoclzholm, Swed. Toxicology, 5(1), 49-62 4-13 (Toxicology) DT mO J TXCYAC PY 1975 LA ll- Eng 5rJ-0r:r--U~,, biological st;.idies 5p-3:°_-B 51-17-2 54:-11--5i.h-49-5 B9--8'-L 90-00-6 9p-02-8, biological 90-12-0 90-15-3 91-1;U-1 91-20-3, studies ***90-05-1*** biologic_a.ll studies (t..oo;ieity of, in aseiteassarcoma cuStures) AN CA82:('.19):119707u TI Aqueous chlorination and ozonation studies. 1. Structure-toxicity correlations of phenolic compounds to Daphnia magna AL1 CS k:opperman,, Herbert L.;, Carison, Robert M.:;C:ple, Ronald Dep. Chem., Univ. M.innesota. LO SQ SC DT CO RY Duluth, Minn., USA Chem..-Bi.ol. Interart., 9(4), 245-51 4-3 (Tox.i.cology) S CLi'INAB 1974 n ,4 LA IT Eng 51-2F3-5.;, biological studies *+*9r7-U5-1*#ar 92-69-ti 95-48•-7 N N 95--57--B 1p0--n2--7 1:n6-4-1-2146-44-5 1rCr6--4B-9 168-95-2,, biological studies 116-79-6. 1:2c'r-BZ-2 15r?-19-6 `-76--26-•1 ~ (toxicity of, to Daphnia magna) W ~ 14 __ _ . .. .. .. ...... _. .._ __.~(
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AN CA1nn; (.1 ): 1.835z TI Ef-Fects of phorbol myristate acetate, phorbol dibutyrate, ethanol, dimethyisul-foxide, phenol, and seven metabolities of phenol on metabolic cooperation between Chinese hamster V--79 lung fibroblasts AU Ma].r-'nlm, A.. Friussell; Mills,Lesiey J.; McK_'enna,Edward J, CS Envi.ron.Res.Lab.., d.!.S. Environ. Prot.. Agancy LO Narr-agara<sett y. RI',L1SA, 7. SO Cc.l l lii ul . Tox.i co1 .. , 1(4), 7fn9-B'. gi~ q._-7 (Toxic:ology)' DT V 7 cC C/,7 CO CITOE^ IS 0742-2091 PY 1905 LA Eng IT 64-17-5:, hioliogical. studies 67--66-5,, bi.oloc3ical studies ###9[Y-6F,-1*>;~ T,[76--`;1-4„ bi,ological steidi.ie%:j 108--95-2„ biologicall studies 16B-95:-.2M,metabolites 124-oU--9, bi.ologicall studies M--.?1--'3, biological studies :i33_7:?._.3 937--'A.- B 16561.-29-)3. 17665--05--1 61557--O8--8 (metabolic cooperation by lung fibroblast response to) AN CA94(7):436u6k. TI Biological transformation of guaiacol by some Micromycetes AU Lacharme, J.:Seigle--Murand.il, F.; Steiman, R. CS Lab. Fi.ol. Veg. Cryptogamie, Unites.. Enseign. Rech. Sc:i. Pi~aarm. Fiol. ~~ L-O Meylan 36:40, Fr. SO C. R. SEaancesSoc. 9io1. Ses Fil., SC 10--2 (Mi[~_robial Biochemistry) DT J CO CRSE+AW 19 0037`-9026 PY 198[7 LA Fr 174(:5), 76•3-E) ~ I'T ###90-n=-..i(metab. oT, by micr-omycetes.) AN CA76.(19):1ti786Sg. ~tpSV7o..GA TI Compounds antagonistic to norepinephrine retention by rat brain AU hamogenates Baldessarini, RossJ. 7 CS Dep. Psychiatry, Massachusetts Gen. Hosp. - ~ ~ LO SD Bo::,ton,, Mass.. , USA C+iochem„ Phar-macol. , 2c)(3), , 1769-GiJ N SC DT 1 (F•harma.codynamics) 3 ~ CO SCPCA6 ~ PY 1971 LA Eng ~ IT 59--92-7 60-16--4, biological studi es 63-91-2, biological studies.. ##~9ci~[75--1#.## 91-16-7 11.1'-86- 4 12U-S[D--9, biological studies. CI 141--43--5, biological studies 156-87-6 TO0-4B-1306-0B--1 2394--2 0 -9 2506-29-4 5796-17-5 I:x325'-10.-5- (norepin_phrine metabolism.by brain response.to) 16- i
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. i(:i3;,.F37' T(.•DF1102039990 MUTAGENICITY TESTINGIN THE SALMONELLA TYF'HIMURIUM ASSAY OF PHENOLIC COMPOUNDS AND PHENOLIC FRACTIONS OBTAINED FROM SMOKEHOUSE SMOKE CONDENSATES Pool. BL.., Li n S'ZInst. Tnxicology and1 Chemotherapy, (`ler-mamt;ancer Res. Center,. Im N4u.=.nheimer Feld •2L'0,. t;'7,Qc'%Heid.eltherg, W. :3ermany FoodChenr. Tnx.i.col:19E'J2 l..anguages. ENGLISH,, L?oc.;ment. T,vpea Journal Article Journal Announcement: 0211 The mutagenici.ty of 10 phenolic compounds (phenol, 3 cresol isomers, :_,4--dimethyiphenoS, pyroca+_echol guaiacol, syrimgol, vanillin, ei-rgenol) from smo!•:ehou.se smokee condensate were tested with the Ames test. None of th~L, compounds tested was mutageni.c. The onlydose-related efflect was toxi.ci.iiy at the highest tlestedcancent.rar_ions of 5,000 ug/plate. Smok:e c::?ndF-.nsat.es and their fractions also gave largely negatilve results with the Amerss test. (2,.3. Refs) k 13719729 ICi}Bl82014482 6/ PYROLYTIC STUDIES ON THE CONTRIBUTION OF TOBACCO LEAF CONSTITUENTS TO THE FORMATION OF SMOKE CATECHOLS Sc-hlotzhauer WS; Martin RM: Snook ME; Williamson RE Tobacco Safety Res. Unit, Agricultural Res. Service, United'States Dept. AgricLnlture,. Athens, GA, 3OA,1T d! Agr-ic Food' Chem;: TO(2)cw72-=74 14B2 Coden: JAFCt~.r ISSN 6ir^1-8661 Languages: ENGLIGH', Documenrr- Type. Journal Article Journal. Amnouneemeni_. g'7it6 Nine varie-ti~es of Bright, and five varieties of Burley type tobacco lea,c, all flue-cured and converted to cigarette shred, werepyrolyzed and'.assayedfor production of catechol (CC), total polyphenol (PR), and chlcrogeni~c acid (CGA) isamers. The tobaccoIea.f components caffeic acid, cellulose, CGA, fructose,, lignin, qr-rercetin, rutin,, and sucrose were also pyrolyzed and fiMeproducts analyzed. Pyrolysis conditions approximated those inn c.;,_garett.esmok.in+g. Pyrolyzate of the L'ri.qhd: varieties averaged 617.more CCC than did the Burley, and 11B-.^71'% more total PP and C3A.:. Phenclicc compo!.xndss in pyrnlv:-ates of' thep{,;recompor.rnds wc,ree from CGA, CC, 4-ethylcatechol, and pheno?y from quercetin, CCC an6 4-methylcatechol; from ru.tin,. CC, 4--ethyl-, 41-methyl-, and 4-propyi.catecho--lg fr-om. !:af+eic acid, CC;; from lignin, Ci_., guaiacol, 4°-methylcatechol, phenol, and isoeugemol. From the car-bohydra'=es mainly fur-furals weree produced. Detection of 4-vinylicatechol from ca.:ffeic acid was possible only when the pyr-olysis.was run under- destractive distillation conditions. Separate pyrelysi=, experimentson four other tobacco.leaf varieties demonstrated that amount of catechol produc,=d increased (n) the higher up the plant sfalk: the leaf wNsfound, and (b) the less nitrogen fer-t_il:izerwas: used from recommended. (2`?' Refs)
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AN CA96(32)_'i:436?69n TI Methods for the determination of harmful substances in guaiacol AU production Kr-{renk:o, L. T.g Mel'na 4.ova, L. 'J. L& 5C) USSR, San.-4:h.im. Metody GipredeleniyaVred'. Ve,ahchsatv, M. 66--70 From.Fof.Zh.,, Khi'rn. 1982,Abstr.No, .4G2`?- . Sr F'Y 59-1 (Air Pollu.tiorm. and Industrial Hygiene) 19431 LA RG!s IT ~#x90-U°r-1P.~## (to>:ic substance f rom manuf. of, detn. of, i;n air)i AN. r_.A96. ( 1 ) :..1.:,79q, TI allJI CS' L& SD SC DT CO 5ome effects of polyphenols on aquatic plants: I. Toxicity of phenals in aquatic plants Stom; D. I.; Foth,, Richard Ftes. Inst. Biol., State Univ. . - Irkutsk.,. USSF:' Burl l... Environ. Contam. Toxicol., 27(3), 3-2-7 4-3 (Ta:,lcology) J EaEC-fA'_. . ? Ifa F'Y LA IT (ii)Q7--4861 1981 Eng x*#9C~-~~S•-i~#+ 1Cr6-51-4, bi-ologi.:cal stud'ies103--4&-T, biological st-udies 1..p8--95-:.', biological studies 120-90-9, biological studies.. AN.. biological studies 1321--.20.-4 69845-49-4 (toxicity of, to aquatic plants). CA95(12):99549g.. TI Organic compounds in kraft bleaching spent liquors. IV. Toxicity AU bioassay of organic compounds from beech kraft bleach effluents Shimada, Kinji CS Div.. For°. Prod. Chem.., For. For. F'rod. Res. Inst. L4 I'baraki W5,,, Japan S0. Moku::ai Bakkaishi, 27(6),. 470-7 SC 4i-6 (Ce1lwl.ose, Lignim,. Paper, and. Other Wood Froduc:ts) ~ DT J 4a CU MK_ZGA7 N IS 0021-4795 - ~ ~ F'Y LA 1981 W Eng C!! IT 50-84-0 67-86-5 87-88-7S8'-06-2 1#** 91-10-1 99--96°-7, properties 1bB-95 -2, properties 112-8U-1, properties CI 1..z1-3:4--6514-10-7. 530-57-4 617-42-5. 617-4%-6632-58-6 944--61-6 1135-24-6 1198-55-6 1740-19-5. 1.825-21-4 2435-53-2 25.T_•9.._1=+--1 2539-17-52539--2h-6 2668-s4~-S ,T.336-41-25438-40-4 :20624-9'6--8 569E• 1 -i 1--8.. 629.'6--23-6 (toxicity of, to activated sl.r-rdge) 10
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Fragrance rnww materials monographs 699 n 25 ;tnd not nlny was :ul- tab- ine. Ihe ant Iro- an tra- 69) ag the nI rlp er, !s. or rg ra d d mardrrdonlics. L('u:umg c14•cls uf exposed hum:rmvital pu!p with ethyl cyanoacrylate. f1rJ/. Tnkru J,•n!. ('1dL 17, 83. Alrrrk lnde.c 119761- An Enrrrluln•dimuf Cbrnrirnlx nndDruys. 9th Ed.. No: 4399..Meruk & Co., Rahway. N1. ktnrcno;.(1_ M.(J9781 Report to RIF'M, II'h1ay. Morioka:.5- 11976). Cliiricn-parhoingic.d smdy of the eRecls of isobutytcyanoacrylate on the exposed human rital pulp tissue. Slnkn Gukuha 76, 197. __:~A4tuakami, H..&Yumafuji, K. (19691 Antitumor activities of polyphcnuts. Sri. Brdl Fuc Alvic Kpuanr Uhin, 24: 191Fcb 28). ONravskiii.M. M. (19(.fl. K IoRsikolugii pnruv gvnyaknlu i ego smuly. Crig. S.urit: 29, 8S. Portolcs,.A., Perez Utnna, T.. Rumos, F.. Lopez, R, & Espinosa, M..11975). Penicillin immunogenicily in the pnsence.of diRerent pharmaceutical adjuvants. Lu. J. Clim Phnrrmrr. Binplwrm. 11, 7. Riley. I'. A. (19691. Hydroxymrixole da+Bigmcnlalion:.in-r•irvr smdics, J. 1'urln 97, 185. Sientonei4 K. D- Zipf, H. F. &Dittmann. E- C. (1966). Unlersuchungen zur endoanaslhetischen and hypno•- tisch;nafkotixhcn Wirkung von 2-Methoxy-4-allyl-phennxyes+igsaure-NlN-diii!hy)amidlG 29505).und vcr- w'undlrn Phenol Ucriva(cn. Aruln inn.P6nrmnurulJ•nThrr. 164, 30. Smythe,.G. A..& Lrvarus,.L. (1973). The use of a hypothetical.receptor-site modelito predict novel pituitary hormone releasing and inhibiting agents..In 1lyprrtlmL llypnplrysiurrrqric Hornr., Proc. CuuJ, 1972, Edited by C..Gual'& E. Rosemberg-.p. 189. Excurplu Medicn, Amslerdam.: Strand, L. P. & Schcline, R. R. H975k The metabolism of vanillin and isovanillin in the rat. XermAiMicu5, 49. Tarasnva, L. S. (1968k.Polifenoly kakpotenksial'nye antisclerotichoskio srttdstva: In Frnofu!rv Sra•din. Ikh Biuf. FwtkG.. Afulrr. 1'ar.v. Sirrrp., le4 1966. p. 377. Taylor. J. M_ Jcnner. P. M..& Jnne.. W. 1..11964). A comparison of the toxicity of some allyl,.propenyl, and propyl compnunds in thc rat. '/b.cir. uPPI. 1'hnrnrar• 6, 378.. U:S..Dispen.wrnrr-rundPhpsirinni P/rurrnurrdn!q•(1967). 261h Ed. Edited by.A. Osol, R. Pran & M. Allschule. p..556. J. B. Lipincou Co.. Philadelphia. Wong, K. P. & Sourkes, T. L. (1966). Metabolism of vanillin and related substances in tho-rat. Cun, J. Biachem. 44.635. ynder, E: L. & Holfmann, D. 119631. . Ein experimenleller Beilrag zur Tabakrauchkanuragenese. Dr. med. WMsr'hr. 88, 623. 07 Addilional rrferences /arrlnred }issurs nnd orynnelles Baldessarini, R..J. H97q. Compounds anlagonistic to norepinephrine retention by rat brain homogenates_ Binrhrnr. Ph'annar: 20,.1769; citcd1from C/iem. Ahsrn 1972, 76, 107865. Evans, B. K.,.James. K. C. & Luscombe, D. K. (19781. Quantitative structurractivily relationsllips and canninative.activity..L.p/wrnr..Soi- 67, 277.; cilcd.from Chrm. Ahsra 1978, 8g, 163898. Evans. B. K.,.)unes. K. C. & Luscombe, D. K. (1979). Ouantitative structure aetivily relationships and carm!native activity. 11: S!criccoruidcrations...L phrrrm. Sut. 68, 370: cnedfrom C6rm. Ah.vr. 1979. 90, 179923. Gee. 1. B. L., Vussallo, C. l., Bell, P- Kaskin- 1., Basford. R. E. & Field. J. B. (19701: Catalzse-dependenl peroxidalive m¢rabo!ism in the alveolar macrophnge.during.phagucytosis. J: r/in. lrrrrsc 49, 1280: cited from Chr•m.. Ahsrr- 1970. 73, 53628. Jobin. F. & Tremblay: F. (1970). Platelet reactions and immune processes. 11. Inhibition of platelet aggregation by complement inhibitors. ThronrH. Dinrh. /ruenurr/r..2Z-466; cited from,Ch.vn..ah.ar. 1970, 73„23561. Lapshin. 1, 1. & Rudinu, T. G. 119741. Ruleof phenols of smoking liquid of the Moscow Inslitutc of the National Economy,during fish smoking. Rph. Khn_. (Moscow) 1974 (2), 75: cited from ('hunr. :IN..v. 1974, 81, 2a3(N. Oshino. N. & Sato. R. (197H. Stimulation by phenols of the reoxidarion of microsomal bound cyrochrome b5 and its implication to fatly acid dkealuratiun. J. Biochrnr., Tnk)n 69, 169; cited frnm Chrm. .4hsrr. 1971. 74, 94329. Paul. B. B. H9791. Human peripheral lymphocytes: biochcmical and antibaoterial activities and comparison with polymorphonuclenr leukucyles. J. Rr•riCUlbeudrnhel. Siu. 25, 49: citcdfrom Chem. Ahsrr. 1979, 9U, 166437. Rilcy- P. A. (1969). Hydroxyanisole depigmentation in rirrosmdies. J. Prnh. 97, 193; cited from Clunr. Ah.rrr. 1969. 70, 113553. Riley, P. A. (1970). ..Mechanism of pigment-cell toaicity'produced by hydroxyanisole..J. Pnth. 101, 163; cited from: Clienr. dhsrr. 1971: 74, 2]688. Rucinska. E. (1968). Myotropic activity of sympathomimetic amines. Arnr pliynio(, pnl_ 19, 417: cited from C7renu d6srr. 1968. 69. 94328:. Sgnrugli, G., Delta Corte. L.. Rizzotli-Conti, M. & Giotli,. A. (1977).. EHects of monocyulic compounds on biomrmbranes. Biurlum; Pliarrnmc 26, ?145; cited from C6em. Absrr..1978, 88, 45655. Cells Pitoni, A., Aneker, K., Arnccnius E. & Enzell. C. (1975): Effects of tobacco and tobacco smokemnstituents on cell mullipllcation irr rirro. T n.cicaloqy 5. 49; cited from Chem. Ahsrr. 1976, 84, 39555. Svauss:, R. R.. Paul,. B. B.. Jucobs, A. A. & Sbarra, A. J. 11970k Role of the phagocyte in host-parasite interactions. XXII. Hydhogen pernxidedependent decarboxylalion and deamination by myeluperoxidase and irn relation to anlimicrobi:J activi!y. J. Rurh'uluendurlml..Sur.7, 754] cited from Chrnr. Absrr. 1970. 73, 10816. Strau>s, R. B., Paul. B. B-, Jaeobs.. A. A., & Sbarra. A. 1. (19711. Role nf dre phagnryle in htKt-parastle interactions. XXVII. Mycluperoxidase IirOr-C-I-mcdialed aldchyde formation artdhts relation lo antimicro- Hiol activity. lnhrr. fmnum. 3, 595; iitalfrom Chrrn. Ahsrr. 1971, 74, 123121.
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POPULATIONS AT SPECIAL ND RISR: C Ct (I AE;SORPTION, DISTRIBUTION MEDICAL EXF'ERIENCE INDSCATES.THAT TOXIC AND EXCRETION QUANTITIES CAN BEASSOf;l3EDTHROUGHTHE SKIN , QUITE READILY. [Patty, F. (ed.). Industrial Hygiene and Toxicoloqy: Volume II: Toxicol.ogy. 2nd e:d. New:Yor-k:: Interscience PuLLisher:, 1963. , p. 1.664] **ACCEPTED** ABSORPTION, DISTRIBUTION METHOXYPHENOL LARGELY ABSORBED FROM AND EXCRETION DIGF_STI'.VE TRACT pc,STORED IN BLOOD, KIDNEYS, & RESP ORGANS. EXCRETED BY RABBITS IN COMBINED FORM WITH SULFATE (15Z') AND GLUCURONIiC ACID (72%). ENatianal Research Counr_i1. Drinking Water & Health Volume1.. Washington, DC: National Academy Press,. 1977. , p. 7403 **ACCEE'TED** ABSORPTION, DISTRIBUTION ABSORFTION WAS DELAYED WHEN IEUFROFEN AND AND EXCRETION GUAIACOL ADMIN. SIMULTANEOUSLY. CCATANESE BET AL; STUDY OF SERUM CONCN OF IbUPROFEN &4 GUAIACOL T.NRATS AFTER.ORAL ADMIN OF I5UPROFEN, GUAIACOL & AF 2<t59; DOLL CHIM FARM 116(4) 232 (1979)7i*#ACCEf?TED** h'{ETADOLISM/METABOL.ITES SEVERAL STRAINS OF ASF'ERGI'LLUS NIGER HYDROXYLATEDANISOLE TO GIVE O~-HYDRUXYAdISOLE AS MAIN PRODUCT. CL+OCKS SM; F'HYTOCHEMIST}-;Y 6(6) 785 f1967)7 **ACCEF'TED*<* METAFtOL..ISM/METAEOLITES METABOLI.r-iM/METABOLITE.~'i O-METHOXYF'HENOL YIELDS,T-•-METHDXYCATECHOL PROBANLYI1\1 RA?IBI'T: DALY ,I'ET AL: MED~ CHEMif B'1;13 (1965), YIELDS O-METHOXY6'HENYL SULFATE PROEsAALYIN RAfirf)IT: WILLIAMS RT:. BIOCHEM ,I; 32 E78 (19343.). /FROMTAFLE/ C6oodwim.,, Es_L.. Handbook afIntermediary Metabolism of Aromatic Compounds. New York::Wiley, 1976. G-7I #*ACr_EPTED**- O-METHOXYPHENOL YIELDSCATECHOIL IN RAT; WONG KF, & SOU"'k:ES TL.;. CAN JB!IOCHEM;. 44 635 (1966). YIELDS~1,.2-DIMETHOXYEENZENE IN MOUSE,. RAEBIT, GUINEA F'IG& RATg. AXELROD 0, DALY J; BIOCHIM.BIOPHYS ArTA;, 159 472' (1966). /FROM TABLE/ CGoodwim, E.11_. H'andhook: of Intermediary Metabolism ofAromatir Compounds. New Yorka Wiley, 1976. G-77 **ACCEPTED** HIOLOG~ICAL HALF--LIFE ND MECHANISM OF ACTION ND INTERACTIONS ND
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AN CA 1 rST-- (.1!9 ): 166 i.:18Tm. TI Influence of water on guaiacol pyrolysis AU La:wson, J. k.y Klein, Mi. T. J CSDep. CYiem... Eng., Univ. Delaware LO Ndwa-r-k, DE 197116, USA SO! Ind. Eng. Chem,. Fundam. , 24 (2) , 203'--8 SC._2--0 (Physical C7rganic. Chemistry) DT 7' Cn: IECFA7 IS 01:.96--4313 PY 1985 LA Eng IT *~~9U--rD5r•L#~~ (pyrolysis of, effect of water on) (I ANI CA94.(,17) : 1133851rJ TI Pyrolytic studies on the origin of phenolic compounds in tobacco sm AU r_hl.otahauer, W. S.; Chortyk,0. T. CSTo6accm. Health L-ab., Sci. Educ. Adm.. LO Athens, GA. 30613, USA S0, TQb. Imt., 183(2), 25-9 SC 4-13 (Toxicology) L7T' 3 CO T&ICIAE IS, 0049-3945 FY 1981 r,A Eng I,,T .***90--0=--1.*** '. 95-48-7, biological studies 95~-65-895-71-6 95--87--4 97--53-O 105-67-9 106-44-5, biological studies it,6--39--4, biologic.:+: sti.rrjies 1t)5-6c^o-9 108---95-.", biologicall studies 12(1-.B0-9', biological .. studies 123-Q7--9 12?-T1--9, bi.ologicaIl st:.rdi~es 452--86-8'4B8.--17--5 1'Qc?-71- 6 1319--77--v 1.2 380.-_1[18.--9 2 6/,:~;8.-i,?:.--q (o+ tobacco smok:e, after pyrolysis, sper_ificc l.ea{constituent as sollr ce af ) AN CA74 ('2 6): 14~3525p. TI Kinetics of thermal conversion of guaiacol and veratrole AUI Kravchenk:o, M I.; N•april.anov, A. I.; Korotov, S. Ya. LO USSFt SO Nauch. Tr-., Leningrad... Lesotekh. Ak:ad.,, 1T5(^), 6ii--4 Fram;. f,tef. Zh., Khim. 1970, Abstr. No. 24P17 SC 41.3 . (Cellulose, Lignin, Paper,, andUther Wood Products) DT J CO NATLAZ PY 197n LA Ru.se IT at.,p~91--16--7 (F:inetics.of pyrolysis oi, as wood tarmode]i compd.) 19
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AN CA70(5,...:17626MTI Effect of doses of mineral fertilizers in soils with varying moisture requirements on the volatile phenolsin tobacco smoke IT ***90--n5;...i.K*r*yp8:-...95--=,.. biological studies 1.319_.-v-3 (in tobacc.o ~mokm, fertil.izerss .in, relatiun to) AN CA69 (23) :: 97760k TI Chromatographic method for the analysis of volatile phenols in tobacco smoke IT 89-•83--8 ***9n-05-1m** 106-44•-5, 108-39-4 1C 8-95-2, analysis 130n--71~-6 (detn. of, chromatographic, in tobacco smoke) AN PA69(2.3):93510d TI Composition of tobacco smoke. VI. Gas chromatographic separation o many lower monohydric phenols I~T 190-00-6 ***90-05--1*** 95-48-795--57-8 95-65--B 95-87-4 105-67-9 106-44--5 1US-39-4 198-68--9 108-95-2,, analysis 123--07-9 150-19-6. 150-76-5 526-75--V~ 576-26-16L•'0--17-7 (detection of, in tobacco smoke, chromatographic) -~ AN CA68(17):75739k: TI Constituents of certain tobacco types. I. Steam volatile phenols of Latak.i a IiT ***90-05-1*** 91-10-1 93-51-6 95-Ei7-4 97-5T-C> 105-67-9 108-95-2, b.ioTogicai stuidies 123-07-915t3-19-61ti0-76-5. 528--75-0 620-17--7 1300-71-6 2785--87-7 2785-89-9 7786-61-0 (in tobacco variety L~;-atakia) 5
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AN;. C:A93.. ( 1) . -:.:'77ov.. TI Isolation and characterization of two isoperoxi:dases from tobacco tissue cultures. IT ~9ScJ--r95r--.1!.*s# 9 .;~...61--5 ::n5-01-1: .. 7-...9.7--9 11-:5.... 24-6 (reactilon of, with isoperoxidasc=_s of tobar-r--n .r-u 1t ,a.re, {::inetics of7 AN! CA92( 1).:.T_•-'507.. TI Rapid pyrolytic method for evaluating effect of tobacco variety, growing, harvesting, and post-harvest treatment on smoke composition IT 54-11--c 5'7-1t7-3,,, analysis ***96-05-1*** iO8-95--2, analysis 120-8ii-9, anal ysi s5cJ4-96-1.. 630-04-6 (,detn. of,., in tobacco smoke, cultivationi and vari:ety in relation ti o ) AN'CA91(15):119308a - TI Tobacco leaf mesophyll peroxidase. Isolation, purification and some properties IT***90-_cn5-1'*+t# (PeroHidase isoenzymes oftobacr_o leafresponse to) AN CA86 (7) :;4E1.T_.72e TI The characterization af cigaret smoke from Cytrel smoking products and its comparison to smoke from flue-cured tobacco. III. Particulate phase analysis IT :,03'2-8, biological studies :,4-1i-5 56-Bi--5, biclogica.l studies 62-75-.9. (35--01.-0,, biological studies 90-Ait--6. *-++r9R--os -1k#~95-4i8-7,- biologicaS, studies. 95--65-f~. 95--87-4, 105-67-9 (of Cytrel and tobacco smoG:e) AN TI CA85 (15) : 1in6779f An i~nvestigati:on of some factors affecting phenol productionin tobacco pyrolyses IT 90-i,0_..6 95-48-795--87-4 105-67-.9 106-44-5 109-39-4 108-95-2, biological studies 123-07.-.9 620-17.-7ry. (forcmation of, by tobacco pyrolysis) / m a N N AN! CA05(9) :591949.'s (ibs04- ~ TI Studies on compositions of smoke components of lamina and midrib & cigarettes. Part I. Comparison of phenols in smokes of lamina and 09 IT midrib of flue-cured tobacco 57-10-3,, biological studies 87--66--1 . 90-0i-r-6 ***9U`-05--1*** 9O-15-: 91-1Ci--1 95-48--7 95--6:;-8 95-;-1--6. 95--87--4,.. 1n5,-67-9' 695--84-111.24..-39-6 2628-17-': 6n5._--0;;-77786--61-0 CI (of tobar-co smoke)
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CE~ (I ( _I :=1N CA98.(7) : S'tr}:J8t TI Mutagenicity testing in the Salmonel.la,typhimurium assay of phenolic compounds and phenolic fractions obtained from smokehouse smoke condensates AU Pool, fi+. L.; Lin, I`. Z„ CS Inst. To>;ico1. Chemo,ther., German Cancer Res. Cent. LO Heidelberg 69t]rj,. F-ed. IRep. Cher. SO Food Chem. To::ir-oP., 20(~40, 7,93--91 vC 17-5 (Food and Feed Chemisiiry) DT J CC] FCTC]D7 IP. 027E3-6915 PY 1982 LA Eng IT **#9n-05--1*** 91-1U-1 95-48-7, biol~ogicall studies97--53-U 105-67-9 D16-44-5:, biological st~-rdies 108-39-4, biological studies 1;08-95-7, biological studi:es 120-80-9, biological! stud'ies. 1 21- 33--5 (of smokehouse smol.:e condensate,., mut-agenicity in re7..ation to), AN CA97('.1)::2044u TI Mutagenic effect of monophenols and diphenols in Saccharomyces yeast strain 15V-P4 A} Pavlenko, V. V.;. Trubacheeva, L. Ya. LO l!SSft 90 Vliyanieffenol. Soedin. na Gidrobiontov, Irp<utsk 40-52 Fr om: Ref. Zh.., Tok.si.kol. 1982, Abstr. No. 375--T•8 SC 4-6 ("Toxicology) DT J Pv 1981 LA Russ IT 95--48--7, biological studies 108-46-3, biological studies 1rr8-95--C, biological studies 120-80--9, biological studies 121-33-5 123-31-9, biological st+-rdies (mutagenicity of, to Saccharnmyr_e=> cerevisiae) AN CA97('1).7.798fTI The use of an alga Chlorella pyrenoidosa and a duckweed Lemna perpusilla as test organisms for toxicity bioassays of spent bleaching liquors and their components AU Rowe,, E. L.; Ziobro, R. J.; '. Wang, C... J. K.; Dence,. C. W. CS Coll. Environ. Sici. For-.., State Jniv. New~ York ~ LO Syracuse, NY 1MCry USA (.1 SO Envir on.. Pollut.,. Ser. A, 27(4), z?S9L-96 N sC 4-~ (Toxicol.ogy). ~ _r,0 EPEBD7 IS 0143-1471 rA PY 19e2 LA Eng IT 88-06-2 ' *#*9t7-n5-1~*+ 1Ct8--95--2, biological studies 12~~--.Bp_9, (toxicity of, to: Chlorel.]la..pyrenoidosa.and Lemma perpusilla) 9
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CRIS/USDA 1975-87/OCT 004471:9 AGENCY ID: ARS 7-624 PROJ'NO: ~62r?-2716r>-010-0UDPROJ TYPE: INHOUSE PERlOD; 02 f7CT78 TO U'?OCf 88 INVESTa POWELL R G; MIIGOLAJCZA: K L; MILLER R.W PERF ORS:. CULTR CONT-.--I!NSCT PATH Q: FEED WESTERN COTTON RES LAB LOCATION: NORTHERN REGIONAL RES CENTER PEORIA I'L..L. 61604 BIOLOGICALLY ACTIVE PLANT CONSTITUENTS FOR PEST CONTROL AND MEDICINE' OCaJECTIVESsDiscover, isol.ate, and characterize biologically active plant constituents with potential utility as pest control agents or antitumor agents. APPROACH: Prepare extracts from NRRC's seed collection and selected whole plants. UseEuropean corn borer larvae fori.n-house screening. Ship extracts to variourscooperators.for screening against a variety of other pests. Also ship extracts to National Cancer Institute contractorsfor srreeni.ng in experimental tumor systems. Also consider plant resistance factors and other allelopathic agents. Fractionate pr-omi.singextracts to concentratee activity. Isolate~ individual compounds from most active fractions and'c.haracterize them structurally by spectroscopic examination, chemical degradation and.interconversions. Synthesize compounds of special interest if required to provideadditional amounts for evaluation. F'ROGRESS:85/41: 85/12Pesticidal activity of a noveli acetogenin from pawpaw has been shown cooperation with Dr. J. McLaughlin, Purdue) to be extensive; its structure has been determined. Ant.if'eedant and toxicity studies of extracts from seeds of MeSiaceae plants demonstrate that a number of them.including those of Aglaia cordata, Chickrassia tabularis, and'Sandoricum koetjape are more effectiven than neem seedle;:tractsin controlling fal.l armyworm larvae and striped cucumber beetle adults in laboratory bioassays. Some of the metabolic products produced by Diabrotica spp. fromcucurbitaci~ns have been il.solated and'. struct.uredeterminatioo is underway. Fractionation of insecticidal materials from. Cornus.s florida, Hydrangea macrophylla, and Dithyrea wislizenii is in preliminary stages. A large-scale fractionation to isolate aLlelochemicals from alfalfa is progressing. The volatile components of peach bark that produce the greatest antennal stimulation in the lesser peachtree borer females have been determined to be guaiacol,, meth~lbenzoate,l and 1-•phenyl-1,2-pr-opanedione. This work, as well as cooperative research on apple maggot fly resistance with Dr. H. Goooewardene (ARS, F'urdue)and rice weevil resistance in corn with. Dr. •3. Rodriguez (University of kentucky), is virtually completed. ' PUBLICATIONS: 85/01 85/12 MADRIGAL, R. V., ZILKOWSKI, B. W., and SMITHy- C. R. 1985. Structure-activity relationships amongmaytansinoids in their effect on the European corn borer, Dstr-ini.a.nubilalis (HUbner).J. Chem. Ecol. 11(4):5r?1-506. - - MIhOLAJCZA6.:, f;.... L., Z.ILk::OWSk:I, B. W., KAHN M. Z. A.,. SMITH, C. R.,, and BURKHOLDER,. W. E. 1985. Attractants for OryzaephiL_rs.surinamensis (L.) from oats: Dimethyl succinate, glutarate,, and adipate. J. Agr. Food 20 8'7119151 „ __........ . - _,- -- . _ .. ... i ;
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AN C4Wv "v 1 Ys 1la67e.. TI Effect of phenols of wastewaters on some energy-dependent parameters of Chaveal alga cells AU Plek:hanov,fi... E.;. Stom, D. I.; Khitrov, Yu, A.; Yasinovsk:ii, V. G.. C5 Mask. Gns. Univ. LO Mosrow, IIISSR SCl Eriol., Nauki (Moscow), (2), 4•'0-7SC 4-' (To>ric.ology) DT J CO BINk::BT I S 030: :-•41 19 PY 19Er> LA Russ IT 1ir6-51-4, biological studies 1c_i8-46--3, biological st.udie<.a 108-95-2, biological studies 12Cr-8r?-9, biol.ogical' studies 123-31-9, bialogical'stUdies (toxicity o,f,, too algae, elec, activity inrel'ation to) AN CA92 (2.;) :209B32s TI Mutagenicity produced by aqueous chlorination of organic compounds AU Rapson, W. Howard; Nazar, Mark A.; Hut.s4,y, Victor V. 05 Dep. Chem Eng. Appl. (lhem., Univ. Toronto. LO Toronto, ON', Can. SO Ltul l. Environ. Cont am. Toxic.ol.., 24(4), ff-90--6SC 4-;T (Tov,irod.ogy) DT J CO BECTA6 IS 0007--4f361 FY 1980 L.A Eng IT 65-85--r7, biological studies67-64--1, biological studies 86-51-1 87-66-`1 88-ci6-2 ***9<:r-(}S_-..1*.rt* , 91-16-7 97--_:-ir ?9--5ir-: 99-93--4 99-..+'6-7, biologticall studies 106-06--i. 1(.)p...5i-:, biological studies106-G1--4,, biological studies 107-22-2 1C?Ei--46--3, bi.olo].ical studies iiag-73--6 108--r'i5-2, biologicaL. sti_rdiFS 110-16-7, biol.'.ogicall studies 1.1n-17-8r, biological. studies. 12Q-ery-9, biolog.ical studies 1?1-_33-5 121i-?:4--612i-71-1 1'L's--qg-<t 1'2;'-1.P--5,, biological studies 12T--31--9, biological'st.udie=,. 1.:.:4r-gtn-..7 45:?-56-5 488--17--5 498-02-2 5]S--7U--4 514-10-3 `,86-.'7--8 6b5-931-0 697-91-6877`-'=-S 1131--6^-i) 1i~6--55.-6 '.-'43,.-S.?-... 2539-17-5 . 3428-i4--8' S(stL5.__-<, biological'Stlld l.eslmutagem.i.-it;. o~f, in cellul'ose pulp, ch.lorinati.on e*-fectt on) I 13
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LIFE SCIENCES COLLECTION 1978-87/OCT C 1'11'2629 82001485762 Hazardous materials: o-Anisidine hydrochloride.. Anon. DANB. PROPF..R.IND. MATER. REP.; 6(5),, pp. 58--61 19B6• Language: English Document Type: Journal article-original research~ Subfile: 24 Toxicology Abstracts o-Anisidine (hydrochloride) is.used chiefly in.the manufacture ofdyesf o-Anisidine is listed as a possible ingredient in permanent oxidation hair dyes; however, as of 1977, it had'not been used'in the U.S. Another use ofo-anisidine is as a star-ting material in the synthesis of ~~ua~.ac~ol. C--methoi, heno . is carcenic to Fis er 344 ra~nd. B6C3- mice. Transi.tional-celll carcinomas.or papillomas of bladder of both sexes, both species; transitional-cell carcinomas of the pelvis of the kidhey and follicular--cell tumorsof the thyroid in male rats.are induced. 1105883 82001467865. The toxicological implications of the interaction of butylated hydroxytoluene with other antioxidants and phenolic chemicals. Presented a.t:Symposic.rm on Food Antiox.idantsc International Perspectives, Washington,. DC (USA), 21-23 Apr 1986 Thompson, D.C.; Trush, M.A. Dep.. Environ. Health. Sci.,.. Div. Exp. Pathol. and Toxicol., Johns Hopk.ins Univ., Sch. Hyg. and Public Health, Baltimore, MD 21205, U5A FOOD AND CHEM. TOXICOL.; 24(1tD-11) Publ: Publ by: , 1986., pp.1189-1195 1986 In FOOD ANTIOXIDANTS: INTERNATIONAL PERSPECTIVES... Gr-i.ce,.. H..fi.. (ed.) l:.anguage;English Summary Language: Engli~sh Document Type: Book-chapter article Subfiles 24 Toxicology Abstracts 5ut latPd hydr-oxyanisole (CtHA) enhanced both the in vitro peroxidase-catalysed cova ent binding of butylated hydroxytoluene (BHT) tomic:rosomal protein and the formation of BHT-quinnne methide. Eugenol, methylparaben,, vanillin, guaiacol, ferulic acid and several other phenolic compounds commonly usedi.n food and cosmetic products also enhanced the. metabolic acti~vation of BHT. BHA was the most effective compound tested. Microsomes from: lung, bladder, kidney medullaand small intestine of various animal'species, including man, were also ableto.support this interaction af BHA and NHT using either hydrogen peroxide or arachidonic acid as the substrate:. These in vitro observations were extended to an in vivo mouse lungmodel.. Subcutaneous injectionss of BHA significantly enhancedl the lung/body weight ratio of mice given intraperitoneal inj.ections of subthreshold doses of BHT. I I
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CHEMICAL ABSTRACTS 1968-87/DEC AN CAI07(15.);.128592^ TI Manganese poisoning and the attack of trivalent manganese upon catecholamines AU Archibald, Frederick S...;Tyree, Curtis. CS Dep. Microbiol. Immunol., McGil:1 Univ. 4.0 SU SC DT CD IS PY lLA I'.T Montreal , F'0,, Can,. Arch. Hiochem. Biophys., 256(2), 63rC-5'4 4-3 (Toxicology) J AFiL+1A4 0003-9861 1987 Eng 65--85-6,, reactions 89--86--i **->t90-Q5-1*+* , Guaiacol 91-16•-7, Veratrole 99-10-5 99-5Cr,-3 99-'36--7, reactions 1r7h_-511 -4, reactions 108-95-2, Phenol, reactions12d-BO-9, Catechol, reactions 12:3--31-9,. Hydroquinone, reactions "'C13-C57-d 303-38-i3: . 490-79--9(ox.idn.. of,, manganic pyrophosphate-mediated, manganese neurotoxicity in relation to) C3 (f AN CA'1.07(:3).: i':9284v TI Antimutagenic effects of phenols on MNND-induced mutagenesis in Escherichia coli AU Kushi, Atsukeg Yoshida., Daisuke CS Cent. Res. Inst., Japan Tob. Inc. LO Yok:ohama 227, ,?apan. SO Agr-ic.. Biol.. Chem., 51(5), 143!7-4ra SC 4-6 (Toxi;cology) DT J CO ABCHA6 IS piu-12-1369 FY i98-,7 LA Eng I'1' o-Methoxyphenol 90-15-3, .alpha.-Naphthol 90-43-7, n-Phenylphen=,1 91-iU--1, 2,6-Di:methaxyphenol 92-b9-'', p--Phenyl.phenol 95--48--7, o-Cresol, biological studies 95-65--8, 3,4-Xylenol 95-87-4, 2,5-Xylenol 97--53-0, Eugenol 98-54--4,, p-tert--Euty1 phenol 99--71--B, p-..ec-Erutyl phenol 1U5-h7-9,. "2,4-Xylenol 146-44-5, p--Cresol, biological studies 198--~9•-4,.' m-Cresol, biological studies 108-68-9, 3,5-Xylenol 108--95-2D, i'henol, deri vs. 123--07-9, 4-Ethyl phenol: 135--19-F, .beta.--Naphthol, biological studies 15U-19•-6, m-Methoxyphenol 15r3-76-5, p'-Metho>:yphenol 526-. 75--0., 2, 3--Xylenol 576°26--1 , 2,,6--Xy1enol 1745-81-9, 2-Allylphenol 7469-77--4., 2-Mei:hyl--1 --naphthol (MNNGmu.tatiomof Escher-ichie. coli response to, structure inrelation to) r F
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MEDLINE 1966-87/DEC 03297125 77199125 Mutagenicity of benzidine and related compounds employed in the detection of hemoglobin. Ferretti JJ; Lu.Wa. Li~u MB . Am JClin Fathol Jun 1977, 67 (6) p326-7, ISBN UCr62'-9173 Journal Code: 3FK Languages: ENGLISH Seven compounds commonly used as chromagens for the detection of hemoglobin and its derivatives have been assayed for mutagenicity employing the Salmonella/mammalian microsome test. Three of these compounds, benzidi:ne, o--dianisidine,, and o-tolidine, were shown to be mutagenic. Since benzidine and o-tolidine are already known to.be carcinogens, there is a high probability that o-dianisidine will also prove to be a carcinogen. Four compounds tested with this system,, o-anisidine,, diphenylamine, guaicol, and o-toluidine, were not mutagenic. 029552.e976136229 Contact dermatitis from antioxidants. Foed-Fetersen ,I; H].orth.N Br J Dermatol Mar 1976, .94 (3) p233--241, ISSN 0007-0963 ~„ Journal Code: AWO f~..lb Languages: ENGLISH In a search for contact sensitivity t.o: antioxidants we patch tested consecutive patients referred with eczematous dermatitis. Six case~t of .allergic contact sensitivity to nordihydroguairetic acid (NDGA) were observed. Three had been sensitized by one brand ofcreamcontaini~.ng 0.1% NDGA, i:n,threepatientsthe source of sensitization could not be traced. In four patientss we found positive patch tests tobutyl.ated hydroxyanisole and:Cor to butylated hydroxytoluene. In two cases theposi.tive patch tests were re.levant,, since both patients remained asymptrma:ti~c when antioxidants were avoided in food. They both had acute flares of vesicular eczema on the fingers after oral administration of small amounts. Gallate esters and Vitamin E (d,l-alpha-tocopherol) each gave one unexplained positive patch test. The present data suggest that f'urther search for hidden sensitizers in topical medicaments and cosmetics.is warranted. A declaratiomof all ingredients in industrial productsshould be placed on.the l.a.bel.. I 1
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DIALOG File 159: CANCERLIT - 1963-57/Nov i;r_-~4m71f3 ICDB/t33Qc757~78 , GUAIACOL. MONOGRAPHS ON FRAGRANCE RAW MATERIALS OpdykeDL., Letizia,CRes. Inst, for Frragrd.nce Mater-i.:,'_s., Incorporated, Englewood Cliffs, NJ, n76?2 Food Chem Toxicol; i0(Bupp.l):b97--701' i982 t.anguages: ENGLISH'.., Document Type: Journal Article Journal Announeement: 17303G~~~ieaia.col. is approved for foodd use and'isincIuded, at a level of 1U.ppm, in the list of artificial flavori.ngg substances that may be added to f'oodstuffs without hazard to publ,_c health. Guaiacol was not mutageni.c in theSal'monr.-lllalmammalian mic:rosome test. Gmaiacol was effective in reducing thewte o-F sarcoma 190 tumors ini mi..ce. The role of guaiacol as co-r_arci,nogen i.n '._obar_co smoke was q+-iestioned.. (9S }:e.fs) 0-314,17:~ 1CDBIEi207;1309 CATALYSIS OF NITROSATION IN V1TRO AND IN VIVO IN,RATS BY CATECHIN AND RESORCINOL AND INHIBITION BY CHLOROGENIC ACID Pignatell..i B, Bereziat JiCp. Descotes G; EartscM.H (c/o. Bartsch) Programme Environmental Carcinogens and Host Factors, Ir?ter-neati.onal Agency Res. Cancer, 150, cours Albert Thomas,. F-69=72 Lyon Cedex 08, .. France Carci'nogenesi s: 3 (9) : 104I5....104'919.g2 Langr-rages: ENGLISH, . Document Type~: Journal Article Journal Announcement: G2'12 Measurements were made of the effects of pnenolic compounds,, some of which are present inthe human diet,, on the nitrosation of proline by nitrite togi,ve N-nitrosoproline CNPRO). Ini vitro, resorcinol, catechin, p--nd trosophenol and phenol were catalysts and chlocagenic acid an inhi.hi.torq guaiacol showed a marginaL catalytic e-Ffect. 'i-:oth the c.atalytic and~the inhibiting effects were dependent or+ pH a.ndlon the c.ancentrati.on of phenoliccompound's, catalysiss tryresor-cinol- and _atechin was increased at optimal ratios ofCniitrite]aCphenoli.c compo+_tnd'.],. Endogenous.nitrosationwas exr.mined in vivo by co-admini~stration of nitrite, pr-oline and a phenolic compo+_ind'tn. r-atrss and by monitea-:ng~ the amor-tnt. NPROexcre±ed in~ theuri~ne.. Under similar es;perimen}_al conditions, the catalytic effEcta observGd' in vivo decreased in.the same order as those observed i.n•-vi~tro: resorcinol greater than p-nitrosophenol greater than ca:techin:.grea~!ter- than phenol greater than or-equal to guaiacol; chl.or-ogenic acid acteH as an ilnhibitor. Catalysis and. .i.nhibitimn of~ N-nitrosation. in rats in vivo appears to occur via mechanisms similar to those in vitro, althou.gh tHre effe,ts in vivowerez smaller. The implications of our findingsfor the endngenor-rs formation of N`-rnitroso compounds and for variations in eiposu.re due t.o di~f+erent dietary constituents in, ..humans are discussed. (Author- abstre;ct): (35 Refs)
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1105754 .82[r01467633Hazardous chemicals: Guaiacol. Anon. DANG. PROPER. IND. MATER. REP.; 6(6), pp. 45-52 1986 Language: English Dor_ument. Type: Journal,article-original research Smbfile: 24 Tox:icology Abstracts Guaiaco.l L.D54] values have been determined for various animal species. The ai.itthorss also discuss human toxicity, symptoms, and the drug's metabolism and pharmaco4::inetics. (I 0711292 82000381934 Catalysis of nitrosation in vitro and in vivo in rats catechin and resorcinol and inhibition by chlorogenic acid. Pignatelli, H.; E{ereziat, J.-C,; Descotes, G,; Partsh, H. Intl. Agency for Res. Cancer, Programme Environ. Carcinog. and Host Factors, 150, Cours Albert. Thomas, F-69372 Lyon Cedex, 08, France CARCINOGENESIS; 3(9), pp. in4S-1049 1982 Langua.ge: English Summary Language: English Document Type: Journal articDe-origi'nal research Su.bfilea 24 Toxicology Abstracts Measurements were madee of the effects pf phenolic compounds, some of which are present in the human diet, on the nitrosation of proline by ni.triteto give N-nitrosoproline (NPRO). In vitro, resorcinol,, catechin, p-ni~.trosophenol and phenol were catalysts and chlorogenic acid an inhibitor; guaiacol showed a marginal eatalyti.c: effect. Both the catalytic and theinhibit.ing effects were dependent on pH and on the concentration of phenolic compounds. Catalysis and iinhibition of N-nitrosation in rats in vivo appears to~ oecur, via mechanisms similar to those to thoFe.in vitro, although the effects in vivo were smaller. The implications of the findings for the endogenous formation of N-nitroso.compounds and'.for variations in exposure due..todifferent dietary constituents in huma,nsar® d.".scussed. 0706861 82000368993-~ Absorption, metabolism, and elimination of selected phenolic compounds included in refined smoke. Part. II. Determination of guaiacol in the urine by gas chromatography. Wchl.anianie, metabolizm i widalanie wybranych zwiazkow fenolowych zawartych w rafinowanym dymie.wedzarniczym. Cz. II. Oznar_zami.e gwa.iakolu w moczu metoda.chromatogra-fii gazowej'. Senr_<uk, W. Zala:. To{::syk:o3. Inst. Bioanal. i Sad'ania, Srodowisk:aAF.ad. Med., F•bznan, F'ol and BROMATOL. CHEM. TOI,SYYOL'.9 14(2), pp~. 185-188 1981 Language: Polish Gt mm_:z .I`_gl.iag Fol i sFi 87119154 Document Type: Journal article-original research A gas chromatographic method for the determination of guaiacol in the urine iss described. Continuous extraction with ether was used to isolate guai~acol from the urine after previous hydrolysis. Variability coefficient of method is .+-. 8%. Chromatographic partition of guaiacol takes about 7 mini
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00.380847 67205867 The antischistosomal and retinotoxic effects of some nuclear substituted aminophenoxyalkanes. folIins RF; Ccnr VA; Davi.=-; M; Edge ND;HiPl ,d; Rivett k:F; Rust MA Br d Pharmacol (England) Feb 1967, 29(y) p24E-5H',, IBGN 00(j7-1188 Journal Code: 200 , Languages: ENrLISH Journal Announcement: 67111 SubfiTe: INDEX.MEDICUS Tags: Animal. Descriptors: *Alk:anes--Fharmacodynamics--RD; *Alkanes--Therapeutic Use --TU; +rGuaiac-ol--Pharmacodynamics----F'D; *Guaiacol----Therapeutic Use-TU; *F'henolsr-Fharmacodynami¢s--PD; *Fhenols-Therapeutic U2e---TU, +Retina ----Drug. Effects--DE; *Schistosoma---Drug Effects--DF; *Schistosomiasis--Drug Therapy---DT; Alkanes---To;;ici.ty--70; Cats; Guaiacol---Toxic_ity--Tn; Mice; Phenol.s--To:;icity--TO 6 I
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041.150071 90161071 iA study on acute toxicity of formalin guaiacol as a root canal medicament (author's tramsl)] Nishikawa T; Ya:suda. H; Kin-io K.; Hirata H; Sakamoto N; Ok:amotoH; Tsujimoto AHirosh~imaDaiga4.u. Shigaku lasshi1978, 10 (2) p213-7, ISSN'. C>046-7472 Journal Code: IDB Languages: JAF~F.SE Journal Announcements. 8009 Siibfile: Dental Tags: Animal Descriptors: *Formaldehyde--Toxicity--TG; *Guaiacol--Toxicity--TO; *Root Canal Therapy--Adverse Effects--AE; Anti-Infective Agernts,, Local--Toxicity ---TO; Mice CASRegistry No.: 50-00-0 (Formaldehyde); 9Q-Cr5--1 (Guaiacol) 04041316 801'52316 Tri- and tetrachloroguaiacol: results of a three and six-month feeding study in rats. Chu I; Villeneuve DC;. Yagminas AP; Valli VE Arch Environ Contam Toxicol 1979, 8 (5) p589-97, ISSN:Q09h-4341 Journal Code: 6YD Languages: ENGLISH Journal. Announcement: 8007 Subfil.ee INDEX:MEDICI7S Tags: Animal; Male Descriptors: ->~8ody Weight--Drug Effects--DE; *Eating--Drug Effects--DE; *Guaiacol--Analogs and Derivatives--AA; Enzymes--Ll.ood--BL; Guaiacol --Toxicity--TO; Kidney--Drug Effects--DE; Liver--Drug F_ffects--DE; Rats CAS Registry No.: 90-05-1 (Guaiacol) 04033565 80144565Intravesical chemical cauterization.and methemoglobinemia. Lebowitz RL Pediatrics. Mar 1980, 65. (3) p63W, ISSN ni-r31-401>5 Journal Code: DXV Languages: ENGLISH Journal Announcement:8007 Subfile: AIM; INDEX MEDICUS Tags: Case Report; Human; Male Descriptors: *Ca!itery--Adverse Effects--AE; *Chlorobutanol --Adverse Effects---AE; *Cystitis---Drug Therap.y--DT; -x-Guaiacol--Adverse:Effects--AE; *•Hematuria---Drug Therapy--DT; *Methemoglobinemia----Chemically Induced--CI; *Phenols--Ad!verse Effects----AE; Age Factors; Drug Combinations--Adverse Effects---AE; Infant, Newbor-mCAS Registry No.: 57-15-8 (Chlorobutanol);. 78457-0.1-9(PCG (combinatiorn:)).; 90-05-1 (Guaiacol) -. 71
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I (1385611679233116 Toxicity studies on chlorinated guaiacols in..the rat. q ChuI;, Ritter L; Mz+rino IA,Yagminas AP;, Villeneuve DC. Bull Environ Contam Tox.icoI Jun 7:979:', r2(3) p293--6, ISSN VDG7-4061Jour-nal Code: DFN Languages: ENGLISH' Journal Announcements 7912 Subfile: INDEX MEDICUS Tags: Animal; Male Descriptors: *Guaiacol--Analogs and Derivatives--AA; Aniline Hydroxylase Metabolism--ME; Guaiacol--Toxicity---TO; Iditol Dehydrogenase--Metabolism ---ME; 8'idney--Metabolism--ME;. Lethal Dose 50; Liver--Metabolism--ME; Rats 03836760 79213760 Preliminary pharmacological and toxicological investigation on 2-guaiacolglycolylamino-3,5-dibromo-N-cyclohexyl-N-methylbenzenemethanamine : a,new bromhexine derivative. Testa R; Gelmi G; Canestrinii C; Cattaneo A Farmaco [Prat] Jun 1979, 34 (6) p243-54,, ISSN CtCri4-027X Journal Code: F-SXLanguages: ENGLISH Journal Announcement: 7911 Subfile: INDEX MEDICUS Tags: Animal; Female; Human; Male Descriptors:. *Antit-ussive Agents--Toxicity--TO; *Li.romhexine--Anal'..ogss and Derival:ives--AA; Antitussive Agents--Chemical Synthesis--CS; Hlood Pressure--Drug Effects--DE; E'rromhexine--To,:ici~ty---TO;. Guai.acol --Analogs and Derivatives--AA; Gucaiacol--Tox.icitly--TO.; Gu~i.nea Pigs; Heart Rate---Drug Effects--DE; Mice; Fats; Stereotyped Pehavior--Drug Effects--DE 0.3801137 79178137 [Infantile thrombopenias and drugs] Piastrinopenie infantili e farmaci.. Carnelli V; Pizzi E; Jankovic M; Sticca M'. Minerva Pediatr Sep 1'5 1978, 30(17) p1429--32, ISSN 0026-4946 Journal Code: NAM Languagese ITALIAhL Journal Announcement: 7909 Subfile: INDEX MEDICUS Tags: Human Descriptors: *Drug Therapy--AdVerseEffects--AE;.. *Purpura, Thrombocytopenic--Chemically Induced---CI;*Thrombocytopenia --Chemically Induced--Cl• Antibiotics--Adverse Effects--AE; Antimalarials ----AdverseEffects--•-AE; Aspirin--AdverseEffects--AE;; Child'; Child, Preschool; Guaiacol.--Adverse: Effects--AEF Histamine Hi Receptor Blockaders--AdverseEffects---AE; Infant; PyrazoDes--AdverseEffecls--AE; Sulfonamides---Ad.verseE f f ec tt s--AE 4
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1195 11-4-85 Flawring 6 172b1a Synthetk flavorlns eubetanree aPi adiuvada. Synthetic flavoring substances and adjuvants may be safely used fn food In accordance with the following con- ditlon& (a) They areused in the mtnlmum quantity required to produce their in- tended effect, and otherwise In accord- ance with all theprinclples of good manufacturing practice. (b) They consist of one or more of the following, used alone or in combi- natlon with flavoring substances and adjuvants generally recoenlzedes aefe[n food,prior-sanctloned for such use, or resulsted by an appropriate section In this part. Acetah aceNldehyde dlethyl .oetal. Acetaldehyde phenethyl Propyl acetal. Acetannole: f-methozYSixtophenone. Acetophenone: methyl phenyl ketone. Allyl anthnn(1ata Ally] butyntte. A11y1 CWumate. Allyl cyclohexanewetate. AByI cyelohelnnebutynte. Ally] cyelohexeneh s. Allyleyelohexaneproprlon.te. . Allyl eyelohesanevalente. Alyl dMU1fMe. AIlyl2cthylbutyHte. dllyl henaw(e; .Ilyl c.proate. Alyl a-lononr, 1-2,&s-trlmethyl-2-cyclo- hexene-t-yl}l,t-heptatltene-34ne. Allyl isotldocyuute:.muefard oLL Allyl Lovalente. AByI mereeptuq 2-pmpentl-thloL Ahyl nemnoete. Allylactaoofte. AByl Phenoxyeoe(.te. Allyt PhePylecetate Ally] proplonate. Allyl eorbatq Wyl s,4-Itexudlenoate.. Allyl wlflde. AB71 tla4lr, el)yl bnw-R-methYl-R-buten- oe4e Ai1y110-undecenoate: Ammonium Isovalente. AnvnonWm sulfide. AmYI alcohol: pentyl alcohol. Amyl butyrate. a-AmY1NnnaulalAehyde. a-AMnYlclmnamaldehyde dlmethyl acetal. a-Amylc/onumYl acetate. a-Amylc/nnamyl alcohoL.. a"tmYtcNufsmyl fotmate. a-AmyldnnumYl IaovaleRte. Am9t (onnate. Amyl heptanoet., Amyl hexenoate- AmYI ucteno.te, Food Drug Coamet[c Lzw Reports Agents 56,497-3 Anirole: methouyhenaene. . Anlqyl acetate. Anisyl aloohoC p-methoxybemzYl elcohoL Anisyl butynte, Atdxyl fo[mate. " Antryl phenylacetate. Anisyl proplontlte. Beeehwood ceauote. Bemaldehyde dimethyl acetal. Benmldehyde (19ceryl aeela): S-phenyl-m- dloxandiol. Benraldehyde Dropylene glycol acelal; {- methyl-2-phenyl-m-dioxolane. Benr.enethlol: thlophenol. - Bemofn; 2-hydroxy-2-pheny)acetophenone. Benrophenone; diphenYltetone.Benzyl acetate. Benryl ecetoetetate. Benzyl alcohol. Benryl benaoate.. Bemyl butyl ether. BenrYi butynte. Benayl °i^^ '^•r^ Benx91 2,9-dimethyleentonate: bePUyl methyl tlalate: Beney[ dlxulfidr, dlbenxyl d1sulBde. Benayl ethyl ether. Benuyl formate. 2-Benayl4-heptanone: bemyl dipropyl ketone. Benzy,l tfobutyrate. Beneyl leovalemte. Benx9l mercaptau: a-toluenethiol. Benapl metkoxyethyl ecetal; soetaldehyde beoxylB-methoxyethylacetal Benzyl phenylucetate. Benxyl propionate: Beneyl Wlcy4te: Birch tar oLL Bomeol: dcamphano). Bomyl ecetate. Bomyl formate. Bomy1 Ieovalerzte. Bertlyl vallRte. 6-Bo rbo1,2,E,De,1bD.4,6A.la$,6ba- decd-hYdro-la-i.aproDYl-7a,-umthyl-& nuethylene-cyclobuta [1,22,U dlcyelopea- tene. 2-Butanot 2-Butanone; methyl ethyl ketone. Butter ecide. Butter eetere. Butyl acetate. Butyl eceto4Ytale. Butyl alcohol: 1-butanoL ButylanthranlLte. Butyl butyr.te. ~ Butyl butyryBeetate; lactic add, butyl eNer, butyrate. a-BUtylctrmemaldehyde. ~ N Butyl emnuuate. N Butyl 2-decenoate. (~ Butyl ethyl malonate. ~ Butyl fonnate. (D Butyl heptanoate. ~ Butyl hexanoete. Butyl p-hydroxYbeonoate. ButYllenbutyrate. Butyllso.valerate. 21 CFR 172.515 1156,514.15 i
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Fragrance rawmateriuls monographs 701 lmds Inst. Ihuse in tbnbns$ 6-phos- _tivrty in on of an '166 64, in the class of nds. J. part of s-t. 829; sferase. iibitory nidase- l:.Aial. cifinty .1.~FBjur. \d` =lians'. c, and halide cited ra of Uzb. and Y7_, Is by rior. 1977, ~nds. and Yrd- ners an 11.1. rv Stom- D. 1. &Beim, A. M. (1976). The action of phcnolson vuriousalgali species. Jlydrohiul. J. (English uansialion of Gidrohiot. ZA.1 12, 43: cited from Ahstrners nn Ifeohh 671i•oro af Enrirummvnnl.l'ullurnnts1v79. g, 1018. Stom, D. 1. & Bnim, A.. M. 119761. Efw of phenols.on sortte spe4es of algac. GiJi'uhinl. Zh. 12, 53; cited front Ab.urmts on. Health EUrt.cuf Enrirurmirrwd Pulhnants 1977. 6. 11548. Slom, D. L-Timofeeva, S. S.. Ilelykh, L. 1. & Bulorov, V. VP. 119781: Rulc of C haraccae algae and other aquuticc plants.in the degradation of phenol cvmpound's. VtahO•e Resurm 1978 141. 105: cited from Chwn. Ahsrr. 1979, 90, 76089; Urs, N. V.. Rama R. & Dunleavy, J...M. (1975). Enhancement of the bactericidal activily ot a peroxidase system by phenolic compounds. Ph)•ropurhuhNly 65, 686; cited from Clieni. dbsrr. 1975, 83, 92168. Weslermark, U. &Erikscon, IC E. (1974k Carhohydrale-dependcnl cnrynticquiitone reduction during lignin degradation. Aetn ehem. xeand.. Sue 11,.28, 204; cited fromChcm. :1f1,tc 1974. 81. 877311. Willard. F..L.& Kodras, R. (19671..Survey~ofchemical compounds teslcd ib oirru against rumen protozoa for possible conlrol of bloat. AppL MicrobiuL 15, 1014; cited from Chem. Ahsrr. 1967, 67, 98799.. Inuertebrates Aldrich, I R-, Blum, M..S. &Fales• fL M. (1979)tSpecic-specifie natural products of adull male leaf-footed bugs (Hemiptera: Heleroptera[ J. chenr• Eral. 5,.53; c11ed from CAem. Ahsrr. 1979, 91, 137452. Burditl, A. K.. Jr. Hinman, F. G. &Balock, J. W. (1963). Screening of fumigants for toxicity to eggs and larvae of the oriental fruit'fly and Mediterranean fruit fly. J. «on. Eut. 56, 261; cited from Chrm. A6srr. 1963, 59, 5712: Duffey, S. S., Blum, M. S: Fales. H..M., Evans S- L Roncadori, R. W:,.Tiemunn. D. L.,& Nakaguwu. Y. (1977)+ Benzoyl cyanide and mandelonitrile benzoate in•.the defensive secretionss of millipesfes. J. rhvm. Eivl. 3,. 101!: cited'from Cherre Ahatr. 1977. 86, 68634. Henzelll R. F. D9701. Phenok an ultraclanl for the male grass grub beetle (Cnsleltvu cerdundreul. N2. J. uyrir. Rrs. 13. 294; cited from Chrnr. Ahsrr. 1970.73, , 76109. Jones- C. G. & Firn, It. D. (1979k Sonte allciochcmicals of Prrridium e,prilinum and Iheir involvement in resistaneelo Piori.e bructicae. Birnhrm. Srs't. Gra1.7, 187; citad from Ch.vn. Ahrtr. 1979. 91. 154676. Kopperman. H. L, Carlson, R. M. & Cuple, R. 119741. Aqueous chlorination and ozonation studies. I. Struc- ture-toxiciry correlalions of phenolic compounds to Daphniamusynm.Chemiro-Biul. Inreraniuns 9,.245; cited from Chem. Abstr. 1975, 82, 119707. Metcalf, R. L, Melealfi.E. R, Mitchell, W. C. & Lee,.L. W. Y. (1979). Evolution of olfactory receptor in oriental - fruit fly Darus dursrdis. Prru. rmm. Acud Sri. U.S.A. 76, 1561:.cited frumChcnr.Ahsic 1979. 90, 20D832. Meyer. H. J. &. Narris, D. M. (19741- Lignin intermediates and simple phcnollis asfceding stimulanls for Sool)msnnrllistrinms: J. Insecr Plusinl. 206 2015; citcdd from Chcrn..Ahstr. 1975, 82,.11696- Neuhauscr, E. F. & Hurlenstein„R. (1978). Reactivity of soil macroinverlebramporoxidases with lignihs and lignin model compounds. Suil Bfu/. Bior'hrun 10, 341; cited from Chwn. .9dsP. 1978, 89, 194198: Nolte, D. J. (1976). Locuslol and its analogs. J. Ltsert Ph)'siul: 22,.833; cited from Chrnr.. Absir. 1976, 85;. 156902. Smith, B. C. & Williams, R. R. (1976}. Temperature relations of adult Culavmeyillu nmeulhm lengi and, C.m. meJialis (Coleoptera; Coccinellidue) and responses to oviposilional stimulants. Can. Emumul. 108, 925;tited from Chem. Ahstr..1976, 85, 18961I. ' . Tusicufuyy Ashirova, S. A.,.Efranova, E. A.,.Pokrovskaya, E: A., Kurenko,.L. T.. Odinlsova, F. P., Mashikhin, E N. & Kogan, V. Y: (1977); Industrial hygiene and health status of workers engaged in the production of guuiacol. Vnpc GlyienpTruda; Professlmml'n Pnoot. RnMrchfkh Khim. Prom-sri:.p. 71: cited from Chern. Ahsrr. 1978, 89, 116995. Zamfir, G. (19721. Air pollution and bronchopulmonarytancvr, Rer. Alyd.-Sue. Med: N'm. lns/ 72. 273; citcd from Abstra•ts un Heulth' Ebios , f ErmirmuneufalPullutrous 1973. 2. 6464.
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E 2-DOdecenal... Eitragole. . . y-Ethoxybenaaldehyde: Ethylacetoacetate. Ethyl 2-aatyl-3-phenylproplonate; ethyl- betuylacetbacetate. - Ethyl aconltate. mlxed estere. . . . Ethyl aerylate. Ethyl pminte. . .. Ethyl anthranilate. . . Ethylbetmoate. . Ethyl benroylacetate, a-Ethyltipmyl butyrate;. a-phenylpropyl bu- tyrate. Ethyl btaesylate•, trldeanedtoic eold Eyelic ethylene glytol diester- cycl0 1,13-ethyl- enedioxytridecan-1,13-dioae. . 2-Ethylbutyl acetste. 2-Ethylbutytaldehyde. . 2-Ethylbutyric acid. Ethyl cinnamate. - Ethyl cretonate:trvna-2-butenoie acidethy- leater. Ethyl cyclohexanepropionate. Ethyldecanoate: 2-Ethylfuran. Ethy12-futahptoplonate. 4-Ethyl9ualacol; 4-ethyl-2-methoxyphenoL Ethyl heptanoate. 2-Ethyl-2-heptenal; 2-ethyl.3-butylacraleln. Ethylhexanoate. Ethyl isobutyrate. . Ethylfeovalerate. Ethyl lactate. - Ethyllautate. Ethyllevulbmfc Ethyl maltot 2-ethYl-3-hydroxy-4H-pyren-4- one. Ethyl2-methylbutynte. - Ethylmyttatate. Ethyl nitrite. Ethyl nonaaoate. Ethyl 2-aonynoate: ethyl octyne catbonate. Ethyl octanoate. Ethyl oleate. Ethyl phenylacetate. Ethyl4-phenylbutyrate. Ethyl3-phenylglycldate. Ethyl 3-phenylpropionate; ethyl liydroein- namate. Ethylpropianate. Ethylpyruvate. . Ethyl salicylate. Ethylsorbate;.ethyl2,4-hexadlenoate. Ethyl tiglate:, ethyl truns-2-methyld-buten- oate. Ethyl..undec3noate.. Ethyll9-undeeenoate. I Ethylvelente. Eucalpytol; 1,H-epoxy-p-menthane; cineole. Eugenyl acetate. Eugenyl benzoate. Eugenyl formate. Eugenyl methyl ether, 4-allylveratrole; methyl eugenol:. Farnesol: 3,7,11-trhaethyl-2,6,19dodeca- trlen-1el. d-Fenchone; d-1,3,3-trimethyl-2-norbonsan- one. Food Drug Cosmetic Law Reports FenchYl alcohoL' 1.3.3-trlmethyl-2-norbur- nanol- Formlcadd - (2-Furyl)-2-propanonC luryl acetooe. . 1-Furyl-2-propanone; turyl acetone. Fusel oiL refined (mixed amyl alcohols). . Geranyl aEetOacetate: trana-3.7-dlmethyl-2, 6-octadien-l-yl acetogcetate.. Getanyl acetone; 6,19dlmethyl-b.9•uadeca- - dlen-2-one. Gerrnyl benxoate. Geranyl butyrate. Geranyl formate. Geranylhexanoate Gennyl Isobutyrate. Geranyllaovaterate... Geranyl phenylacetate. Geranyl proplonate. Glucose pentaacetate. . . lycerylmonooleate.. *QualacaC p-methmqphenoL Gvalacyl acetate: p-methozyphenyl acetate. Gualacyl phenylacetate. Gualene; 1,4-dtmethyl-7-leopropenyl-a9,10- octahydtmazulene. Gualol acetste; 1,4-dimethylR[a-hYdroxy- laopropyl}p9.19actabydroasulene aee- , tate y-Heptalactone; 4-hydroxyheptanole aeld, y- lactone. Heptanat; enanthaldehyde, Heptanal dimethyl acefsL Heptanal 1.2-glyceryl aeetal. 2.3-Heptanedione; acetyl valeryL 3-Heptauol. 2-Heptanone; methyl amyl ketone. 3-Heptanone: ethyl butyl ketona 4-H.pt•rwma:dipropylketone; cis-4-Heptenal: cia-,I-hepten-l-eL Heptylecetate. Heptyl alcahol; enanthle alcohoL Heptyl butyrate. Heptyl cinnamate. Heptyl formate. Heptyligobutyrate. Heptyl octSToate. 1-Hexadecanol: cetyl aleohol- m-e-Hexaderenlactone: 16-hydroxy-6-hexa- decenoic acld;: r-laetone: ambrettolide. y-Hexalactone; 4-hydroxyhexanoic.ac(d;C y- lactane;.tonkalide.. Hexanal; caproic aldehyde- 2.3-Hexanedione; acetyl butyryl. H exano/c acid;; cap roic acid. 2-Hexenal. 2-Hexen-I-ol. 3-Hexen-lol; leaf alcohol. 2-Hexen-l-yl aeetate. 3-Hexenyl Csovalerate.. 3-Hexenyl 2-methylbutymte. 3-Hexenyl phenylacetate; ctt-3-hexenyl phenylacetate. Hexyl acetate. 2-Hexyl-4-acetoxytetFahydrofuran Hexyl alcohol. Hexyl butyrate. a-Hexylcmnamaldehyde... Hexyl formate. Hexyl hexanoate. 21 CFR 172.515 456,519.15
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i ( I . AN CA85 (7) : 4 1921 -3b TI Determination of odor components in.tobacco smoke: design of mixture to simulate odor IT 66- -25-P. 79--70--6 76-79-5, tiiolog~ical studies 78-83-1 **-M-9r.-n51-1*:*x93--6n-7 95--1.3'-6 95--48'-7 93-<D1-1'., hiol.ogicali Studi es. ^r8-E6-2, biolog.ical studies 100-48-1 Pi?0--52-7, hiological studi~es iq0-69-61i04--87-() i()41 .-1..q6-2^:-9 107--.11...5' 107--9~.2-6, h.ir,);ogical studies 1i.~0-0.2-1 110-58L7 11n--66-7 110-86-1, biologica1 s,tudies 110--72-9' 123--35-3 142-83-6 30i-s--57-?`4.31-0'-8' 500--22--1.. 516-74-T 620-02-0 6'L:i-84--3872-85-5 1003-29--a 1'19^r--62-7 2459--o9--8: 2B47._3<}-5 :2 882'-2."--6 436 i-47-8 W 2-56--65--5 5989--27-5 1,.5707--2T-0 29797-i19--9 (of simulated tobacco smokeodmr formulation) AN CA84r.'.9Y.:56680Y TI Analysis of the monophenolic fraction of tobacco smoke condensates by a combination of chromatagraphic methods IT ***9O:-05-1*** 95-48-•795-65-8 95•-87-4 1Q;.;--67-9 106-44-5 1c78-~,T•9--4 526-85--2 527-54-8527--60-6. il76-26.-.1 697-82-5 2416-°94-6 32:58--.T_•8-8 (of tobacco smoke) AN CAB2(3):14151e ~ n\U TI Essential oil of tobacco. Ptt~nol fraction IT 9V-00-6 #**9n-05-1*.** 91-10-i 93-51-6 95-4£3-7 9.J-65-8 95--87-4 99-96--7, biological studies 105-67-9 106-44-5 108-39--4 1(-:8-68-9 108-95-2, biological studies 121-34--612'-i)7-9 150-19-6 526-75-0 527-60-•6 (of toba~-co) AN CA78(9):55475z TI Acids and phenols in the semivolatile fraction of smoke fromblended, Virginia, Burley, and Orient cigarettes IT 57-10-3, biological studies 65-85-0, biological studies 105-43-1 1n8-95--^<,, biological stUdies 120-8G--9, biological st!;dies503--74-2 1319-77-3 (of tobaccosmoke,, of cigarette blends) AN CA72(3)a9996y ~ TI Constituents of certain tobacco types. III. Further volatile ba phenols of Latakia tobacco leaf IT 90-On-6***.90-05-1*** 91-10:-1 9:3-51-6 95-48~7 95-65-8 ~ 95--87--4 97--5?;-0: 97-54-Y' 1[_~5-67--91n6--44-5 PA8-39-4 7786-61-0 20578-'97-6~ (in tobacco) 4
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AN CA1i)7(5);36PJ4.Is TI A new method to determine the relation between the particle size and chemical composition of tobacco smoke particles L-'IfiLI£]GRAPHIC DATA NOT AVAILABLE AT THIS TIME AN CAIiLg.( 1'9) : 169166f TI Studies on phenolic compounds in Thai flue-cured tobacco DT' t'-*#9G--05--1**w 106-44-5, biological studies 108-46-3, biological st.udies108-95-2~.,. biological studies 120-80'-9, biological studies 15 ='-18--4 T _^•.7--97--9 .^•- 785-87-7 (of Thaiflue-cured tobacco) AN CA99(15);119523g TI Qualitative analysis of the hydroxyl fraction of cigarette smoke IT 50-21-5, biological studies 54-1i--5 56-81-5, biological studies 57-55-6, bi'ological studies 79-14--1, biological studies 80-71-7 80-72-8 37-6a1-1 87-99-0 95-4;8-7, biological studies 95--65--f3 95-71-6 96-26-4 102---76-1 105-67-9 106-44--5, biological studies 107-21-1, biological studies 108--39--4, biological studies iO8-68-9 108-95-2, biological studias 109-00-2 1i<rr 7x-9, hialogical studies 12: Bu 9, hiologicall st-udies 123--07--9123--31'-9,. biologicaS. st!-rdi.es 149--=::2-6452--86-8 488'-1'..7-5 498-(-)7-7504--96-1 :i."-77---~ 620-17-7 1198--69-2 :'177-78-8 2349--70-4 5989-27-5 6338-41-6 7425-74-3 7493'-9ra--5 14L60~--85-1 1767.5--99--9 2f-E:35-01--3 25596-9ir-1 28564--f33-2 26930-2i-a--:y 31257-96-2 67965-48-4 7 3-229-7CF-6. (of tobacco smoke hydhor.yl fraction) AN CA97(15)c1218766: tA/CiLGVT TI Effects of tobacco smoke.compounds on the ciliary activity of the embryo chicken trachea in vitro IT 51/-32-3, biological studies. 51-17-2 54-11-5 56-55-3 57-55-6, biologi~cal studies59-67-6, biological studie s 60-12-8 62-53-3, biological studies 64--18-6, bio:logdcal studie s 64-19-7, biologica7l studies65--85-0, biological studie s 66-25--1. 67--47--0 67-56-1:,, biological studies 67-63-0, biologic al studies 67-64-1, bi..ologi'..cal studies.. 71-.-4?..._2, biological studie s 7t5-:75-..8, biological studies 78--G?'1--9 78--92-07f3-84- 2 78-85-_._: 78:-93-', biological studies78--94-4, biological studie s 79-77-6 Bir-5-h-8 87--.3z~.9 g3_ B'-34--1: B4--66-2 84--74_2 BS_G1--8, biological studies S5-4~4-9 B6-2B-2 86-53-3 86-74-8 07--.°J1--4,, biological atudiesB7-59--2 Es7--621-7 E7-66-1 BB-05-1 f39-81--6 9p-c7u-6 9Q-02--8, biological studius ** k9(?-q5-1.>t_>F* 90-12-0 9ir-15--=' (ciliary activity of tr-achea response to, tobacco smoke in ~ relation to) ~ « F+ CD rA OS 1 ~
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effect_ The mu.sclerelaxant effect was.l4nger for glycidy;. ethers than for glyceryl ethers. Glycidol ethers were less effective than glycerol ethers on strychinine spasms. Clycerol ethers had a=_-tronger influence on the cataleptic effect b!rtt the effect of glycidol ethers was more persistant. The ar.ithorsconcludethat gllyc.erol ether is the actual active form of glycidol ether,. (German). 0000466 N10SH-00013645 The Determination of Toxic Substances and their Metabolites in Biological Media by Gas Chromatography-III. The Presence and Measurement of Guaiacol in the Urine Sedivec, V., and J. Flek: Pracovnii lekarstvi, Vol. 22, No. 5, pages 176-181, B references June 19711 Detailed description of a gas chromatography techniquee for the measurement ofguai.acol, . and other phenol-conta.iningsubstancesin.the uri.ne.. The urine sample is mixed in equal proportionswith 80 percent phosphoric acid and heated for 15 minutes in a water bath. After cooling, the guaiacol which is liberated from its conjugated form, is extracted' with carbon disulphide to which nitrochlornbenzenehas been added as a standard. The extract is anal.yzedchromatograph.ically in polyethylene glycol at 150 degrees centigrade with the aid of a. flame ionization detector. A calibration curve is presented. This technique was used to determine the guaiacol present in the urine of personsexperimentally exposed for 8 hours to ceiling vaiueconcentrations of benzene. Comparison with the normal urinary gu~aiaco~ level (1,024 mg/liter) confirmed that gaaiacyl1 is a metabolite- of- benzene and that its level in the urine can be Xsedas a secondary inidication of benzene poisoni.ng. (CzPchJ 26
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0692f3£37 820rj0337765 Mutagenicity Testing in the Salmonellatyphimurium Assay of Phenolic Compounds and Phenolic Fractions Obtained From Smokehouse Smoke Condensates. Pool, Lr.L_.; L_im,.. P.Z. Inst. Toxicol. Chemoth,er., German Cancer Res. Cent.,. Im Neuenheimer Feld 28p,, 69rJG Heidelberg, FRGFOOD CHEM. TOX ICOL. ; 20(4), . pp. -'_•B'j'-ti91 19P32. Language: English Summary Langua.ge: English Document Type: Journal article-original research Subfile: 24 Toxicology Abstracts; 07 Genetics Abstracts Smok:ehouse smoke, which is used for flavouring meat products, was investigated for iflsmutagenic activity in the Salmonella typhimurium assay. The authors weree chiefly concerned with the fractions free of polycyclic aromatic hydrocarbons but containing phenol compounds,, which are responsible for the preservative and aromatizing properties of the smoke. The most abundantly occurring phenol compounds (phemol, cresols, 2,4-dimethySphenol, brenzcatechine, syringol., eugen'ol,vanilline andguaiacol) gave negative results when they wereteshed for mutageni.city att five concentrati.ons up~to5ntr0 .mu..g/plate, with and without 5-9 mix, using five strains of S. typhimurium . When smok:ehouse smoke was condensed and'fractionated thee majority of the variousS phenolic fractions.alsogaves negative results when tested at five concentrations using five strains of S. typhimurium . However therewa-s a slight increase in the number of revertants in a few cases. C-i 481497 800E12B.1707The effect of treatment with some phase II substrates on hepatic xenobiotic metabolism and the urinary excretion of metabolites of the D-glucuronic acid pathway in the rat. Lak:e,Ci.G. ; Longland,R.C. .; Harris,R.A. ; Collins,M.A. ; Herod,.I.A'. ;Gangolli,S.D. (ErIBFA., Woodmansterne Road, Carshalton, Surrey, SM54D5., Uh;) Toxi.col. Appl:. Pharmacol. ; SL(T), 371-378 1980 Language: Engli.sh Summary Lang!!age;. English Investigationshave been conducted to study the relationship between the acti~.vitiss of certain enzymes of Phase I and Phase lIhepatic xenobioticc metabolism and the urinary excretion of somemetabol'.ites of the D-g.lucurnnir. . acid.pathway. Male Sprague-Dawley rats were treated for 7 days with daily i.p. injections of either di'phenyla.ceti.c aci,d, guaiacol, 4--hydroaybiphenyl,. 2-naphthol, 1-maphthylacetic acid, paracetamol (acetaminophen), 2--phenylpropionic acid, n-propyl gallate, or phenylacetic acid. Nbne of the compounds administer-edi had the ef'fect on aa number of parameters of hepatic Phase I xenobiotic metabolism, namely mixed function, oxidase enzyme activities, cytochrome P-450 and themir-rosomal content of protein. 4-Ibwever,, some of the compounds diid stimulate hepatic micro.somal UDF-glucuromyltrans'Ferase activity. While none of thee compounds st.i.mulated the urinary excretion of either D--glucaric acid, L-gulonic acid, or •.ylitol, all of the compounds except phenyLacetic acid increased the urinary excretion of total (-freea.ndconjugated) D-glucuronic acid. 'Theresults indicate that when hepatic xenobiotic conjugative activity is increased without the stimulation of mixed functi~on oxidases and'cytochrome P--4:5G, thenr-inary excretion of certain metabolites of theD-glucuronic acid pathway, other than D-glucuronic acid,, remains unchanged. 8'7119155 i
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Mechanism of pigment-cell toxicity produced by!hydrouyanisole. Riley PA J Pathol (.F-.ngland) Jun 1970, 101 (2) p163--`'+', ISSN iii>2,'?--Y417' Journal Cod'e: •:JLCiLangu.ages: ENGLISH Journal Announcement: 7101 Subfile: INDEX MEDICUS Tags: Animal Descriptors: *Chromatophores--Metiaboli:sm--ME; *Gwaiacol---Toxicity--TO; *Melanins; Ascorbic Acid--f-'harmacodynamics--PDpAutor•adi~.agraphy; Catechols --Pharmacodynamics--PD;. Cell Membrane--Drug~ Effect.s---DE; Cytoplasm--Drug Ef-Fectls--:DE: Dipept-.ides--Pharmacodynamica--Pfi; Diphenol Oxidases --Metabolism--ME;. Dopa; Electron Spin. Resonance; Free Radicals• Glutathione ---Pharmacodynamics---F•D:; GLiinea.. Pi~gs;. Melanocytes--Enzymology--EN; Melanocytes---Metaboli.sm--ME; Phenols--Pharmac.odynamics--PD; Skin --Drug Effects---DE; Thiocarbamates---Pharmacodynamicsr-PP; Ti.s=sue. Culture; Tritium; Ubiqui.none--Pharmacodynamics--PD G 01438962 70283962 [Lethal allergic and toxic reaction following common drug administration] Letalna.alergickotoxick:a.reakcia po 4e<ny=h liekoch. Zuffa MI Vni.tr Lek (Czec.hoslovak:ia) Sep 1970,. 16 (9) p9n9---11'., SSSN 0042-773X Journal Code: XFY Languages: SLOVAK Journal Announcement:7Q12 Subfile: INDEX MEDICUS Tags: Female; Human Descriptors: *Drug.Hypersensitivity; *Phenylbutazone--Adverse Effects--AE ;, Death~.,. Sudden; Diabetes Melli.tus--Drug Ther.xpy--DT;English Abstract; Guai acol --Adverse Effe_ts--AE; Guaiacol Glyceryl Ether----Adverse Effects--AE ;, k:etones--Adverse Effects--AE; Middle Age; Shock, Septic---Etiology--ET Descriptors: *Carcinoma., Squamous Celi---Chemically Induced---CI; *ConnectiveTissue---Drug Ef'.fects---DE.s.. *Guaiacel---To;:i=ity---TO~; *Neoplasms, Experimental--Chemically Induced---CI;:. +rPhenols--Toxicit..y--TO; *Skin--Drug Ef+ects--DE; Connective Tissue---F'athology--PA; Hamsters; Neoplasms, Experimental-°Pathology--PA;. Sk:in---Pathology---PA - 0101223i. 69157231 The effects of 4-hydroxyanisole on hamster cheek pouch. Woods DA; Smith CJ Exp Mol Pe+thol (United States) Apr 1969, SCi. (2) p107-14, 0014-4800 Journal Code: E05 Languages: ENGLISH Journal Announcement: 6907 Subfile:. INDEX MEDICUS Tags: Animal ~ O 5
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u59[ i3651 86204651 CCytotoxicifly of root canal filling materials. 2. Preparation of calcium hydroxide (Cal~vital), paraformaldehyde (Triozinc paste), epoxy resin (AH 26), guaiacol formaldehyde resin (FR), and other root canal filling materials (N2, Nogenol)] Nakamura H , Shigaku Feb 1986., 73 (6) p1619-56, ISSN i>029-8484 Journal Code: OEX Languages: JAp[!= Subfile: Dental. Descriptors: *Root Canal Fillinq Materials--Tox.icity--TO; Bismuth ---Toxicity--TO; Calcium Hydroxide---Toxicity--TO; Cells, Cultured; Drug Combinations--Toxicity--TO; Eugenol--Toxicity--TO; Fatty Acids--Toxicity --TO; Formaldehyde--Toxicity--TC; GuaiacoS--Taxicity--TO; L Cells--Cytology --CY; L Cells--Drug Effects--DE; Methenamine---Toxicity--TO; Resins ---Toxicity---TO;: Salicylic Acids--Toxicity--TO; Silver--To>:i~city--TQ; Thymol ---Toxicity--TO; Titanium--Toxicity--TO; Zinc Qxide--Toxi~city--TO CAS Registry No.: 1QU-97-0~ (Methenamine); 1305-62-0 (Calcium~ Hydroxide); 1314-i.3•-2' (Zinc Oxide); 50-00-0 (Formaldehyde); 55599-25-2 (AH 26);: 7440-22-4 (Silver); 7444-3^c-6 (Ti~tarn,ium)..; 7440-69-9 (Bismuth.); 7859D-82-6. (Nogenol); Bp6B-83-5 (N-2); B9-83-B (Thymol); 90•-05-1 (Guaiacol); 97-53-9(Esxgenol) F 04575330 82118330 Acute toxicity of thioguaiacol and of versalide in rodents. Butterworth KR;Mason FL Food. Cosmet Toxicol Dec 1981, 19 (6) p753-5,. ISSN 0015-6264 JcaurnaI Code: F3W Languages: ENGLISH Tags: Animal;, Female; Male Descriptors: 4-Flavoring Agents---T oxicity--TO; *Guaiacol' --Analo s arrd g Derivatives--AA; *Naphthalenes--Toxicity--TO; KPerfume---Toxicity--TQ; *Tetrahydronaphthalenas---Toxicity--TO; Guaiacol--Toxicity---TO;, Rats; Rats, InlDred Strains 8ex Factors . ; CA8 Registry No.: 7217-59-6 (thioguaiacol); 88-29-9 (versalide); 90-05-1 (Guai aco.l ). 04206301 B1034341 A toxicological and pharmacological study of ibuprofen guaiacol ester (AF 2259) in the rat. Cioli V;. Futzol'u.S; Rossi V; Corradino C Toxicol Appl. Pharmacol Jun 30 198(1, 54 (2) p332-9, ISSN r]041-0Cr8X Journai~ Code; VWO. ENGLISH L anguages: - Tags: Animal; Comparative Study;. Female; Male Descriptors:*Guaiacol--Analogs and Derivatives---AA; #.Ibuprofen--Analogs and Derivatives--AA; Analgesics, Ant.i-Inflammatory; Behavior, Animal---Drug Effects--DE; Edema--Drug Therapy--DT; Fever---Drug - Therapy--DT; Gastrointestinal Sy.,tem -Drug.. Effects--D£; Guaiacol--Tox.icity--TD, Guaiacol ^, --Therapeutic Use TU; Ibmprofen--Toxicity---TO;. Ibupr-ofen-Ther-apeutif: Use j? --TU; Lethal Dose 50: Mice; Rats - s CAS Registry No.: 15687-27-1 (Ibuprofen); 66332:-77--2 (AF 2259), (£ 90-q5-1 (Guaiacol) I,.13 2 " 8"l11915 ! . _. _ ............., i
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File 166rSmoking and Health - 70-87/Aug 0035979 870019 In Vitro Studies of Biological Effects of Cigarette Smoke Condensate. II. Induction of Sister-Chromatid Exchanges in Human Lymphocytes by Weakly Acidic, Semivolatile Constituents. Jansson, T.; Curvall, M.; Hedin, A., et al. Mutation Research. 169(3):129-139. 19B6 LanguagesEnglish. The weakly acidic semivolatile fraction of cigarette smoke condensate and its individual weakly acid'components were.tested~for induction of sisterchromatidi exchange (SCE) in human lymphocytes. The weakly acidi:c fractionwas separatedd by preparative gel chromatography into. 1.1 subfractions (F1-F1i). All fractions except F11 induced SCE in a dose--dependent way (pt0.05-R.0(?1). The effects of the subfractions were normalizedusimg the total weakly acidic fraction as the reference. Fraction F4, containing mainly al6:y1-2-hydroxy-2-cyclopenten-1--ones, wass the most active sub+raction and the only one showing cl'ear' enrichment of the activity compared with the total fraction. Of 23 individual components tested, mostly alkylphenols, 7 induced SCE. Three were potent inducersa catechol (p;n.01), 2-(1-propenyl)phenol (p;0.0q1), and vanillin (pCO.c,i1). The other fouu- had moderate effects: cyclotene (p<.0.05),. guaiacoli (peG.05), isoeugenol (p:O,C75), and maltol (p<,U.PjS). Many are important flavorants intobacco, tobacco smo4.e,..food,, candies, beverages, and perfumes. 0032697 850640 Quantitative Analysis of Cigarette Smoke Condensate Monophenals by Reverse-Phase High-Performance Liquid Chromatography. Jeanty, G..; Masse, J.; Bercot, P. Beitraege zur Tabakforschung International. 12(5)z245-25Ci. November 1984 Language: English A rapid quantitative analytical method for determination of monophenols present in tobacco smoke condensate using reverse-phase high-performance liquid'chromatogra.phy (HPLC) i's proposed and demonstrated with analysis of five different fil'ter-ti.ppedci~garette brands from the French market. Levels of phenol, guaiacol, methylphenol, and dimethylphenoS were determined in comparison with anintermal' standard, 0-chlorophenol. The method' was also used' to compare the phenolic retention efficiency of different cigarette filter tips. It is concluded that this me+hod, which is simple, rapid,, and selective,, shows good reproducibility. (Auth. Abs. Mod.) OD ~ N ~ ~}14 N ~ 11 i
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( 10 0:Sn1'('4 84001." The Distribution of Cigarette Smoke Components Between Mainstream and Sidestream Smoke. Sakr.ima., H.; kLtsama,. M.p: Munakata., S.; Ohsumi,T..; Sugawara, S. - Beitraege zur Tatiakforschung International. 12(2)0 6'i-71. June 1983 Language: English. , Analysis of yields in sidestream~and mainstream smoke and sidestream to mainstream distribution ratios (SS/MS) of acidic components, showed (.1) major phenol.ic components were phenol, catechol, and hydroquinone; and (2) major acids were formic, acetic, lactic, glycolic, and succinic. The four cigarette types examined were made from Bright, Burley, Turkish, and domestic (c.v. Matsukawa) tobacco. Compounds tha.t gave SS/MS ratios >1 for all types of cigarettes tested included: phenol, cresols, xylenols, guaiacol, formic acid, and acetic acid. Formic and aceticc acids were analyzed by gas chromatography following n-butyl esterif's.cat.ion; acids other than formic and' acetic acids and phenolswere analyzed after trimethylsi.lylatione (Auth. Abs. Mod.) 0027673 821492 What's Been Added'to Your Cigarette? Foss, W. S. keader's Digest. 1<1(727): 111-114. July 1982 .. Language: English The advent of the low-yield cigarette with its altered flavor hasbrou.ght about aa new concern that manufacturers might be putting in more harm with additives than they are removing. Representatives of the. Office of the Assistant Secretary for Health havebeem negotiating with thetobacco.industry for the release of lists of additivesused in U.S. cigarettes.: Although many of these substances are derived from compounds.consideredsafe to humans,, alterations may occur upon burning. An,30al Gk•in +ests have shown tumorioenic.activity for the following substances: coumarin, caramel, invert sugar, eugenoli, guaiacol, angelica root extract, cocoa, licorice, glycerol, glycol, dodecan-5-oli.de,, and nonan-4-olide. Acrolein, formed by the pyrolysis ofgdycerol,suppiresses theacti~.on of microscopic cilia that force irritants from the lungs. As a.resuQ~,t, the riskk of chronic bronr_hit.is and emphysema is increased and the lungs are exposed to toxins and carcinogens. It is concluded that.t if additives pose no threat, that fact should be known; if there is a threat, the surbsi:ances should not beused.a 12
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I Scientific Literature Review of Phehols in Flavor Usage. volume 1. Introduction and Summary, Tables of Data, Bibliography Flavor and Extract Manufacturers' Association of the United States Washington, DC Prepared for Food and Drug Administration, Rockville, MD Ci Sep 85 t I'1 U.S. Deoxdnent of CanNTWce t Netionel TesMnial Marmetbn Serrke PB86-155868 I
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File 6:NTIS - 64-97/ISS24 1146263 PB85-225530 Effects of Activated Carbon on the Reactions of Free Chlorine with Phenol s (Journal article) Voudri~as, E. A. ; Larson, R. A. ; Snoeyi..nk, V.. L. Illinois Univ. at Urbana-Champaign. Corp. Source Codes; 0:754597006r Sponsor; Environmental Protection Agency, Cincinnati, OH. Water' Engineering. Research Lab. Report No.: EPA/600/J-95/071 c19B5 10p Pub.. in Environmental Science and Technology v19, n5 p441-449 May 95. Languages: Engli~sh Document. Type:. Journal article. NTIS Prices: Not available NTIS Journal Announcement.: GRA10522 Country of Publi:r_ation: United States Contract No.: EPA-R-B05293 The use of prechlorination in drinking water treatmentt resulits in contact of free chlorine with activated carbon which..has been added to remove organic compounds from water. The chlorine then reacts with the carbon and'd adsorbed compounds. Free chlorine reacts readily with a group of phenolic compounds(phenoL, guaiacol, ca*echol.,, 2,6-dimethoxyphenol, and: p-chlorophenol) in dilute aqueous solutions (.00001 M) to produce mono-, di-, ortrichloro derivatives, but when it reacts with phenols adsorbed onn granul.ar activated carbon (GAC), many additional productsa.re -Formed. GAC exposed tochlori.ne becomes capable of promoting reac.tionssuch ass hydroxylation of the aromatic ring, oxidation to~ quinones, chlorine substitution, car'bo::ylati,on, and or,.idativecoupling (dimer formation). The formationn of chloro-hydroxybiphenyl's (hydro,;ylatedPCE+s) (in vivo metabolites ofPCBs) is particularly important becauseoft.heir potential tox.icity.,~'iuch.compounds are the main reaction products from chlo:'ophenols, but they are also formed in smaller amounts from nonchlori.natedphenols (phenol and.guaiacol). (Copyright (c)1995, American Chemical Society.) 1015430 AD A133 053/9 Role of Oxygen Radicals in Biology and Medicine Packer, Lester : Pryor, William A. Gordon Research Conferences, Inc., Kingston,, RS. Corp. Source Codes: 011704000; T97..7,06 1983 .i24p Languages: Engli,sh NTIS Prices: PC A02/MF A01 Journal Anmouncement: BRAIB4tr1 Country of Publication: United States. Contract No.: NC>r7014rar-B--U1Ci7 No abstract available. m ~ N A N ~.. i Ma N 9 0 ©
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AN C1-196d:?5, TI Fractionation and capill~ary gas chromatographic-mass spectrometric characterization oo-F the neutral components in marijuana and tobaccosmok:e.condensates d I.T (detection of, in marijuana.and tobacco smoke condnrrsates by gass r-hromatog..-massspectroscopy) 4~ * :* 9C) --1*:*M- i.::--0. 91-57-6 4i5-4,8-7, analysis.. 106-44-5, analysis 1U8--Z.9--4., analysis 108-95-2, analysis 112-40--'± 1.20--72-.9, ana.lysi=-, . 1.2(-)-80r--9, analysis 13Ct--86-=: 5p4-96-1 59:3--49--7 6.^:-9-62-9 63c3-02--4 6.38-68-6 112o-21-4 18794-84-8 26447-...28--9 29828--'.='.8--2 60976--7.3._n AN CA96(.21) :178173d TI Analyses of mono- and di!hydroxybenzenes in tohaccoo smok:eand pyrolyzates by glass capillary gas chromatography I'1- 541-11-5 56-81-;7., bi.ol::,gical studies 88-18r6 88--69--7 89-72-5 #**9u--05--1+.** 95....48-7, biological studies 95--57--G 95-65-8 AN TI i-T 95-71-6 95-87-4 98-..54-4 99--89-8 W5--67-9 W6--44-5, biological studies 10c;-39-4, biological studies1i,8--46-3, biological-studies1U8-68--9 1Ct8--'95.-2,, biological studies 12ii-Bp-9, biol.ogicalstu.d'ies 123-07-9 123-31-9, biological stt.idies 4.:i2--86-8' 468-17-5 526-75--0 5.27-54-3 527-.E6p--6 585-.34.....^ 6117--45-1 644-.5-9' 645--56-7 697-82--5 2416..94._6 (in tobacco smoke) CA97(1 ) : 4089d Application of gel chromatography to characterize more completely the phenols of cigarette smoke 526-75--n527-54-8 527-6Ci-6. 576-26--1 585--34..,-2 618-45-1 620--17--7 620-18-8. 621-27-2 645--56--7 695-f34-1 697-82-5 2197--57-1 2416-94--62628--17-T 591i2-86--75932-68-3 7469--77-4. 7786-61-'0 2q49e>-.42-0 '. 26761-75-1 29760-89-2 ';f-1^30-52-5 4ii529-54-2 50851-'69.... 9 5n9f-s4--45-7 56631-57-3' S9534---=,5-9 `.i9534-'6-CI' S9632-9Q-- 5 62744-'64-3 7 3 85Q-i)1-8 73850--02_9 d~ 7=:i85CO--q::-<a 73850--04- 1 7'3B5f l•-( i5-2 7:2:850-06-3. 'r' :535i i-07-4 732,.cJ/l'-(78-..=,i 7385C1-1.19- 6 73850--10--9 738.50--1 1 _o 7._8e:i.- 1-•-1 I 7385O-1.3-273850-14- 3 7385Cr-1 °J-4 7385C?--16--.°.,.. 73850--17--6 7~3850--18-773850-E.9- 8 73'85@~-20-1' S.-27-5 88- -18--688--69-.7 89-72-5 90-00-6 9n--15_z. 41.._16-7 95-48-7„ hiological studies 95'-6g--8 95--87-4 97'-53-.<l 98--54-4 99-71-8 99-89-8 i'.n5-67--9 10<-.-44--5, hining:icals:ia_,dies 1p8--39-4,, biological stLidies. 1UL-68-9 i.t]r.~~--95-2 , biological studies 121-33-5. 123-6f7-9' 49fr-78-6 (of tobacco smoke,, gel f'iltratiomn of) 2
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0021220 NIOSH-°00061115 On the.ExpectorantAction of Creosote and the Guaiacol.s Stevens, M. E., A. Y=.F(onan, T. S. Sourkes,., and E. Mi, Boyd Canadian Medical Association Journal, Vol. 40, pages 124-127, 14 references Febuary 1943, ,. Creosote, guaiacol, guaiacol carbonate and guaiacol glycerol ether were found to incr-ease the output ofrespiratory-tract fluict when given by stomach tr.rbe to cats andd rabbits.irr doses up.to 5 g or cc per kilo body weight. Wfhilethe increased output occurred with~.reasonable consistency, the average output was seldom increased more than 50% over 44 hours orso. Evidence was obtained that these drugs act (a). by a reflex from the stomach and(b)afterabsorption into the blood stream. Guaiacol glycerol ether given in daily doses of 1.2 g to medical students significantly reduced the number of coughs per day during~colds from the number occurring in other medical studies receivi~ng a placebo in a "blind test". The historical evolution ofc.reosote and, guaiacol as.drugs is alsod~iscussed. . (3 . (I 0002622 NIOSH-00003564 Toxicology of Ouaiacol Vapors and Resins f Ostrovskii, M. M. Hygiene and Sanitation, Vol. 29, pages 105-100, 4 references March 1,.964 Toxicological study of the short-term inhalation of guaiacol vapors by mice to determine the ha:ard to the skin caused by the gums or resins of guaiacol. The study involves a.two-hourexposurea of guaiacal vapors in a concentration range from 1.98 to 17.31 milligrams per liter (mg/1).. Results indicate that permissible concentratiom of guaiacol vapors is 0.02 mg/1g maximum tolerance dose:i~s 2.03 mg/1.. To study the local effect of guaiacol resins an the skin,, ointment containing 0.5, 5 and 10 percent resin is applied to the abd.amem.of rabbits, and left on thee skin -for 4 hours over a two-week test period. The resin is found to possess neither an irritating nor a: blastomogenic effect. 0074154 NIOSH-00102172 Studies on the Pharmacological Action of Aromatic Slycidol Ethers Sollner, K., and K. Irrgang Ar'zneimittel•-Forschung,. Vol. 15, No. 11, pages 1'1.355-1'1.357, 17.references 1965 The pharmacblogi.cal effects of the.gIycerolethers of phenol(5:'8432), guaiacol (9T1,41) andd o-cresol (59472) were compared with those of the corresponding glycidol ethers toxicity; spasmolytic action,, muscle relaxant effect, cataleptic effects,, and effect on strychnine induced spasms were tested', in laboratory animals, andd splitting of o-cresyl glycidol' ether was tested in viflra. The lethal dose of glyceryl ethers wasgemer-all.y lower than t.hatt gl..ycidyl ethers,, except for phenol derivatives. The conversi~on of diglycolss to epoxideswasaccompani.ed by reduced toxicity: Spasmolytic effectt on spasms induced by barium chloride, acetyIcholi.ne and histamine were stronger with glycidol ethers than with the analogues glycerol ethers. Epoxides were moree effective than the corresponding~.diglycols in their musculotropic/spasmolytic effect and their atropine likee and antihistaminic. 25, 8'7119165 ., _.. _ _,. _.., _ _.....-. ~:
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:nedl.r"tl.ary centers and thern peri'.pher-alP.y again to the respiratory tract. L•'xperimeritsx in, . cats given i~ntramusc!ul.ar guaiacol-gl, ' !cerol'-ether suggested~ that the guaiacols may also act after absorption i:ntm.the blood'stream. In medical stu.demts treated for colds with either placeboor guaiacol--glycerol-et:-her tablets (1.2g per day)., t.ha tahlrets were {ound toreducet-he number of coughs per day. The authors conrrl'.udethat although the.e t.es•t drugs i~.ncreaaed output ofrespi ratory tract flui.d,, although.h theincr-easewas not asmai-ked asthat which has been found for other e;:pectorants. 0069272 tot 4L NIOSH-nCrQ97?b1 Mutagenicity of Henzidine and Related'Compounds Employed of Hemoglobin Ferretti,, J. J., 4J.. Lu, and M-E+. Liu in the Detection American JournalC.li~nical Pathology, Vol. 67, no. 6, pages 526--527, 13 references 1977 CODEN!: AJCF'AI. TheSalmonell.a mammalian microsome test system was used to assess the mutagenicity of benzidine (92875) and related compounds. Three of the compounds tested, benzidine, o-di~anisidine (119904), and o-tolidine (119937), resulted in a higher number of revertants per plate in the presence oc microsomal activating enzymes. Four other compounds, o-anisid'ine (9nCrRU.), dipehnylamine (.122394), guaicol (90051), and o--toluidine (9sW34) were not mutagenic even in the presence of the microsomal activating enzymes. The authors conclude th,:,tt since the c.ompo!rnds that were mutagenic in this study have been extensively used by technologists and r-esearclherss in clinical chemistry laboratories, a potenti:.al hazar-d'+..o these workers exists, and reevaluation of the use of hemoglobin chromagens is suggested. n022828 N I OSH>-{J0062 iJ 5 Toxic Solvents; A,Review Erowning, E. British Journal of Industr-ial. Medicine, V'ol pages 2:'~,-,9, 116 6 references 1959 Aliteratua-e review of toxic solvents is presented. General developments in experimental physiology, metabolism and detoxification, and methods for measuring: atmospheric concentrations ared'zscussed briefly. The main part of the paper-discusses thee experimental and clinical aspects.of poisoning by individual solvents.. The physiological effectsof the:fol.lowing solvents are disc:ussed~r tetrahydroAurfuryl alcohol; butyll alcohol; butylamine; 1-chloronitraprnpaneq cresol; di'ethy'_ sulfaxce; dimethyl sulfate; d!ioxane; ethylenediarnine; toluene: trichloroethylene; per-chloroethylene; carbon tetrachloride; trichloromethane; isophorone; ethylene gl!ycol;propylene glycol; methyl cellosolve; butyl cellosolve; l.etrahydrofuran; cycl..ohexyl'..amine, carbon dxsul'.fi.de. m N to N 24 wp. R
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File.161:Occupational Safety & Health (NIOSH) -73-87/Aug 0106519 NIOSH-00130362 Influence of Germ-Free Status on the Excretion of Simple Phenols of - Possible Significance in Tumor Promotion Bakke, 0. M., and T.Midtvedt . Experientia, Vol.'26, No. 5, page 519, 11 references May 15,1970 CODEN: EXPEAM The excretion of phenol (108952) and simple phenol compounds from natural plant diets was studied in rats. Four germ free and four conventional CDF-rats were fed commercial pellets beginning 10 days before the collection of urine. Urine of germ free rats was collected from isolators every- 24. hours, stored, -thawed, filtered, and di uted; while urine from. conventional rats was collected at 4e hour intervals and stored in solid carbon-dioxide. Samples were hydrolyzed with beta-glucuronidase, extracted, and analyzed by gas chromatography. Less than 0.005 milligrams per 24 hour (mg/24 hour) period bff phenol was found in _the urine of the germ free rats;' and no , other simple phenols were found. The urine of conventional rats yielded means of 0.09mg/24 hour phenol,'0.2Bmg/24 hour p-cresol (106445F, 0.38mg/24 hour 4-ethylphenol (123079), and smaller amounts of catechol (120809), 4-methylcatechol (452868), resorcinol(10846?),. and guaiacol (90051): The authors conclude that, with the exception of phenol which arises from tissues, microflora are necessary to produce the simple phenol compounds. The promotion of tumors by simple phenols should be investigated in the context of the normal production of simple phenols within the body. Differences in susceptibility to hydrocarbons such as dimethylbenzanthracene (57976) might be related to tumor promotion by simple phenols produced through the action of intestinal microflora on precursors present in normal diet. . _ 0104968 NIOSH-00128858 - On the Expectorant Action of Cresote and the Guaiacols Stevens, M. E., A. K.. Ronan, T. S.Sourkes, and E. M. Boyd Canadian Medical Association Journal, Vol. 48, pages 124-127, 14 references Febuary 1943 The expectorant action of creosote (8021394) and several guai~acols (90051) were investigated in cats, rabbits, and medical students. Anesthetized rabbits were administered 5.0 milliliters (ml) per kilogram (kg) gua,~ iacol (90051) or S grams (g) guaiacol-carbonate (553173). Anesthetized cats were administered 0.1 or S.Om1/kg creosote or O.lg/kg guaiacol-glycerol-ether (93141). The output of respiratory tracflfluid from a tracheal cannula was measured under condi~tions in which the inhaled air was warmed to body temperature and saturated with water vapor. At L.Oml/kg, guaiacol increased output of respiratory-fluid -from 20 to 40 percent in B of 9 rabbits; the 5.Om1/kg.dose resulted in slight and occasionally marked increases in respiratory fluid output. Similarly, guaiacol-carbonate increased output.t of respiratory tract fluid, but increases were not significant. Creosote increased output ofrespiratory tract fluid in.cats, with greater increases at the higher dose. Guaiacol-glycerol-ether at 1.Og/kg consistently increased output of respiratory tract fluid. When administered'to rabbits in which the gastric branches of the vagus nerve had been severed, S.Crg/kg guaiacol--carbonate had no effect on respiratory tract fluid output, suggestingt.hat the drug acts_as an expectorant by virtue of a reflexfrom the stomach via the afferent gastric nerves to the-
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001'5393 006888 Ein Beitrag zur Beurtei'lung von 'Oatbakzusatzrstoffen Pyrolyseprodukte. 2. Stark:eund;Ihre Derivate. auf Grund Ihrer ~ 3 [Evaluation of Tobacco Additives on Products. 2. Starch and its Derivatives.] 1::roller, E. the Basis of Their Pyrolysis Liundesgesundheitsblatt. 9(8): 106-107. Language: German April 15, 1966 (I Carbonization tests were carried out on starch and.its derivatives, suchh as.carboxymethyl-,oxy--, and'.methyl starch, used ass adhesives or thickening agents for tobacco products. Identification was based upon their Rp and Rn values, fluorescence colors and maxima, and their characteristic reactions; 3,4-benzopyrene was d'etermined' fluorometrically. Fluoranthene and -',4-benzopyrene were always present. Anthracene, phenanthrene, and their derivatives were alsofound frequently, while only isolated instances of picene, pentacene, fluorene, and'terphenyl were observed in carbonizationn pr-oducts. Of the quinones, anthraquinone was always present, phenanthrenequi~none occurred often, and toluquinone only in isolated cases. Cresols, phenol, and pyr-ogalloll were most frequently present in the acid portion of pyrolysis products, while gallic acid, guaiacol, o-ethylphenol , and pyrocatecPool were seldom encountered. While starch.h yielded 7 mcg/14C?q ofL,,4-benzopyrene, its derivatives yielded considerably larger quantities, apparently as a result of a loosening of the molecular structure in substitution. 0615295 006782 Ergebnisse von Schwelversuchen an Farbstoffenzu.r Far-bmattierung von Tabakwaren. 2. Mitteilung (Kreuzbeerene>:trakt). [Results of Carbonization Tests on Dyes for the Color Matting of Tobacco Products, 2. Communication--Buckthornberry Extract.] R'roller, E. E<undesgesundhei.tsblatt. 6(24): -717-318. 1963 " Language:CnermanAn investigati~on was made of the carcinogenic content of the pyrolysis products of buckthornberry (. Rhamnus cathartica )'.) lake which was being considered as a.coloring agent for cigar wrapper leaf. Two color components of the lake wereknown,e rhamnetin and rhamnazideni the mineral content of the commercial lake was about 50 percent. Fifty grams of the test substance in 3 g portions were heated.in a porcelain boat pliaced in a quartz tube wi.tlhh a combustion zone temperature of 7Ur:r^'C. The.pyrolysis products were separated by paper chromatography and fluorescence-spectrophotometrically ildentilfied. In additi.onto water, acetic acid, and paraffin hydrocarbons, the phenols, pyrocatechol, guaiacol, m--cresol, alpha- and beta-naphthol, and pyrogal.liol, as well as the aromatics, pyrene, fluoranthene, and the carcinogen a,4-benzopyrene,, the latter in quantities of 2 gammas per 100 g of lake were found. In view of the high concentration of 3,4-benzopyrene in tobacco smoke, the test material'~ was considered unsuitable on the grounds of health. - r. 13
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D..L.1.Ornrxsand~~C.Lcrrz1A (iuaiacull applical 6illstrcugtlt lo inlac'I or abrndcd raBbil sklln for 24hr under ncclusion, was moderately.lo scvcrely, irritating (Moreno. 1978). Tested ul 2" in pelrol:num. it produced no irritation after a 48-8rclosed-palch test on human subjects (Kligm:m, 1978), Srrnirircilinn. A maximization test (Kligntan; 1966: Kligman & Epstein. 1975) was.carried out on 25 volunteers. The material (RIFM no. 77-199) was tesle& al a concentration of 2",' in petrolatum and produced no sensitization reactions (Kligman. 197R). MurnprJdcyrs,5/1'rGuaiilcul. a chromagen used for Ihe detection of hacmoglobin.derivatives, was not Inutagenic in.lhe SrrlinoneflnJmammalian mlbrosomc test (Ferrelti, Lu & Litt. 1977G Anrluariifir%rJ/2~i'i.e: Guaiolcul was eRective in reducing lhe wei~it of sarcoma 180 tumours in mice (Murakami & Yamafuji. 1969). . Curciurylrnfc r(fi•rta. Wynder &Hoffmann (1963) questioned the role of guaiacol, one of the many componenls in tobacco smoke, as a ancarcinogcn.. Me81bn'7fsmkWhen guaiacol was administered iv to rats in a dose of 50mg/kg, catechol was identified asihe principal urinary metabolite (Wong & Sourkes, 1966). The administration ot disul- firam Priorto guaiacol blocked this formation of catechol. Ttaces of guaiacol, among other metab- olites, were identified in the urine following oral administration of vanillin to rats (Strand & Scheline, 1975).. - ~- Phaffials"I ye, Guaiaeol exhibited local anaesthetic effects when tested on the rabbit cornea, onn the lung stretch receptors of guibea-pigs, and in white mice (Siemoneit et nf. 1966). The central depressant . and general anaesthetic eBectsof guaiacol were manifested at toxic doses. RaGserum prolactin levels were depressed in animals receiving ip Img guaiacol in 0-05N-hydro- chloric acid (Smythe & Lazarus;. 1973). Administrationn of guaiaeol to rabbits and rooslerson an atherogenic diet was associated with inhibition of alimentaryy atherosclerosis (Tarasova, 1968), Guaia- col administration did not aRecl pcntobarbital sleeping time in rats (Jorii Bianehelli & Prestini. 1969) or liver regeneration in partiallyhepatectomized rats (Gershbein, 1977): Ciliary motility in the frog ocsophaguswas stimtd:ded byadministralion of guaiacol (Das, Rathor. Sinhat &Sanyal, 1970). The antigenicity of benzyl penicillin eonjug:nes in the rabbit was unaffected by the addition of guaiacol (Portoli:a Perez Urena. Rantos ri n!: 1973). .. . Tfir%iipgWj'VuscSGuaiacol has hecn used internally chiclly as.a .timul:ning expectorant: when given orally i0 increases respiralory-Iract Buid (U.S: 1)icPevnnrart•, 1967)...Its expectorant activities are appar- enf in both hulnans and anim:ds (~1/Brvi bufex- 1976). Latrge doses arc capahle of increasing the output.ofrespiralory-tracl Ilnid in.animals byg actinglhrough a gastric rcOcx and also after absorption (DrilC.r,Plmrmuenlrn)r ur Medirinr: 1965). Incumbination witli z-cy:musryliuc deriv:uives. guaiacol exhibited cicalri7ing elfeclss and .xdative. analgesic and disinfectant clfects on human dental pulp (Kuroda. H.uuyamu. Asai & Sckine, 1976: Morioka. 1976). References . Arctander,. S. (1969(. Pnfume and Ffnmt Chemiru/x.lAmmu Chemirnlsl. Vol. I,: no. 1472: S. Arclander, Monlcclair,.N1. Cioc M. & vomSchilling:.H.11965).-Ch:mges.in muscular Iiisue,prmluced byy inicstion of someknown drugs. dir.ir. app?. P6nrrrmr.7. 179: CIVO TNO (19776.Ifnlnrilr ('imipomrd.. iii 17nx1.' 4th Gd. EdBcd byS. Van Stralcn. Ccnlraal'.Instituur Voor VrndingsondenocklTNO. Zciai The Nelherlands. Council of Europe 1197411 . N:uural Flacoming Suhst:mccs.. Their Sourccs% and AJdcd Artifiiial I'lavnuring Suh.tances:.PartinliAgreemcnt in thcStxi:d and Puhlic Hcnlth Fi<Id..Lisl t. no. 173. p. I6(I. Strashaturg. Das.P. K.. Rathor.R..S.. Sinha, P..S. & Sanyal. A. K. (197(1). ERrxt on cillary movements ofhsome agents ahich aromc.in contact wlth thc respiratory Irnrl. ln.lirm J. 1'hRCiu6 I'Itnrrrnlr. 1-1.'_97: DrilP:r Plirvmnwdrylr in Afedfcfrre(19654 3rd Ed. Edited by J. R. Dipalma..p. 789, McGraw-HiII,.New York. FenrrrnlileHundhrmk rJFluro. brgrerlient.t (1975)..Edited by T. E. Furia &N. Bellanca. 2nd Ed. Vol. II, p. 225. CRCPress, Cleveland, OH. ---jD.Ferreni. 1, 1.,,Lu, W: & Liu:M..B. (1977). Mutagenicity of benzidine and related compounds employed in the detection of hemoglobin.,Am. J. dln. P.uli. 67, 526. FlavorinRExtract Manufacturers' Association p9651. Survey of flavoring ingredient usage levels. No. 2532. Fd . - Ter/mnL, Chnmpmtpr 19.121-par02, 155. Cicrshhein. 1_.I..,II'177k Regerretation of rat liver in dlrprc.renec of essenli:d ndc and their cnmpnncnlx.,Fr1 . . Cnwire. 'hr virvrl 15. . 173. Gosxlin, R. 6.,. I lodgc. 11. ('.-. Sinirh.. R. fl. &. Glcasnn- M. N. (1976). Clmi.vl 'frnirrrlrryJy n! Cimunr.riW PrrrJlCrc Arrnr I'ni.4minp..41h Ed,p. 126. Tlte Williams & Wilkins ('o.- Baltimore, MD. Hake. C. L. &Rowo.V. K.11967[ Ethers. In fndratriid 11p9imr mx! Tn.ciinlrx/r. 2nd lid. Ediled by. E. A. Patty. Vol. II. p. 1681. Irncr.cicncc Pu6lishcm New York. Jori. A.- Biunncelti. A. & Prestini. P. E:.119691. ERecr nf esscnti:d oils on drug neclaholism. 8irrrinvu. 1'hnnrrar. Ig.'_IIRI. Kligimm.A. M. (19fi61.1Lc idcntificatinn of cvwtact :dlergcn.'b,v Irunumass:n': Ill. Thc maximi~ulinn tcsl. A pnn:ednrr for screening and r:nin} contact sensiri~crs...J. /1rrr..t. Dernl. 47, 391. Kligman-A. M.p97af Report to RIFM.'_7'February. Kligman..:1. NI. & Hpsleia W.I19751.: l lj,dating the masimiznrinn Ic.c for, idcntifvingcnnlacl allergens. Canrmt DYlmuriti. I. •-11. Knnda. M.. Hunrymnrz. Y..dsni. V. & Sekinc. N..(1976). ('linim-pathoingic:d ohsertations.on o--cyanoacni:rtc
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TOXLINE 196G-67/DEC AU -- JANSSON T ;~, CURVALL M ; ZECH L ,._,~,y,r,~.~; TI - INDUCTION OF SI5TER-CWRqPZAT.IArEXCHANGES IN HUMAN LYMPHOCYTES-SY NEUTRAL,AND'•PHENOLIC: COh~5T-I~TIJENTS FROM THE SEMI-VOLATILE MATERIAL OF CIGAREETTE SMOKE CONDENSATE J SI -- EMIC/85/n(,n585 SO - MUTAT RES; 147:299,1985 AE - EMIC/OFRNL KW - EC,SCE ; CIGARETTE SMOKE CONDENSATE,SEMI-VOLATILE FRACTION'; CIGARETTE SMOKE CONDENSATE,NON-VOLATILE FRACTION ; CIGARETTE SMOKE CONDENSATE,SEMI-VOLATILE FRACTION,FRACTIONATED ; PHENOL ; GUAIACOL ; EUGENOL ; CYCLOTENE ; FURFURYL ALCOHOL ; 2,E--DIMETHOXYPHENOL.; MAMMAL,HUMAN CELL CULTURE ; SOMATIC CELLS ; HOI`10 SAP'IENS. ;. LYMPHOCYTES AU - NESTMANN ER ; LEE EG TI - MUTAGENICITY OF CONSTITUENTS OF PULP AND PAPER MILL EFFLUENT IN GROWING CELLS OF SACCHAROMYCES CEREVISIAE SI - EMIC/83/048958 SO - MUTAT RES; 119c273-28U,1983 AB - EMIC/ORNL SEE: CA 98-174537 KW -- EC,GENE CONVERSION-TRP ; GM,REVERSION TEST-TRP ; GM,REVERSION. TEST-HIS ; GM,REVERS'.ION TEST-HOM ; AC£TOVANILLONE ; 3-CHLOF'tO-CIS-MUCONIC ACID ; 4,5-DICHLOROCATECHOL ; 4,5-DICHLOROGUAIACOL ; NEOAHIETICACID o 7-OXODEf4YDR0A8IETIC ACID ; 1,1,2,3-TETRACHLOROPROPENE ; 1,1,2,3,3--F'ENTACHLOROPROPENE ; EMS ACENAPHTHENE; TRANS-ANETHOLE ; 3,4-DIMETHOXYACETOPHENONE ; ETHYLBENZENE ; 4-HYDROXY-3-METHOXYPROPIOPI4ENONE ; TRIMETHOXYCHLORODENZENE.;. DICHLOROSTEARIC ACID ; CIS--9.,IQ-F-POXYSTEARIC ACID; GLUCOISOSACCHARINIC ACID ; HOMOVANILLIC ACID ; VANILLIC ACID ; CHLORODc.HYDROABIETICACTD'; DEHYDROAB'IETIC.ACID~; DICHLORODEHYDROABIETIC ACID ; IS(lE•IMARIC ACID ;. LEVOPIMAfiIC ACID ; PIMARIC ACID ; SANDARACOPIMARIC ACID ; O-CHLOROBENZALDEHYDE ; VERATRALDEHYDE ^ 1,f,2,2-TE'TRAL"HLOROETHANE ;, ACETOSYRINGONE ; 2,b-DICHLOROPHffNOL ;, EUBENOL ; GUAIACOL ; ISOEUGENOL ; SYRINGOL ;. 2„3,4.,5-TETRACHLOROCATECHOL ; 3,4,5-TRICHLOROCATECHOL..; 2,4,5--TRICHLOROPHENOL ; CHLORANILLIC ACID ; 2,5-DICHLORO-3,6-DTSULFOHYDROQUINONE s TETRACHLORO--O--BENZOQUINONE ;, FUNGUS,YEAST : SACCHAROMYCES CEfnEVLSIAE,.D7 ; SACCHAROMYCES CEREVIS.IAE,XV185-14C 1
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CI F 001.41 £0.4 008720. Ergebnisse von Schwelversuchen, an Farbstoffen zur Farbmatt.ierung von Tabakwaren. 4.Mitteilung (I-iuminsaure). [Results of Carbonization Tests on Dyes for the Color Matting of' Tobacco Products. 4. Report. Humic Acid.] Kroller, E. Bundegesundheitsblatt. 6(22):348•-349. November 1,. 1.R63Languagea German The carbonization of humic acid (as sodium huminate) at 70t1^C under smoking conditions yieLded a number of less hazardous pyrolytic products such as carbon dioxide, water, carbonic acid, aliphati~c hydrocarbons,' pyrocatechol guaiacol, m- andp-cresol', beta-naphthol, phenol, pyrogallol,anthraquinone,, pyrenequinone, 1,2-benzopyrene, coronene, fluoranthene, and perylene. In addition, there were 27 micrograms of 3,4-bemzopyreneand.ate times 1-2 micr-ogramsof 1,2:.5,6-dibenzanthracene, 2r1-methylcholanthrene and2,'-•benzofluorene,. based upon 100 g sodium hmminate. Such relati',ve1y large quantities of' fhazardouspyrol.yti.c products require further examination regarding the use of humic acid in Tobacco VO.The hydrocarbons were identified on the basis of their Rp-values, fluorescence colors., and fluorescence maxima. 0013363 007622 .. EinBeitrag zur Beurteilung von Tabakzusatzstoffen auf' Grund ihrer Pyrolyseprodukte. 1. Cellulose und'ihre Derivate. [Evaluation of Tobacco Additives on the Basis of Their Pyrolysis Products. I. Cellulose and Its Derivatives.] Kroller, E. Bundesgesundheitsbl'att. 7(24): 375-378. November 27,, 1964 Language: German - Carbonizationtestswe-e carried out on cellulose and its.derivatives, such as acetyl-methyl-hydroxyethyl-, and carboxymethy cellulose, used as adhesives or thickening agents for tobacco. Separations were made chromatographicaPly,identi•fications were established on the basis of their Rp and RN values, fl'uorescence.colors and maxima, and their characteristic reactions. Fluoranthene, 3,4-benwpyrene, and anthracene were alwaysformeds in the pyrolysis. Frequently present were phenanthrene and its 2-methyl or 4,S-methylene derivative, d'.imethyl-benzanthraceneand 1,2-benzanthracene, picene, naphthacene, and 2U-methylcholanthrene. In isolated instances, naphthalene, fluorene, tribenzpyrene,, and perylene were also observed. Of thequi'nones,e anfihracene was always present, and phenanthrene lessfrequently.. In isolated instances, toluquinone, naphthenequinone and pyrenequinone could be identified. In the.acid portion of the pyrolysis products, pyrogalToli, a cresol and other phenols, as well as gallic acid and guaiacol wer-eal.ways present. Ali~.phatic_ hydrocarbons, aldehydes, carbonic acid, and,carbon.dioxide were always found in r_he.condensate. The lowest quantity of 3,.4-benzopyrene, 8 mcg, was present i~n the carbonization of the pure cellulose, but rose to much higher levels in its derivatives. Presumably, the nature of the substitution resulted in a.greater or lesser relaxation of the structural bonds, which led to a greater condensation in pyrolysis. . 14 I
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of the urinary metabolities revealed zingerone (54X), Zingerol (12X), 4-(3,4-dihydroxyphenyl)-2-butanone (6X) and 4-hydroxy-3-methoxyphenylacecic acid (8x) (Monge, et 'al. 1976. Ref. No. 1108). The latter two metabolites arise by well characterized in.vivo reactions of aromatic ketones (FEMA. 1978. Ref. No. 467a]. Although the metabolic fate of p-vinylphenol has not, as yet, been studied, a related report on styrene metabolism indicates that this phenol is excreted in the urine as p-vinylphenyl glucuronidate. Measurable quantities (1-10 ng/100 ml urine) of the phenol.wereidentified in the urine of workers exposed to an atmosphere containing approximately 130 ppm styrene for 8 hours [Pfgffli 1981. Ref. No. 1257a). The formation of p- vinylphenol via arene oxidation of styrene represented a minor metabolic pathway (<0.3Z) compared to formation of mandelic acidy the product of in vivo vinyl group oxidation. It is therefore, reasonable to expect that administered p-vinylphenol may be excreted either unchanged as the glucuronide conjugate or as the corresponding mandelic acid derivative. p-Vinylphenol has also been detected in the urine (enzyme-hydrolyzed) of rats maintained for five days on plant enriched diets. The phenol is probably generated Via decarboxylation of p-coumaric acid, a common plant constituent [Bakke, 0., 1969. Ref. No. 78I. TOXICITY The acute oral LD50 values reported for these phenol derivatives indicate . a low to moderate order of toxicity (see Figure I-1). Values range from 121 mg/kg for o-cresol (No. 2) in rats to 13,200 mg/kg for butyl -1-hydroxyben- zoate (No. 21) in mice. These values are significantly influenced by the type of vehicle used for administration of the compound. Higher toxicity is normally observed when the compounds are administered neat rather than in an oil or water vehicle. Since studies which use oil or water solutions moree closely parallel conditions for flavor use of these phenols, accurate estimations of acute toxicities should be based on these experiments. Multiple-dose feeding studies have been conducted for 14 substances in this group (see Figure 1-2). Phenol, the parent compound and most extensively OD N N Imb t O D I~ W
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I 0002 107 71 tJ822 Tabak:onoK:emuri Seibr.rn ni H:ansurtr Kenkyu. VII.. Tabako Kemuri no. K:ihatsusei Fenorurui no Sosei.. (Studies on Composition of Tobacco Smoke. VIT. Composition of Volatile Phenol Fraction of Tobacco Smoke.) Kaburaki, Y.; h'usakabe.,. H.; Shigematsu, H:. Nippon Senbai Kosha Chuoh:enkyusho Kenkyu Ho4::oku. 1~1Cr:121-125~. 1968 Language: Japanese The compositionn of volatile phenol fraction fromthe smoke of various unblended cigarettes was elucidated by a gas chromatographic method. The phenols separately determined were phenol, all isomers of cresol, xyl~enol, ethylphenol and methoxyphenol, and 3 isomers of trimethylphenol. Although the ratio of each phenol to total phenol in smokee was not so altered with the type of tobacco, minor variations were found i~n some types of tobacco. The ratib.of p-cresol to total phenol was slightly lower for the flue-cured tobacco smoke,, and that of phenol was lower and that of p-substituted phenols washi.gher f'orMitoNo. 3 (Burley) tobacco smoke. The majorvol.at.i~le phenols in smoke: were phenoL,3 isomers of cresol, guaiacol, p-ethy3phenol and 2,4-xylenol, which amounted to: about 90 percent of the total vol.a;ile.phenol estimated. Concentration of volatile phenols in crude condensate of smoke was higher in that of Bright Yeri ow and Izumir A tobacco than native and Burley tobacco.. (Auth. Abs. Mod'.) 0002043 71'6720 Effect on Ciliary Movements of Some Agents Which Come in Contact With the Respiratory Tract. Das, P. K.; Rathor,.fi. S.; Sinha, P. S.; Sanyal, A. K... Indian Journal of Physiology and Pharmacology. 14(4):297-303. October 1970 Language: English The effect of a number of agents which are either inhaled or excreted through the respiratory tract or are used for the treatment of inflammatory r_onditions of the respiratory tract was studied on ciliatedepithelium of frog`s esophagus. All the agents studied weredisso.lved in normal Ringer solution.. The ci~liary motility was found to bee stimulated by very weal: solution of ethanol (kr.Ca2 percent), ether, pare.ldehydle,oil of peppermimt, menthol, guaiacol, creosote, oxygen, carbon dioxide and cigarette smoke. Relatively higher concentrations of ethanol (C).1 to1.4percent), chlorofor,n, thymol,, petrol gas, potassium iodide and potassium:.bromi.dewere found to depress ciliary motility. ThestiRnulant actions didmot appear to be mediated' by neuro~humoral mechani.sms. The action of carbon dioxide was not dueto~ any change in hydrogen ion concentratiom., and'.the effect of cigarette smoke was possibly not due to the presence of nicotine. Of the three anions,, (chloride,bromide and iodide), the iodide ion was the most depressant. (Au±h,.. Abs. Mod'.) 16
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0 01 001'.3044 007179 Tabako, Rigunin rro Netsu.bun4:ai.. [Thermal Decomposition of Tobacco Lignin.] Kato, M::. ;, Sakai, F. ; et al, Nippon Senbai Kosha Chuo.L:enk:yushu Kenkyu Hbkoku. (107): 171-175. 1965 LLanguage: Japanese The following phenols have been isolated from the thermal decomposition product.ss of tobacco alkali lignin, prepared. from the stalk, as their 3,5--dinitrobenzoates: phenol, guaiacol, and pyrogallol 1,3-dimethylether. (m)-Cresol, p-cresol and p-creosol have been identified by paper chromatography of their phenyi-azo-benzenesulfonic acid-dye derivatives. In the case of stem lignin, almost the same phenols as that of stalk have been identified by paper chromatography. Vanillin, p-hydroxybenzaldehyde, vanillicc acid, and p-hydroxybenzoic acid have been identified inn the cigarette smoke from~~ solvent (ethanol, benzene, and water) extracted tobacco. (Auth. Abs.) 0013008 007141 Determination of the Steam-Volatile Phenols Present in Cigarette-Smoke Condensate. Part II. Determination of Phenol, The Cresols and Guaiacol by Thin-Layer Chromatography. Smith, G.A.L.; Sullivan, P. J. Analyst. 89: 312-318. May 1964 .Language: English A method is described for determining phenol, o -, m.-, and p-cresols. and~ guaiacol present in tobacco-smoke condensate. The phenols are coupled with diazotised, p-ni+_roanili.ne under specified conditions and extracted into ether. The ethereal solution is streaked on to a plate of k:ieseiguhr- G impregnated with formamide and developed with either a benzene--cyclohexane-dipropylenee glycol (30 + 70 +_ by volume) or aa benzene-cyc:lohe:<ane-diethylamine (5'. + 5 + 1' by volume) solvent mixture. AA good separation is obtained, and the individua7l bands aree scraped off and dissolved in ammonia.sol!rtion, sp.gr. 0.88. The individual phenols are then determined col!orimeflrically by using a Unicam SP500spectrophotometer. As little as 0..Smugg of each phenol can be determined quantitatively by this method. 15
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0020622 801323 Lipophilic Constituents of Natu Tobacco.: II. Phenols. Nagaraj, G.; Chakraborty, M. K. Tobacco Research. 5(2): 147-148. December 1979Language:EnglishNatu tobar_coo was extracted with hexane,, methanel, and diethyl ether. Phenols were separated by steam distillation unde:r nitrogen, saturatediwith sodium chloride, and extracted into diethyl ether°. Individual phenols wereseparated and identified by thin-layer chromotography. The phenolsi.dentified were resorcinol, catechol, p -cresol, phenol, mi -cresol, quaiacol, eugenol, p-metho:cy phenol,, andm-metho::y phenol. It was found'that phenol'~, eugenol, quaiacol, and p-metho;ay phenol occur in higher quantities than the other phenols. Quaiacol and eugenal' along with resorcinol, p--cresol, an6 m -methoxy phenol are responsible for the aromatic flavor of Natu.tobacco. G 6e013850 008295 Tabak:o1Eemuri Seibun ni Kansuru f'enkyu (V9 k;emurichu no Kakushu. Kihatsusei Fenoru no Firuta e no H'oryu. EStudies on Composition of Tobacco Smoke (V) Retention of Various Volatile Phenols in Cigarette Smoke by Fiiters.] Kaburaki, Y.; Kuniyoshi, H'.;. Shigematsu., H. Nippon Senbai Kosha Chuo I+;enkyushokenkyn Hokoku_ (109): 129-1T2.. 1967- Language: Japanese The selective retention of 17 volatile phenols in..r_igarette smoke on cellulose acetate Filter, Neo--filter (a bonded'd pulp filter), and polypropylene filter wasevaluateds by a gas chromatographic method. Although almost al.lthephenols were unselectively retained by polypropylene filter, the lower phenols showed higher seIectiveretenti~.on in general' on acetate filter and Neo-filter. o-Methoxyphenol alone was selectively retained by polypropylene filter showing the special characteristics. These resuitswere discussed on the basis of the vapor pressure of phenols and their affinity to the filter materials, indicating thee different affinity to therespect.ive phenols of acetate filter and Neo-fi,lter. (Auth. Abs.) 17
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I (TI studied substance, has'been added to the diet of rats at 0.10% (daily intake of 8.62 mg/kg) for up to four weeks with no significant adverse effects [Ind. Sio-Test Lab, Inc. 1969. Ref. No. 721]. Nor were adverse effects observed when phenol was added to the diet of rats for six months at levels resulting in daily intakesof 0.05 or 0.5 mg/kg [Kostovetskii and Zholdakova. 1971. Ref. No. 893]. However, at 5.0 mg/kg levels rats exhibited minor blood (decreases in free - SH group levels and serum cholinesterase activity) and hepatic (in- creased cholinesterase activity) changes which, for the most part, became more pronounced at higher dose levels. Rats also exhibited dose- dependent reductions in erythrocyte levels and liver weights when maintained for four months at daily intake levels of phenol above 2.3 mg/kg [Remenar, et al. 1964. Ref. No. 1326]. These minor effects of phenol on blood chemistry and liver function were not seen in longer-term feeding studies. Rats were maintained for one year on drinking water containing phenol at / levels calculated to result in daily intakes of 70, 100, 163, 224, or 239 mg/kg. The only significant effect was a depression in body weight at the two highest dose levels (Deichmann and Oesper, 1940. Ref. No. 354]. No adverse effects were detected when phenol was administered to rats in the same manner at daily intake levels up to 30 mg/kg for five generations or 155 mg/kg for three genera- tions [Heller and Pursell, 1967. Ref, No. 667]. Intake of 218 mg/kg/day in the five generation study and 310 mg/kg/day in the three generation study were neces- sary to produce significant effects on growth and reproduction. For four weeks the cresol isomers, o- (No. 2), m- (No. 5), and p- (No. 6), were added to the diet of rats at levels up to 7.0, 46 and 4S mg/kg, respectively, with no adverse effects. However, a 23 mg/kg intake level of o-cresol did re- sult in increased adrenal.weights [Tnd. uio-Test Lab, Inc. 1969. Ref. No. 721]. No effect levels of these cresols are at least 400,000 times the PCI. Other alkyl phenols, the p-ethyl (No. 131 and o-propyl (No. 14), derivatives, were administered to rats in daily oral doses of 0.2 mg/kg for 90 days [Posternak, et al. 1969. Ref. No. 1285] or 0;02mg/kg for six months [Veldre and Norman. 1974. Ref. No. 1723], respectively, lb both studies no adverse effects were noted at levels which are approximately 15,000 and 1,200 times, respectively, the PCI. 6
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SECTION II.B. ALPHABETICAL CAOSS-REFERENCE LIST OF NANES AND SYNOMlNS TABLE 11-2 This table lists alphabetically the principal names (in capital letters) and synonyms (in lower case letters) of the substances in this review. All synonyms are listed after each principal nsme as well. The numbers refer to the structural listing in Table 11-1 and indicate the ordering of the sub- stances in all other tables. This cross-reference list includes those synonyms most commonly fo md in the literature and synonyms, including some trade names, frequently used in the flavor industry. In a11 cases the reader is referred to the capitalized name which is the name used throughout this review. I 34 . 1.
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SECTION II.A. NIg4vRICAL DESICNATION AND CHEr:ICAL IDENTITY TABLE II-1 This table lists the primary names, chemical structures, molecular formulae and molecular weights of the substances included in this review. These compounds are listed by chemical structure as explained below, and'are aasigned a number by which they are referred throughout the review. In order to more easily make comparisons between metabolically related substances, the compounds are organized and numbered according to chemical structure D-o- gressing from simplest to more complex. A cross-reference of the ssbstances and their synonyms is listed alphabetically in Table 11-2. Additional inclusions in Table 11-1 are: a) the r.uanber assigned to the substance by the Flavor and Extract Manufacturer ' Association (FEMA) in its publications [467] listing substances judged by the Expert Panel of FEMA to be generally recognized,as safe under conditions of intended use; b) the sec- tion of 21 Code of Fedesal Regulations in which the substances have been listed by the Food and Drug Administration (FDA) ; c) the Chemical Abstracts registry number. 25 s~~~~~ti T ii
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11 SECTION I.B. SUNMARY fa urine are approximately 10 and 50 mg per day, respectively. Their presence derived from hydrolysis of theseesters are known to occur naturally. Endogenous levels of phenol (No. 1) and p-cresol (No.. 6) in human esters have as yet been identified in food,~.all the corresponding phenols .Twenty-eight of these materials are natural components of food. Eleven. - are present in a wide variety'offoods. Although none of the eleven phenyl',l isobutyrate (No. 4) to 89 ug/day for 4-(p-hydroxyphenyl)-2-butanone (No. 22) (see Table IV-2 and its introduction for details). estimated per capita intakes (PCI) range from 0.03 ug/day for o-tolyl 2-butanone (No. 22; 8890 ibs)having a volume greater than 1100 lbs. The level of use is reflected in low annual volumes with only 4-(p-hydroxyphenyl)-. with none having average maximum use levels`exceeding 55 ppm. This low. This review presents information relevant to the safety evaluation of phenol and fourty-five substituted phenols and phenyl esters used in food flavors. Low levels of use in food are characteristic of these substances is.a result of the intestinal-breakdown ~oftyrosine [Tompsett. 1958. Ref. No. 1672; Tompsett. 1958. Ref. No. 1671].~ol"~(No. 25) is also present in human urine at a level.of $mg/day which exceeds the PCI from ~ flavor use by about 1000 fold [Dawson, et al. 1964. Ref. No. 344]. Both phenol (No. 1) and cresol have'been reviewed elsewhero [NIOSH.1976. Ref. No. 1150a; NIOSN. 1978. Ref. No.7150b]. In these reviews, standards for acceptable occupational.exposures were recommended; for phenol, a maximum time-weighted-concentratiori in the air of 20, mg/m3; for cresol o-(No. 2), m-(No. 5),p-(N'o:$), or a mixture thereof, the limit of 10 mg/m3. Assuming a normal intak'e`volume of air of 10 m3 in 8 hours for an adult human and the conservative'.`figure of 5.6% absorption, these levels are equivalent to an acceptable daily exposure of 100 mg for phenol and 50 mg for cresol. These values are over~'600,000 and30,000 times, respectively, the estimated per captta daily intakesfrom.flavor use. Additionally, the greatly exceeds (approximately 200,00 txmes;the PCI) the flavor exposure to medicinal use of phenol in.throat lozenges'alone (approximately 32 mg/lozenge) this substance. a V M~,
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28 Ilydroquinone monoethyl other 29 2-Hethoxy-4-nethylFhcnol sgtst~g Ip STRUC71)RE oa OCI12C113 FORHUTA NM 0 CP 124 14 8 2 . C9H1003 166.18 C151t 1403 242.27 C8111002 138.17 C8111002 138.17 FF?01 NO. FDA STATUS C.A. REGISTRY HUMBER 2532 172.515 90-05-1 2687 172.515 613-70-7 2535 172.515 4112-89-4 3695 622-62-8 2671 172.515 93-51-6
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i FICURE'I-1 (cont) No Substance Vehicle Species LDS m/k , 16 2,5-Xylenol (cont) Oil Mouse 1140 Water Rat 444 Oil Rat 1270 Water Rabbit 938 17 2,6-Xylenol Water Mouse 479 Oil Mouse 980 Not specified Rat 400 Water Rat 296 Oil Rat 1750 Water Rabbit 700 18 3,4-Xylenol Water Mouse 400 Oil Mouse 948 Water Rat 727 Oil Rat 1620 Water Rabbit 800 19 Thymol Cottonseed oil Mouse 1800 Water Mouse 640 Propylene Rat 980 glycol Propylene Guinea Pig 880 20 Carvacrol glycol None Rat 810 21 Butyl p-hydroxybenzoate Olive oil Mouse 13200 22 4-(p-Hydroxyphenyl)-2- Propylene Rat 1360 butanone glycol/water (1:1) 24 Resorcinol Water ' Rat 301 Not specified Rat 980 Water Rabbit 750 25 I:GuaiaCol7 None Rat 725 F Corn oil Rat 1500 32 2-Methoxy-4-propyl- Corn oil Mouse 3071 phenol 33 Propenylguaethal Propylene Rat 2400 38 2,4-Dihydroxyacetophenone glycol Not specified Rat 2830 '12 . ~ T~-__ ,M7-._R - Ref. No. 1705 991 1705 991 991 1705 1337 991 1705 991 991 1705 991 1705 991 1058 740 753 753 753 1374 541 721 1337 - 1237a 1628 1237a 495 922 1337 V
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It should further bp emphasized that the data and description of sub- stances contained within this table are those listed in the referenced source and no attempt has been made to clarify these listings. For example, although some substances are listed as colorless, it should not be assumed that com- pounds not so designated are colored. As another example, the difference between soluble and miscible was not always clearly defined. Abbreviations Used in This Table w • water a1c • ethyl alcohol eth • ethyl ather s • soluble ins • insoluble 315 • slightly soluble mis • miscible mp a melting point bp - boiling point spg • specific gravity d - density dec - decomposes 47
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At higher levels a decrease in liver metochondrial alanine-transaminase ac-. tivity was observed. At 20 mg/kg, alterations in blood chemistry (decreased erythrocyte, leucocyte, and hemoglobin levels) and major organs (functional alteration of liver parenchymal cells and edema of kidney and liver) were re- corded. 2,6- (No. 17) and 3,4-xylenul (No. 18) administered orally to rats in daily doses of 0.06 and 0.14 mg/kg, respectively, for eight months resulted in no significant adverse.effects [Maazik. 1968. Ref. No. 991]. These in- takes are approximately 30,000 and 18,000 times, respectively, the PCI. A 100 fold increase in these intake levels resulted in significant dystrophy of heart, liver, and kidney tissue as well as increased serum SH group levels [Naazik. 1968. Ref. No. 991], ,Studies exist for one phenolie ester, butyl p-hydroxybenxoate (No. 21), and one phenolic ketone, 4-(p-hydrozyphenyl)-2-butanone (No. 22). The ester and ketone added to the diet of rats at levels resulting in intakes of 0.25 ~ mg/kg (40,000 times the PCI] for 1S weeks [Ikeda and Yokoi.1950. Ref. No. 715] and SO mg/kg (I50,000 times the PCI) for 13 weeks (Gaunt, et al. 1970 Ref. No. 541], respectively, resnlted in no detectible adverse effects Higher intake levels of the ester (50 mg/kg/day) resulted in vacuolar de- generation of liver cells and hyperemia of the spleen while increased ketone intakes (200 mg/kg/day) produced increased liver and kidney weights. Thymol (JVo. 19) was administered to rats orally for 19 weeks at intake levels up to 500 mg/kg/day (350,000 tines the PCI)' with no significant ad- verse effects [Hagan, et al. 1967. Ref, No. 63l]. Nor were any adverse effects detected when rats were fed diets containing propenylguaethol (\o. 33) or 2,6-dimethoxyphenol (No. 36) at intake levels (S00 mg/kg for 15 weeks [Lab. Ind. Hyg., Inc. 1951. Ref. No. 922] or 6.0.(?D and 6.8(F) mg/kg for 90 days [Posternak, et al. 1969. Ref. No, 1255], respectively) at least one million tines the PCI. A11 substances in this review have been individually reviewed by the Expert Panel of the F.E.M.A. and were judged to beGenerally Recognized as safe (GRAS) under conditions of intended use [F.E.M.A. 1965-198 . Ref. No. 467] taking into account the data summarized above and the reasonable analogies the Panel judged could be drawn. 9 - 10 Revised 8/85
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The reference number assigned to each summary refers to the biblio- graphy number assigned to the article summarized. A copy of each of the articles summarized, witH English translations of all foreign papers,, is included only for those substances not appearing in the original review or accompanying supplement. Articles covering substances in the original review were included in Volume III (1/79) while articles relating to substances in the First Supplement were included in that supplement (4/79). 61A ~- -. '" ""~ ' T ?q 77 7777 "~ ~ ~
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i FIGURE 1-2 (cant) Dose No. Substance S eg cies m/k ) Duration 19 Thymol Rat 50 and 19 weeks 5002 Rabbit (pregnant) 300 6 days 21 Butyl p-hydrox- ybenzoate Rat 0.25 15 weeks Rat 50 15 weeks 22 4-(p-Hydroxy- Rat 50 nd 13 weeks phenyl)-2-but- anone 100 F+ 2002 13 weeks ~ 5002 13 weeks 24 Resorcinol Rat 25.9, 4 weeks 87.4 and 260.7 25 ~aia2oT~ Rat 240 4 days 33 Propenyl- Rat 100, 250 IS weeks guaethol and 500 36 2,6-Dimeth- Rat 5.99 (M) 90 days oxyphenol and 6.85 (F) Z49L6LZZ+g Commenta. Ref. No. No adverse effects 634 No abnormalities of organs of dams or fetuses 1419 Estimated per cap- ita daily intake 0.0087 mg (1.4 x 10'4 mg/kg) No adverse effects 715 0.00038 mg (6 x 10'6 mg/kg) Vacuolar degeneration in the 715 liver, swelling of the glomeruli and hyperemia of the spleen - No adverse effects 541 Iricreased kidney and 541 liver weights Increased adrenal (M), 541 kidney and liver weights; reduced growth (M) 0.089 mg (1.5 x 10'3og/kg) Increased adrenal weights 721 4 0..001 mg at all dose levels (see Summary (<2 x 10-5 mg/kg) comments) Weight loss; mortality (2/6); mild 1628 0.0025 mg `~ liver lesions consisting of slight discoloration, mottling and blunting of lobe edges observed in sacrificed animals No adverse effects 922 0.011 mg (1.8 x 10-4 mg/kg) No adverse effects 1285 0.00048 mg (6 x 10-6 mg/kg)
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AU - F•OOL BL ;LIN PZ TI - MUTAGENICITY TESTING IN THE SALMONELLA TYPHIMURIUM ASSAY OF PHENOLIC COMPOUNDS AND PHENOLIC FRACTIONS OBTAINED FROM SI SO AE{ SMOKEHOUSE SMOKE CONDENSATES -- EMIC/S2/048476 - FOOD CHEM TOXICOL;,20:W83-391,195^ EMIC/ORNL SEE: CA 98-52458 ktW - AROCLOR ; GM,,AMES TEST ; MS,COMUTAGENESI.S ; MICROSOMES,RAT LIVER,SPRAGUE-DAWLEY,S9;. PHENOL ; 0-CRESOL ; M-CRESOL ; P--CRESOL ; 2,4-DIMETHYLPHENOL ; PRENZCATECHINE ; GUAIACOL ; SYRINGOL ; VAN'ILLIN'; EUGENOL ; SODIUM AZIDE ; 2-NITROFLUORENE ; 9-AMINOACRIDINE ; 2-AMINOANTHRACENE ; NORHARMAN ; SMOKEHOUSE SMOKE CONDENSATE,TARFRACTION ; SMOKEHOUSE SMOKE CONDENSATE,AQUEOUS FRACTION ; SMOY.EHOUSE'SMOKE CONDENSATE,NEUTRAL COMPOUNDS ;, SMOk'EHOUSE'SMO4:E CONDENSATE,MONOFUNCTIONAL PHENOLS;. SMOKEHOUSE SMOKE CONDENSATE,PHENOLIC FRACTION ; SMOKEHOUSE SMOKE CONDENSATE,BIFUNCTIONAL PHENOLS ; BACTERIA ; SALMONELLA TYPHIiMURIUM,TA98 ; SALMONELLA TYPHIMURIUM,TA100 c SALMONELLA TYI-'HIMURIIllM,TA1535 ; SALMONELLA TYPHIMURSUM,TA1537 ; SALMONELLA TYPHIiMURIUM,TA7538 AU TI - ANONYMOUS - GUAIACOL SI - EMIC/82/047984'. ~ ~ SO AB 4::W -- FOOD CHEM TOXICOL;. 2U(SUPPL):697-701,1982 -- EMIC/ORNL -- GUA.IACOL. . AU - NESTMANN ER ; LEE EG ; MATULA TI ; DOUGLAS GR ; MUELLER JC TI - MUTAGENICITY OF CONSTITUENTS IDENTIFIED IN PULP AND PAPER MILL- EFFLUENTS USING THE SALMONELLA/MAMMALIAN-MICROSOME ASSAY 51 - EMIC/86/0375t74 SO - MUTAT RES; 79':<<?3-21.2,198O - AEs -- EMIC/ORNL.SEE: CA 94-816 KW - AROCLOR 1254 ; GM,AMES TEST ;, MICROSOMES,RAT LIVER; (f DICHLOROMETHANE ; CHLOROFORM ; 1,2-DICH-OROETHANE ; 1,1,1-TRICHLOROETHANE ; 1,1,2,2-TETRACHLOROETHANE ; 1,1,.^.,_-TETRACHLORO--2-PROPENE ; 1,1,2'„T.,7~-PENTACHLOROPROPENE ; ACENAPHTHENE ; TRANS-ANETHOLE ; 3,.4-DIMETHOXYACETOF'HENONE ; ETf-fYLBENZENE ; 4-HYDROXY--:3-METHOXYPROPIOPHENONE ; TOLUENE ; TRIMETt-IOXYMONOCHLOROREN'ZENE ;. TRIMETHOXYTRICI-ILOROBENZENE ; ACETOSYRINGONE ; ACETOVANILLONE ; M-CRESOL ; P-CRESOL ; 0-CRESOL ; 4,5`°DICHLOROCATECHOL ; 3:,4,5-TRICHLOFiOCATEGHOL.; .^•_,.3,4,.5--TETRACHLOROCATEChIOL ; EUGENOL ;ISOEUGENOL. ,. GUAIACOL ; 4,6-DICHLOROGUAIACOL ; TRICHLOROGUAIACOL ;, TETRACHLOROGUAIACOL ; 2,6-DICHLOROPHENOL ; 2,4,5-TRICHLOROPHENOL ; SYRINGOL ; O-CHLOROBEN2ALDEHYDE ; YERATRALDEHYDE ; CHLORANILLIC ACID ; 2,5-DICHLORO--3,6-DISULFOHYDROOUINONEg TETRACHLORO-O--BENZODUINONE ; BETA-CHLOROMUCONIC ACID ; DICHLOROSTEARIC ACID ; CIS--9,10-EPOXYSTEARIC ACID ; TRANS-9,10-EPOXYSTEARIC ACID ;. GLUCOISOSACCHARINIC ACID ; HOMOVANILLICACID ; VANILLIC ACID ; 2
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4421 Griseoviridin (1977). Review and evaluation of studies of caminogenic action in laboratory animals: JARC.Monographs 10, 151- 161 (1976). Review:, Grovc Quars Reu~ 17. i(1963): Huber in Antibiotics vol. 3,. J. W: Corcnran. F:.. E. Hahn- Fds. (Springcr-Verlag• New Ymk. 1975):pp.606-613: Compre- hensive description: E. R. Townley in AnalyrienlProfrlesrofDrug Suhvances voL e. K. Florey:. Ed. (Academic Press. New York.1979) pp 219-244. . cx2p Stout octahedra or rhombs from benrene, mp 22P. [o]bT +37U (satd CHCIr soln).. uv max: 286, 325 nm. Soly in DMF at 25:12 to 14.gl.IOOml. Slightly sol in ethanol-methanol, aeetoney, benxene. CHCIy elhyl aceute. acelic I acid. Practically ihsol in water- petr cther. TREMP CAT:. Antifungal. I TREnAF CAT (VE[h Antifungal anlibiotic 4421: Gllsroviridin, OV. CyHnN~ 'O.tS: mol wt 477.55. C 55.33%, H 5.70Ya N 8.g07a O 23.43'~, S 6.72%. Antibiot- ic substance obtained together with Enmycin (viridoglisein). from S(Mpmnrysxag.iseuK Ber1z st aL. Antibfot.Ann..1954-~ 195g,.777. Characterirahon: Ames et aL J.. Chem Soc 1955, 426RAmes. Bowmen: tbid. 1955, 4264. Structure studies: FWona etaL. J. Am.. Chem. Soc..e4, 4162 (1962);. eldem. Can. J. C6em.42. 3714 394 (1964).. Revised strueturc G- I. Birnbeum, S. R. HAL. J. Am. Chem- Soc 98, 1926 (1976).. Absqtute oonGguration: B. W. BycrdL T. J. King. J. Chem. SoePerkin Tronc 1 1976, 1996. Production of griseoviridih and viridogriaein: BsrR et aL-,US. pat. 3,023,. 204 (1962 to Parke. Davis). Synthetic sludy: A. 1. MeyerA R. A Amos, J. Am. C.hem. Soc 102, 870 (1990). i 933 Polymorphic crystals. dec 158-166-, or 194-200-: or 230. 240' depending on the crystal modifipNon.. Ials -23T (c 0.5 in metianoU. uvmsa (methanol):. 221 nm (Et~*870):. Sol in pyridind moderately aal in lower dcoho sparingly salin waler and nonpolar solvsnts. J1;acetate C,.HsrNsO,S,.neeNes from methanol + other + petr ether. dec 137-I4(1- [018 -230' (e - 0-44 immetha- noUn uv mut (dhanol)-. 218 nm (a 41.g00).. M22. Grumeerite. 7FeO.8SiOpHrO-iron silicatc hydrate. 4423, GufLe. Gum guaiac;. «ein guaiac. Resin frumwood of Cruapcum offieinute L. or G mncrum:L, Zygophyl- Jaceaa CnnstJL Aqout 7U% a- and 9-guaiaennic aeids, about 11%guaiacic acid, related eompds and gualare0c add. 15% vanillin, gualac yellow, gusiac seponin (8uaiadn). Brownor greenish-brown, irregularr hunps. mp 85-90-. Insol in water,fraly sol in alcuhhol. chloruform, ether. creo- sote-soln 9f ehloral hydrate,:alkalies; slightly sol in benzene, carbon dlsulfule. InmmpaC Of liquid preparations: Min- eral acids, aeaeia, ferric chloride.gold chlo.ide,perman- ganstes., sp:At nitrous elher,r water. 1Dle~ orally in rats: > 5000 mglkg,. P. M.Jenner er aL. Fmd CaemeE Toxirol 2, 327 (1964). USE: Aa reagent in testing for occult blood. TxExeP CAT: In vilro diagnnstlc aid(bbod or hemm globin). 4424. GUaiu-Copper Sulfate.Paper. Schunbdn-Pagen- stecher's paper.. White filler paper impregnated first with an ale soln of guaiac resin, then after d.ying• with an aq soln or eupric sulfate. Used for detecting HCN, a trace.nf which plots the papcrblue. _ 4425. Guatiatol. o-Methoxyp6enn/;methylcatecholi o-hy- droxyanisole; 1-hydrory-bmcthoxybmiene. CrffsOr: mol wt 124.13. C'67.73%, H 649%w 0 25.78%. Isolated from guaiac resin: Sobeero. Ann 48, 19 (1843); from hardwood tar:. McGinness er al.. Tappi 43,. 1027 (1960). Prepd by mercuric oxidec oxidation of lignin: Lcwis, Prul. U.S. pat. 2,431,227(1947to Sulphite PnrdJ: by oxidation of anisolc with Mlluoroperoxyauetic aeid: McClure, Williams, J. Org. Chem. 27, 627 (1962); from acetovanillone + ZnCtr; Read. USS pat. 3,057.927 (1962 to Ontario Res. Found.)4:from the diaLOniumsall of o-anisidine: Herbsl..Gec.pat. 1,148,216 (1963 toHoechs0. White or slightly yellow cryst muss or colorless to yeA- lowish- vcry re@active liquidt characteristic odon Derkens omexposure to air and light. d(cryslals) 1.129; d{Oq) aboul 1.112. Solidif 28-, but may remain liq for a long time even at a much lower tqnp.. bp 2D4-206^ bp. 53-59. One gram dissolves in 60-70 ml water. 1 ml glycerol: miscible with aluohoh chlorofonn. ether,oils,, glacial acetic acid- Slightly sulin petr ethcr; Sol ih NaOH saln; with moderately wncd KOH itt forms a sparingly aol compd. Prorect j.um liBhL mg/kg, Taylor a aL, TozicoL.Appf. ha rmorolL 6, 378 (1964). P THERAPCATc ExpeetorenC TREM1AP CAT (yET): ExpCetomnl- "'4426. Guaiaeol BeazUate. 2-MeehoxyPhenoJ6enzoate; tienxoylguaiacN; Berlrasol. C.eHn0C¢.Hm~olwt 228.24. C 73:67%, H 5:309a O 21.03%. CHrp~ OOCCeHS. Prepu: Lynch,.Moore. Cnn. J. Chem.40, 1461 (1962). Odorless, almost tasteless, cryst powder. mp 57-58-. Slightly Sol in water;. Sol in hot dcohol. ether, chloroform. THERAP CAT: Expectoranl. 4427..Gltaiatwl Cacbonate. Carbonic acid bis(2-mRh- oxyphenyl) ester, guaiscalcarbonic acid neutral ealer; ear- bonic acid gueiacol ethcr; Duotal. CuHr,Op mol wt 274.26. Cb5.699q H3.14%, 0 29.17%. Prepd by the action of phos- gene on a concd s91b ofguaisml in aq NaOH: Oer.. pat.. 58,129 (to Bayer); S. P. Schotn Syntfretfe Organic Cem- povnds (London, 1925) p 137; 1. Schwyur, DkFubrikanbn Pharrnaeeutischer und C7cemischtectinischer Produkre (Berlin, 1931) p 212; H. P. Kauhnann..Arnimine!-Synrhese (Sprin- ger-Vetleg, 1953) p 607. Odorkss nee4les from ethsnol, mp 88a'. Faint taste of guafacol. Practically insol in water. At roomn temp I g dis- solvee in 60 ml erhanol, 1 m1 chloroform. 18 M etker. Much moresol in hot alcohol;.hm henrene, hot chloroform. Slight- ly soDin liq fatty acids. At 20-,.I00 g ot glycerol (d 1.2612) will dissolve 0.043 g of guaiatul earbonate. Difficult to liydrolyze. Upon ingestion much is excrotaduneWngedl THE6AP CAT: Expe<torant.. TNERAP CAT (r'ETI: EtrpeCtorant. ' 4428, Gutliacul Phosphate, CitH rO1P; mol wt 416.36. C 60.57%, H 5.08%, 0 26 9~..P 7.44~. (CHaOCtHtO)IPO. Obtaibed by. the action of POCfs upon gueiacol sodwm: Kucflerov. Zh. Obshch Khim 19, 126 (1949). Page 654 Consult rbe erm ihder beforc using this srctfon.. Crystals, mp 91' cthen Solin hot. Sodium salp A Rend. 146, 1131 l THERAP CAT: E: 4429. Gatlaco 69.21%. H 7.75% anlibacterial prn Forsrh 5, 224 (H Yellowish hq. . FreelY'sol in alce Use. Detstion 248 (1962). 40.10- Gttaiap rrsy)ethoxl/ethyl). NOA; mo1 wt 3: 19.79%n. Basicetl simi. F. Ravenn: 1,120,254 (1965, al. J.. Med. Chea eal studies: 0. 1 lyeical study: S. (1969). 1]q. bpas 190 Carissimi et aL. TnERAP CAT. 4431: GaW. tene; 7-irepruP. A!-ralon;Euear -reHle:; mal wt. chamonule oil: fi26 (i951J; fro:"(1956). CA.51 CoIL Geth. G Hehe Cbinr..A,20, 2821 (1964 251 (1979). Rlue oil. hl Trinitroben: from ethmd- 7stFJtAP CAT 4432. G.e did;. giyoerol methoayphen acyl efMr,. i o-methuxyph oxypbenoxy)1 gusiacaran; 1 Colrex Fsps ~ eaina; Myoc ~ Reludl G; R §,mlh Tenneuss:.Tr ~ 7.1256. O 32 ~ (1912); Yale viralts Auc ~ (1955)- irrg
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1800). The thymol isomer, carvacrol (No. 20), is expected to follow a simi-. " lar excretion pathway even though in vivo studies in rabbits indicate an un- usually slow rate of intestinal absorption. After 22 hours, about 30'% of an initial 1.Sg oral dose was present in the gastrointestinal tract of rabbits [1Villiams. 1959. Ref. No. 18001. In at least one compound, 2,6-dimethoxyphenol (No. 36), p-hydroxylation precedes conjugation. This is not unexpected since tha presence of both methoxy and hydroxy functions should activate the molecule toward further substitution. Administered intravenously to cats single 20 or 40 mg/kg doses of 14C-ring labelled 2,6-dimethoxyphenol (No. 36) results ,painly in urinary excretion of conjugated (90% as disulfate).and.unconjugated (3%) 2,6-dimethoxyhydroquinone. Also detected were free, (5%) and conjugated (2% as giucuronide) forms of the parent alcohol [Aliller, et al. 1976. Ref. No. 1084). In contrast, 2-methoxyphenol o(No. 25), is excreted withou'F-1 1:( of a 0.1-0.2 mg/kg oral dose of resorcinol was excreted unch.nged in the urine of rabbits. Also present were the monosulfate (13:5%);and the monololucuronide (52;) derivatives[Garton.and Williams. 1949. Ref.:No. 536]. As expected, the presence of a second labile functional group provides for alternate metabolic pathways. For example, butyl c-hydroxybenzoate is excreted by rabbits in the urine as either the free acid (40+), conjugates of the carboxyl group (30% hippurate and 7; glucuronide) or conjugates of the phenolic-OH group (14% glucuronide and 9% sulfate),[Tsukamoto and Terada. 1964. Ref. No. 1684]. .Zingerone (No. 34), given to rats in single doses (100 mg/kg), either orally or intraperitoneally, was excreted in the urine mainly as glucuranide and/or sulfate conjugates (95°s and 97%, respectively). Enzyme hydrolysis prior hydroxylation (73% of a 50 mg oral dose present in the hydrolyzed urine of humans within 14 hours; [Sedivec and Flek. 1970. Ref. No. i457]). ~ Since the simple esters of these alkyl and alkoxyphenols are expected to undergo rapid in vivo hydrolysis (o-tolyl acetate (No. 3) incubated with pancreatin was 60.•hydrolyced after,.2 hours;:at1'37°C [Grundschaber. 1977. Ref. No. 6161), it is reasonable to.assmne+;that these phenols and their corresponding esters are excreted principally;as the sulfate and glucuronide conjugates. Although resorcinol (No. 24) is mostly excreted in much the same manner as other phenols, evidence indicates appreciable amounts of unconjugated ma- terial is also excreted in the urine. Within 24 hours, approximately 11% 6 yN)f3: 3~~3H3f~11f3~;~~~?1j~i,3 .~I fl R
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SECTION II.C. PHYSICAL PROPERTIES TABLE 11-3 The data in this table were gathered.fram a variety of sources. The following sources were searched in the collection of data for this table; the initiali in parenthesis are used in the table following each entry to identify the source of that entry: (C) - "Food Chemicals Codex"' [494] (PC) - Private Communication, submitted by a manufacturer or user of the substance (ARC) - Arctander's "Perfume and Flavor Chemicals" (F) - Fenaroli's "Handbook of Flavor Ingredients" (40] [526], (EOA) -'"Essential Oil Association Specifications and Standards" [445] (HCP) - "Handbook of Chemistry and Physics" [1771] M - ^Merck Index'" [1065] (G) - "Givaudan Index'" [566] (GE0) - Guenther's "TheEssential Oils" [622a] The most reliable information on the properties of the substances reviewed, as used in flavors, are the data taken from "Food Chemicals Codex" and'from the Private Communications. qost of the additional sources searched are pub- lications oriented primarily towards the use of the substances as food additives. No attempt was made to judge the accuracy of this data. Therefore, in cases where data from different sources did not agree and was not clearly in error, all values are presented for comparison. This results in some com- pounds having two or more melting points, specific gravities, or other proper- ties listed. It should also be emphasized that the data presented may involve only typical specifiutions, and therefore might not be the same for different manuf acturers . I 46
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Name of Substance: GUAIACOL (25] Reference No.: 1628' (Taylor, J.H., et a1. 7bxicol. Appl. Pharmacol. 1964) Species: Rat (Osborne-Mendel or Sherman) Route: 0rai (intubation) No./Group: 6(SM 6 3F) Vehicle: None 1V/. Duration: 5 days Control: Not specified Rats received daily oral (intubation) doses of 240 mg/kg of guaiacol for 4 days and were sacrificed on the fifth day and their livers examined for groas lesions. Two of six animals died as a result of dosing. A number of animals lost weight and were in poor condition at the end of the test. Mild liver lesions consisting of slight discoloration, mottling and blunting of the lobe edges were observed in the sacrificed animals. Reference No.: 1237a(patty, F.A. Ind. Hyg. Toxicol. 1963) Species: Rat Route; Subcutareous r No./Group: Not specified ' Vehicle: Not specified Duration: Single Control: Not specified g/kg. The subcutaneous fatal dose for guaiacol in rats was reported to be 0.9 • Reference No.:. 1237a (Patty, F.A. Ind. Hyg. Toxicol. 1963) Species: Guinea pig Route: Subcutaneous No./Group: Not specified Vehicle: Not specified Duration: Single Control: Not specified The subcutaneous fatal dose for guaiacol in guinea pigs was-reported to be 0.9 g/kg.
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.RE'PORT DOCUMENTATION "m FEMAUS - 85/04 L PAGE 5 8 1. Ta1..nC iuMltl. • Snientifie Literature Feview ef Phenols (C12) e. 11.CaN Ca. • . . 9/85 a ~ 7. YWtna(U L hherIw11K Orf.Matbn Mnt Na :i... i Flavor and Extract Manufacturers' Association f, MmnnlK OMniptlan N.ro...d ALAY.n . 10. nq.RRaa/Wwk Unit Na. F].dWr and FyCtrdCt Ma73Uf7+c^?*'°ms AsSOC13ti071 , 900 17th Street, N. W., Suite 650, 11. Cenu.n(C) w G.a.Nm N,; Washington, D. C. 20006 (Q (w 22. fsnwwAN On2.nir.ww N.....r AaY.a. t7. TYw aY N.Pnn a P.dae Ce:..ad . F18VOr aACd FSCtr2Ct 1•SanlfaCtt7LES5( ~AS80C3atiOn . . :... ~ 900 17th Street, N. W., Suite 650 Revised washington, D. C. 20006 7~ ;asl/oa...na.aNoa.. . .. .. i SUpe=*des PB87 141224 2a Ab.tnct [UMC 2ao rs.W . . . .. ' .,': ., . . . . This Scientifie Literature Review includes specifications; safety data, poundage data, natural occurrenceinformation,.chemical identities and other data on Phenols. 1 1 . . 17. Oau.ro An.ryYS. .. O.seaqas .. , ' .. . • Id.MNMn/OWn•EnC.d T.nns ... ' .. e COSATI/f.W/Gw.p i< A..IbdWIBItlaMn:' . .. . /1/. S.wMtY.CI.u (This IbpuO. 27. Na. o/ Pa... - NTIS . - 20.S.eu.1tY Class RNg hPl 2L Mc. w,R...e.. . . OPTNNAI iORN Z72-C.-77) , (FMnyp.NT1iJ51 ~: ~ .py./tw+Mte.CSinm.rc. ~
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AU - RAPSON WH ; NAZAR MA ;; BULSKY VV TI - MUTAGENICITY PRODUCED BY AQUEOUS CHLORINATION OF ORGANIC COMPOUNDS SI - EMIC/8»/SJ37307 SO -BIJLL ENVIRON CONTAM.TOXICOL; 24:590-596,1980 AB - EMIC/ORNL GEEi CA,92-'209832 KW - GMI,AMES TEST ; MS,COMUTAGENESIS ; CHLORINE ; CHLORINE. DIOXIDE ; ACETOVERATRONE ; VANILLIN ; ACETOVANILLONE ; SYRINGALDEHYDE ; PHENOL ; O-CHLOROPHENOL ; M-CHOROPHENOL ; P-CHLOROPHENOL ; 2,3-DICHLOROPHENOL ; 2,4-DICHLOROPHENOL ; 2,5-DICHLOROPHENOL ; 2,,6-DICHLOROPHENOL ; 3,4-DICHLOFOPHENOL ; 3,5-DICHLOROPHENOL ; 2,3,4-TRICHLOROPHENOL ; 2,3,5-TRICHLOROPHENOL ; 2,4,6-TRICHLOROPHENOL., 2,6-DIBROMOPHENOL ; GUAIACOL ; 2,4-DTCHLOROGUAIACOL ; TETRACHLOROGUAIACOL ; RESORCINOL ; PHLOROGLUCINOL ; PYROGALLOL ; ABIETIC ACID ; EUGENOL ; CATECHOL ; 4-CHLOROCATECHOL ; 4,5-DICHLOROCATECHOL ; 3,5-DICHLOROCATECHOL ; LI~GNIN ; BACTERIA ; SALMONELLA TYF'HIMURDUM,TA100 AU - FLORIN I;'RUTBERG L; CURVALL M; ENZELL CR TI - SCREENING OF TOBACCO SMOKE CONSTITUENTS FOR MUTASENICITY USING THE AMES' TEST SI - EMIC/80/037208 SO - TOXICOLOGY; 15:219-232i.1;G0 . AB - EMIC/ORNL . KW - AROCLOR 1254 ;, 3-MC ; GM,AMES TEST ; BENZALDEHYDE ;, 2-METHYLBENZALDEHYDE ;.'3-METHYLBENZALDEHYDE ;4-METHYLBENZALDEHYDE.; 2-HYDROXYBENZALDEHYDE; 3-HYDROXYBENZALDEHYDE ; 4-HYDROXYBENZALDEHYDE ; ANISALDEHYDE ; 3,4-DIHYDROXYBENZALDEHYDE ; VANILLIN ; VERATRUMALDEHYDE; 5-METHYLFURFURAL ; 5-HYDROXYMETHYLFURFURAL ; 3-INDOLECARBOXALDEHYDE ; 3-PYRIDINECARBOXALDEHYDE ; PHENOL ; O-CRESOL.; M-CRESOL ; P-CRESOL ; 2,3-DIMETHYLPHENOL ;. ;- 2,4-DIMETHYLPHENOL g 2,5-DIMETHYLPHENOL.; 2,6-DIMETHYLPHENDL 3,4-DdMETHYLPHENOL ; 3,5-DbMETWYLF'HENOL ; 2„3,5-TRI.METHYLPHENOL ; ~ 2,4,5-TRIMETHYLPHENOL ; 2,4,6-TRIMETHYLPHENOL ;s 2-ETHYLPHENOL ; . 3-ETHYLPHENOL ; 4-ETHYLPHENOL ; THYMOL ; 3-ETHYL-`.;-METHYLF'HENOL ; .'.. 2-METHOXYPHENOL ; 3--METHOXYPHENOL ; 2,3-DIMETHOXYPHENOL ; SALMONELLA TYPHIMURIUM'.,TA98 ; SALMONELLA TYPHIMURI'.UN,TAI0O; SALMONELL~ TYPHIMURIUM,TA1535 ; SALMONELLA TYPHIMURIUM„TA1S37 ' i AU LEVAN A ; TJIO JH . TI SI INDUCTION OF CHROMOSOME FRAGMENTATION BY PHENOLS EMIC/48/010652. SO HEREDITAS; 34:453-484„1948 . AD EMIC/ORN_ . KW CE,.CYTOTOXIC ; EC,CHROMOSOME ABERRATIONS-FRAGMENTS ; MM,C-MITOSIS ; M-NITROPHENOL ;,0-NITROPHENOL ; P-NITROPHENOL ; M-CRESOL ; ; ORCINOL ;. THYMOHYDROQUIhIONE~.;RODI.NAL; GUAIACOL ; HYDROQUINONE ~ MONOMETHYL ETHERC ANILINE ; METHOXYHYDROQUINONE ; - ~ 2,4--D.IAMINOPHENOLDIHYDROCHLORIDE ; PICRIC ACID ; ~ ~ 2,6-DI.CHLOROPI4ENOL ; TRIBROMOPHENOL ; GALLIC ACID PURPUROGALLIN ; PLANT,ONION; SOMATIC CELLS'~ROOT'TIPS ; ALLIUM CEPA. . N 3
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AU -- LAKE L.`6 ; LONGL.AND RC ;HARR3SRA ; COLLINS MA ;. HEROD IA ; GANGOLLI SD TI - THE EFFECT OF TREATMENT WITH SOME PHASE II SUBSTRATES ON HEPATIC XENOPIOTICMETABOLISM AND THE URINARY EXCRETION OF METABOLITES OF THE D-GLUCURONIC ACID PATHWAY IN THE RAT. SI --Eh'IDEM/Cr022.:51 SO TO'X:ICOL. APPL. PHARMACOL. 1980, 52(3) 371-378 AB - ElSa Epidemiology I!nformation System k:W -- ACETAMINOPHEN ; PROPYL SALLATE ; GUAIACOL : 2-NAPHTHOL ;. DI'.F'HENYLACETIL ACID; CYTOCHROME P-450 ;, PHARMACODYNAMICS ;. ANIMAL ; FtIOTRANSFORMATION ; ANALYSIS ; DIPHENYLACETIC ACIDS ; ENZYME INDWCTI'ON ; GLUCURONATES,; METABOLISM ; URINE ;, LIVER ; ENZYMOLOGY ; MALE ;: METABOLIC DETOXICATION''., DRUG ; MIXED FUNCTION OXIDASES ; NAPHTHOLS ;.. PHENYLACETATES; PHENYLPROPIONATES ; RATS ; UDP GLUCURONOSYLTRANSFERASE G AU - Sjoblad RD ; Minard RD ; Col'lag .7M TI - Polymerization of 1-naphthol and related phenolic compounds by an extracellular fungali enzyme. SI -- PESTAD/77/G159 SO - Pestic. Biochem. Physiol. 6(5): 457-46-3; 1976. (12 references) AD -F'ESTAB. Attempts were made to characterize compounds which were -Formed by the partially purified enzymatic fraction from Frthizoctonia.praticol'a. Anextracellular phenol oxidase was iso3atedifromthe culture filtrate by gel fittr-ati.on on SephaJexG-2r90. The enzyme catalyzed the polymerization of 1-naphthrol to dimeric, trimeric, tetrameric, and! pentameric compounds determined by mass spectrometry. Identified cnmpou.nds included: 4,4'-bis--1-naphtihol,, phenol, o-methoxyphenol (guaiacol), p-meth'ylphenol (p-cresol), 2',6-dimethoxyphenol,.., andand 1,5-dihydr-oxynaphthalene. The results.ofthis study with 1-n:.aphthol,, an estatlslished intermediate.in carbaryl degrad'ation,, focusattentien on possible polymerization of >;enobiotic compounde by phenol oxidases. Possible impact on environmental pollution throughh the formatiomn of molecules whichi are very resistant to bio.'ogical attack must be considered. 4
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Name of Substance: GUAlACOL [25] Reference No.:1800 (9/illiams, R.T. Detox. Mech. 1959) Female rabbits were administered guaiacol via oral intubation of a single 325 mg/kg dose. Urine excretions were pooled for 2 days prior to and 4 days following treatment. Analysis of 48-hour (i.e., days 3-4) urine samples for sulfate excretions revealed 17.S-18t of original dosage present in the urine as the ethereal sulfate conjugate. Reference No.: 1457 (Sedivec, V. and J. Flek. Prac. Lek. 1970) Four human subjects were administered orally SO ml of water containing 50 mg of guaiacol. Urine was collected for 48 hours, hydrolyzed and analyzed. Approximately 55% of the administered dose was excreted within the first 2 hours. An additional 15% of the administered dose was excreted during hours 2-4 and 73.4% was excreted by the 11-14th hour. At this time (11-14 hours), the guaiacol content of the urine of test subjects was equal to that in nor- ma1 human urine (2.2 ug/ml)- Reference No.: 344 (Dawson, J. A., et al. Bull. N.H.O. 1964) The normal concentration of guaiacol in human urine was reported to be 0.1- 3.0 Yg/ml.
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103TaBOLISM The primary route for phenol excretion involves conjugation of the,hydTOxyl group as either an dthereal sulfate or glucuronide [ Nilliams. 1959. Ref. No. I800). Minor metabolic pathways utilize side chain oxidation and/or ring hydroxy- lation which normally occurs ortho or para to the existing phenolic group. In bolic pathways (Daly, et al. 1965. Ref. No. 329; Grimes. 1959. Ref. No. 611; Sato, et al. 1956. Ref. No. 1414; Sato, et a1: 1956. Ref. No. 1415; Spencer. 1960. Ref. No. 15301. vitro incubation of many of the phenols with liver preparations support these meta- (No. 19) in studies with humans, dogs and rabbits [Nilliams. 1959. Ref. No. was converted to a minor extent (less than 0.29c) to 4- and 5-methylguaicol by hydroxylation and subsequent o-methylation (Bakke. 1970. Ref. No. 79]: Incorporation of a second alkyl substituent does not seriously alter this general conjugation pathway. As`withthe'cresols, single 0.4 g/kg oral doses of either 2,5- (No. 16), 2,6- (No.; 17) or 3,4-xylenol (No.18) given to rabbits resulted primarily in.urinaryexcretion of glucuronide and sul- fate conjugates in an approximate ratio of5:1(Bray, et al. 1950. Ref. No. 172). The same type of conjugation mechaniam has been observed for thymol bolites (less than 3:) formed by o, p-hydroxylation were detected for all three cresols whereas only the p-isomer underwent side chain oxidation (10, of dose excreted as p-hydroxybenxoic acid). In rats;a.100 mg/kg oral dose of p-cresol and para-cresol (No. 6; 61% and 15%) were the principal urinary metabolites isolated from the urine of rabbits given single 0.2 g/kg oral doses of the respective parent compounds (Bray, et al. 1950: Ref. No. 1731. Minor meta- animals. The glucuronide and sulfate conjugates of ortho-cresol (No. 2; 72'; and 15%, respectively, of administered dose),`meta-ciesol (No. 5; 60% and 10°: by results obtained for the metabolism of numerous phenol derivatives in other et al. 1972. Ref. No. 230aJ. Data collected on the metabolism of phe'nol in man is closely paralleled. ~ Phenol conjugation and excretion occurs readily in other species as we14'[Capel, than 10% of the administered dose [Seutter<and-Sutorious. 1972. Re,f. No. 1469]. 14C-phenol (No. 1). Traces of hydroquinone conjugated both as a glucuronide: and as a sulfate were also detected [Capel, et al. 1972. Ref. No. 230a). A second report shows the extent-of hydroquinone fornation in humans to be less detected in man within 24 hours following a single oral dose of 0.01, mg/kg o~f . Urinary excretion of phenyl sulfate (774) and phenyl glucuronide (168),Was `
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700 . D. L..J. Ornvlae and C- Lttlzm Eicrmcr wrd'prurrirr. Andrejew, A. (19671. Peraxrdaue xctiviiy in mycobucreria.tluring Ihe selective inhibition of catalasc. Anrds Lrst. Pasrruc, Puris 112,.733: cited'from Chnn. Ab.vr. 1967, 67,.5126)'.. . Bo.u- If: C'.. Ram:utathan. N. & Navud:mtma, Y. (1962). EHeen nf dl0irenL chemicals includingHtose in vegettrble Ianning,malerials nn Ibu optical properlics.of cnllagen fiber. Bull..ceur. Lemh..Re.c. frut, Modrns & 263: uilavl from Chrvn. Ahxrr: 1962:57, 2556. . Bclficld,.A. & Goldberg, D. M. (19(r91. Enxyme diversion applied tu.the kinetic estimation of glucose 6-phos- phulasc aelivily..Li/i• Sr'f.8, 129: cited from Clrenr. Absrr. 11170, 103107. Levil.kii;,A. I'.. Gukevich, E..K. & Barahnsh, R. D. (19791: Method for determining peroxidase activity, in biological mmcrials. Ukce hiok/renr. Zh. 51. 289: cited from.Chem: Ahxrr. 1979, 94.34584.. Muhoney. C. P. & Igo, R. P. (19661. The hiosynlhrsis of lhyroxine. II. SolUbilization and characterization of an iodide peroxidase from thyroid lixsue. Bfrxhfm. hiuplrys. Aaa 113, 507; cited from Chern. Ahsrr. 1966, 64, 13027 Marshall. M., &. Chism, G. W. (1979): A comparisonn of the suitability of three hydrogen donors in the determination of pnoxidase activity. J. Fd Scf. 44, 942; cited from,C/rem. Ahsir. 1979. 91, 34573. Nikodejevic. B.. Scnoh. S.. Daly. J. W. &Creveling, C. R. U9701. Cmectiol-o-methyltransferasc. fl. Newclass of inhibilors of catechol-o-methyllntnsferase 3,5-dihydroay-4methoxylxnzoic acid and related compounds. J. Plrnrnurc, e.vp. Ther. 83. 174; ciled,from Chem. Ah.srr. 1970. 73, 84107.. Ohki. S.. Ogino. N,. Yamamoto. S. &. Hayaishi. 0. (19791. Prostaglandio, hydtoperoxidase,, an integral part of proslaglandin.endoperoxide synthelase from bovine vesicular gland microsomes- J. hinh Cltem. 254, 829; cited from Chem: Alisrr. 1979, 90, 147506. Ozawa. H...& Kawashima, H. 1J967A Inhibiloryy effects of tropolune analogs on catcchol-o-melhyllransferase- YaAur(nka Zrtcshi.87, 345; cited fromClrenr. .4Hsrr. 1967; 67, 61175. Panganamala- R. V:..Miller, 1. S., Gwebu- E: T., Sharma,.H. M. & Cornwell, D. G. (1977/. Differential inhibitory effects ofvilamin E and other unlioxidanls on prostaglandin synOrclase- platelet aggregation and lipoxidase. 1'rr.naglmrdin.r.14. 261; cited from ('Irent. Ahxrr. 1977;R7, 193682. Pnmmicq,l. & Cahnm:mn. H. J. (1979J. Inlcractinmof laclopcroxidasc with Ihiols and diiodolyrosine. J. hinl. (Chrnr. 254, 30f16; cilcd from ('lu.•ur..Absrr: 1979. 91, 34756. Roasch, R.I1966L Intracellular polyphenol oxiduscsofbrown rot fungi. IV. Occurrence and substrate specificity uf tyrosinasc and inlmocllulan luctnse., Arrh. M!k'mhio/. 54,.80: cited from Chem. Ahsl.. 1966- 65. 9390. Siegel. B. Z. & Siegel. S. bf. I19701.. Anomalous substrate specificities among the algal peroxidases. Am. J. Bnr. 57,.285: cilcd4rom Cluml. Ahsa. 1990; 73• 32272:. Taurog. A. (1976). The mcehanisrn of action of Ihe thioureylene anlilhyroid drugs..F.nrJnrrinnlapt' 98. 1031: cited from Chvru. .4ftmr. 1976. 85, 40532.. - Tenovuo..J. (19781. Inhibition by thiocyanate of lactoperoxidase-catalyzedoxidatiomand iodination reactions. Ar'efrs oral einl. 23. 899: cited from Chrm; Abstr: 1979. 91, 15754. Thomasa. A..Morris• D. R.,& Hager. L. P..(1970). Chloroperoxidase. VIL.Classical peroxidatic'catalalicc and halogenating forms of the enz.y'me. J. hfnL Chrm. 245. 3129: citcd from Che.rr: .4her. 1970. 73. 42020., Thomas. J. A. Morris, D. R. &Hager,.L. P. H970/:Chloroperoxidase. VIII. Formation of peroxide and halide complexes and their relaliomto the mechanism of the halogenation reaction. J. binl.. Clrem. 245, 3135; cited from Chem: A6.vr. 1970- 73, 42065. Stont D 1. (19710 - DelCelion of Iydroxy-and metllnxy9uinones in aqueous solurions./'rikf. Bink'him, rtlikrnhiul., citcd from Chrvn. ahnrr. 1972. 76. 95161. gcrmination and apPressnrial funnation in CnllrranrriairmrJidi rmr. Prne. InrfiM 4rnd..S,i.. Srcl. B. 74, 81:. Srini.vsam. K. V. & Narasimllan. R. p1971f Hllecl ofcerluin phcnnlie cmd relatcd cnntpounds on spore hiul., 4rh, Gorehorq:.S•'eden 1964, 3: cited from.Chem.. Ahsrr. 1966, 64, 11761. Saturu, O. (19641. Mechanism of antimicrobial effect of various food prcservalives. Inrern. Spmp: FrmdMirro- cxtrnctivc>:md related campnnntls. IInL-Inrs,lnorg 17, 54: cited front ('hnn. :Ih.rr..1963. 59. 5712. .Ihxrr. 1972. 77, 43920. RUdman. P. (1'/63f Cmrsc+ if natural durubllty iir timlicr. XI. Suntc Icsrv nm Ihe limci Inxicily rd woaVf Micruarganfsms. Abdullaev;, Kh. 11978) Anlitrichomonas activity of different groups of substances isolaled from the flora of Central Asia. In Furnrnknfngiya Prfrrrdnvkh Veelrelresla Edited by B. S Sadrildinov. p. 103. Izd. "Fan" Uzb. SSR, Tashkcnl. USSR:<iled from Chem. Ahsr.. 1979, 91, 49295. - Burlnn. 1). [:. ('Iarkc. K. & Gray. G. W. (1964); Mcchanism of Iltc nntibaclcrial action of phenols and salliyHdddrydes. 11. Snbslituled phcnols. J. rh.arr_Srn'.,1964• 1.314:',eited from Clmru: i6em..19(W. 611, 14872. Engelhardtl Ci.. Ras1.. H. G. & WullnoeRr. P. R. H9791.Cometuholism of phcnrd ;md suhstiluled phenols by Nn<vudimspre. DSM 43251. FFMS Slfcrehinf. Lrn. 5. 377:cilcd7rom Cl1r•nr ~11 1979, 91, 1, 52404. Fox: M. E. 11 r)761: Fatc uf sclccled orgunic compounds in tltc discharge of Kraft PuPtt mills.into Lake Supcrior. IAenril. Airnl. Orll. Pnlhrc. IVnrer (Chem. Cnn4r. Narlh Am. Cfnrr.) Lsr:,1976: 641 : cited from Chrm.,Ahsrr. 1977: 86, 33770. Hucllcr,.H., Lucss. K. D...Wnllmann; H. & PnhloudekFabini. R. U968k Org:uric Ihitxyatwgen, compounds. XXXII. funginmic properties and skin Ioleranec of some thiocvanophennls..lhloccnnophenol elhers. and Ihinewmuphenol kutones.l'hrrrnrech- 23. IRR; cilod from ('heur. :Ihxrr. 1968. 69, 4JG3]. Kirk. T: K. & Kclmnn. A. H965/, Lignin dcgradiuion as related In lhc phcnoloridhses,nf selected uood- ducayingbasidiomycclcs.PJrrrupndm/rryr 55, 739; cited from Chrur. Ahac 190. 63,.5774: Kloccking. R.. Tliicl, K. D.. Wolrler. P.. Hclhig. H. & Drabkc..Rl197Xt. Anliviral'aetn iquf phenolic polymers againsl.lypc I Hcrpcs virus bominis: 1'Irurrrnlcir33, 539; cited from (7irnr. :Ihur. 1975. 89, 15%(W9. Naravamrrno, A.. M:mihhuehanrao. K. & Sunyanarayamm. S. (19721. Eflccl nf snnta Pbvnntic cnmpaunds on s(vrrulatibn and growth of Plrioulrrriospedbs Prrat.Irrdinn Acnd Srf...SrrC.li. 75, 1411: cited from Chcm. 12, 121: eilerl frnm('henr: Ahsrr. 1976. 91 197575. Slom tr:~ 8. Stun AI Sloa pl: 90. Urs. bv West dr Wilf: Po Inoer Aldrr, bu Burd or 57 DuBc Ber citr Hena Re Jones res Kapl tar fro Mete. fru Mcyr Sri Neuh figr Nolte 15( .` Smill: ' mr, fror -' Toxft Ashirr I' Ko : Vr9 116 - Zamli - fror
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Name of Substance: GUAIACOL (25] Reference No.:1237a (Patty, F.A. Ind. Hyg. Toxicol. 1963) Species: Human Route: Oral No./Group: Not specified Vehicle: Not specified Duration: Single Control: Not specified x A single oral dose of guaiacol to humans reportedly resulted in irrita- tion, burning pain with v_omiting and bloody diarrhea. A tolerance reportedly developed following repeated~small doses: Death occurred in a 9 year old girl foliowing ,inge§t,i., on bf~SJoT guaja¢ol""
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. . . . .. .. . . MELr1R0 MO yECIPIC 4MrqY - -- t101uS6. AND SINSTANl:6. OC4d111T/0M SOUq1urIF5 POIMfS 711 OEP7KTIVL INOS2, 131D4GL IISSAf DIxMm c- O.lerless pri.rrlc crys- wsls (MC)(P) rp 26•(MC) SpS 1,125/2U• (PC) !pn (PL7 ~ . . . tels (MC) (1ICP) . .p 27.f• (F) Sp0 1.13 (AOC) . . ' - Yrllurisb lar"ltlel. °• ~bp 203 (ASC)(P)(IOS) SpS 1.7395.(P)' cr7stels ar l4(ula (p) ;Ep lo)'!2d .+"(P) ' ai2l 1.1=11 (1Sa•) bp 203-206' ePC) ap '0 LS361 (P) ' Ilasosansl pr4m (IICP) . . . . . . . • CA PowerGr/, swko-Ilb,. . .c7_ eefawh.t ..Jicln.l edor qMC) - Cuarc7l .cat.te 1261 Culorles.ar pels itrer- v-sls (AIIC) . .p 31-32' (IICP) SpS 1.16(MC) >_9S,St (P[) rnlared 1lyrrld rith r-lM (R) . ' bp 241' (MC) SpS 1-156/0' (p) wody, ar7. sllchtly . bp i35-240• (P) .. !}een cdor (MC) .. ' .. bp 123-12d'/13 r. (F) - , bp Ild-13S1/It w, (ICP) ' . NeproCaceE Innm ' . - r-~~w~~ . e+ Cea aval.El. eopy. _ . . - ' _ .. . . . - . .. '
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SECTION III PHABMACOLOGICAL.AND TOXICOLOGICAL EFFECTS ~ TABLE III r literature and private industry sources are also summarized and included herein. The table is organized by substance with the simplest chemical structures preceding the more complex, as in other sections of this review. The numbers after the name of each substance are the same for all of the other tables in the review. Within each substance group, the entries are organized from smallest test animal to largest with metabolism abstracts preceding the pharmacological and toxicological effects abstracts. This table contains information on the biological properties and metabolism of the substances included in this review. For compounds presented in the original review, the bulk of information was derived from articles obtained from a comprehensive survey of the scientific literature published between 1920 and 1977. This survey was carried out by an independent organization, Informatics, Inc., of Rockville, Maryland. Informatics, Inc. used the chemical names, common names and synonyms of each substance as key words to locate references to the published literature pertaining to the pharmacological and toxicological data on these substances. Those additional substances (Nos. 40 through 46) which had obtained FEMA GRAS status after completion of original review were searched by a different method. Computerized data-based search services (CAS Online, TOXLINE, TOXBACK, ATECS and TDB) available through national scientific organizations (American Chemical Society and National Library of Medicine) searched the scientific literature for data on these new substances. In this more recent search Chemical Abstract Registry numbers as well as common names and synonyms were used to find relevant literature references. Portions of the scientific literature not included in available databases were searched manually. The combined search methods provided a comprehen- sive survey of the literature from 1920 to 1983 on those phenol derivatives recognized as GRAS by the FEMA Expert Panel between January, 1980 and June, 1983. Summaries of all thE pertinent articles from this survey were then written and are presented in this table. Other data collected from earlier Hs
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Abbreviations Used In This Table GLC • gas liquid chromatoYraphy IR  iafrared spectroscopy UY w ultraviolet spectroscopy MS • mass spectroscopy ?!Wl • nuclear magnetic resonance spectroscopy TLC • thin layer chromatography PC • paper chromatography ~ 210 ~... I il
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Name of Substance: GUAIACOL [25] / Reference No.: 1812a(Norig, K.P. and T.L. Sourkes. Can. J. Biochem. 1966) Albino rats of the Sprague-Dawley strain were administered guaiacol via intraperitoneal injection of a 50 mg/kg dose and urine excretions were pooled for 24 hours. Chromatographie analysis demonstrated the presence of catechol in the urine, indicating 0-demethylation. !S Reference No.: 66 (Axelrod, J. and J. Daly. Biochim. Biophys. Acta. 1968) Guaiacol (O.S14moles) was incubated with rabbit liver microsomes and S-adenosyl[methyl- C]methionine. The substrate was examined for its ability to accept a methyl group from S-adenosylmethionine. The activity of phenol-0-methyl- transferase was reported to be 11 mumoles/g and with phenol the activity was re- ported to be 16 mumoles/g of liver. Reference No.: 119 (Benoit, W. Z.G'tlsamte Exp.Med. 1928) One rabbit was administered 3.0 cc guaiacol via subcutaneous injection. Urine excretions pooled for up to 18 hours exhibited a blackened appearence when exposed to normal atmosphere. Analysis revealed levels of combined guaiacol, while no levels of free guaiacoi were detected. Sedimentation examination showed /, no granulated cylinders, no erythrocytes and only a few leukocytes. Analysis o ~[lf urine excretions 42 hours after treatment contained no detectable levels of either free or combined guaiacbl. Sedimentation analysis showed more leukocytes than previously noted, some cylinders and no erythrocytes. The authors concluded that guaiacol is quickly eliminated and the principle amount is excreted within the initial 24 hours after injection. Reference No.:32g (Daly, J., at al. J. Med. Chem. 1965). A microsomal hydroxylating system requiring NADP and glucose-6-phosphate and 14 utilizing the enzyme eatechol-0-methyltransferase, with 5-adenosylmethionemethyl-C as methyl donor, was assayed. The system reportedly hydroxylates various phenol substrates to catechols then methylates them to extractable radioactive methoxy- phenols. Guaiacol (0.5 umole) was incubated with a rabbit liver supernatant (i.e., microsomal and soluble fractions) at 3Y in a medium (pH 7.9) containing both NADP and glucose-6-phosphate for 1 hour. Chromatographic analysis demon- strated the presence of the corresponding 0-methoxyphenol with a formation rate of 2.8 umole/g of liver. `
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[ Name of Substance: GUAIACOL [2S] Reference No.: 119 '(Benoit, W. Z. Gesamte. Exp. Med. 1928) Species: Rabbit Route: Subcutaneous No./Group: 3 Vehicle: Neat Duration: Acute Control: Not specified Two rabbits were administered guaiacol via subcutaneous injection of a single S.0 cc/kg dose. Toxic symptoms noted were immobility, convulsive spasms, difficulty in breathing and paralysis of the hind quarters. Death occurred in one animal within 1..5 hours and at 7 hours in the second. Urine excretions, collected via a catheter, exhibited discoloration and the presence of albumin, granulated cylinders and hemoglobin. Macroscopic examination ~r revealed congestion of the visceral organs, especially the kidneys, and a small white liver cyst in one animal. Microscopic analysis of the kidney revealed the absence of erythrocytes in some glomeruli loops which also exhibited swelling or hyaline transformation. The loop lumen showed col- lapsed walls. Alterations at parietal and viscerable capsular epithelium were not detected. A presence of roundish clotted albumin and tubule epi- thelium was noted in the capsule space of the glomeruli. In addition, hernia- like projections of epithelial cells were detected in the capsular space. Tubuli epithelia were swollen and displayed vacuoli in the protoplasm. Roun- dish bodies, hyaline cylinders and hemoglobin cylinders were detected in tubuli lumens. A third rabbit was administered a 1.4 cc/kg dose of guaiacol / subcutaneously. Death occurred within 8 hours. Hematological examinatio /n showed an increase in red blood eells and a decrease in the coloring index of erythrocytes. Urine samples displayed a slight presence of albumin, hemo- globin and granulated cylinders. Microscopic analysis revealed similar con- ditions as noted above with the exception of the hernia-like projections into the capsular space: Reference No.: 119 (Benoit, W. Z. Gesamte Exp.. Med. 1928) Species: Rabbit Route:Subcutaneous No./Group: Not specified Vehicle: Not specified Duration: Acute Control: Not specified X Rabbits were administered guaiacol subcutaneously in doses ranging from 1.0-4.0 cc. Temporary convulsive spasms, varying in degree from slight to strong, were observed. Recovery was noted. Q:ronic administration of un- specified, but very much reduced, amounts of guaiacol subcutaneously repor- tedly resulted in a considerable reduction in weight. 193 - -.--- .........m Wisti.~;....... ~. ..,... . .
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r Rabbits were administered guaiacol via oral intubation of either a 0.1 or 5.0 ml dose and the effects on the output of respiratory tract fluid were determined. Measurements were recorded at half-hour intervals for 3 hours prior to and up to 4 hours following treatment and the output calculated as Name of Substance: GUAIACOL [2S] Reference No.: 1561 (Stevens, M.E., et al. Can. Med. Assoc. J. 1943) Species: Rabbit Route: Oral (stomach tube) No./Croup: 11 Vehicle: Water Duration: Acute Control: Prior to treatment or non-treatment y ml per kilo bodyweight per 24 hours (i.e., ml/kg/24-hours). is a list of measurements obtained: The following Dose (ml/kg) Hourly Output of Respiratory Tract Fluid Before Drug After Drug 1 hr. 2 hr, I hr. 1 hr. 2 hr. 3 hr . 4 hr. Control 1.2 2.0 2.1 2.4 2,2 2.4. 2.3 0.1 1.3 2.1 2.2 2.6 2.5 2.9 2.7 5.0 1.4 1.8 1.8 3.2 -- ..- -- In addition, at 0.1 ml/kg, the increase in output of respiratory tract fluid reportedly was not marked, ranging from 20-40% of that prior to dosing. Slightly greater effects and occasional marked increases in the output of the respiratory tract fluid were observed at 5.0 m1/kg. Reference No.; 252 (Chabrol, E., at al. C. R. Soc.. Biol. 1931) Species: Dog Route: Intravenous No./Group: 3 Vehicle: Glucose solution Duration: Acute . Control: Not specified Three dogs were administered I g(ca 0.05 g/kg) guaiacol in 100 cc of a glucose solution. Within one hour bile flow increased threefold and then returned to control levels one-half hour later. Bile concentration is lower per unit volume but the voltmme of fluid excreted is greater. 19 5
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SUBSTANCE NO. FOOD SOURCE DETECTED DY P.P.M. (8)-ESTIMATE GUAIACOL (continued) 2S Crispbread, rye Filberts, roastedi Fish, smoked Grape, Concord Ms, BtC MS, GLC Paper Chromatogiaphy GLC 0.03(8) lV W d:• Herring, smoked aple syrup onion, roasted Passion fruit, yellow Peanut, roasted Paper Chromatography IR, GLC MS, GLC NS, GLC NS, GLC 3.25 Pork belly, smoked MS, GLC 35.6 Characterized by GLC retention tiwe, spectroscopy and comparison to authentic sample Characterized by GLC retention time, spectroscopy and comparison to authentic sample Characterized by paper chromatography and comparison to authentic sample - Characterized by GLC retention time Characterized by paper chromatography, comparison to authentic sample and colorimetry Characterized by GLC retention time, spectroscopy and comparison to authentic sample Characterized by GLC retention time and spectroscopy Characterized by GLC retention time, spectroscopy and comparison to authentic sample Characterized by GLC retention time, spectroscopy and comparison to authentic sample Characterized by GLC retention time, spectroscopy and comparison to authentic sample REF. 1747 851 1843 IS57 1844 476 945 iBOS 1762 731 sozstiZe
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Name of Substance: GVAIACOL [25] Reference No.:1213 (Ostrovakii, M.M. Gig. Sanit. 1964) Species: Mouse (white) Route; Inhalation No./Group: 10 Vehicle: Air Ouration: 2 hours Control: Not specified WISte mice were subjected to inhalation of guaiacol vapors in concen- trations of 1.98-17.37 mg/1 of air for a single 2 hour exposure period. I f Initially, mice at all levels of guaiacol inhalation showed inflammation of the mucous membranes of the eyes and respiratory tract. At concentrations up to 4 mg/1 toxie'symptoms are reversible. Higher concentrations (not speci- fied) caused hair standing on end, poor reflexes, shaking walk and inter- rupted breathing. At levels above 7.74 mg/1 clonic and tonic convulsions occurred. Most animals died during or within one hour of the exposure. The maximum tolerated dose (LC) was reported to be 2.03 mg/1. The LC50 and the LC100 were reported to be 7.57 mg/1 and 17.19 mg/l, respectively. Reference No.: 1237a(Patty, F.A. Ind. Hyg. Toxicol. 1963) Species: Rat Route: Oral (intubation) No./Group: Not specified Vehicle: 20% solution in corn' Duration: Acute oil Control: Not specified The oral LDS for guaiacol in rats was reported to be approximately 1.5 g/kg (95§ C.L. ~ Q:C-2.0 g/kg). When observed, death occurred within several hours. Autopsy of surv3ving animals at 24 hours and 2 weeks following treatment revealed no significant lesions. Reference No.: 1628 (Taylor, J.M., et al. Toxicol. Appl. Pharmacol. 1964) ~ within one hour to four days. Toxic symptoms included marked depression and a high dose-related comatose condition. Species: Rat (Osborne-Mendel or Sherman) Route: Oral (intubation) No./Group:. 10 (SM 6 SF) Vehicle:. Neat Duration: Acute Control: Not specified The oral LD (Litchfield-Wileoxon) for guaiacol in 8 hour fasted rats was reported to §2 725 mg/kg (95% C.L. - 302-1740 mg/kg). Death occurred 191 Y
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. .~~. t SUUSTANCe NO. FOOD SOURCE DETECTED aY P.P.N. e -esTINATe COl4@ffS REF. GUAIACOL 25 Rum 11S, GLC, Odor -- Characterized by GLC retention time, 399 (continued) spectroscopy and comparison to authentic sample Ru. lL4, IR, GLC -- Characterized by GLC retention tiae, 1664 spectroscopy and comparison to authentic sample Rum, Jamaican MS, IR, GLC 0.25 Characterized by spectroscopy 961 Sesame seeds, H4, GLC -- Characterized by GLC retention time, 1021 roasted spectroscopy and comparison to authentic sample Soybeans, deep MS. GLC -- Characterized by GLC retention time, 1790a A3 fat fried spectroscopy and comparison to W authentic sample Soybean protein 15 -- Characterized by GLC retention time, 1019 . spectroscopy and comparison to . authentic sample Soy protein, HS, IR, GLC -- Characterized by GLC retention time, 1020 hydrolyzed spectroscopy and comparison to authentic sample Summer sausage, NS, GLC 1.0 Characterized by GLC retention time, 731 smoked spectroscopy and comparison to authentic sample oZZSZZ14e
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No Subs->•ce SynonYms • 6-Ethoxy-•n-anol (PROPryYLCJAVIt:OL--no. 33] 1-Ethoxy-•t-hyd:o:cfxnrene (hYDRCQUl\O%e }SCh'OE'HYL ET??EP: -no. :8] 1-Et:oxy-:-hyd:oxt-4-prope.Vlbm:ene [PROP:NYLGUAZ'NOL--r.o. 33] 23 o-(ETHO7CYMET.yYL)PHE,7JL p-Etqoxyphenol {HYDROQUIYONE MQNOE7HYL ETHER--no. 28] 43 4-EYHYL-2.6-D11•Q:THOXYPHENOL . 2,6-Dimechoxy-4-ethylphenol ` - . 4-Ethylsyringol - 4-EThYLGUALACOL . 4-Ethyl-2-methoxyphenol Hosocreosol 1- Hydro xy- 2-e:e thoxy-•t- et'rylben_ene 2-Methoxy-l-ethylphe^.o1 4-Ethy1-2.-methwypher.oF [4-ElHYLGUAIaCJL--no. 30] 4-Hydroxyet.`.ylben.er.e 4-Ethylsyringol`(4-ETHYL-2;6-D1METHOXYPHEHOL--no.~43] . a-Hydroxfanisole' ,,I--Hydroxy-2-ciethoxyben:eae t/o-Methoxyphenol "-Methylcatec:tiol - vMethylcateahol a-1-c=.l-2-methoxybencene tRyrocatecF.ol~nonoae:.*.y1 ether L-Pyrogeaiac acid- Guia.ol phenylacetate [GIL:IAC.L PI•EhYLACTA:E--no. 27] 1-Aceto)c;-2-cet7:oxyben:ese- ace-,vl guaiacol o-)!ethoxy^yhenyl acetat: 27 GL'AIAC.L P!nYLA[° ;.T_ . . . Guaiacol ;.henyLce:atr o-F1ei^ox'~nenyl pne'lylace:a:e o-Mechylca:echol phenylaceca:e PaaocatecFsol oonoaethyl ether [2-)~Y307(Y-S.)g?,y1pYE,yCL--qo. =?j „mr I ~ .,;3 u , qp ~ ~ . .
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I SECfION IV.A. NATURAL OCCURRENCE TABLE IV-1 / Table IV-1 su®arizes references in the scientific literature to the natural occnrrence of 1khe substances in foods. For each substance which was found to occur in foods, the table indicates the food source, method of detection, level of concentration in the food (if reported), and other pertinent comments. The reference number following each entry in the table refers to the bibliography number assigned to the article in which that data was found. Copies of the articles cited with English translations of foreign.language articles are in- cluded in Volume III. As previously stated in Section I, the natural occurrence of a substance in foods indicates that the substance has been consumed by humans for centuries. Where quantitative data on the level of concentration are available, the level of cbnsumption can be e'stimated; where no quantitative data were found, the occurrence of a substance in a variety of foods usually indicates that more than trace quantities have been consumed. The data in this table were gathered primarily from information provided by the flavor industry. Although the tables do not necessarily represent a com- plete listing of all references to natural oecsrence which exist in the litera- ture, an attempt was made to include references to natural occurrence of each substance in as many different food sources as possible. Where it is stated that a substance was detected by isolation, this means that the actual substance, or a simple derivative, was isolated in a relatively pure form. The term "chemically characterized" means that identification was obtained by an appropriate chemical detection method. For convenience in pre- paring the table, methods using physical means of detection, such as mized melting points, were included under this heading.
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Name of Substance: GUAIACOL (25) Reference No.:1237g(Patty, F.A. Ind. Hyg. 7oxicol• 1963) Species: Rabbit Route; Oral or subcutaneous No./Group: Not specified Vehicle: Not specified Duration: Single Control: Not specified The•oral lethal dose for guaiacol in rabbits was reported to be 4.0 g. The subcutaneous fatal dose was reported to be 2.5 g. vx Reference No.: 1361 (Rovati, L. and E. Galante. Boll. Soc. Ital. Biol. Sper. 1960) via intravenous injection. Exhaled air was collected for 6 hours and a total; ~ of 10.2 ug of guaiacol recovered. Species: Rabbit Route: Intravenous No./Group: 8 Vehicle; Not specified Duration: Acute (6 hour observation) Control: Not specified Rabbits were administered two 7S mg/kg doses of guaiacol 3 hours apart Reference No.: 1550 (Stefano, B. and P. Quirico. Arch. Farmacol. Sper. 1939) Species: Rabbit Route; Intravenous No./Group: 8 • Vehicle: Aqueous solution Duratioa: Acute Control: Not specified Rabbits were administered guaiacol intravenously in single injections approximately equivalent to 60-598 mg/kg. A 60 or 90 mg/kg dose produced slight tremors for10-15 minutes followed by complete recovery. A 299 mg/kg dose resulted in tremors and immobilization for 1 hour, and recovery 1 hour later. Levels of 449 mg/kg also produced the above symptoms in addition to impaired reflexes, tachycardia, rapid breathing and hypothermia for a period of one hour. Recovery was observed the next day• Doses ranging from 464-598 mg/kg produced various symptoms including impaired or abolished reflexes, hypothermia, severe body tremors, reduced or increased respiration, sporadic to violent tonic-clonic convulsions, tachycardia or bradycardia and occasional discoloration of urine excretions. A dose of 464 mg/kg produced death in about 28 hours, while 494 mg/kg resulted in death within 4-5 hours, A level of 525 mg/kg also resulted in contractions of facial muscles and 598 mg/kg resulted in a slight bloody salivation while both doses produced death in 4-5 hours. Histopathologieal examination revealed alterations of hepatic canaliculi and congestion of visceral organs, particularly in the kidneys and lungs. The latter also exhibited sporadic hemorrhages. The minimum lethal dose was reported to be 464_ mg/kg. 124
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.....:. ..~....-. L_<Y y"JYCY. ~:.~......~ . '.: :"' ....... . ...... . .. ~ SUBSTANCE NO. F'UUD SOURCE DETECIED BY P.P.A. E -ESTINATE YbNlD]ITS REP. CARVACROL 20 Thyme lR, GLC -- Characterized by spectroscopy and 600 (continued) comparison to authentic sample Thyme oil CLC -- Characterized by GLC retention time a 1067 comparison to authentic sample Thyme oil IR, GLC -- Characterized by spectroscopy 137D Thyme, wild TLC, Column _ ChraatograPhY -- ------- 1442 4-(p-NYDROItY- 22 Raspberry Isolation -- Chemically characterized 1806 PNENYL)-2- BUTANOFtE Raspberry --- -- . Data from review article, no 1184 ei[perimental.data - : R13SORCIN[)L 24 Barley, roasted MS, IR, GLC -- Characterized by spectroscopy and 1484 iV comparison to authentic sample W -.--iG. .. . . Cane molasses CLC -- Characterizedby'6LC retention time 6S3 and comparison to authentic sample Coffee --- -- Data from review article 1763 .A_ - 2S Butter oil MS, OLC 0.00001(Ii)- Characterized by GLC retention time. 498 ., - 0.01(E) spectroscopy and comparison to ~-- authentic saaple '~ . Cassia oil NS, IR, CLC, -- Characterized by CLC retention time, 1636 TLC : spectroscopy and comparison to authentic sample ~ Celery' ~`iR,-CLC _- -- ': Charaeterized by,6LC retention time, 572 " - spectroscopy and comparisonn to . _.. , . _,... . , . :~ . authentic sample .... . _ ,
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Of a substance in cheese blintu , it would be assumed that the substance was in all bread, cakes, cookies, etc. in the baked goods category at the same level. Secondly, it is assuned that the average person eats food from all food categories every day. 'lhis would result in a daily intake of over 5,000 Calories, obviously an exaggeration. fie foregoing discussion is only a brief swmmary of the significance of the usage data and the inherent inaccuracies that exist in the table. To fully understand and appreciate the limitations of this data, the Addendim should be reviewed carefully. 1 i 1
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SUBSTANCE NO. FOOD SOURCE DETECTED BY P.P.M. M-RSTIMATg - COlOffiMTS REF. GUAIACOL 25 Whiskey, Scotch MS, GLC 0.012 Characterized by GLC retention time, 1176 (continued) spectroscopy and comparison to authentic sample Wine, red MS, CL4 -- Characterized by GLC retention time, 398 spectroscopy and comparison to authentic sample -ME778]XY-1- 29 Anise -- No experimental data S . E71 L Cocoa Paper -- • Charactesized by paper chromatograpA 4BS Chrommtography and comparison to authentic sem ~ belly, smoked MS, CLC 27.0(E) Characterized by CLC ention time, 731 ~ speetroscapy a mparison to authentic W saaiple ~ R~a MS, IR, GLC -- scterized by GI.C retention time, 988 spectroscopy and cosparison to authentic sample Nun, Jamaican MS, R, GLC 0.05 Characterized by spectroscopy 961 Sausage, summer MS, CLC <7.0(E) Characterized by CLC retention time. ~31 _ spectroscopy and comparison to authentic sample Nhiskey, Japanas .~~ MS, CLC -- Characterized by CLC retention time, 1176 ~ ectroscopy and comparison to authentic sa h7~iske , Scotch S, GLC 0.005 Characterize LC retention time, 1176 spectroscopy and com . on to authentic sample ziz6zi~s
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w R- ~.: _ _ ~~-~ - SUBSrANC6 NO. FOOD SOURCE DBTSCTBp BY P.P.l1. B -BSrIMATB CUNlIDiTS .. . . REF. : CUAiACOL 2S Cloudberry, oil MS. CLC 12,000 Characterized by spectroscopy 689 (continued) Cocoa M8, GLC -- Characterized by CLC retention time, 1123 spectroscopy and comparison to authentic sampze Cocoa Paper -- Characterized by paper chromatography 4BS Chromatography and comparison to authentic sample Cocoa beans GLC -- Characterized by CLC retention time and 1716 ' spectroscopy Coconut oil GLC. Paper 1.2 Characterized by CLC retention time, Chromatography paper;chromatography and comparison to 1642 W sothentic-sample ' w Coffee MS, CLC -- Characterized by spectroscopy 1739 Coffee MS, IR, GLC - Characterized by CLC retention time, 1571 . , spectroscopy and comparison to authentic sample - . . . : . . Coffee, aroma IR, GLC -- Characterized by GLC retention time , spectroscopy and comparison to authentic 1731 : . sample Coffee, ground 1R, GI.C -- Characterized by GLC retention time IS67 and spoctroscopy - . Coffee, roasted IR, CLC Characterized by GLC retention time, . 560 , . , spectroscopy and comparison to authentic ssmrle . . . . . . .offee, rossted' Isolation -- Characterized by compaiison to authentic 1289 ~ sample and chemicallycliaracterized. - 8UZ6TiO
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Substance Nae / Fir.s Reighted MeaD of s Possible Daily Intake Me. Survey No. Food Reporting Level of Use. PPM 1970 Lb R a d -_ . eoorte Cateaory Usage Usual Maximss S) A N ~ CD . 6-11 YD. .e161 .04Sa .0301 ~_12-2].ra._.a=3 -05t6_.045a. /~ __ 2-654 Ya _, .0444 __ .101 .0e11 .._.~..._ ~. .. , „ _ _~ , 6-11 YO,0207 .Oiia - .Oa= ,~1 , ' IS 23 NC.0302 .O1tf .09C6 :3biYd.._.0164 _.t3 ~__.275. 1 a-$ NO.-~~-".000613 ~ ~.00613 ~~".00117 _ ' 6-ll r0.~.00677_.0209.__.Olia . 7 12-2] Y0. .O107 .Oie3 .0705 9 Note 2 S• Ya. .0171 .oi4 .033/ ~' _OJ17.., .o02aa~.00367 _ ___ 1'I Nate 3 - -._ _~ ae .eragag Righ A Ni~h g _ -~ ... .. aLL_GikyDR s. 0'§ NQ._s0{7.p-~eo]570331 [l~ . _ . .. .. . . . . . . . .... . ....... . . _ 6-t~ .0. .ISZ .3a .303 ' 12-23 Ya. 2a7 57 .563 IS LBS..,--.---- ?-Ui.YEu2N 1~~ ' ~.19.. OAKEO GCCCSINI • 3.07 5.E3 1 0-S KO.. .0104 .013/ .019a Note 4 -.0253-_ J37< Note 5 Note 6 9ZZ6Li148 e ~ •-i1,.I0._.0779_.15! 140_ _. _ . i i . .. ._ . _ _,_. , . .a _ FaC2N OAIaYIaI 0-S R0. .00177 .007t1 '.0031i
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SECTION IV.B. USAGE LEVELS FROM SURVEYS BY FEMA AND NAS TABLE IV-2 (I This table is primarily a collection of the most pertinent results published in a series of tables by the National Academy of Sciences and available from the National Technical Information Service. These tables contain results calculated from data obtained from several sources, princi- pally from the 1970-71 surveys of usage conducted by the Flavor and Extract Manufacturers' Association (FEMA) and the National Academy of Sciences, National Research Council (NAS/NRC). The methods of compilation and computation are explained in detail in € the Addendum to Table IV-2 and in soueces referred to in that Addendum. j These doauments are available from the National Technical Information Service. } E For substances not inciuded in these surveys, the data come from the food industry source or sources that originally submitted the substance to i the F.E.M.A. Expert Panel for GRAS evaluation. In these cases the usual and maxdmm usage levels and the poundage figures are those.which the com- pany or companies had intended to u_va in flavors, as published by F.E.M.A. [467]. These publications are r:ferred to in the Table as GRAS 5, 6, 7, 8, 9, 10, or 11. Possible daily intake figures were estimated from these usage figures in the same manner as the "possible average daily intake" figures were calculated from the results of the surveys (see the Addendum to Table IV-2). It should be esQhasized that although these figures have the same multiple exaggerations built into them as do those calculated from the sur- vey results, they are directly comparable only if the data submitted by the company or companies requesting GRAS evaluation are the same as the weighted, . average values that would be calculated from a survey of the industry. While this is not likely, the variation should not be.so large as to pre- ' clude comparison entirely. It should further be mentioned that all of these i substances are currently being surveyed by NAS/NRC and the results will be m I available in late 1978. W j 2Q6 ,,
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SECTION V. BISLIOGAAPRY ( I I r 7he bibliography presents a eamplete listing of all literature articles and references used in the preparation of this scientific literature review. The entries in this section are listed in alphabetical order, by name of the principal author. The bibliography includes all articles located in the literature search, whether or not information from those articles was included in the literature review. Those references denoted by a single asterisk (*) are articles cited within the literature review, and references denoted with a double asterisk (••) are articles cited within the Summary portion of the review, in Section I. i Volumes II and III contain copies of all cited articles, including English translations of foreign language articles. 304 P OD Ij
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r The notes below which explain the entries in this table refer to the sample page that follows. Note 1- This column lists the name of the substance and in brackets, the number by which the substance is listed in all other tables in this review. Beneath the name is the total annual poundage used in flavors as reported on both the FENA survey and the NAS/NRC survey. The NAS/NRC sub- committee estimated that the data reported represented between 60% and 70% of the actual poundaye added to the nation's food supply annually. To esti-, mate psr capita daily intake in mg for any substance, move the decimal point of the poundage figure five places to the left. This takes into account conversion faetors and an estimated 60% coverage. Thus, an annual usage of 1S lbs for phenol is approximately equivalent to a daily per capita intake of 0.00015 mg. Note 2 - The food categories for which usage was reported are listed in this column. See the Addendum to this table for further explanation. Note 3 - The number of firms reporting usage in each food category is listed here. An•aaterlsk (*) indicates 3 or fewer firms reported usage. Note 4 - These columns list the weighted means of the usual and maximum levels of use reported on the survey of usage. Usage levels were weighted according to the annual potatdage reported as being used in foods. Each usage level reported by•each firm on a given substance was multiplied by a ratio caiculated by dividing the total pounds reported by the fixm by the total reported by all firms. The levels thus weighted were summed to obtain the weighted means. Note S - This lists the age group for which food consumption data were used to calculate the possible daily intake values. Note 6 - These colimn.s list the possible and potential intakes of the designated substance for each category by age group. The total for all categories is presented at the top of each column on a level with the name. of the substance. These intake levels were calculated, as described in the Addendum, from usage levels and food consumption data obtained from surveys conducted by the Narket Research Corporation of America and United States Department of Agriculture and are highly inflated as a result of the multiple exaggerations built into the calculations. Fint, the assumption is made that if a substance is used in one food in a category, it is used in all foods in.that category. For example,, if one manufacturer reported the use 247 aa:
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322. Dachav, B., and P. Botushanov. 1975. Laboratory study of the saaling qualities of materials for filling root canals. Stomatologiya (Safia) 57(4)z251-257. 323. Daere, J. C., P. A. Dans, and T. H. Kennedy. 1956. The metabolism of butylated hydroxy-anisole in the rabbit. giochem. J. 64(4)s777-782. 324. Daenens, P., and L. Larualle. 1973. Column chromatographic cleanup and gas-Siqaid chromatographic determinacion of hydroxybensoic asters in food. J. Assoc. Off. Anal. Chem. 56(6)s1515-1517. 325. Dalgliesb, C. E., E. C. Horaing, !1. G. Horaing, &. L. Tuo, aad L. Yarger. 1966. A gas-liquid-ebromatographic procedure for separating wide range of inetabolites occurring in nrine or tissue extract. Biochem. J. 101:792-810. 326. Dalin, B. K., and S. xristoffersson. 1974. Physiological effects of a sublethal concentration of inhaled phenol on the rat. Aan. Zool. Penn. 11(3):193-199. 327. Dallemagne, R. J., and B. Philipott. 1961. Anticurare action of adrenaline and some sympathomimeties: influence of sensitization agents on adrenaline. Agressologie 2(3)s269-274. 328. Dal Poazo, A. 1969. Analysis of some comoonly used therapeutic associations. I. Analysis of balsam mixtures. Boll. Chem. Farm. 108:47-56. Daly, J., J. x. Inacoe, and J. Axelrod. 1965. The formation of o-mechylatad catechols by microsomal hydroxylacion of phenols and subsequent enzymatic catechol o-methylatioa. Substate specificity. J. Ned. Chem. 8:153-137. 330. Daly, J., D. Jarina, and B. Witkop. 1968. Migration of deuterium during hydroxylation of aromatic substrates by liver microsomes.. Arch. 8iocbem. Biophys. 128:517-527. "331. Daly, J. N., J. Axalrod, and B. Vitkop. 1960. Dynamic aspects of enzymatic o-methylation and demethylation of catechols in vitro and in vivo. J. Biol. Chem. 235:1155-1159. 332. Damarijian, E. 1963. Nerve block for pain of fractured ribs. Ned. Trial Techn. Quarc. 10:75-77. 333. Dangelmajer, C. 1975. Toothpaste. Ger. Offen., 1975, p14. 334. D'asuto, F. 1951. Recent advances in the field of chemogenetics: Products of inetabolism as swtagenic agents. Caryologia 3(2): 249-259. 335. Danz, M., H. Urban, K. J. Stiller, and E. Amlacher. 1975.. Proliferative activity of the adrenal cortex as influenced by carcinogenic and noncarcinogenic substances. Exp. Pathol. 110-0:142-147.; 325
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z Substance Name I Firms Weighted Mean of Poesible Dai19 Intake, MR. Survey No. Food Reporting • Level of Use, PPM _ 1970 Lbs. ReOorted Cstegory Usage Usual Maximum A e Avere e Hfgh A High B dJAIACAT..~[75]._...-----6tLC..TEGCRtEi-_.15. CBEESEIRI, . ... ._ ..., _., ' bwT rxaoStRl - ._...R-S..M.- .o130_=o2a7. 6-!! ra.• .I4e .309 .292 -- -- •-- -- - - ---• .. 1t-23 IO. .0919 .ISI •Ie5 2-65. Y R 231 34 • . ( i4 64 I 0-5 Ra., . _ _~. _..-._.._____ 6-/l MO. „ .00027 .0001? .0002T It-21.Ma._ .00070_.00222 . .0007a , _ 2-63. YR. .00094 .OOf36 .00094 - •._F.g9lp-Qdlg7lRl.__.11- ' Orl.lR.~...0006R.L.00202,..06143_ 4 !SOFT ClNOY/RI 9 3.29 6.31 1. .12. 23.1'0..._.0026 a _.00731 ___ .00633-, _ ~ . ..___ .~.. ~.__..-~-_ . ._ . cEU71K~OPIRl~ •-..._ -S~4 . . 1.e4 R S.Ka._~00179 . •' ,oo2a1_._.ao364_~ 0EV TYPE IIRf 7 .756 __ 12-23 PO..__,041 _-_. .123 . .0767 __ 2-05! YR. ,0760 2l .151 __. _.. .... .....---•-._ OFV~7YpE IIIRI • 1.45 3. 6-lt MO._,00653 .O161 _ .0145 ~12-23 FO,,__,009W,~.0232 , .022 . , _.*-61 LtR v 9116 . . 0-5 R0. .00762 .00955 .00694 ._l011 __.12-23 PO.._e099}_,171_ ,- .19 _ _.2-65. YR, ...25a. •429. .494 . 0-S R0. .000162 .00162 .OOOly3 6-11 h0. .00176 .00949 .0041 6 ll Y0. .0114 .036T .0235 12-23 R0. .0123 .03. .6254 _ .2-65• YR. .0162 .0469 .07/5 0-5 MO. .o01e1 .00272 .00349 ,6-11.P0.___.0172__.0507.__.033 - -o-s hc~ -.ou0ooo~.ooaooo:oooaao- -'I 6.I1M0. ......• .0091 ....... -__ , .~_ .09T6 -_-- ~ ... (R ~__-.Ri ULi ~ 0 S ra. ~~...0.~! ~....... .0..... I _~_ ; '/ 6-11 Fo. ,000001 .000001 .000001 . 12-23 r0. .000001 .000003 .00C091 2"67l.YP...~.OOnVOX_.000004. .090002 P u i ~ fl -+1
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t A+trand, I., A. Kilbom, P. Ovrum, I. Wahlberg, and 0. Vestarbarg. 1974. Exposure to styrene. I. Concentration in alveolar air and blood at rest and during exercise and metabolism. Work Environ. Health 11(2):69-85. Aatrovskii, H. lI. 1961. Working conditions and their improvement in guaiacol and vanillin industry. Cig Sanit. 26s92-94. Attavay, J. A., A. P. Pieringer, and L. J. Barabas. 1966a. Origin of citrus flavor components. I.Analysis of citrus leaf oils using gaa-liquid chromatography, thin-Layer chromatography, and mass spectrometry. Phytochemistry 5(l):141-151. Attavay, J. A., A. P. Pieringer, and L. J. Barabaa. 1966b. Origin of citrus flavor components. II. Identification of volatile components from citrus blossoms. Phytochemistry 5(6):1273-1279. Attaway, J. A., A. P. Pieringer, and L. J. Barabas. 1967. The origin of citrus flavor components. III. A study of the percentage variations in peel and leaf oil terpenes during one season. Phytochemistry. 6:25-32.. Auerbach, C. 1949. Chemical mutagenesis. 8io1. Rev. Cambridge Philos. Soc. 24(3):355-391. Auerbach, C. 1950. Differences between effects of chemical and physical mutagens. Pubbli. Stn. Zool. Hapoli 22:1-21. Angier, J., and N. H. Henry. 1950. About bromine in the Hhodophyciae. Bull. Soc. Bot. Pr. 97(1/3):29-30. Aurbach, C. D. 1959. Eztraction.of parathyroid hormone with phenol. Areh.Biochem. and Biophys. 80(2):466-468. Auterhoff, 1. 1972.: Constituents of the essential oil from Turnera diffusa var. aphrodisiaca. Arch. Pharm. (Weinheim) 305(6)1455-462. Axelrod, J., and J. Daly. 1968. Phenol-o-methyl-transferase. Biocbim. Biophys. Acta 159:472-478. 67. Azous, W. H., D. V. Parke, and H. T. Williama. 1953. Studies in detoxication. 51. The determination of catechols in urine, and the formation of catechols in rabbits receiving halogeno- .benzenes and other compounds. Dibydrorylation in vivo. Biochem. J. 55(1):146-151. 55.: 56. 57. 38. 59. • 60. i 61. 62. 63. 64. 65. Astrand, I. 1975. Uptake of solvents in the blood and tissues of man. A review. Scand. J. Work. Environ. Health1(4):199-218. F aW m 68. Babkin, H. P., and A. E. gpitsin. 1965. Determination of phenols a in water by diazotized p-nitroaniline. Izv. Vyssh. Ochebn. N Zaved. Ehim. i Ehin. Tekhnol. 8(3):521-522. ~p C_~ N N O I 307 s
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14. Altes, E., C. Bornmam, and C. Banke. 1968. The excretion of (p-tolyl)methylcarbinol in bile. Naunya-Sctimiadebergs Arch. Pbarnakot. Exp. Pathol. 259(5):427-432. 15. Aloisi, M. and D. Cavallini. 1943. Activation of succinic dehydroganase-oxidase system. Arch. Fisiol. 43:261-266. 16. Aly, 0. H. 1968. Separation of phenols in vaters by thin-layer chro.atography. Water RAa. 2(8)t587-595. 17. Aeanot T., T. Takeuchi, and R. Toshii. 1967. Shoyu antiseptics. V. Determination of butyl p-hydrorybenzoate in shoyu by an ultraviolet absorption method. Chomi Lagaku 14(1)t5-9. 18. Amano, T., L. gazuo, T. Takeuchi, and H. Yoshii. 1967. Preserva- tives for shoyu (fermented soy sauce). VII. Determination of preservatives by gas chromatography. Nippon Shokuhin Rogyo Gakkai-Shi 14(11)t499-S03. 19. Ambasta, S. S. 1975. A rare ureteral injury secondary to lumbar sympathetic ganglion block with phenol. Br. J. Drol. 47(3):276. 20. Ames, S. B., M. L. Ludwig, W.. J. Swanson, and P. L. Harris. 1956. Effect of DPPD, aethylene blue, BBT, and hydroquinone r on reproductive process in the rat. Proc. Soc.Exp. Biol. Ned. 93(1):39-42. 21. Anasava, I., and L. Getake. 1960. Colorisutric,dete:mination of eresola.. I. Colorimetrie determination of er andp-eresols as nitroso derivatives. Rora Tarn 12=72-76. 22. Andersen, R. A. 1973. Carcinogenicity of phenols, alkylating agents, urethane, and a eigaret-smoke fraction in Nicotiana seedlings. Cancer Ras. 33(10):2450-2455.. 23. Anderson, S. C. at a1. 1948. I. Inactivation by RDE and by viruses of the influenza group, of the serum inhibitor of haemagglutination. II. Isolation and characterization of the serum.smcoid inhibitor ofheated virus.,by J. F. McCrea. III. Inhibition of virus haemagglutination by glandular mucins. IV. Inhibition by purified sucoid of infection and haesug- - glutination with the virus strain WSE. V. The destruction of "Francis inhibitor" activity in a purified mueoid by virus action., by F. N. Burnet. VI. General discussion., S. C. Anderson, F. M. Burnet, S. patakas de St. Groth, J. P.. MeCrea, and Joyce Stone. Australian J. Esp. Bio1. Ned. Sci. 26(5):347-411. 24. Andrieu, 0., F. Caujolle, C. Franck, and L. Girard. 1946. The pharmacodynamic study of carvacrol and its ethers. Compt. Rend. 223:755-756. Angel, A., and R. J. Rogers. 1968. Convulsant action of polyphenols. Nature 217(5123)s64-85. I 304
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( „ 0 MOUSEINHALATION.LC/S0l 7.57 MG/L CSTROVSKII 64 (continued on next page) GUAIACOL (2Sj PYROCATECHOL MONOMETHYL ETHER 1-HYOROXY-2-METHOXYBENZENE METHYLCATECHOL 1-OXY-2-METHGXYBENZENE C-METHOXYPHENOL 0-METHYLCATECHOL 0-HYOROXYANISDLE PYROGUAIAC ACID CFR 172.515 , USE LEVEL /PPMI MAX 0.01 TO 6.30 USUAL 0.01 TO 3.29 FEMAN 2532 VOLUME ILBI PER CAPITA INTAKE (MG/OAY) 251. 0.00251 r. I", NATURAL OCCURRENCE IN FOOD BIOLOGICAL DATA. . RAT ORAL LOISO) REFERENCE PATTY 63 1.5 G/KG RAT ORAL LOI50l TAYLOR J. 64 725MG/KG. RAT ORAL TAYLOR J. 64 240MG/KG/DAY FOR 4. OAYS. FECT ON GROWTH L LIVER. •RAT iNTRAPERLTONEAL METABOLISM EF- WONG 66 CATECHOL DETECTED IN URINE IN- OICATES 0-DEMETHYLATION 475
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GUAIACOL (25J (cont.) IN VITRO DALY J 65 HYOROXYLATION G 0-NETHYLATION BY RABBIT LIVER SUPERNATANT. t 477
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GUAIACOL 1253 (coat.) r RABBIT UNCLASSIFIED BENOIT WZ 28 1.4(5.OCC/KG.TOXICITY G DEATH. RENAL INJURY. RABBIT UNCLASSIFIED BENOIT wZ 28 1.0-4.0CC SC.CONVULSIONS WITH RECOVERY. WT LOSS IF C}iRGNIC. RABBIT UNCLASSIFIED PATTY FA 63 ORAL LE-THAL DOSE - 4.0 G SC LETHAL DOSE - 2.5 G RABBIT UNCLA4SIFIED ROVATI L 60 '75 MG/KG IV TYICE' IN 3 HOURS. 10.2MCG FROM 6 HR EXHALATIONS RABBIT UNCLASSIFIED STEFANO B 39 60=59"G/KG LV CNS EFFECTS. MLD 464MG/KG . RABBIT ACUTE ORAL STEVENS ME 43 0.1 L.15.OML/KG.INCREASEO RES- PIBATORY TRACT FLUID OUTPUT. RABBI,T ORAL METABOLISM 325MG/KG TO ETHEREAL SULFATE . CONJUGATE 117.5-18T1. RABBIT METABOLISM 3.0CC SC TO qOMBINED GUAIACOL. COMPLETE ELidTNATION IN 24 HRS -GUINEA PIG UNCLASSIFfEO'.` SC LETHAL DQSl~- 0.9 G/KG WILLIAMS BENOIT NZ PATTY FA 59 28 63 DOG 0.05 G/KG IV INCRE,ASEO BILE FLOW CHABROL E. 31 HUMAN UNCLASSIFIED PATTY FA 63 TOXIC EFFECTS W/"TOLERANCE. ' DEATH IN 9-YR OLD AFTER SkL. HUMAN OCCHRRENCE DAWSON JA 64 NORMAL CONCENTRATION IN URINE OF 0.1-3.0 MG/L/DAY HUMAN ORAL METABOLISM SEDIVEC V 70 73X OF DOSE IN URINE IN 14 HA IN VITRO AXELROD 68 RAD LLV HICROSOHr METHYLATION FROM.C-14-S-AOENOSYLMETHIONINE (cantinued om.next page) 476
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336. 337. Das, P. [., R. S. Rathor, P. S. Sinha, and A. K. Sanyal. 1970.. Effect on ciliary movements of some agents which come in contact vith the respiratory tract. Indian J. Physiol. Pharmacol. 14(4)t297-303. t 338. Das Gupta, V., and R. A. Bomer. 1975. Quantitative determination of phanol in phenolated calamine lotion USP. J. Pharm. Sci. 64:1199-1200. 339. Das Gupta, V., and S. F. Chan. 1975. Quantitative decermination off phenol in ointments. Am. J. Rosp. pharm. 32(5):522-523. 340. Da Silva, L. C., A. DeGodoy, S. T. Lurban, D. P. Neves, and J. F. Pontes. 1958. Critical study of flocculation tests: • Mechanisms and practical value. Rev. Assoc. Med. Brasil. 4.(4): 303-314. 341. Davidson, J. T., S. Rubia, Z. Eyal, and A. Polliack. 1974. A eomparison.of the pulmonary response to the endotracheal instillation of 0.1 N hydrochloric acid and Eartmana's solution in the rabbit. Br. J. Anaesth. 46:127-132. * 341a. Davis, D. V., and R. G. Cooks. 1982. Direct characterization of nutmeg constitutents by mass spectrometry-mass spectrometry. J. Agric. Food. Chem. 30(3):495-504. 342. Davson, 8., and J. F. Danielli. 1938. Studies on the permeabi:ity of erythrocytes. V. Factors in cation permeability. Bioehem. J. 32(6):991-1001. * 343. Darby, W. J., B. H. DeBsio, M. L. C. Bernheim, and F. Beraheim. 1945. The metabolism of phenolie compounds by normal and scorbutic guinea pig liver slices in vitro. J. Sio1. Chem.. 158(1):67-69. Davydova, R. Z. 1972. Chromatographic determination of phenols in urine. Hed. Zh. Uzb. 9:77-78. Davson, J. A., D. F. Haath, J. A. Rose, E. X. Thain, and J. B. Vard, 1964. Excretion by humans of the phenol derived in vivo from 2-isopropoxyphenyl N-metholearbamate. I. Detersization by gas chromatography. Bull. RBO 30(1):127-134.. 345. Day, H. A. L944. Thymol in cavity sterilization. J. Am. Dent. Assoc. 31(9):605-615. 346. De Barbieri, A. 1965. Protective and antitoxic effects of liverextracts. Boll. Chim. Fara. 104(6):33'r-341. ' 347. Debrawere, J., and :S. Yerzele. 19:5. :fe+ constituents of the oxygenated fraction of pepper essential oil. J. Sci. Food Agric. 26:1887-1894. 348. Decker, C. C., and C. J. Drake. 1940. Pre1!=inary studies on the use of dinitro-o-ereosol dusts ia grasshopper control. Iova State Coll. J. Sei. 14(4):34d-351. ' 349. De Croate, V., C. Lambotte. 1960. A case of fatal phenol poisoning in a newborn infant. Ann. Xed. Leg. (Par.) 40:288-290. 326
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t 113. Beer, U. D. 1959. Toxicity of phenols and phenol-coctaining industrial wastes to aquatic and terrestrialorganisms. Visa. Z. Rarl-Harx-Univ. Leipsig, Math.-Naturviss. qeihe 8:67-96. 114. 115. 116. 8ahm, R. C., and,J. M. Nelson. 1944. Aerobic oxidation of phenol by means of cyrosinase. J. Am. Chem. Soc.. 66:711-714. Belova, I. H. 1961. Detatmination of monohydric phenols by partition chromatography and ultraviolet spectrophotometry. Zh. Anal. ghim. 16:229-236. Banazeeh, J. P., A. Moisan, G. Pitet, and Mrs. Caillard. 1970. Identification and determination of p-hydroxybenzone esters used as drug preservatives. Trav. Soc. Pharm. :lontpellier 30(4):293-300. ~117. Benezet, L. 1945. Contribution to the study of the oil of lavander. Parfumeria. 1:153-157. 118. 120. 12I. 122. i t. 122a. 123. 12A. 125. ~ 126. Ct Bennett, I; L. Jr., D. F. James, and A. Golden. 1950. Severe acidosis due to phenol poisoning; report of two cases. Ann. Intern. Had. 32:324-327. Benoit, V. 1928. Poisoning of rabbits by guaiacol with special consideration on kidney changes. Z. Cesamte Exp. Hed. 62(5/6):585-613. gent, H. E. 1938. The movement of water from concentrated to dilute solutions through liquid membranes. Science 88(2292): 525-5'26.. Berei, K. and L. Vasaros. 1967. Separation of certain aromatic compounds by thin-layer chromatography. Magy. Kea. Foly. 73(7): 313-317. Sergan, T., and A. Lyscad. 1972. Antitubercular action of disinfectants. J. Appl. Bacteriol. 34(4):751-756. Berger, P., E. H. Sanders, P. D. Gardner, and.N.C. Negus. 1977. Phenolic plant compounds functioning as reproductive inhibitors in 7h:crotua mmttm:us. Science. 195:575-577. 3ergstrom, S. 1942. Autoxidation of sterols in colloidal aqueous solution. III. Quantitative studies on cholesterol. iV. The influence of esterification and of constitutional 'actors. J. Biol. Chem. 145(1):309-333. 3ermn, M. L., andJ. T. 3oehantia. 1972. Evidence of er.haaced sicroaomal enzyme activity by preservatives _n volatile anesthetics. Ca11. Biol. Toxicity Anesth. ?roc. 3es. Symp. 267-274. 3ernheim, F., and H. L. C. 3ernheia. 1943. Conjugation in vitro of phenol by guinea-pig liver. J.: Pharmaeol. 78:394-399. Bernheim, F., and M. L. C. Bernheim.1944.. The metabolism of tyramine, 1-tyrosine,, and phenol by rat tissues in vitro. J. 3io1. Chem. 153(2):369-373. 311 t
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1204. Ordal, E. J., J. L. Wilson, and A. F. Borg. 1941. Studies on the action of wetting agents on microorganisms. I. :he effect of pN and wetting agents on the germicidal action of phenolic compounds. J. B:et. 42(1)t117-126.. 1205. Organon, N. V. 1957. Choleratic substance from p-tolylmethylcar- binol. Dutch 85,756. ' 1205a. Oser, e.L. and R.L. Hall. 1977. Criteria employed by the Expert Panel of F.E.M.A. for the GRAS evaluation of flavoring substances.Food Cosmet. Toxicol. 15:457-466. 1206. Oshino, Z., and 8. Sato. 1971. Stimulation by phenols of the reoxidation of aderosomal bound cytochrome b and its . implication for fatty acid desaturation. J. Bioegem. (Tokyo) 69(1)3169-180. 1207. Oaintseva,'P. P., U. C. Pogasyan, and 0. D. 6olbasovs. 1967~ Experimental rasarptive effect of small concentrations of acetone and phenol on the organism. Sb. Tr. Inst. Epidemiol. Gig. Arm. S5A 5:358-362. '~1207a. Oso1,A., P. Robertson, and H.D. Altschule. 1967. The United StstesDispensatory and Physicians' Pharmacology. 26th ed. J.B. Lippincott Company. Philadelphia g Toronto. 1208. Osterhout, W. J. y. 1937. The action of guaiacol as affected by p8. J. Cen. Physiol. 20(5):685-693. 1209. Osterhout, W. J. V. 1939. Calculations of bioelectric potentials. VI. Some effects of guaiacol on Nitella. J. Cen. Physiol. 23(2):171-176. 1210. Osterhout, W. J. V. 1941a. Positive potentials due to aniline and the antagonistic action of ammonia. J. Cell. Comp. Physiol. 18(2):129-134. 1211. Osterhout, W. J. V. 1941b. Effeots.of nitrobenzene and benzene on nalonia. J. Can. Physiol. 24(6):699-702. 1212. Osterhout, W. J. V. 1943. Diffusion potentials in models and in living cells. J. Cen. Physiol. 26(3):293-307. Ostrovskii, :f. X. 1964.. Toxicology of guaiacol vapors and of its resin. Gig. Sanit. 29(3):85-88. Otsuka, li., and Y. Bonoa:ura. 1963. Attidn of phenotie substances on motor nerve endings. J. Pharxcol. Exp. Ser. 140:41-45. Otsuka Pharmaceutical Co., Ltd. 1982. Electropreparation of 3,5- dialkoxy-4-hydroxybenzaldehyde. Jap. Pat. 57/2881.. Ozaki, S. 1958. Comparison.of various eolori=etric.deteminations of phenols. Bunseki Zagaku 7:275-283. 1216. Ozanne, P. G., and 1. `J. W. 8owes. 1971. Preference of grazing sheep for pasture of high phosphate content. Ausc. J. Agric. Res. 22(6):941-950. 1217. Ozawa, Shahei. 1929. The seat of elisination of drugs in the kidney. Proc. Imp. Aead. 5(8)t396-398. 1218. Paee, D. M., and A. Elliott. 1962. E:6ects of acetone and phenol on established cell tines cultivated in vitro. Cancer Res. 22(1):107-112. 385 -i
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SECfION VI. DATA GUIDE r The following guide provides a summary of various data included for each substance in this Scientific Literature Review. The principal name and FEIM nuober of the substance is followed by its synonyms. The Code of Federal Regulations (CFR) reference is given for those sub- stances published by the Food and Drug Administration. The range of average usual and average naximum usage levels of the sub- stance is shown next, as well as annual volume in pounds. These data are taken from the 1970-71 NAS and FENA surveys, if the substance was included in the survey. For substances not included in the survey, the data come from industry sources as explained in the introduction to Table IV-2. The per capita daily intakes are calculated as described in the intxvduction to Table IV-2 from the annual volume. If the substance has been found as a natural component of food, this is noted after the usage data, with the number of different foods in which the substance is found indicated in parenthesis. Finally, a stmmar}' of the biological data available for the substance is listed. The references are to the surname of the principal author and the year of the publication for each study. The bibliography can be consulted for the complete journal reference. This guide has been generated by computer and is therefore printed in capital letters, with the brief descriptions of the biological data being limited to 60 characters. Where Greek letters should occur, these are printed as their English equivalent followed by an ampersand; i.e. a. As and A - D8. 431 i
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1543. Stanier, R. Y. 1948. The oxidation of aromatic compounds by fluorescent psaudomonads. J. Baet. 55(4):477-494. 1544. Stanley, P. E., A. C. Jennings, and D. J. D. Nicholas. 1968. Ultracentrifuge studies of proteins in mixtures of phenol, acetic acid and vater. Phytochemistry 7(7)s1109-1114. 1545. Stark, i7., C. Urbach, O. 11. Jacobsen, and J. S. Hamilton. 1970. Recovery of added acids, lactones and phenolic compounds from milk fat by cold-finger distillation. Int. Dairy Congress 1E:82. 1546. Starkey, R. J., Jr., J. J. Shall, and E. D. Orr. 1971. General Electric phenot analysis equipment. AIAA Tech. Event 1970:203- 212. 1547. 154$. I 1549. 1551. Steck, W., and S. H. Aender. 1965. Paper chromatography in benrene-acetic acid-vater as an aid in estimating free hydroxyl groups in phenolic compounds. J. Chromacogr. 19(3):564-571. Steele, R.. H., and D. L. Nilhelm.. 1966. The inflammatory re- action in chemical injury. I. Increased vascular permeability and erythema induced by various chemicals. Br. J. Exp. Pachol. 47(6)s612-623. Stefanko, S., et al. 1968. Histopathological picture of neural roots and spinal cord after injection of phenol into the spinal canal to remove spasticity. Reurol. Heuiochir. Po1.2:19-23. tefano, B., and P. Quirico. 1939. The pharmacological action of guaiacol. Arch. Parmacol. Sper. 67:190-206. Stein, C. D., and D. W. Cates. 1932. A further note on the action of phenol on the virus of equine infectious anemia. Am. J. Vet. Res. 12(47)s195-197. 1552. Stein, C. D., 0. L. Oateen, L. 0. Mott, and Y. S. Shahan. 1944. Action of chemical and physical agents on the virus of equine infectious anemia. Am. J. Vet. Res. 5(17):291-302. * 1552a. Steinke, R. D., and H. C. Paulson. 1964. Volatile phenols in cooked corn. J. Agric. Food Chem. 12:381. 1553. Steinber, R. H. 1970. Oral medi.ation for relief of sinus congestion. O.S. 3,515,781. 2 pp. 1554. Steiabrecher, X. 1973. Collabor.ive study of the Leterainat:cn of ar®onia as an index of decooposition in crabmeat. J. assoc.. Off. Anal. Chem.. 56(3)s598-501. 1555. Steinigen, 11. 1975. Analytical stud7 of liquid precaine preparacions. Pharm. Ztg. 120(31)s1119-1123. 1556. Sterling, Okunievski, and Rosnovski. 1930. Ccntribution to the study of epidemiology and diagnosis of trichinosis. J. Physiol. Path. Gen. 28(2):387-390. 408
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•1232. Parke, D. V., and 8. T. Williaos. 1953b. Detoxication. L::. Yetabolism of benrene. (a) For3~tioa of phenyglucuronide and phenylsulfuric acid from C-benzene. (b) !lecatiolisn of C1°-phenol. giochem. J. (London) 55:337-340. 1233. Parmentier, &. 1969. NoCa on the antiaitotic action of several phenals and aromatic smines in the mouse. C. R. Soc. 9io1. 144(7/8):585-586. 1234. Parmentier, R., and P. Dustin. 1951.. Experimental production of a particular anomaly of metaphase in malignant cells (TSree-group metaphase). Caryologia 4(1):98-109. 1235. Paatuska, I. C. 1961. Silica gel layer chromatography of phenots and phenol-carboxylic acids. Z. Anal. Chem. 179:355-358. 1236. Patakova, D., and M. Chladek. 1974. Antibacterial activity of thyme and vildthyme (essential) oils. Pharmarie 29(2):140-142. 1237. Patkovski, J. at al. 1967. Pbarmacologic properties of certain 1238. r derivatives of phenylathylamine and phenylmethane. Arch. L-mun. ther. Exp. l5:88-96. Patty, P.A. 1963. Industrial Hygiene and Toxicology. Vol. II. Toxicology. ~ohn Wiley and Sons, Inc. New York. 1546 pp. Paul, K. G., and T. Stigbrand. 1970. Four isoperoxidases from horse radish root. Acta Chem. Scand. 24(10):3607-3617. 1239. Paulson, C. D. 1975. Metabolic fates of herbicides in animals. Residue Rev. 58:1-111. 1240. Pavlova, Z. V., S. P. Sviridova, and N. E. Isakova.. 1971. Analgesia for incurable oncological patients by.chemical danervation. Ehirurgiya 47(8):93-96. 1241. Pavlaczyk, 1!. 1965. Effect of glucose and urea on the rate of phenol degradation (purification of phenol vastes) by Pseudomonas fluorescens. Acta Microbiol. Pol. 14(2):207-214. 1242. Paya, D. S. 1960. Routine analysis of phenals by gas-liquid chromatography. Chem. Ind. (London) 1090. . 1243. Pedersen, E.., and P. Juui-Jeusen. 1962. Intrathecal phenol in the treatment of spasticity. In: First Scandinavian syoposium on multiple sclerosis, 1962.Acta Neural. Scand. 38(Suppl.. 3):69-77. 1244. Pellegrini, C., A: Visea, and I. Papo. 1969. 7Secaanism of the analgesic action of intratbecally injected phenol solutions. Acta Neurol. 24(1):85-93. 1245. Pellerin, F., and R. Chasset. 1969. Decera?nation of phenols by ultraviolet speetrophotometry. II. Following conversion ?nto brominated derivatives. Ann. Phara. Fr. 27(9-10):57'.-574.. er 1245a. Pellmont. 1973. Unpubl. Rep. 1246. Peluffo, A. 1929. The lipasic action of saliva. C. R. Soc. 9io1. 100:113-116. Pemberton, J. N. 1971. Automated method for the deteraination of phenol in biological products. Appl. Microbiol. 21(4):763-765. 1247a. Pepper, J. M., and R. W. Fleming. 1978. Lignin and related compounds. V. The bydrogenolysis of aspenwood lignin using rhodium-on-charcoal as catalyst. Can. J. Chem. 56(7):896-8. 1247. 387 -
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* 1557. Stern, D. J., A. Lae, V. 8. McFadden, and K, L. Stevens. 1558. 1967. Volatiles from grapes. Identification of volatiles from eoncord assenea. J. Agric. Food Chem. 15:1100-1103. Starn, C. G., S. Land, R. R. Drever, and A. J. Carlson. 1559. 1950. Further chronic toxicity studies of hydroquinone. Fad. Proc. 9(1):121. Sterner, J. S., S. dmea, and D. W. Fasaect. 1949. Comparison 1560. of the acute toxicity of hydroquinone and same related di and trihydrozy b.nsenes. Fad. Proc. g(1):334. Stevens, R. L., J. Bomben, A. Lee, and W. S. McFadden. 1966. Volatiles from grapes. Muscat of Alexandria. J. Agric. Food Chem. 14(3):249-252. evens, 8. E., A. K. Ronan, T. S. Sourkes, and E. K. Boyd. 1943. On.the expectorant actiom of creosote and the guaiacols.. Can. Med. Aaaoc. J. 48s124-127. 1562. Stewart, F. G. 1930. Tha excretion of phenols in mental patients. J. Heat.-Sci. 76(313)r277-283. 1563.. Stawart# W. A. et al. 1963. An experimental evaluation of the effeets of sabarachnoid injection of phenol-pantopaque in catai a biatologf-.aI study. J. Keuroaurg. 20a64-72. 1564. Steyn, D. G. 1929. A note on the symptomatology of phenol poisoning in sac-tp induced by certain dips. Union S. Africa Dept. Agric. 15:.. Annu, Rep.$ Dir. Vet. Serv. 2(5/9):657-658. 1565. Stackdale, li., and H. J. Selwyn. 1971. Effects of ring substituents on the activity of phenola as inhibitors and uncouplers of mitochondrial respiration. Eur. J. IIio-Chem. 21(4)s565-574. 1567. Stoffelsma, J., C. Sipoa, D. R. IIettenes, and J. Pypker. 1968. New volatile eomponents of roasted coffee. J. Agric. Food Chem. 16(6)r1000-1004. 1568. Stoichev, St., G. Zolotovich. 1C.. Hachev, and K. Silyanovska. 1967. Cytotosic effect of phenols, phenol ethers, furan derivativea, and oxides isolated from essential oils. C. R. Aced. Dulg.. Sci. 20(12):1341-1344.. 1569. Stojcev, S., C.. Zalotovitcb, T. 8achev, and R. Ljaaoveks. 1967. Investigations on the cytoxic effect of phenols, phenol ether, furan derivatives and oxides. C. R. Acad. 8ulg. Sei. 20(12)t1341-1344. 1570. Stokinger, S. E. et al. 1963. threshold limit values for 1963. J. Ocoup. Ksd. S(10)s491-498. ! •1571. Stoll, H.., H. Winter, F. C.utscbi, 1. Flameat, and H. Villhalm. C_{ 1967. Research oa aromas. ZIII. Coffee aroma. 8elv. Chim. Acta 50(1):628-694. 40 9
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TABLE I THE EFFECT OF TREATMENT WITH SOME PHASE 11 SUBSTRATES ON SOME PARAMETERS OF RAT HEPATIC XE]OBIOTIC METABOLISM TreatmenPa Diphenyl- 2-Phenyl- Phcnyl- Control acetic 4,Hydroxy-. I-Naphthyl- Paraceta- propionic n-Propyl acetic Parameter values acid Guaiacol biphenyl 2-Naphthol acetic acid mol acid gallate acid Relative liver weight (g liver/l00 g body wt) 4.7 105 100 100 100 105 100 100 105 105 Microsomal protein (mglg of liver) 32 100 105 100 105 110 95 100 90 100 Cytochrome P450 (AE 450-490 nm/mg microsomal protein 0.075 100 95 100 105 110 95 110 100 95 Ethylmorphine N-demelhytase (µmollhr/g of liver) 24 110 95 90 95 95 100 105 95 95 Biphenyl 4-hydroxylase (Fmollhr/g of liver) 5.7 95 105 110 95 95 95 105 90 95 Aniline 4-hydroxylase (pmoVhr/g of liver) 2.1 100 115 105 105 n00 , 105 110 100 95 4-Mcthylumbelliferyl glucuronyltnnsferase (µmoVhrlmg microsomal protain) .9 40" 45, 5 50° 65` 65' . 15 45° 5 I-Naphthyl glucuronyl- transfense (µmoVhr/mg microeomal protein) 4.3 110 125r 115 110 120, 1'_0` 95 125, 103 • AII compounds were administered by daily ip iNections for a period of 7 days at dosages described under Methods. • All reaults are expressed as percentages or control values, each group comprising eight control and six treated animals. ^ p< 0.05: 8u.deni a r teM. W A
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248. 249. 250. 271. 253. 254. 254a. e 255. 256. *257. 258. 259. '250. 260a. 261. C~.I 262. F Cerruti, C. G. 1929b. Coccidia, the bovine plaque. Ann. Soc. Helge Nad. Trop. 9(4)t403-417. Carve, L. 1953. The effect of disinfectants to cysts of Lamblia intestinalis. Cask. Parazitoi. 2s17-21. Chabaud, A. 1942. Alteration of Hansen's batillus by fixatives. The protective role of phenic acid. Ann. Iast. Pasteur (Paris) 68s106-113. Chabrol, R. 1930. Cholagog action of phenols. C. 1. Soo. Ziol. 104:43-45. Chabrol, E., R. Charonnat, H. N.ximin, and 8. Waitz. 1931. Cholagag action of guaiacol derivatives. C. R. Soc. 8io1. 10731240-1243. Chabrolin, C. 1940. Chemical function of certain compounds versus their selective toxicity to small phanerogam plants. C. R. Aced. Sci. (Paris) 210s262-266. Challis, 3. C. 1974. Rapid nitrogacion of phenols and its implications for health basards from dietary nitrites. Nature 244s466. Chambers, D. L. K. Ohinata, N. Fujimoto, and S. Kashivai. 1972. Treating tephritids vith attractants to enhance their effectiveness in sterile-release programs. J. Econ. Entomol. 65(1):279-82. Chamburkar, P. E. 1975. Effect of flavoring eils on preservative concentrations in oral liquid dosage forms. J.Pharm. Sei. 64(3):414-417. Chang, I. W. 1972. Study oh the threshold limit value of benzene and early diagnosis of benzene poisoning. J. Cath. Yed. Coll. 23s429-434. Chang, S. 1976. "21eat Flavor" Iat. Flavores Food Addit. 7: 77-80, 86. Chaterjee, H. N. 1946. A further 5iochenical study of the blood of cholera patients. Trans. R. Soc. Trap. ]Sed. Hgy. 39(4)s321-326. Chen, L., and P. Issenberg. 1972. Interactions of some wood smake cnmponents vith e-maino groups in proteins. J. Agric. :ood Chem. 20(6)s1113-1115. Chea, T. C., W. W. Nauar, and 1- E. Levin. 1974. IIdentification of major high-boiling volatile compounds produced during refrigerated storage of haddock fillets. Appl. Nicrobiol. 28(4)s679-680. Chenicek, J. A., and W. L. Cox. 1965. Antiozonants for rubber. Cer. Pat No. 1,206,150. Cherkasovs, L. S., and L. A. Liseako. 1938. The effect of ordinary and chemically preserved ensilage on metabolism in the animal organism. LI. Siochem. J. (Ukraine) 211:213-222. Chihara, :1. 1953. Types of drug syneryism classified by the concentration-action eurve. polia PharmacoL. Jpn. 51:531-544. '32u
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0 .. 1800. 1801. ' 1802. 1803. 1804. '1805. '1806. 1807. ~ 1808. 1809. 1810. 1811. Williams, R. T. 1959. Detoxication Meehanisms - The Metabolism and:Detoxieation of drugs, toxic substances and other organic compounds. 2nd ed. Chapman and Hall ltd. London. 796 pp. Wills, E. D. 1969. Lipid peroxide formation in microsomes. Relation of hydroxylation to lipid peroxide formation. Biochem. J. 1~13(2): 333-341. 1 Wilson, t. A., and tats. 1972. Review of literature on chicken flavor and report of isnlati0n of several nev chicken flavor components from aqueous cooked chicken broth. J. Agric. Food Chem. 20(4):741-147. Winter, H. 1961. Odor and eonstitution. ZZC. Homologues and analogues of p-hydrox7-phenyl-1-butanone-3(^raspberryketone"). Helv. Chia. Acta 44(7):2110-2121. Winter, H. 1965. Flavoring composition. 9.S.,3,222,187. 3 pp. Winter, H., and R. Floati. 1972. On the flavor of the yellow passion fruit. (Passiflora edulis f. flauicarpa). Helv. Chim. Acta. 55(6):1916-1921. Winter, H., and E. Sundt. 1962. Analysis of the aroma of raspberries. I. Volatile carbonyl constituents. Helv. Chim. Acta. 45:2195-2211. Withell, 8. R. 1942a. The effect of change on concentration on the relative activity of.phenol and paraehlorovetacresoL.Qpart. J. Pharm. Pharmacol. 15(3):301-313.. Withell, E. R. 1942b. The evaluation of bactericides. J. Hyg. 42(4):339-353. Witter, R. F. 1944. The metabolism of moaobronoben:ene, benzene, benzyl chloride and related compounds in the rabbit. 186 p. Woidich, H., H. Cnauer, and 6. Calinoviky. 1967. Thin-layer chrosutographie separation of some food preservatives. Z. Labensm.-Onters. Forach. 133(5)s317-322. Wojeik, J. 1975. Application of the method ofmotor accivity measurement in white rats as a test for toxicological assessmenc of phenol. Rocm. Panstv. Zakl. Hig. 26(1):21-33. Wolcott, C. Y. 1943.. Phenol as a te=ite'repeller.t. Pests 13(9):26. I3TJa. Wong, X.P. and T.L. Scurkes. 1966. Heta:oliam ai•:an:llin and re?a:ec substances ?n the rat. Can. J. 9iochea. =1:65c-6}1'• 1813. Woo, C. H., S. K. Ris. and S. H, 8in. 1967. Com,nlex interaction oE acacia and sodium alginate with certain presecvacifes (spectrophotomecric studies). Yakhsk 'oe;i 11(3-4):27-32. 1814. Woodard, C., E. C. Hagm, and J. L. Radomski. 1949. Toxicitl of hydroquinone for laboratory animals. Ped. proc. 8(l):348. 1815. Woods, M., K. Right, J. Hunter, and D. :urk. 1953. E_fects of insulin on nelanoma and brain metabolism. 3iochim. Biophys. Acca 12:329-346. 1816. Wrabets, I., and W. Sassanberg. 1961. Cas-chronutographic analysis of phenol-cresol-xylenol mixtures with special attention ta the o-cresol content of tritolyl phosphate. Z, Anal. Chem. 179:333-342. 425
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v .... .P:...~-. ~ae.~.... ..- .. .~... .. . . ... - ....... .............. . ....... --.. e.ne.. ....:.e~.... ~ '!~'...:~..~u.V....~3 . .. . I Y~ ~~SOBSTANCE ~ . :. NO. . - . ..~._ . . . PODD SOURCE . 'DBTBCTED BY - ~ P.P.N. B -BSTINATE _ ~ ~ QO!@ffiITS ~ RBP.-~ - GUA7AGOL -~. (continued) -;~~~ = 25 Tea leaf oil ~ ~ GLC ~- ~ Characterized by GLC retention time and co l ' 571 ~ . . spar son: to authentic sdiple -.--"`-_-~ Tca~oil GLC 45;100 ~ Characterized by GLC retention ti.e and 571` comparisontoauthentic-sample Tea oil, black ~ - GLC -- Char a cterized.by~Gl.Cietention ti.o and 571 .- " ~ conparison ~to auttienticzdsaple -- - ~~~ .. . ~ .. i _ Tea ~oig'rEen~ ~ ... GLC~. --- ~.~ti `Cha efer?sad by CLC retent~3o n tie-end 7i _ complr ison -~ to authentic sample - ' . . Tomato . ~ 615, IR, GLC ~ -- - Charaeterizad by:'spectioscopy end-:. 1732 - ~_.... . . . . - comperison=to euthentic sa.ple ~ Tomato ~ . ~ . 1R, GLC Characterized by C[,Cretent on_ti.e, 829 ~~. ~ .. ~ . - . . ~speettoscopy andco'.parison"to C) enilEdample , ~. .. . Tomato ~ - . NS, IR, GLC, . , Characterized by GLC retention ti.e .,2i6 . Isolation - . -: and comparison to spectrosco py . _ eui~~nticsflmple , -- . - ~ ~ . Vanilla pals, ~ tsolation~ ~ .. . -- Ciia racterized by.cmparison to authentic ~155- . Bourbon ,~ ~ . . semple and.chemically chardcterized ~ii~skey, dapanese UV;~ GLC ~ . ' 0.001 Cfiarec terlxed by GLC retenfion tine, 1pq2-. -. s- .. 3pectroscoPy end'coeparisonfo authentic - Sample-- . . . - - - - ~~ Wisk"ey Japanese ~, Grf 0.006 ..,.. , ~ Charatiterized by GLC,retention tidc, :., 1176~~ sspec,ttr+aoseopy and eoparison to.outheniic dt- g - ~ ~ ` , t ~. . ~ ~ . . . e.. w . = tizsii~s ~ ~ ~ _- ' . In
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a i ..._..._ ,..-.....-::::-__:::.. ` 8'7119252 ~ C_~
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1456. Sedivec, V., and J. Flek. 1970a. Determination of toxic substances and their metabolism in biological fluids by gas chromatography. It. Phenol Cresol ia urine. Collect. Czech. Chem. Cox,un. 242-230. •- 1457. adivec, V., and J. Flek. 1970b. Deteraination of toxic substances aad their metabolites in biological fluids by gas chromatography. III. Occurrence and detetaination of guaiacol in urine. Prac. Lek. 22(5):176-181. 1458. Seeboth, H. 1963. Thin-layer chromatographic analysis of phenols. Monatsber. Dent. Akad. Visa. Berlin 5(11-12):693-698. 1459. Seaboth, 8. 1965. Thin-layer chromatographic analysis of phenol (mixtures). Fortschr. Vasserchem. Ihrer Creazgeb. 2:128-133. 1460. Seeman, P., S.: S. Chen, 14. Chau-Wong, and A. Staiman. 1974. Ca1- eium reversal of nerve blockade by alcohols, anesthetics,tranquiliters, and barbiturates. Cam. J. Physiol. Pha~col. 52(3):526-534. 1461. Segal, 8. L. 1953. Kinetic studies on the phenol-sulfate con- jugating system of rat liver. J. giol. Chem. :13(1):161-170. 1462. Seglen, P. 0. 1973. Preparation of rat liver cells; III. Enzymacic requirements for tissue dispersion. Exp. Cell Res. 82(2):391-398. r 1463. Seide, C. 1941. Blood poisoning effects from catalase. Siochem. Z. 308(3):175-186. 1464. Seki, T. 1960. Chromatographic separation of derivatives of phenol on cation-exchange resin. J. Chromatogr. 4:6-10. 1465. Semenchenko, L. V., and V. T. Kaplin. 1967. Determination of monohydroxyphenols in waste waters. Zavod. Lab.. 33(7):801-804. 1466. Semenenko, A. D. 1963. IIsa of functional electroencephalography in studying the effects of unnoticeable concentrations of atmospherieimpurities on humans. Gig. Sanit. 28(7):59-55. 1467. Sentein, P. 1958. The effeet of phenol on oitosis in the segcent- ation of amphibiam eggs. Acta Anat. 34(3):201-234. 1468. Seoane, E., and A. Carnicer. 1967. Identification of natural phenols by microseala reactioas of t5ei: functional groups. Yic:o- chem. l. 12(3):291-306. 1468a. Seto, T. A., and W. Keup. 1969. Effects of alkylmethoxybenzene and alkylmethylenedioxybenzene essential oils on pentobarbital and ethanol sleeping time. Arch. Int. Pharmacodyn. Ther. 180(1):232-40. •• 1469. Seutter, E., and A. 3. :4. Sutorius. 1972. Quantitative analysis of hydroquinone ?n urine. Clin. Chia. Acta 38(1):231-232. 1470. Sezerac, A.1961.. The detection of phenol.by paper chromato- graphy with the aid of.phenitrazole. Bull. Soc. Chim. Fr. 1193. 1471. Shank, 6.. E., and C.: L. Eoagland. 1946. A modified method for the quantitative determination of thettymol turbidity reaction of serum. J. 8io1. Chem. 162(1)t133-138. 4132 Cb N ..~, . .,r: •. -------- ---------
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1354. Hosenstein, C., and I. Levin, assisted by H. Levin. 1935. The bactericidal and antiseptic action of preservatives frequently used in biological products, and the effect of these preservativeson the potencies of the products. Am. J. Hyg. 21(2):260-279. 1355. Hosentbaler, L. 1956. Identification of organic compounds. Pharm. Ztg. 101s150-152. 1356.. Hotenbarg, T. S. 1974. Correlation between the toxicity of chemical agents and their inhibitory action on isolated mitochondria. Byull. Eksp. Biol. Hed. 78(7):65-68. 1357. Roth, S., and P. Sasaun. 1972. 1l:mbrane concentrations of neutral and positive anesthetics (alcohols, chlorpromazine, morphine) fit the Heyer-Overton rule of anesthesia. Negative narcotics do not. Biochim. Biopbys. Acta 255(1);207-219. 1358. Rothbaecher, H., and P. Suteu. 1975. Hydroxyl compounds of caraway oil. Planta Med. 28(2):112-123. 1359. gothbaacher, H., A. traus, and P. Sutau. 1967. Phenol components in Pamanian Mentha piperits oil. Herba Hung. 6(2):171-176. 1360. Roubal, J., and V. Vasak. 1966. Tentative hygienio evaluation of technical-grade trierphosphates. Cesk. Hyg. 11(4)s221-225. Rovati, L.# and E. Calante. 1960. Pulmonary elimination of guaiaeol and some of its derivatives. Bo11. Soc. Ital. Biol. Spar. 36: 1658-1659. 1362. 8.ovensky, J.,et al. 1964. Study on the assessment of the barrier following damage with phenol. Cesk. Derm. 39:94-99. 1363. Rovesti, P. 1961. Aromatic therapetucs of thyme essences. giv. Ital. Bssenza Profumi, 43(3)t117-125. 1364. goshin, J., 8. L. Soderstrom, and S. C. Brooks. 1974. Specificity stndies on bovine adrenal estrogen sulfotransferase. J. Biol. Chem.. 249(7):2079-2087. 1365. 1366. 1367. * 1368. 1369. 8osin, 8., F. Magora, C. C. Robin, and A. Magora. 1970. Treatment of spasticity by phenol block. Isr. J. Med. Sci. 5(4):555-560. Hucinska, 8. 1968. Myotropic activity of sympathomimetic ®ines. Acta Physiol. Pol. 19(3):417-428. Budol'fi, T. A., B. I. Sharapova, and V. I. Lushehik. 1964. Gas chromatography of cresol isomers. Zh. Anal. g.him. 19(7):903-905. Ruedemann, A., and il. B. Deichmann. 1953. Blood phenol Level after topical application of phenol-containing preparations. J. Am. Med. Assoc. 152:506-509. Rusch, H. P., D. Bosch, and B. K. Boutwell. 1955.. The influence of irritants on mitotic activity and tumor formation in mouse epidermis.. Acta Unio Iat. Contra Cancrum 11:699-703. 395 : 11
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Ilyud''kovu. A'.M.,:und N.N. Medvrdrv - VuprosyOokologii, Nn,2: I 951i. 201. Cnlobcv A-.A.. coKl E.1. Lyubli'na. --pigicna'1'ruda, No.4:26. 1962.. Shabad L.M. Ocherki eksperimental'noi onkolbgii (Outlines of Experimental Oncology). Moskva. 1947. Received 27 October 1962 108
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b „ U-GLIII'UatINIC Al'IU I'A"I'IIwAY Rli(R11.A'1'I(IN stimulated 4-methylumhcl I i feryl glucuronyl- Iransferase; 4-hydroxybiphenyl, 2-phenyl- propionic acid,.and phenylacetic acid were without any effccl.(Tnble I). Urinary Swdi<'.r None of the compounds investigated had any significant effect on the urinary ex- cretion of either D-glucaric acid,. L-gulonic acid, or xylitol after either 2 or 7 days of treatment (Table 2). However, with the ex- 375 ceplion of phenylacetic acid, which is not conjugated with D-glucuronic acid in the rat (James et al., 1972;. Dixon et al., 1977d), all of the compounds studied signifrcantly stimulated the urinary excretion of total (free and conjugated) D-glucuronic acid after 7 days of treatment (Table 2). We measured the urinary excretion of D-gluc- uronic acid only as total acid and not as separate values for free and conjugated acid in order to avoid errors due to the break- down of labile glucuronides in the urine prior to analysis which would increase the TABLE 2 THE EFFECT OF TREATMENT WITH SOME PHASE II $OBSr11ATES ON THE URINARY EECRETION OFCOMFONENTS OF THE D-(JLUCUAUNIC ACIO PATHWAY IN THE RAT Compound 2-Naphthol 1•Naphlhylaceticacid Paacetamol (acetaminophen) 2~Phenylpropionicacid a-Propyl gatlale PAcnylacctic acid Urinary excretion u perccntage orcontrol• Days of Ireatment n-Glucaric acid L-Gulonic acid Xylitol Total o-glucurooic acid 2 70 110 103 135' 7 85 80 100 135' 2 120 90 110 190• 7 115 90 115 165' I1t1 IIRI 71/ N5• 7 80 75 80 240' 2 60 80 85 240' 7 105 80 95 285• 2 125 90 125 170` 7 90 loo 125 15Y 2 100 105 85 190. 7 105 110 110 170' 2 60 90 103 120 7 75 90 115 190• 2 95 125 120 200' 7 115 90 IOo 160t 2 105 85 90 110 7 100 85 l00 110 65 270 45 1,200- • AB results are expressed as percentagef of control values, each group comprising eight control and six Ireated animals... • p c 0.051 Studeni s r lesl. ~ In units of µg metabolite per mg of crealinine.. 87119242 i
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r Tara, S.., and Trouard-Riolle. 1948. Phenolic marasmus. Arch. Mal. Prof. Mad. Tray. Secur. See. 9:288-292. 1618. 1619. 1620. Tarr, $. L. A. 1947. Chemical disinfection and corrosion pre- vention. J. Fish. Eas. Board Can. 7(3):101-115. 1625. Tamhane, g. 0. 1971. Estimation of thymol•dn analgesic balm. Indian J. Pharm. 33(4)t70-72. Tank, T.. M., at a1. 1963. Imtrathecal injections of alcohol or phenol for relief of intractable pain. Cleveland Clin. Q. 30:111-117. Tanker, M. 1963. A colorimatric method for the determination of the total phenol content of thyme oil. Istanbul Univ. Tip Fak. Mee. 26(1)126-31. Tasaka, II. 1957. 8istamine release by chemical substances from skin of the dog. Okayama Igakkai Zasabi 69:2853-2868. , Tatematsu, A., T. Nadai, E. Yoshizumi, and T. Coto. 1969. Analysis of mixed drugs by mass pactrometry. II. Food additives 1. Analysis of preservative agents. Shitsuryo Bunseki 17(3)s773-786. Tavberidze, A. I., M. I. Coryaev, A. D.. Dembitakii, C. I. Krotova, and E. B. Chelishvili. 1969. Dynamics of the accumulation of essential oil from Saturaia spicigera. Tr. Sukhum. Optyn.Stan. Efirmaslichn. Tul't. 8s151-157. 1626. Tavberidza, A. I., N. Z. Yakobashvili, E. B. Chelishvili, M. I. Coryaav, and A. D. Dembitakii. 1969. Use of the essential oil in spike-bearing savory. Masl.-Zhir. Prom. 35(2)224-25. 1627. Taylor, H. D., and J. E. Austin. 1917. Toxicity of certain widely used antiseptics. J. Exp. Med. 27:635-646. Taylor, J. M., P. M. Janner, and W..I. Jones. 1964. A comparison of the toxicity of some allyl, propenyl, and propyl compounds in the rat. Toxicol. Appl. Pharmacol. 6(4)s378-387. 1629. 1630. 1631. 1632. 1633. ~ ~. E i Tedeachi, C. G., and A. De Cieco.. 1953. Action of me-cresol, p-cresol, and xyleaols against vita®in E. Quad. Nutr. 13s166-173. Tedeschi, G. G., and A. Da Cicco. 1954. The antivitamin E action of thymcl, carvacrol, and guaiacol. Boll.. Soc. Ital. Biol. Sper. 30:727-729. Teisinger, J., and V. Fiscrova-Bergerova. 1955. Phenol in blood of vorkers handling benzene and phenol. Prac. Lek. 7:156-160. Telegina, i. A., and V. I. Boiko. 1972. Vitiligo-like changes of the akin in workers producing phenol-containing additions. vestn. Dermatol. Venerol. 46(l):31-34. Tenta, L. T. 1976. Topical composition for treatment of eborrbeic keratosis. U.S. 4 pp. 413 ~~:
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~I. volume 14 Drugs on Ciliary Movements 303 Number 4 .. 9. Linton, C.S. Resistance of the upper aespitatory mucosa to infection. Ann. Otol. Rhinol. and Laryngol., 42 : 64,1933. 10. Lommel, F. Zur Physiologic and Pathologic des Flimmerepithels der Atmungsorgane. Deutsches Arch. Xpn. Med., 94 : 365, 1908. 11. Perrine, R.L., A.H. Ibrondson and M.L. Tainter. Effects of dental detergents on ciliary activity. J. DenralRes., 18 : 81, 1939. 12. Prootz, A.W. The effects of certain drugs upon living nanl ciliated epithelium. Ann. Otol. RhTnol, and Laryngol., 43 : 450, 1934. 13. Proetz,A.W. Further experiments in tho-action of drugs on the nasal mucosa. Arch. OtolaryngoL, 30 : 509,1939. 14. Proetz, A.W. Essays on the Applied Physiology of the Nose. Annals Publishing Company, St. Louis, 1941. 15. Proetz, A.W. Essays on theApplied Physiology of the Nose. 2ndEd., Annals Publish- ing Company, St. Louis, 1953. 0~ hation i i f hlor te for b[ f ti d d li d h i i i eon nue a m n s o c a a es a oo c potent t 16. Ric ardson, A.P. Tox - and tissues. J. Pharmac. Exp. Ther., 59 :101, 1937. •
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respiratory arreat:. The maximum t 1 rance dose LC jwas 2.03 1 of the guaiacol vapors; t e ahsolutely lethal dose (LC4q . 7.19 mg/1. As a result of treatment of the experimental data by the Kerber method (M. L. Helen'kii) the concentration of gualacol vapors causing the deaths of 50% of the experimental animals (LC50) was determined. It proved to be equal to 7.57 mg/l. Acute toxiehy of gualacol vapon by inhalatlon (2-hour exposure) Number of mice which died Concentrallon. Average Numbaof total ibc PerhN folliwing range cmcenmarlun ml« In ew JurlnB the experimem n2 which (mg/1) . (in mg/I) expedmena t0e died esp<timen[ 1 hmur 18 haun 1-4 days I,BB-'j06 2,09 10 - - 0 9.2 - 60 8.6 10 - - _ 1 ` - 1 8.13- 6.7z a.42 1o - s 2. - s 7.74- 7.78 1.76 10 - 6 - - 6 9.78- as 9.57 10 - 7 - - 7 12.48-12L89 12.64 10 3 6 - - 9 17.07-17,81 17.19 10 1 9 - - 10 In the study of the morphological changes caused by guaiacol vapors, * disorders of vascularization of the internal organs and small hemorrhages in the meninges and brain matter, heart muscle, lungs (subplcurally and intra-alveolarly) and kidneys were found. Degenerative changes were rarer: cloudy swelling of the heart muscle and (in some preparations) focal fatty degeneration of the liver parenchyma were found. An analysis of the pathological changes found in the dead animals permits us to suppose that a disorder of innervation of the vascular system playss the leading part in the pathogenesis of the circulatory disorders described, although permeability disorders of the blood vessel walls cannot be ruled out. Since the permissible concentration has not been determined for guaiacol vapors, we, on the basis of experimental data, calculated, its approximate value from A. A. Golubev's and E. I. Lyublina's formula: where PC Is thepermisslbleconcentration (in mg/1); LC50 is the lethal contentration for 50% of the mice after inhalation of the substance for 2 hours (in mg/i);- M ie the molecular weight. Calculation by this formula showed that the approximate value of the permissible concentration of guaiacol vapors is 0.02 mg/1. We made a study of the local effect of guafacol resin on 5 rabbits. The ointment containing 0.5, 5 and 10% resin prepared in vaseline and lanolin was applied to 3 areas of the abdomen of the rabbit (fromwhichthelmirhnd been shaved) and left on the skin for 4 hours. Over a 2-wcek period such applications weremadc three times, but no visible sktn changes were found. Forthepurposeofstudytngtheblastomogeniceffectoftheresin, twoseries of experimentswere performed (each series on 80 3-4-month-old mice,of the • The author Is gra[cful to Pm(cnu S.S. Vail'ior hh eunsuilatlve asshnnce Ihdhe evaluaNen of the pathologlcalehanges 106 8f7119256
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af ,a o. eartocic0lo p:xl studies iR Jene chloride incorporated in the or adminixtered by repented inhala- 1r.-.drJl Pr..,prrL 21, is_43. NRINr, p. J.. ANaPARa, C. N:(197g); risk c*imation for chlorofurm: an p.PA'a pnrednres. pund Cnamrt. 1-514. . L. (1969)..Tumors promoting agents rcinogenesis. Prng. Erp..Tamor Re.r. \ND CAPUTO, A. (1977)- CarCino- s on vinyliden<chloride. Environ. r. 21, 45-47: OLErT. W., AND FORS'rER, W. (1977)- pharmacokinetic pammetcn an the+t 1. Clfn. PGarmacol•.15, 572-575. IO]ICOLOGY AND APPI IP.n. PIIARIIACOI.OfI' 52,.37{-j7g (1980) (5v~ C, ~J re1N ~ 1996' The Effect of Treatment with Some Phase II Substrates on Hepatic Xenobiotic Metabolism and the Urinary Excretion of Metabolites of the D-Glucuronic Acid Pathway in the Rat BRIAN 0. LAKE, RONAI D C. LONGLAND, ROSALYN A. HARRIS, MICHAEL A. COLLINS, IRENE A. HEROD, AND SHARAT D. GANGOLLI TLe Britirh Industrial Biological Research A}sotiation, WoodmanstrrnFRoad, Cars.4ahan, Surrey, SMS 4DS• England ' Rreeiltid May 10, 1979, accepted Ortn6rr, 110. 1979 The Effect of Treatment With Some Phase 11 Substrates on Hepatic Xenobiutic Metabolism and the Urinary Excretion uf Metabulites of the o-Glucuronic Acid Pathway in Ihe Rat. LANE, B. G., IANOLAND, R. C.,AARRIS, R..A., COLLINS, M. A., HERUD,I. A.,.AND GANOOLLI, S. D. (19xa1. Tr..riFal. Alq•!. Plamnnrul. 52, J71-17g. InvcstigalionsIrave 6een conduaed to study the relationship between the activities of certain enzymes of Phase I and Phase II hepatic xenobiotic metabolism and the urinary excrelion of some Inetabolites of Ihe o- glucuronic acid pathway. Male Sprague-Dawley rats were treated for 7 days with daily ip injections of either diphenylacetic acid,.guaiaco4 4-hydroxybiphcnyl. 2-naphthol. 1-naph- thylacetic acid,paracetamol (acetaminopheN, 2-phenylpropionic acid, n-propyl gallate, or phenylacetic acid. None of the compounds administered had anyatfect on a number of parameters of hepatic Phase 1 xenobiotic maabolism, namely mixed functionoxiduc enzymc activities, cytochrome P-450 and the microsumal content of protein. However, some of the compounds did stimulate hepatic microsomal UDP-glucuronyltransferase activity. While none of the compounds stimulated the urinary excretion of either lucaric acid; L-.SulDaic acid,orx ' allafthecompoundsexceptphenylacedcacidincreasedtheurinaryexcretion of total (free and conjugated) D-glucuronic acid. The results indicate that when hepatic xcnobiolic coq'rygative activity is increased without the stimulation of mixed funetion oxidascs and cytochrome P450, the urinary excretion of certain metaholites of the n- glucuronic acid pathway, other than n-glucurunicacid, remains unchanged. The enzymatic processes by which xeno- biotics are metabolized in the mammalian liver may be broadly divided into two categories (Parke, 1968; Williams, 1967). Frst, xenohiotics.are subjected to Phase I biotransformations, which include the re- actions catalyzed by enzymes of the cyto- chrome P-450-dependent mixed function oxidase(mfo) complex (Mason ef al., 1965),, and then the resultant metabolitess may be conjugated by Phasell reactions withh various endogenous acceptors such as o- glucuronic acid, sulfate, acetate, gluta- lhione, amino acids,.etc. In the case of the important Phase II pathway of glucuronide formation the microsomal UDP-glucuronyl- transferase enzyme (EC 2.4.1.17) catalyses the tnmsfcr of n-glucuronic acid from thc coenzyme UDPGA to the exogenous ac- ceptor. UDPGA is synthesized from either D-glucose or D-galactose by enzymes of the D-glucuronic acid pathway (Burns ef al.,. 1960; Conney ePal., 1961). In recent years a number of studies have been devoted to the development of non- invasive methods for the detection of in- 371 0041-OOgX/80/030771 -08S02.0010 c^py^Da o NUly Acadsek Fn.. Inc. AII lyhk of rtpoavclion's .ny rarn rt.meYL
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:r~- Mus 7,57g NG - R-19 an- ' t~UD- /kR. licc, vns Ibits inen tion CIIAIAIK)1  ACUTE TOXICITY: TERRESTRIAL LIFE-ORAL LD50 Sµrin (Jrnl 1a)ftl (u,J,d[y~J lirf Rat 725 R-~ TEST CONDITIONS Administered by gavage. RESULTS Tox symptoms included: marked depression,, comatose on higher doses. rime of death after doeing ranged from I H to 4 D. Dose range 302-1.740 mg/kg. Rat 1,500 . R32 Abl. 4AggR32  ACUTE TOXICITY: TERRESTRIAL LIFE-INHALATION LC5D ' sprr"vr LC10(mglw' ') Dsrnl~'m. (H)Ref  ACUTE. TOXICITY: AQUATIC LIFE-FRESHWATER FISH LC50 Sp.dn n,nn6,n. (11)rASn pxxgRJ (.Kll J. Rr/ Perch 48 7g-8g R3  ACUTE TOXICITY: AQUATIC LIFE-FRESHWATER MACROINVERTEBRATESEC50 Sprr/n. )ImxduuillP F.CS/1.turi,dlJ 115('~ C.J.IrnXs/J /iq Water Re¢ (Daphnia magna) 48 25.9 R-23 TEST CONDITIONS Static test. Total obmrratio¢ time of RESULTS Data reported aa 2.I18F. 4 M. 96 H.  ACUTE TOXICITY: AQUATIC LIFE-OTHER FRESHWATER STUDIES Spttitt Duration (H) Effid Lrurl.(uvg/L) Ref flea (Daphnia magna, NG 5-15 R-I6 Water pulex) Dophnia RESULTS Daphnia magaa showed much less adaptation than D. pu)exlb 5-IS mglL ao¢u of guaiacol. as indicated by the survivability of the orgoniama. 0/ the 8@xperi- me¢tawilh D. pulex, theorganisms showed reduced num- bers in 4 experiments while a stimulating etlect of the guaiacol on multiplication was noticed in the other 4. No additional in/ormatimtwas given, 87119248 115 tng/L (when used asa fumig,lut). (R-25) N Oriental MUTAGENICITY: GENE MUTATION- NEGATIVE REF- i Guit fly: 3rd instar larvae LD50 = > 115 rng/L. (R- t1tENCES R-8, R-17, R-24. 25) PHYTOTOXICITY: TERRESTRIAL PLANTS 'freatment CARCINOGENICITY: NEGATIVE REFERENCES oP nondormant Winter Rye (Vyatka variety) seeds with NOV7DEC I%6 47 I • p
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376 LAKE ET AL. apparent excretion of free D-glucuronic acid. Furthermore, certain glucuronides may have beenn partially hydrolyzed by the J3-giucuronidase (EC 3.2.1.31) of the kid- ney prior to excretion in the urine.. DISCUSSION The results described in this paper demon- strate that the administration of a number of compounds which did not stimulate certain hepatic Phase I biotransformation reactions (i.e., levels of cytochrome P-450 and mfo activities) had no significant effect on the urinary excretion of o-glucaric acid, L- gulunic acid, or xylitul. I lowcvcr, all of the compounds studied, with the exception of phenylacetic acid, stimulated the urinary egcretion of total n-glucuro Wt. majontyof ChE Com-poun~ds employed in this study are known to form glucuronide con- jugates in the rat (Berenbom and Young,. 195 1; Dixon et al., 1977a,b,c; Jollow et al.,. 1974; West et al., 1956)) at least part of the increase in urinary total D-glucuronic acid may be ascribable to enhanced excretion of glucuronide conjugates. Indeed, phenyl- acetic acid, which is not conjugated with D-glucuronic acid in the rat (James et al.,. 1972; Dixon et al., 1977d), had no effect on the urinary excretion of total D-gluc- uronic acid. Of the compoundswhichs stimulated thc urinary cxcretion of u-gluc- uroniracid neither 4-hydroxyhiphenyl nor 2-phenylpropionic acid stimulated hepatic microsomal UDP-glucuronyltransferase ac- tivity. The observed lack of increased ac- tivity of the conjugative enzyme by these two agents may be due to a number of factors. For example, both xenobiotics may be guod substrates of the enzyme and thus the endogenous levels of the transferase may be sufficient to rapidly convert thecompotmds ton glucuronidcs in Ihe liver, Alternatively, they may be less potent stimulatorsof microsomal UDP-glucuronyl- transferase., measured in thiss paper as the unactivated enzyme, than the other xeno- biotics studied. In previous investigations the adminis- tration of inducerss of hepatic xenobiotic metabolism to the rat has been shown to stimulate the urinary excretion of both Ir glucuronicacid and other metabolites of the D-glucuronic acid pathway (Aarts, 1966: 8urnss et al., 1960;. Conneyer al., 1961;. Hunter, et al., 1973; Lake ef al., 1976; Marsh and Reid, 1963; Notten and Hender- son, 1975). However, the present data, in agreement with the observations of others (Hunter and Chasseaud, 1976), wouldsug- gest that hepatic conjugative activity may be increased without any overall stimu- lation of the D-glucuronic acid pathway. Tlius, allhough.lhere was no increase in the urinary excretion of either n-glucaric acid, L-gulonic acid, or xylitol, more UDPGA was presumably made available for the UDP-glucuronyltransferase catalyzed for- mation of glucuronides. Furthermore, the lack of stimulation of the above three o- glucuronic acid metabolites correlatedwith the lack of effect of the Phase II compounds administered on the hepatic mfo measured. In conclusion, these observations on the short term administration of Phase 11 sub- strates lend support to the validity of ineas- uring a spectrum of certain urinary metabo- lites of the D-glucuronic acid pathway, e.g., D-glucaricacid, L-gulonic acid, and xylitol, as an index of the activities of hepatic microsomal. cytochrome P-450 de- pendent mfo in the rat. However, these tindingsshoulds be substantiated by studies on thestPectof chronic treatment with Phase H substrates on rat hepatic mfo activities and on the urinary excretion of D-glucuronk acid metabolites. ACKNOWLEDGMENTS 171is work fnrnu pan uf a fen¢arch project s{wmsant by the UK Ministry of Agriculture. Fisheries sd Food to whom our thanks are due. The resulas d the researchare the property of the Ministq d Agriculture. Fisheries and Food and are Cro.f Copyright. The skilled technical assistance of Wr. C. R. Stubberfieldis gratefully acknowledged. 87119243
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378 I,AKfi !:1' AI.. MARSU, C. A. (11x,)b), MalaMdivn of o-gl6curnon- lactum: in mammaliun syxtents. 3. Furthcr studies of i,glucurumdla:lonedchydrngcnase ur ml liver. /lindlrnr. J. N9,.IIIR-I Id. MARSH, C. A.. nNn Rlan, L. M. II9611. Chungcm in D-glnraric Icid exeretion induced hyslimululrNs ur axcPrbic arid hinsynlhcs{z., ilirrrlriNL BirrPlrrtc Arrn 78, 726-72N. MASON. H. S., NORTH, J. C., AND VANNESTE, M. ( I965). Micrusomul mixcJ-fnneiton oxidaliunx: The metabolism.ofxenobiotics: Fed. Prw•: Fed. ANlrr. SNr. ErA. Biof: 24, 1172_I IeO. NAKANISHI S., MASAMUNI. E., TSUKADA, M., AND MATSUMUSA, R. (1971). Kinetic studies for aniline hydroxylase after prolonyed ethanol treatment. Japan J. Pllarmnrvd. 21, 303-309. NnAl.r:,M. C., ANI) PANKa,. I). V. II973). N:ITecls of pregnancy on the metabulism of drugs in the ret and rabbit. BincherN: Phnrmorrd. 22, 1451-1461. NnnI:N'. W. R. P., AND IlnNnnRV)N, P. Th. (1975). Ahcml iunx in IKr ghn:urunic acid Ivnhw:ry canzcd hy various drugs.lrrr.J. f!luclrrm. 6, I I1-119. NNrT'eN- WI R. 1"., IIE:NIl1iNU1N, p. Th.. ANP KuYIY:IL L'Ir. M.A.I 10751. Stiimdalinn rA.lhc gluummic:a:id pulhwuyy ih.isnlnted rat liver cells by phemrh;IrAiDd. lnr. J. 11irNirem: 6, 713-71N. n.uiNA. 'I.. ANn: SAIN, K. (IWW). 'fhe carSon monoxWc-binding pigment of liver microsomes. 1.. Evidence for its hemoprotein nature.l.Btd. Chr/a. 239. 23711= 37R. PARKE,.D. V. (196N). 77/r BirN-hrulixrr,c nfFbrrfxn ('unqNrund.r. Pergamon, Oxronl.WEST,. H. D., LAWSON, 1. R.., MILLER„ 1. H., AND'MATNUaA, G.R. (1936). The rate of diphenyl in the ml. ArrG. Binrhem. Binp/rys: 60, 14-20. WILLIAMS, R. T. (1959): Drtoxil'urion Mrdmnianu,. 2nd ed.- Chapman & Hall.l.ondim. W D.uAMS, R. T. (1967). Comparative patterns of dru5 metabolism.Frr1..Prw': Ferl, Amrr..S.N, F_rp. BtiA. 26, 1029-J/D9. !k
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300. Datet at. Octntier 1970 Ind. J. PhysioL & Phamfac. Ringer was' 9.0, normal Ringer solution with its pH adjusted to 9.0 with sodium bicarbonate _ was used for comparison. - The hydrogen ion concentration of cigarette ash-Ringer was also adjusted to pH 7.5 using dilute hydrochloric acid and tested on ciliary movement. Normal Ringer of pH 9.0 produced a greater depression of ciliary movements than that of ash-Ringer of pH 9.0, while ash-Ringer of pH 7.5 produced slight stidulation of ciliary movements. DISCUSSION The ciliated-epithelium of frog's oesophagus is a very. simple, convenient as well as a very sensitive preparation for studying the effect of drugs on ciliary activity.: It has been shown to respond to changes in temperature, osmotic pressure, hydrogen ion conceotration,.diffetent anions andcations(3). In thapresent study oxygen and carbon dioxide have been found to-stimulate ciliary activity. The action , of carbon dioxide was direct and not due to change in hydrogen ion concentration. Carbon dioxide is known lo stimulate the respiratory eentre directly. It appears, therefore, that inhalation of fresh air in heslthyindividuals and of carbo- gen in diseased states is conducive for the health of respiratory tract. With the extensive use of gas as fuel for domestic as well as industrial purposes and with theincteasing use of gas driven vehicles, the inhalation of petrolgas has become common. Weak solution of petrol gas was found to have no significant effect on ciliary activity but in high concentration it was a depressant. This fact is of special significanceto those who are engaged in such industries or vocations where the personnel are constantly exposed to petrol gas. The relationship of cigarette smoking with the incidence of respiratory infections and bronchogenic carcinoma is well known. Hilding (6,7) reported that cigarette smoke stopped ciliary beat after 11 min of exposure. He thought that stoppage might have beenn produced by a substance toxic to ciliated cells, probably nicotine. In the present study, however, cigarette smoke was found to stimulate ciliary activity. The difference in the results is probably because Hilding (6.7) studied the direct effect of smoke, while in the present study a dilute solution of smoke was used. There is no doubt that cigarette smoke affects'ciliary activity, and it appears that weak inhalation may act as excitant while deep inhalation may act as depressant of ciliary activity. It is possible that prolonged heavy smoking for a number of years induces respiratory diseasesbecause of its damaging effect on ciliated epithelium. The excitant principle in the smoke aswelt as the ash of cigarette does not appear to be nicotine as suggested by Hi[ding (6,7) beea- use nicotine actioa"could be blocked by a,ganglion blocking agent while cigarette smoke as well as ash effect remained'unaffected. Some other constituent of tobacco, viz. pyridine bases, organic acids,aldehydes, hydrocarbons, etc., may be responsible for the activity on cilia. Etfectsof a fewanaesthetic gases and vapours on ciliary aetivityhave been reported (8,15). Nitrous oxide even in high concentrations was found to be without any action on ciliary activity: But chloroform has been reported to be a marked depressant this has been confirmed in the present study. Ether vapour, however, has been reported to•have no specific effect on ciliary movement. Any depressant action wass thought to be due to the cooling effect of ether. 87119262 c n:.
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119 Pceudoionone 120 p-Ionone` 121 Acetophenone 122 1-Phenyl•1•butenonee 123 1-Phenyl-2•butanone 124 1-Indanone 126 9-Fluorenone 128 2,3,B-Trimethyl•1,~A naphthoquinone 127 (8-Pyridyl)-1-propanone Aldehydes 128 Propetul 129 Butanal 130 Pentanale 131 Hezanal 132 Heptanal 188 134 136 136 137 188 139 140 141 142 143 144 146 146 Octanale Nonanale Decanele Undecanaled Doderanal 2•Methylpropenele 2•Methylbutanal 3-Methylbutenele 2-Methylpentuul 2,2•Dimet1191propanal Olycenldehyde (H,O-solution) Propenal 2-Methylpropenal 2-Butenal Time (min) to c9io.tuis at 6 mM concentration No. compound Time (min) to ciliostasis at 5 mM concentration 23 147 2-Methyl-2-pentenale > 60 22 148 2-Hexenal 4 > 60 149 2,4•Hessdienal 6 12 180 P-cyelocitral` 9 > 60 151 Salrenelc 6 > 60 152 Phenylethanal` 18 >60 168 8•Phenylpcopanale 4 154 8-Phenylpropenal 4 166 Benzaldehyde 21 > 60 166 2•Methyibenzaldebyde 8 157 8-Methylbenzaldehyde 8 158 4-Methylbenaldehyde 8 > 60 169 2,4•Dimetbylbenzaldehyde 4 43 160 2,6-Dimethylbenzaldehyde 21 43 161 2,4,6 "lYimethylbenzaldehyde 6 11 162 4-Methozybenzaldehyde >60 8 163 8,4-Dimethozybenzaldehyde >60 7 164 2-Hydrozybenzaldehyde 5 14 166 8-Hydrozybeuzaldehyde 18 3 166 4•Hydrozybenzaldehyde a 11 167 8,4-Dihydrozybenzafdehydee 10 2 168 4•Hydrozy-8•methozy- > 60 > 60 benzaldehyde 29 169 5•Methyl-2-Lurlural > 60 >60 170 6•Hydrozymetbyl•2-[urfural >60 10 171 S•Pyridine earbozeldehyda > 60 >60 172 8•Indole cerboxeldehyde 6 60 > 1 Phenols 7 178 Phenole - > 60 6 174 2-Methylphenol 9 .1
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372 LAKE ET AL. duction of hepatic microsomalmfo enzymes in both experimental animals and in man. Toward this end the measurement of the urinary excretion of metabolites of the D- glucuronic acid pathway such as D-glucaric acid (Marsh, 1963a) has been proposed (Aarts, 1965;. Hunter ef al., 1973; Hunter and Chasseaud, 1976). However, any re- lationship between the urinary excretion of D-glucuronic acid metabolites and hepatic mfo activities must be indirect as the D- glucuronic acid pathway is independent of cyluchromc l'-45t1. Vurlhcrroore, in Ihcntlthe.measurement of the excretion of a single o-glucuronicacid metabolite does not al- ways appear to provide a reliable index of the stimulation ofhepatia xenobiotic me- tabolism. For example, the pesticide dichlo- rodiphenyltrichloroethane induces rat he-patic mfo but does not elevate urinary o-glucaric acid excretion, while stimulating that of L-gulonic acid, L-ascorbic acid, and xylitol (Aarts, 1971; Lake ef al., 1976). Similarly, the activity of the hepatic enzyme catalyzing the synthesis of D-glucaric acid, namely D-glucuronolactone dehydrogenase (EC 1.1.1.70,.Marsh, 1963a,b), is not always stimulated by the administration of known inducers of ratt hepatic mfo enzymes (Aarts, 1966; Marselos and HSnninen, 1974; Marsh and Reid, 1963). Indeed, in studies with a number of inducers of hepadc mfo enzymes we found that it was necessary to measure a spectrum of D-glucuronic acid metabolites in order to obtain a urinaryindox of induc- tion of hepatic xenobiotic metabolism, in- dicating that there was no common. mecha- nism of stimulation of the D-glucuronic acid pathway by the inducing agents (Lake et al., 1976). As most inducing agents stimulatethe activities of both Phase I and Phase II xenobiotic metabolising enzymes, the con- comitant stimulation of the D-glucuronic acid pathway with mfo activities may only reflect an increased synthesis of UDPGA for glucuronidation reactions. Indeed, a number of inducing agents are known to stimulate the hepatic concentrations of both U DPG and U DPGA (Notten and Henderson, 1975). Similarly, phenobarbital has been shown to increase the synthesis of UDPGA in studies with isolatedd hepatocytes(Notlen et al.. 1975). In this paper we have-at- tempted to probe the relationship between hepatic xenobiotic metabolism and the en- zymes of the D-glucuronic acid pathway where Phase I mfo biotransformation en- zymes are not induced as a consequence of the administration of the xcnobiolic. The compounalschosan Gu~ludy were diphcnyl- acetic acid (Dixon rt af., 1977a),.guaiacol (Williams, 1959), 4-hydroxybiphenyl (West et al., 1956), 2-naphthol (Berebom and Young, 1951), l-naphthylacetic acid (Dixon et al., 1977b), paracetamoF (acetaminophen) (Jollow ef al... 1974). 2-phenylpropionic acid (Dixon et al., 1977c), and N-propyl gallate (Booth ef al., 1959; Dacre, 1960), all of which have been shown to be either directly conjugated and/or metabolised to products which are substrates for Phase 11 enzymes of mammalian liver. We also investigated the effect of phenylacetic acid which unlike the above compounds is not conjugated with o-glucuronic acid in the rat (Dixon ef al., 1977d; James et al., 1972). A preliminary communication describing part of this work has appeared elsewhere (Lake et al., 1978)- METHODS chrmiralr NADP', nt: ix.a:itric acid. UDPGA, 4-hydroxy- IdphenyL.a-propyl g.dlate. Tris, 1-naphthyl-p-o-gluo uronide, 4-methylumbcllifcryl-li-o-glucuronide, isxi- tratcdchydrogcnasc (EC 1.1.1.42, from pig hean): and 8-glucuronidase (EC 3.2.1.31, from bovine liveqrrcre purchased from the Sigma Chemical Company. Pook. Dorset, England. Guaiacol '(2-methoxyphenol),. 7- naphthol, andphenylacetic acid were obtained from nDH Chemicals. Ponle, Dorset, England, and di- phcnylacctic acid I-naphthylacctic acid and 2-pherey- propionic acid from Aldrich Chemical Compmy, Gillingham, Dorset. England. Paracemmol (N-aeelyl- 4-aminophenel,, acetaminophen) was B.P. gr.de sad all.mher reagents were of the highest purity available. S'7119239 Male 5pr; in roomf ,~ humidity:.ofy to food' aq. to the expe~ pounds wer-, period of 7 e day for: 2-i dipnenylace lhylacetk,:, phenyl, par gallate.Ccirt lies-ofthei rurr prta.cr:. NaCI (Suml corn uil:ycl lherc weie, n, the conUOl.'. described 9t controlgrou shown~in Ts Rats wer, livers immc conutining. . . subsequent Whole liver ~ prepared:e'm -ddvenhom, at 10,01I0g„ supernatant ethylmorphi biphenyl 4-, aniline4hy, of the 10,0( centrifuged . microsom9l~ in fresh hon at 10S,U00} anrye8far, protein- (t.ovansfeasc--: (Nesle and I u agtycohe. . vated by tn ~~ glucuronyltr
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.!, \ 302 Das er aL October 197 Ind. J. Physiol. & Plu It is evident that ciliary activityy is affected by a variety of drugs, and as nermal funetioof the ciliated epithelium is necessary for the health of respiratory tract, it is necessary that al drugs which are expected to come in contact with the epithelium of the respiratory tract sho be investigated for their toxicity on ciliary activity. . !, SUMMARY lheeffect of a number of agents which are either inhaled or excreted through therespira '~ tory tract or are used for the treatment of inflammatory conditions oI respiratory tract wass st died on ciliated epithelium of frog's oesophagus. All the agents studied were dissolved in normal Ringer solution. The ciliary motility was found to be stimulated by very weak solution of etha- nol (0.02%), ether,, paraldehyde, oil of peppermint, menthol, guaiacol, creosote, oxygen, carbop dioxide and cigarette smoke. Relatively higher concentration of ethanol (0.1 to 1.0%), ehloro- form, thymol, petrol gas, potassium iodide and potassium bromide were found to depress eiGary i motility. The stimulant actions did not appear to be mediated by neuro-humoral mechanisms. The action of carbon dioxide was not due to any change in hydrogen ion concentration, and the effect of cigarette smoke was possibly not because of nicotine. Of the three anions chloride, bro- '3IF mide and iodide, the iodide ion was the most depressant. The results have been discussed. 7' REFERENCBS 1. Boyd, E,M., W.F. Perry and M.E.T. Stevens. The effect of damage to the tracheal mucosa upon the drainage of respiratory trict$uid.. Am. J. Phyriol.; 140: 467, 1944. 2. Das, P.K., A.K. Sanyal and P.S. Sinha. Studies on ciliary movements: Part I- Meahanism of ciliary movement in frog's oesophagus. Arch. Int. Pharrnacodyn. Ther., 150:348, 1964. 3. Das, P.K., P.S.: Sinba,R.K. Srivastava and A.K. Sanyal. Studies on ciliary move• ments: Part II-FJCects of certain physical and chemical factors on ciliary movement in frog's oesophagus. Arch. lnt. Pharmacodyn. Ther., 153: 367, 1965. 4. Hilding, A.C. Ibe relation of ciliary insufficiency to death from asthma and other respiratory diseases. Trans. Am. Acad. Opl:rhalmol: Otolaryngol., May-June, p. 3,1943. - 5. Hilding, A.C. The role of ciliary action in production of pulmonary atelectasis, vacuum in the paranasal sinuses and in otitis media. Ann. Otof. R6inoL. and laryngol. 52: 816,1943. 6. Hilding, A.C.An cigarette smoking, bronchial careinoma, and ciliary aetion. t. Smok- ing habits and measurement of smoke intake. New Eng. J. Med., 254:. 775, 1956. 7. Ht7ding,A.C. Possible relation of the manner of deposition of cigarette smoke in the bronchial tree to catcinoma.: AetaOro-Laryngol., 48: 26, 1957. 8. Hill, L The ciliary movement of the trachea studied in iitro; a measure of tosi- city. Lancer., 215 (2) : 802, 1928. 87119264
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~ ._ .. _ ..r~ -._:_ <W.~_ .... ... . . .. . . . . & TABLE ]I (Continued) 00 No. compound Time (min) to ciliostasis at 5 mM concentration No. compound Time (min) to ciliostasis at 5 mM concentration Furons ond thiophenes 274 7-Methylindole° 2 247 Tetrahydroturan > 60 275 1,2-Dimethylindok° 5 248 Furan >60 276 1,8-Dimethylindole° 12 249 2,6•Dimethylturan >60 277 2,8-Dimethyllndole° 7 250 2,8•Benzo[uran 7 278 2,6•Dimethylindole° 18 261 Dibensofuran° 42 279 8,6•Dimethylindok° 6 252 Thiophene >60 280 1,2,3•Trimethylindole° 26 281 2,3,4•Trimethylindole 8 282 2,5,6•TrlmetbylindolehA > 60 N•Heterocycles 288 2,8,6-Trhnethyllndole° 14 253 Pynolidine >60 284 2,8,7•Trlmethylindolee 8 254 S•Pyrroline >60 286 Cubazole (EtOH/DMSO > 60 286 Piperidine >60 solution) 256 Pyrrole >60 286 9•Ethylearbazole° 20 257 Pyridine >60 287 Benximidatole > 60 258 2-Methylpyvidinee >60 288 Nornicotine > 60 259 2,6-Dimethylpyrldine >60 289 Nicotine > 60 260 3,6-Dimeihylpyridine >60 290 P•Nicotyrine > 60 261 2,3,6 •Trimethylpyridine > 60 291 Anabssine > 60 262 2,4,6 •Ttimethylpyridine >60 292 Cotinine > 60 263 8•Ethylpyridine >60 293 Norharman° 4 264 3-Hydroxypyridine >60 294 Harman° 9 265 Q i li u no ne 28 266 Pyrenine >60 Amtnes 267 2-Methylpyrazine >60 295 n-Butylunine > 60 268 2,6•Dimethylpyn.zine > 60 296 n-Hexylemine > 60 269 lndole 6 297 n•Decylsmine 8 270 1•Methylindole 6 298 2•Methylpropylamine > 60 271 2-MethyBndole° 6 299 1-Melhylbutylemine > 60 272 S-Methylindole° 4 300 2-Propenytamine > 60 278 6•Methylindole 2 801 ' B°nzylamine > 60
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142 2,2-Dimethylpropanal 143 Olycereldehyde (H,O-wlution) 144 Propenal 145 2-Methylpropenal 146 2-Butenel > 60 > 60 1 7 5 Phenola 178 Phenol° 174 2-Methylphmol 60 9 175 3-Methylphenol 15 210 2,4-Hendiena > 60 176 4-Methylphmol 8 211 Cyclohexane 14 177 2,8-Dimethylphmot 8 212 Cyclohe:ene 14 178 2,4-Dimethylphenol 7 213 Limonene > 60 179 2,6-Dimethylphenol 10 214 e-Pinmee > 60 180 2,6-Dimethylphmol 12 216 0-Pinene° >60 4-Dimethylphmol 181 3 9 216 Benzme - >60 , 182 3,6-Dimethylphmol 16 217 Methylbenzene 11 183 2,8,6-7Ylmethylphenol 5 218 1,3-Dinrethylbenzene 10 184 2,4,6-14imethylphenol 4 219 1,4-Dimethylbenaene 12 186 2,4,6-4timethylphenol . 6 .220 1,3,6-Ttimethylbenzeneep > 60 186 2-Ethylphmol 3 221 Ethylbenxene 12 187 8-Ethylphenol 8 222 n-Propylbenzene 17 188 4-Ethylphmol 8 223 I.opropylhenzene 11 189 8-Ethyl-6-methylphmol 2 224 Styrene 10 190 2-hopropyl-4-methylphenal >(Up 2-Methoxyphenol 4 >604'f i.'~l(e..n ,}, 225 226 Phenylacetylene Indan 9 18 192 8-MethoxyphmoP >60 41 cS4c.sts cJ 227 Tetralin , 20 193 4-Methoxyphenol' ' > 60 228 Indene 11 194 2,6-Dimethoxyphenol >60 5 . M cone. 229 Diphenyhnethane° > 60 195 Eugenol 6 230 Azulene° > 60 198 leoeugenol 6 281 Naphthalene° 20 197 Catechol >60 232 1-Methylnaphthalenee 22 198 8-Methylcatechol 15 238 2-Met6ylnaphthalene° 24 199 3-Lopropylcatechol 2 234 1,4-Dimethylnapthelenee 39 200 Resoreinol 7 235 Anthracene° > 60 201 Hydroquinone > 60 286 Phenmthrene° >60 202 Pyrogallol 16 237 Benzla)anthncene° > 60 203 2-Hydroxyacetophenone 12 23B Chryeene° >60 204 8-Hydro:yacetophenone 4 239 Pyrene° > 60 205 4-Hydroxyacetophenone 8 240 Benz[o)pyrenee >60 206 1-Naphthol 2 241 Perylene° >60 207 2-Naphthol 2 242 Pieene > 60 243 Coron ne >60 Hydrocarbons 244 e Acenaphthenee >60 208 n-Hexane >60 246 Acenaphthylenea 22 N 209 2-Hexenee >60 248 Fluoranthme` >60 UIz6it11a
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Toxtcology, 23 (1982) 41-88 Elsevier/North-HoBend Scientific Publishea Ltd. 13808V. ~.~ . V • RFPR)Na . EFFECTS OF TOBACCO SMOKE COMPOUNDS ON THE CILIARY ACTIVITY OF THE EMBRYO CHICKEN TRACHEA IN VITRO BERTD. PEPPERBSONe,1dAROARSTA CURVALLb and CURTR. ENZELLb *Diaision of Cellular Toxicology, Environmental Toxicology UniG Wallenberg Labo,n. tory, Uniuersity of Stockholm. S-106 91 Stockholm aad bReseorch Department. Swedish Tobacco Company, P.O. Box 17007, 8-104 62 Stockholm (Sweden) (Received October281h,1881) (Accepted January 16tfi,1882) SUldld_aRY The ciliotoxicity of 316 individual compounds representative of the gaseous and semivolahle phases of tobacco smoke has been investigated using chicken,tracheal organ cultures. When examined at 5 mM concentra- tion and measuring the time to complete ciliostasis, 36% of the compounds were found to cause ciliostasis within 15 min, while about 50% had no visible effect on the ciliary activity during a 60-min exposure. The majority of the ciliotoxic compounds were either alkylated phenylethers, ben2oni- triles, benzaldehydes, phenols, bensenes, naphthalenes and indoles, or e,fl-un- saturated ketones and aldehydes or C6--Cte aliphatic alcohols, aldehydes, acids and nitriles. Most of the compounds classified as benzoic acids, esters, polyaromatic hydrocarbons, amines and N-heterocycles, except indoles, were found to be inactive. INTHODUCTION The importance of an intact "mucociliary escalator" for the clearance of the respiratory tract is well documented [1,2]. Since inhibition of the ciliary activity reduces the clearance capacity, which allows airborne particles to remain in the air-passages and increases the risk of acute and chronic dam- ages, further knowledge about the inhibitory effects of individual organic compounds 1s of considerable interest. Address cornspondence to: Curt EozeB, Professor, Research Department, Swedish Tobeceo Company, P.O. Box 17007, 8-104 62 Stoekholm, Sweden. Abbreviations: DA1SO, dimethylaulfoxide; HMEM, Hank's minimum auential medium. 0800-488X/82/0000-00001502.75 O 1982 Elee.3er/North-HoBeud Scientific Publishers Ltd. 41 I
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269 270 271 272 273 Indole 1-Methylindole 2-Methylindolee 3-Methylindolet 5-Methylindole 6 6 8 4 2 267 298 299 300 801 n-1/ecylunwe 2-MethylpropYlamine 1-Methylbutylunine 2-PropenYluoine Eenzylam{ne > 60 > 60 > 60 >80 802 AnBine >60 310 2-Ethylaniline > 60 308 2-Methylaniline > 60 811 4-Ethylan8lne >60 304 3-Methylan8ine >60 812 N-Ethylaniline >60 306 4-Methylaniline >60 313 Diphenylaailne` 88 806 2,8-Dimethylaniline > 60 814 1-Naphlhylamine 7 307 2,6-Dimethylan8ine > 60 816 2-Naphthyl.mine° 19 308 2,6-Dimethylan0ine > 60 316 2,6-Diendnofoluene > 60 309 2,4,6-7tihnethylan83ne >60 317 8,4-Diuninotoluene > 60 All compounds are ezamined as ethanol solutions when not Indicated otherwise. eReduced ciliary activity after 60 minutee. bNot reproducible results. ePrecipitatee, . dAdded as DMSO solutions. ~ eo p46LuB
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Most studies of the ciliotoxicity of tobacco smoke, which have involved different organisms and different ways to monitor the response, have con- cerned the effects of different cigarette filters on the whole smoke or on the gaseous phase of the smoke [3-6]. Since only a limited number of the more than 3000 compounds identified in tobacco smoke has been examined for their ciliotoxic properties [7-14], more detailed insight into this area was deemed to be of importance and a necessary prerequisite for meaningful selective elimination studies. In the present investigation, 316 compounds representative of the con- stituents of the gaseous and semivolatile phases were therefore examined by exposing embryo chicken trachea in vitro to the compounds individually at the 6 mM level. The time to complete ciliostasis was measured using a microscope equipped with a TV-camera, which was linked to a TV-monitor and a videotape recorder. MATERIALS AND METHOf7eg Prepwation of organ cultures . Chicken tracheal organ cultures were prepared aseptically from 16-day to 17day old chicken embryos, essentially as described by Cherry and TaykmEobinson [16]. Aftrs dissection, the trachea was placed in a plastic petri dish containing minimum essential medium with Hank's salts (HMEM), Hepes (20 mM) and L-glutamine (2 mM). This medium was used througbout the investigation. The trachea was rinsed free of extratracheal tissues, and medium was flushed by means of a pasteur pipette, through the trachea in order to remove mucus and debris within the trachea. The trachea was subsequently cut transversely with a scalpel to give rings approx. 1 mm thick. The rings from one trachea (25-35 rings) were transferred to a second Petri dish containing the medium and stored in a C03-gassed in- cubator (6% CO, in air) at 37°C and 80% relative humidity. Under these conditions the ciliary activity persisted for more than 4 weeks. However, the rings were normally used for experiments within 6-10 days after pre- paration. Testing principJes The ciliary activity was studied at 37°C by means of inverted microscopy using a megni5cation of 250. One tracheal ring was placed in a perspes testing chamber (vol. 3.1 ml), containing the medium admixed with an ethanol or dimethyl sulfoxide (DMSO) solution of the compound to be tested. After addition of the tracheal ring, the chamber was closed to ea- temal air. The microscope was connected to a TV-camera, a TV-monitor and a videotape recorder allowing automatic recording of the ciliary activity. The ciliary activity was displayed continuously on the monitor during the entire exposure time (maximised to 60 min) and recorded on video tape for 10 s every min. The tape was then replQyed and the time to complete cessa= tion of the ciliary activity determined with the aid of a video-timer. All com- OD 42 ~ N 0 ~ O! %I
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erWaO i uS TABLE I (continued) No. Compound `%c inhibition at level (milf) No. Compound `kinhibitionatlevel(nulO" 0.1 0.01 0.001 1 0.1 0.01 0.001 222 2,6-Dimethylpyridine . 3 239 Benzylamine 13 223 3,5-Dimethylpyridine 3 240 Aniline 30 224 2,3,6.Trimethylpyridine 3 241 2-Methylaniline 1 225 2,4,6-TrimethylPy?idine 2 242 3-Mathylaniline 0 226 3-Pyridinole 0 243 4-Methylaniline . 34 227, _ Quinoline 36 9 244 2,3-Dimethylaniline 16 228 Pyrazine 0 245 2,5-Dimethylaniline 12 ~ 229 2-Methylpyrazine 0 246 2,6-Dimethylaniline 10 230 2,6-Dimethylpyrazine 5 247 2,4,6-Trimethylaniline 37 231 Nicotine 2 248 2-Ethylaniline 14 232 y-Nicotyrine 28 249 4-Ethylaniline 2 233 Anabasine 24 250 N-Ethylaniline . 28 23-1 Harman 100 66 251 Diphenylamine . 100 15 252 a-Naphthylamine 100 8 Arnines 253 Q-Naphthylamine 72 14 235 1-Aminobutane 28 254 2,6-Diaminotoluene 0 236 1-Amino-2-methyl-propane 28 2553,4-Diaminotoluene 87 8 235 2-Aminopentane 4 Other compounds 238 3-Aminopropene 100 100 52 4 256 Dimethylsulfoxi.ie 3 ' Values n0t statistically significant (P > 0,001) are given in italics. b Compoundsadded-onlyasDMSOsolutions;allothercompoundswereexaminedasethanolsolutions,eitherexclusively orinbothaolvents. ° Compounds not completely soluble in DMSO and therefore counted manually. d Compounds not yet found in tobacco or tobacco smoke.
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aphthol k acidic did not m-toxic. ohexenee u-pinene rpenoids Fas IIon- ic. How- )r size of )hthalene the other inactive. c furanes nes were 34, 266- . All the :58-263) :ic to the 38-292), and the s include ibundant bstituted )rs of the despite tives nor- a. Of the ity. Most aromatic ; system, ciiiotoxic I DISCUSSION The techniques used in this study were developed to meet several require- ments. The chicken embryo trachea was selected because the organ cultures could be prepared aseptically and the ciliary activity of the tracheal rings remained active for several weeks, thereby allowing exposures of exceptional duration. TV-monitoring and time-lapse recording were chosen in order to reduce the total observation time, which was not only an unconditional prerequisite for the screening of the present set of compounds, but also necessary when using the extended exposure time. The time to complete ciliostasis was used as a measure of ciliotoxicity [231 to avoid evaluations based on changes of ciliary beat rate, metachronal coordination or similar parameters, which tend to be more subjective and of lower reproducibility. Since dose-response studies were beyond the scope of the present investiga- tion, all compounds were examined at one concentration (5 mM), and the exposure time, based on preliminary studies of a limited number of the com- pounds, was fixed to 1 h. In several cases the test compounds precipitated on addition to the medium (Table II) and the true final concentration of these substances could not be determined. As evident from Fig. 1, the compounds were well separated as 49% of them, inhibited the ciliary activity within 30 min (36% within 15 mn), while 48% did not cause ciliostasis within 60 min. It seems therefore that this system can serve as a rapid and reproducible A s0+ 17 v 7 3_141 31 n 5 10 15 20 25 30 35 40 45 50 55 60 151 il Time to ciliostasis (min) Fig.1. Thediegrun illustrates the frequency with which cilimstasis oceua within a given time interval for d1 compoundss in this study.. N 53 ~ F~ . 1.~ ~ s> ~ h. ~
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HAZARLDOUS CIICMICALS STORAGE AND HANDLING INFUNMATION HAZARDS Moderate fire hazard when exposed to hral or Il:uue. Can rcacl willt oxidizing materials. (I2- ,)\ , ADl: AGENCY RESTRICTIONS. STANDARDS. OR CRI- TERIA USSR (pl-oposed) = 0.02 mg/L. (R-19) STATE Solid or liquid. (R-2) COLOR White or slightly yellow (solid); colorless (liquid). (R-4) ODOR Arom:uit'. (R-5). MELTING POINT IC) 28. (R-2) :)•2. (R-31) 27.9. (R- 5) BOILING POINT (C) 205. (R-5) FLASH POINT (c) 82.2 (OC). (R-2) DENSITY (:rystals = 1:129; liquid = 1.112. (R-4) 1. 1395. (R-5) SPECIFICGRAVIIY 2l/4 D= 1.12871 (R-li, R-31) 25/25 D = 1.097. (R-2) SOWBQ.ITY: OTHER SOLVENTS Slightly .wtl in water. (R-81) Sol in :dcohol, ether, organic solvents and chlu- rnform.(R-31) - THEORETICAL-0XYGEN DEMAND 2.01i. (R-/i). BIOCHEMICAL OXYGEN DEMAND BOD5 = fiNtYuThOD. (R-fi). CHEDIICALOBYGGEN. DEMAND 100% ThOD (0.05 N R._(ll:2O;).. (R-(i)  AQUEOUS SOLUBIWTY .tiolxbilip (filnu) Tcs y ((.7) 15,964.6 16.000:0 15 16,500.0 20  VAPOR PRESSURE Vrtpnr Pres.uro (x:m Hg) Texp (C) 0.103 25 1.000 52 100.000 144 46 CONVERSION FACTOR I mg/m' = 0.197 ppm;. I ppm = 5.07 mg/m". (R-(i) ODOR THRESHOLD: WATER 0.(1(12f} mg/L C<0 ti(1 C. (R-9) 0.002 ml;Pl. (m 30 C. (R-9).0.021 ppm, 0.013 ppnl. (IL83), TASTE THRESHOLD -L'h reshold conc for tainting of trout is 82 ppb. (R-34) OTHER PHYSICAUCHEMICAL PROPERTIES ReFrac- tive index = L5429/20 C. (R-31) __ CHEImCALSYNTlESIS METHODS Prepared by wcruu'ic oxidc oxi(L-:liou nf liguin;. by oxidatinn of :misole with trilluorolxroxyacetic acid; frum acctu- vanillune +%nCl.,; I'rom the diaronium sdt of o-an- isidine. (R-4) •TD%[CI7T ACUTE TOXICITY: TERRESTRIAL WFE-OTNF1t STUD- IES I Rabbit, route not specified, LD50 = 3.74 mg/kg. (R-7) III R:us, parental LDLo = 50 mg/kg. (R-7) a Mice, oral LD =0.4 g/kg. (R-7) I Scu injettionn to pigeons hacl an LD50 of 0.2 g. (R-32): EScu injection to rabbits had all L050 of 2.5 g. (R-32)t Scu injection to guinea pigs had an LD50 of 0.9 g/kg. (R-32) . Scu injection to rats h:Hl ;ut LDfi0 of 0.9 glkg: (R-32) ACUTE TOXICITY: HUMAN STUDIES 0 The oral hu- man probable lethal (lose is 50-500 mg/kg (I tsp-I oz). (It-11) 0 The derntal lethal dose is estimated to be 2 g/man. Exposure produces chills, reduced body temp, collapse, and death due to respiratory Failure. (R-32) , TOXIC EFFECTS: TERRESTRIAL LIFE-tHACRORIYFA• TESRATES:III Oriental fruit fly: naked eggs LD50 = > Mo.JC'A MNliwf R`J MrrulF.#MeUmr! 8t711924'7 I F 0
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Tx7 ~ M~ n,.rf~ LYaI 1,~~ 1,•f,+l .,.1„1 I+Y. f+KY 1.~ C11 ,n alltr 1.'S„Y,al. I,Y.11 IIIInIM•rol 1lrl1,(OCJ- l „n ..111 I\I+phylu 1,630 Nm 12 1,975 Nvle 9,2a5 4 11,020 Nww 16,620 4 12 32,450 4 8 3,330 16 Ncne 3,650 26 12 4,215 R •1,7n1 Y 15,430 16 i 13v7o 24 10 .lii/bru. 'I?hl•' Inlat unulhrr uf indicaturs of sanitary cm,di- Itduringthelessons. As a rule, e detected in the air before ~f children participated, cocci groups of children, strepto- vere more than double those The differences in bacteria iren were confirmed by sta- Iteceived'14 Urcember 1962 `I"JC.P,to`Y,a/1 lP,,-(lA'~..'J OX -Za: (os'fDB. I ~t (.P`4 4AN 5 d ? ~ TOXICOLOGY OFGXAiACUL-VAPORS AND RESINS (K toksikologii parov gvayakola i ego smoly) 14. M. Ostrovskii, Junior Science Fellow LEMNGRAD CNEMO•YNARMACEVTICAL INSTITDTE Rae I Data on gualacol in the literature are incotnplete from the toxicological . vIL`WpOillt, because the prnln•raLLrntion has not been given to the effect of : inhalation of its vapors. It should also be noted that i'n the production of guatacoL (as a production waste) aguaiacolgumisobtained, of which the effect on the body has not at al l been clarified. Nevertheless, from each tonn of the ' finished product 220 kg of resin of an undetermined chemical composition . remains: this must be removed from the surface of the reaction mixture . and fliNlx)NIRI of outsidc the Industrial premises. During these operations, performed by hand, the resin soils the skin, special suits and . equipment. We set before ourselves two tasks:: first of all to determine the toxicity of the guaiacol vapors when they enter the respiratory tract+ and secondly. to determinewhether the solidified resin is a hazard to the skin and whether it possesses any local effect. ' ' The toxicity of guaiacol vapors was studied in short-term experiments / on 70 white mice. In a two-hour exposure the substance was administered under dynamic conditionss in a concentration range from1.98 to 17.31 mg per liter; these concentrations were created by means of heating a goose- neck flask over a water thermostat at 70 - 97°. The survival of the animals was the criterion for evaluating the toxicity. In the initial period of the effect a moderate irrita ion of the mucous membranes of the respiratory tract and eyes was noted in all mice, these phenomena were accompsnied by brief motor restlessnessfollowed by inactivity. With concentrations less than 4 mg/1 these phenomena were .. chiefly of aa reversible nature, but at higher concentrations other symptoms also appeared:ru~ff ing.of the hair _rMurtion of e citabilitv _el.ltLti,tlg_[rom a tapping noise, dtlatation of the blood vessels of the aural.conchae, an unetead gait and dysppPa. -~- ~ n the cnncentration range from 6.13 - 6.72 to 9.26 - 9.8 mg/I some , mice fell ov r nnto their sides, which heralded their approaching deaths. With concentrattuns from 7.74 - 7.78 to 9.26 - 9.8 mg/1 clonic and tonic convulsions were observed, e u s o e ezperiments are shownin the table. - - From the data presented It is evident that during the period of inhalation of the poison the mice rarely died; in the majority of cases this occurred within 4 hours after the end of the experiment. Death occurred from 105 817119255
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r, . 298 Das et al. October 1970 ind. 3. Physiol. & Phsmme agents were dissolved in Ringer solution. Effects of potassium iodide were compared with \ equimolar eyncentrations of potassium bromide and chloride. Oxygen, carbon dioxide and petrol gas were bubbled through 50 ml of Ringer solution for I andfor 10 min and the solutions were then used for study. The smoke from one cigan;tte (Panama brand) was bubbled through 50 mlof Ringer solution by smoking it the hookab fashion. This was then used for study. Thee ash from the cigarette was carefully collected and put in another 50 ml of Ringer solution. The ash was shaken thoroughly and filtered. The filtrate `_ . was also used for study. TAua 1 Effed of acne o6ents on tAe dJlary murrmentr ABeen md connnrnuiw~ l. Alcohol 0.02% „ 0.1% 0.5% 2. C3iorroform 03'/. 3. Ether 10.0•J. 4. PantWryde 1.0% 5. Oilof 0.2% Peppemmt 6. Men0alr• . Guaixbl 113'/.. 8.Crrooote 067% 9. Thymol 0.1%10. Oxygen 10 m1n 11. COr(pH 6.3) 10 min Rmeer (pH 6.3) 12 Pdrol6af (bubbled for 1 min in RinBcr solution) Pevol gas (bubbled for 10 min in Ringer solution) 13. PoWsium iodide0.1 Potassium chl6ride' Poassium brrnoide' 14. A6Uylcholine 10'* Untreated PrdrNled wW+ atropine 1" Nomber TrovrllGn P Number Tra.el ttnm P of (%olcoN.a '.adre ol (%ofeontral .otrre rspnfmmtr memrrS&) 64.1.4.1 <0.001 150.7.6.3 <0.001 183.2.16.2 <0.001 209.6.15.1 <0.001 246.7.43.1 <0.01 61.8-2.3 <0.001 47.1 t2.5 <0.001 58.2+3.8 <0.001 10 71.9+3.0 <0.001 4 S..3.2 <0.001 4 50.8r5.8 <0.001 8 172.6+7.3 <0.001 rspnimener mean..SE) 4 57.0r5.7 <0.001 . 9 59.7r2.6 <0.001 7 110.2. 3.1 <0.01 5 103 . 0-3 . 5 > 0 . 05 ' 10 160.8+5.3 <0.001 6 4 4 4 22.0+3.8 <0.001 25.3.3.1 <0.001 22.7a0.8 <0.001 29.5+9.8 <0.001 4 37 9~7 5<0 00 ~ 4 19.4.3.0 <0.001 ~ ` . 4 54.3.3.0 <0.001 I 4 105.1 r 1.0 <0.05 5 192.5+19:4 <0.005 4 107.4a7.9 >0.05 4: 143.5+11.1 <0.01 8 60.8r5.2<0.001 7 ~oneeafration equ'uooler to potadum iodide 0.1'f,. ••Satugatedsolution. 8'7119260
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TABLE IIS PER CENT CILIOTOXIC COMPOUND6IN THE 12 DIFFERENT CHEMICAL CLASSES Chemleal clea % Aetive compounds No. of compouads Alcohols 50 20 Ethers 83 12 Acids E.tezs and anhYdrides 29 18 17 - 13 NitsOa and amida 66 33 ICetomy 52 81 Aldellydes 73 45 PheooL 80 35 Hydroc.rbon. 46 39 Furanes.ud thiophenet a3 6 N-Haterocydea 45 42 Am{nr 17 23 qualitative test for identifying ciliotoxic compounds. Moreover, it is poten- tially, useful also for quantitative studies and for studies involving recovery of the ciliary movement. An account for the correlation between chemical functionality and inhibition of ciliary function is given in Table III. The ciliotoxic compounds were alkylated phenylethers, benzonitriles, benzaldehydes, phenols, ben- zenes, naphthalenes and indoles, or a,$-unsaturated ketones and aldehydes or Cb--C,o aliphatic alcohols, aldehydes, acids and nitriles. In contrast, most of the benzoic acids, esters, polyaromatic hydrocarbons, amines and N- heterocycles, except indoles, were found to be non-toxic. Additional toxic- ity data for these compounds are available, such as inhibition of cell growth on Ascites sarcoma cells (24], inhibition of noradrenaline stimulated oxi- dative metabolism in isolated brown fat cells [25), damages of the plasma membrane of cultured human lung fibroblasts [26] and mutagenicity in the Ames'test [27]. It may be concluded, that tobacco smoke contains a number of ciliotoxie compounds that could cause acute or chronic damages, on the ciliated epithelia. As the basic biochemistry and mechanisms behind the motility of ciliated epithelia probably are the same for all organisms [16], it may be assumed that most of the compounds found ciliotoxic in this investi- gation should have the same effect in other species, including man. ACKNOWLEDGEMENTS The authors are indepted to Ms Marianne Andersson, Ms Eeva Kazemi- Vela and Ma Eva Neihoff for skilful technical assistance. This work was financed by a grant from the Swedish Tobacco Company. 54 I
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HAYJtRUUUS CI ILMICALL'T  SUBACUTE-CIIItONIC EFFECTS: TERRESTRIAL LIFE S^rrir.. Rmde U,w.r. 1)a.iuA.1'dvrlnG Nrf r{as Orl ,. 0.15%. NG R-7 0.6% RFSULTS A 0.15%soln aaused paralysis of the hean mus- cie, and a 0.6%soln caused paralysis of intestinal smooth muscle. Rodents NG NG NG R-7 RESULTS 2-Methoxyphenolinduced leukopenia thatled to leuhocylosis in rodents. -- ftat Orl 7.40 mglkg OncelD. for 4 D R-22 TEST CONDITIONS The capacity of 2-methoxyphenol to produce macroscopic liver lesions was determined in groups of 3 male and 3 female Osbome-Mendel rats by using a doee (viagavage) of approximately I theLDS0. Rat Parenteral 50 mg/kg RESULTS The rals were sacrificed on the SIh D. and the Uvers were examineddor gross lesions. The mortality ra- tio during the test =%i6. Three out of 6 animals exhibited matroacopic liver tesions. NG R-7 RESULTS 2-Methoxyphenol was determined to be embry- otoxic in rats. When it was injected into pregnant ra1s, it was fatal to the fetus. When similar doses were injected into maleanimalse serious disorders of the lestes and destruction of the germinaDepilheUum were observed.  SUBACUTE-CHRONIC EFFECI'S: HUMAN STUDIES lteule lN.vn /MsingSrlwtutr Ref NG NG  CARCINOGENICITY: ANIMAL STUDIES Sp-us Runtr. lh.uc Rat Scu I-N% in olive oil guaiacol,, in a Ig'e conc and with fresh potato juice (from dormant tubers), suppressedd sprouting. A low conc (0.05%) stimutatedgrowth of rye spr<ruLs. OTHER 1tDYEBSEEFFECTS:  2-Methoxyphenol enhanced the bactericidal activity of a reaction mix- nu'e bactericidal to Xanl$mnonas /tluueuG sojcnsiq a 48 AESULTSin most clinicaP poisoninge, 2-methoryphenol was taken as wood-tar creosote. Percutaneous absorption is dangerous. RESULTS The author determined that 2-methoxyphsnd was not carcinogenic to rats at this dosage. 8'71192q,q pathogen of soybeans. The mixture consisted of horseradish peroxidase. KI, and hydrogen.pernxide. (R-12) E Acute administration of 2-methoxyphenol in rats (dosage not specilied) su ppressed serum prolactin levels. Chronic administration resulted in decreased pituiWry prolactin Icvcls but increased pituitary weigM. (R-1 i) IN 2-Methoxyphenol in vitro inhibited human p t s) as ea ol m hi ga 111 tic dr QI ttll in in: sys tes m6 cul for an( obt hib
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_ r ILC100). :.. 1? mg/t. ua by the Kerber method vapors causing tAe.deaths :.Qetermihcd. It proveti to. lcDerjjgjotlovlnx me expedmem 16 haun 1 2 1-4 dayr 0 1 5. 6 T 9 lo ]sem~guaiacol vapors, • r ins and small hemorrhages, lungs (subpleurally and nerative changes were (in some preparations) were found. in the dead animals :tion.of the vascular system circulatory disorders re blood vessel walls een determined for al data, calculatedits . I,yublina's formula: igJl); LC50 is the lethal n of the substance for iproximate value of the0~02 mg/1, 1 resin on 5 rabbits. The ed in vaseline and lanolin Mt (from which the hair had ver a 2-week period • visible skin changes tfect of the resin, two series 3-4-month-old mice, of the untt In Ih<cvalaadm uf,h< C57 black line, hoih sexcs equally represented). Mice of this line, as is well~know'n~ are resistant to sponlaneous Lumors (A..M. 17yad!kova, NL N. Medvedev). In the first series each, . animal received 50 applications of a 1%a guniacol resin solution in benzene for 6 months (I drop twice a week on theInterscapularregim]), In the majority of the mice, epilatedareas,.dryness of the skih, and slight branny desquamation appeared at the application sites, which apparently were the result of the effect of the solvent on the skin (L. M. Shabad). Vo other skin changes were found. True, 5months aftcr the beginning of the experiment a superficial erosion measuring0.5 x 1 cm appeared at the site of application. Thee epithelizationprocess in this mouse lasted over 3 months and ended in the recovery of the skin and restoration of the hair. Our single observation of this kind does not give us the right to consider guaiacol resin the cause of erosions andd delayof Epithelization. During the.entice observation, period 9 mice died;.; as has been determined, the direct causes of death In all cases weree intercurrent infections.. In the second series, conducted at the same time as the first, 8.7 $, solution of technical guaiacol in benzene (50 applications for 6 months) was used for the application. The symptoms noted above (loss ofhair, slight desquamation or partial loss of hair): were first found only at the end of the second month in 14 mice and at the end'of the third month in 20 mice (of 70 surviving mice). However; subsequently a gradual restoration of hair was observed. Therefore, in contrast to the first series, the local effect of guaiacol resin was noted later and only in a comparatively small group of experimental animals. No other skin changes were found. During this period 20 mice died; in 10 which died during the first: ten days hypostatic congestion of the parenchymatous organs was found macroscopically. This gives us the basis for considering the resorptive effect of guaiacol the probable cause of death. The cause of death in the other mice, asautopsyshowed~ consisted of intercurrent infections,, chiefly pneumonia. Conclusions 1. On the basis of experimental findings on the toxicity of guaiacol vapors a calculation of an approximate value was made for thee permissible concentration of these vapors., On the basis of the LC50 (7.57mg/I) itt was found equal to 0.02 mg/l. 2. In the study of the effect of guiacol resin on the skin it was determined that it possesses neither an irritating nor a blastomogenic effect,. BIBL.IOCRAPIIY B e l e n' k ii M. L. Elementy kolichestvennoi otsenki farmakologicheskogo effekta (Elements of a Quantitative Evaluation of Pharmacological Effect). Riga. 1959. .. 107 8"J11925"1
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106 3-Phenylpropanalo 3-Phenylpropenal 108 Benzaldehyde 109 2-Methylbenzaldehyde 110 3-Methylbenzaldehyde 111 4-Methylbenzaldehyde 112 2-Hydroxybenzaldchyde 113 3-Hydroxybenzaldehyde 114 4-Hydroxybenzaldehyde 115 Aniealdehyde 116 3,4-Dihydroxybenzaldehyde 11 i Vanilline 118 Veratrumaldehyde 119 5-MelhylfLLrfural 120 5-Hydroxymelhylfurfural 121 3-Indolecarboxal dehyde 122 3-Pyridinecarboxaldehyde Phenols 123 Phenol 124 n-Cresol 125 in-Cresol 126 p-Cresol 127 2,3-Dimethylphenol 128 2,4-Dimethylphenol 129 2,5-Dimethylphenol 130 2,6-Dimethylphenol 131 3, 4-Dimelhylphenol 132 3,5-Dirnethylphenol 133 2,3,5-7Yimethylphenol 134 2,d,5-Trimethylphenol h inhibition at level (m-t!) No. Compound % inhibition at level (m-N) 1 0.1 0.01 0.001 1 0.1 0.01 0.001 100 83 1 135 2,4,6-'ltimethylphenol 81 6 100 100 4 136 2-Ethylphenol 77 5 40 137 3-Ethylphenol 38 100 19 138 4-Ethylphenol 91 22 100 5 139 Thymol 100 2 57 7 140 3-Ethyl-5-melhylphenol 81 2 100 79 1 ® 2-Methoxy henol. 74 8 _ 42 142 - e oxyp eno 35 28 143 4-Methoxyphenol 93 79 15 19 144 2,6-Dimethoxyphenol 50 0 100 98 16 145 Eugenol. 98 16 30 146 Isoeugenol 96 13 23 147 2-Hydroxyacetophenone 36 25 148 3-Hydroxyacetophenone 14 19 149 4-Hydroxyacetophenone 21 81 7 150 a-Naphthol 100 17 63 18 151 (3-Naphthol. 85 13 152 Catechol 88 55 6 153 Resorcinol 32 25 5 154 Hydroquinone 100 96 1 56 7 155 Pprogallal 100 39 ` 31 5 156 3-Methylcatechol 100 91 0 , 93 13 157 33sopropylcalechol 100 100 6 78 99 11 Hydrocarbans 74 0 158 Isopentane 12 79 5 159 n-Eicossne 10 75 5 160 2-Methyl-l-butene 0 43 7 161 2-Methy I-2-hu tene 10 81 U 162 Isoprene 5 88 r 163 cis-2-Pentene 0
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TABLE I (continued) 9c inhibition at level (mM) No. Compound . 1 0.1 0.01 0.001 164 1,3-Pentadiene (cis/fran.s) 6 . 195 165 Limonene 100 11 196 166 Neophytadiene 2 197 167 Cyclohexene 3 198 168 a-Pinene 100 3 199 169 13-Pinene 100 10 200 170 Benzene 4 171 Taluene 9 _ 172 m-Xylene 46 4 201 173 p-Xylene 32 10 202 174 Ethylbenzene 43 3 203 175 n-Propylbenzened 100 3 204 176 Isopropylbenzene 100 3 205 177 Styrene- 25 206 178 Phenylacetylene 34 179 Indan 100 5 207 180 Tetralin 100 7 208 181 Indene 100 9 209 182 Diphenylme thane 100 17 183 _ Azulcne 85 7 210 211 184 Naphthalene 100 6 212 185 1-Methylnaphthalene 100 10 ?13 186 2-nlethylnaphthalene 100 /2 214 187 1 4-Dimethylnaphthalene 100 35 , 215 188 Anthracene` 12 216 189 Phenanthrene 22 190 Benz[aJanthracenee 27 217 218 191 Chrysene` 9 192 Pyrene' 33 219 220 193 Benzo[a]pyrene` 6 221 194 Pervlenec 0 Picene` Coronenec 3-Methylchelanthrene`'d Acenaphthene Acenaphthylene Fluoranthenel' Furanes..INiophenes Tetrahydrofuran Furan 2, 5-Dimethylfuran 2,3-Benzofuran . Dibenzofpran Thiophene N-Xelerocycles Pyrrolidine 3-Pyrroline Piperidine Pyrrole Grhazule° 9-Eth-vlcarbazole Benzimidazole Indole 2-Methylindole 3Melhylindole 2,3-Dimethylindole 2,3,5-Trimethylindole Pyridine • 2-Methylpyridine 3-Etlrylpyridine 9o inhibition at level (rrrAf) 1 0.1 0.01 0.001
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a a IIA%AI[I. NI:1 , 'III:MU'AI:: CIS (:1'l:l'. Chcm'fuxii-oluRy of (:omnlcrcial 14rxl- ucts: 4511: (:IS. FRSS (Feclel.d Register Sea:ch Systent). (11S. CESARS NON-CIS REFERENCES EPA. ~I:SCA Invemnry l.ist F:PA, Reslicides-Rcgistcred Active Ingredicros. 025005 Mrr.k Indcm:.1.125 NI'A. I'aslici8ct-Analytii'al Rrl: Studs.: 17i65 tl,S. Imcrnaliunat'IYadc Conimissinn 1'&I Statixticx: I M I'C I'llti-77.1'K( )D! I YNi-77,1'ROtN I'Oli-711,1 MIM1'12- 7R.1 hl I'F I' 12-79 h:urulw:m P:rnnumiF G,uuunuib' luvrmuly: 23908 h:ROI(:A-l)rpanic 1'rulxnics: 211(1121[IML 1'I:IS-I19'(::nriu.,Xrui,'A<'livily: RIIIi7Rd _'9'f,D~l31i NIOSIi, NOHS M2R4d ORNL. F.MIC C:IS, II(titi ' USAN & USP Dictionary of Drug Names NLM. CHEMLINE: '1'OXLINE,TOXBACK Aldrich.CV:Jbg/Handhook: G010903 (:tde of Federal Rcgtdations (CHEMLAW) Japan. PI'WI'lly'LI51 IOr A\iC%snlCm in P:nvimnmrnu 7325 Diniou:uy ntUrgauic Conywunds (UO(:S): M-INl/ili t I(OI)ARI:Ibnralorv Chc•mirals: 14'S.1'14:i N'IP, iluwr.d I'I:lu: ItFlali-U (builrusrvl (ln•Inir:d Diclianary.  6-PROPYL-2- THIOURACIL CAS RN 51-52-5 NIOSH # YR 1400000 See N. I. Sax. 1984. Dangerous Properties of Industrial Materials,61h ed. New York: Van Nostrand Reinhold, p. 2315. SYNS propylthiouracil; 2,3-dihydro-6-propyl- 2-thioxo-4(1H)-pyrimidinone (9CI); 2-mercapto-4- hydroxy-6-n-propylpyrimidine; 2-mercapto-6- propyl-4-pyrimidone; 2-mercapto-6-propyl- pyrimid-4-one; 4-propyl-2-thiouracil procasil; propacil; propycil; 6-propyl-2-thio-2,4-(1H,3H)pyri- midinedione; propyl-lhiorit;.6-n-propyl-2- lhiouracil; prothycil;.6-n-propylthiouracili 6- propylthouracil; 6-thio-4-propyluracilt propyl- lhyracilt propythiouracil; prothyran;PTU; 2-thio-4- 52 oxo-6-propyl-1,3-pyrimidine; 2-thio-6-propyl-1,3- pyrimidin-4-one; 6-thio-4-propyluracil; thyreostat; propilthiouracil; propyl-thiorist; prothiucil; prothiurone; prothycil; protiural; T 72; thiuragyl; thyreostat IP. COMMON USES Propylthiouracil is used as an anti-thyroid agent (R-14) and as a metabolic depressant to promote fattening in animals. (R- 14) Uaed in medicine. (R-13) - STRUCTURAL FORMULA H CHsCHzCH. N \ / I SIY NH 0 ml CtHION2OS mw 170.25 STORAGE AND HANDLING INFORMATION DISPOSAL AGENCY RESTRICTIONS. STANDARDS.OR CRl'tERIA RCRA 1S(H)1-:11104: designates propylthiour- ncil as an Appendix Vl ll chemical. (R-96) STATE Sulid, Iwwlltv' Anul cryaalliot•. (Id-Ia)COLOR Wltitc. (R-13) MEL'rD(G POINT (C) 21g-221. (R-1'3) sOLUSILITy: OTHER SOLVENTS Sparingly sol in al- cohol, sol in ammonia and alkali hydroxides. (R-19) OTHER PHySICAL/CHE[cIICAL PROPERTIES Has a bit- ter taste; is sensitive to light. (R-13) Forms complexes with metals and reacts with sulphydryl-oxidizing agents. (R-1.4) The ntaximunt chlorofornt/watercoelTtcieno was 0.92 at pH 6.0. (R-26) CHEMICAL SYNTHESIS METHODS Propylthiouracil is derived by the condensation of Ixta-oxtxaproate with thiourea. (R-1f1) TOXICITY ACUTE TOXICITY: TERRESTRIAL LIFE STUDIES • Charles River male rats (100 g)) were given ipr injeo tions ol' PTU at I, :i, and 5 mg/11)0 g body weight.. Half the rals'were injected with [12511 15 M later, then after an additional 15 M were sacrilicedd andd iodination was determined. The rest of the animals were sacrificed 30M after PTUlinjection, and thyroid peroxitlnse activity was detenniued. Nurmal levels nf 87119254 DANGEROUS PROPERIIRS OF INDUSTRIAL MATERIALS REFORf
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49 Benzylcyanide 50 Nicotinonitrile . 51 Indolyb3-acetonitrile Kelones 52 2-Propanone 53 2-Butanone 54 6,10-Dimeth,vlundecan2eone 55 2,3•Butanedione 56 2,3-Pentanedione . 57 3-Buten-2-one 58 3-Methyh3-buten-2-one 59 3-Penten-2-one 60 6-Methyl-5-hepten-2-one 61 6-Methyl-3,5-heptad ien-2one 62 Solanone 63 Pseudoionone 64 (3donmre .. 65 Cyclopentanone 66 Cyclohexanone 67 2,2,6-'ltimethylcycluhexanone 68 Piperitone 69 Carvenone 70 Carvone 71 Acetophenone 72 1-Phenyl-2-btrtanone ° 73 1•Phenyl-l-hutanune 74 1.Indanone 75 9-Fluorenone 76 2,3,6-'frimethyl-l,4-naphthoa hrpTT p quinone % inhibition at level (mbl) No. Compound % inhibiton at level (ml!) 1 0.1 0.01 0.001 1 0.1 0.01 0.001 5 77 .(3-Pyridyl)-1-propanonc I 4 71 Aldehydes 78 Methanal 100 lu 79 Ethanal ' 92 34 I1 80 Propanal 98 27 (1 81 Butanal 100 26 100 51 3 82 Pentanal 95 4 100 37 83 Hexanal 100 69 0 100 34 84 Heptanal 100 69 3 3 85 Octanal 100 29 100 100 9 86 Nonanal 100 73 3 100 99 9 87 Decanal 87 60 0 96 9 88 Undecanal . 86 2 100 100 77 3 89 Dodecanal 81 1 100 33 90 2-Methylpropanal 38 0 100 100 9 91 3-Methylbutanal 100 13 100 20 92 2-Methylbutanal 100 0 93 2-Methylpentanal 100 1 0 94 2,2-Dimethylpropanal 28 22 95 Glyceraldehydea 97 10 31 96 2-Oxopropanal 100 6 33 1 97 Propenal 91 94 20 100 13 98 2-Methylpropenal 100 100 34 9 99 2-Butenal 95 86 45 6 100 2-Hexenal 100 96 66 7 9 101 2,4-Hexadienal d 100 100 44 9 102 2-Methyl-2-pentenal 100 38 0 63 51 2 103 /3-Cyelocilrnl . 82 6 100 100 22 104 SaCranal 100 100 6 105 Phenylethanald 100 76 0
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0 TABLE I EFFECTS OF TOBACCO AND TOBACCO SMOKE CONSTITUENTS ON CELL CULTURE GROWTH RATE No_ Compound %inhibition at level (mM) No. Compound % inh ibition at level (mM) 1 0.1 0.01 0.001 1 0.1 0.01 0.001 Alcohols 24 Indolyl-3-acetie acid 21 3 1 Methanola 3 25 Xynurenic acidb 4 2 Ethanol 10 26_ Terephthalic acid6 15 3 2-Propanol . . 7 Esters and anhydrides 4 1-Octadecanol 8 27 Diethyl malonate 5 . 5 a-Terpineol 37 - - 28 Ethyl stearate 7 6 Gcraniol 99 84 7 29 Vinyl acetate 3 7 Solanesol 0 30 Methyl acrylate 2 8 Farnesol 100 6 31 , . Benzyl acetate 6 9 Propylene glycol 2 32 Benzyl benzoate 100 22 10 Fur(urylalcohol 28 6 11 Benzvlalcohol 6 33 Benzyl cl nriamate 73 35 12 2-Phenylethanul 7 34 CDumarin 34 7 35 Diethyl phthalate . 44 4 Ethers 36 Di-n-propyl phthelate 100 68 0 13 Anisole 6 37 Dibutylphthalate 45 14 Diphenylether .100 17 38 Dioctyl phthalated 32 15 4-\leth lanisoie 43 39 Phthalic acidanhydridea 14 16 y 3-Alethylanisole 40 Amides and Nitrites 17 2->lethoxynaphthalene 46 13 40 Nicotinarriide 5 18 2-Ethoxynaphthalenc 100 9 41 N-Methylnicotinamide 0 19 1,2,3-Trimethaxybenzene 14 42 Acetonitrile . . 0 Acids 43 Propionitrile 4 20 4-Oxononanoic acid 44 Butyronitrile 4 21 2,5-Dihydroxybenzoic acid 45 Isobutyronitrile 4 22 3,5-Dimethoxy-4-hydroxybenzoic 4 46 Valeronitrile . 44 acid 47 Isovaleronitrile 12 23 Nicotinicacid6 48 Acrylonitrile 6
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organclles Ifaveobvious advantages. Although not directly applicable to the human situation these systems provide more clear-cut information on the mechanisms involved in the interaction between specific smoke constit-uents and distinct biochemical cell functions,.thus fulfilling a necessary con- dition for a prcrper understanding of the more complex situation. The aim of the.presentstudy was to obtain such basic information with- out either taking into account such effects o8 penetrallion and distribution as are often encountered in more complex systems, or evaluating the addi- tive, synergistic or antagonistic interactions frequently associated with com- plexmixtures. However, the very large number of constituents in tobacco emdd tobacco smoke. prevents a thorough studyof the effects of all of thesee on the various organelle functions. A pre-screening procedure isthus re- quired for the selection of those constituents which affect at least one organelle function. Ascites tumour cell cultures were used for this pur- pose; thcy were exposed to a number of different substances at various concentrations. No specific cell function was studied and: thee toxic effectwas measured only as the ca aci m.studie com owtd to inhibit the growth rate o a cell culture. Although for reasons still to be discussed this syss my- s pinpoints every biologically active substance, the infor- mation obtained nonetheless provides a basis for further studies of the ef-s fects produced on subcellular organelles by a limited number of selected tobacco and tobacco smoke components. METHOD Tesfsystem. The cell culture system used in the present experiment is.a further devel- opment [11 of an immunological cytotoxic test system developed by Mun- roe et al. [ 2 ] . 'l'he cell strain Ascites sarcoma 13 I'8 used here offers certain experimentali advantages allowing the handling of largere.rperimental series. Thus these cellscan be grown for many generations in testtubes without intermediate -inoculation of mice,.and:they are therefore easily handled in in vitro systems [21. Moreover the cells can easily be reinoculated in the abdominal cavity of mice,, providing the possibility of comparing results obtained in vitro with those obtained in vivo, where the cells are simultaneously exposed to the. metabolites of the studied compound produced within the mammalian host (cf. ref. 3). The property of these cells not to adhere to the surface of the in- cubation vessel provides a further advant'agee as with high reproducibility they can be suspended by shaking and counted with the aid of an electronicc particle counter. Stem cell cultures originating from inoculated C3H mice were grown in test! tubes in Hams F 10 medium sterilized by filtration (through Millipore 0.45 M), with fetal calf serum (15% w(w); penicillin (100 IU) and st'repto- mycin (100 IU) added. The test tubes were gassed with sterilized air con- 50
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Volume 14 .. Drugs on Ciliary Movements 299 Number 4 aFSULTS The results are summarised in Tables I and 11. Ether, paraldqhyde, oil of peppermint, menthol, &M'ag1l creosote,, oxygen, carbon dioxide and weak solution of alcohol (0.02•/,) lieni- ficantly inaeased cili_T arv~tivity. In order to find out whether theaxcitant action of these agents was due to direct action or mediated through a local refiexatc (2), their effects were also studied on atropinised cilia. Atro ine retreatment did not block the excitant action gf these agents wh;le ;t enmnlrtely locked that of aeetykholine indicatln¢_that they all-acted as eet -s ofsilia,. . The hydrogen ion coneentntionn of carbon dioxide Ringer being pH 6.3 the effect of normal Ringer with pH adjusted to 6.3 with the help of dilute hydrochloric acid wes also studied. Acidic Ringer was found to be depressant indicating a specific stimulant actionn of carbon dioxide. T*sta II EffecroJciaarene smoke md uk on eiBnrT a+o.emmrs Unrnafed Prerreoled wlrL pauolD*m 2 s.10-r No. of Trcxew rinr P No. oJ Tmvd r7nw y e:pertments (%qf .due experimertn (%ol +dre ecmrnt . wmrol. mmx•S.N)meatts&) 1. SmoheRinBer - 9 59.7a5.7 <0.001 4 22.8r1.7 <0.001 Nicotine 10"s 4 64.4.2.4 <O.00I • 4 102.1+2.5 >0.05 2 AahRinBer(pH9A) 6186.3a6.9 <0.001 RL1gar (yH 9.0) ' 4 253.1.5.9 <0.001 I AshRiaBer(pH7.5) 4 85.3.2.0 <0.001 5 59.3.4.6 <0.001 Chloroform, thymol, petrol gas and bigher concentrations of alcohol (0.1 to 1.0 %) caused signifleant depression of ciliary activity. Potassium iodide in the concentration of 0.1 % ;6.024 mM) produced a marked retardation of ciliary movements. To assess whether the effect was due to the anion or the cation, the effect was compared with those of equimolar concentra- ionsofpotassiumehloride andpotassium-bromide.-Potassium chloride had no significant The effect of cigarette smoke-Ringerand ash-Ringer are summarised in Table II. Since tine is one of the constituents of cigarette smoke, the eHect of smoke•Ringerwas compared aotion while potassium bromidewass moderate depressant. . with that of nicotine.. Nicotina and cigarette smoke stimulated ciliary activity to nearly the bame extent. Pretreatment of the epitheliumwith pentolinium tartarate completely blocked the nicotine response. Smoke-Ringer on the other hand markedly stimulated the ciliary Aotivity which was significantly depressed by pentolinium treatment. Since the pH of ash- a ft
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?EYqIN F ra:,;<araxr;,5 ( 1975) .fs-uz.. 10553 -0 V IuvierfNorth-Holland, Amsterdam-Printed in The Netherlands 30f: . EFFECTS OF TOBACCO AND TOBACCO SMOKE CONSTITUENTS ON CELL MULTIPLICATION IN VITRO AKE PILOTTIb, KLAS ANCKER°, ERIK ARRHENIUS" and CURT ENZELLb 'F.nvrronmen/ Toxicology Crnup, IYalfenberg Laboratory, Stockholm Unfaersily, J.lUlu3S(ockAobn, mid"L7remical Resrarch Oepurpnenl, Swedish Tobacco Co., S-f 0462Sfockkalm(Sweden)(Ruceived Octa ber 14 tli, 1974) (dtcvisiortrcccivedMarck 34th, 1976) (,AccePted April 10th, 1975) SUMMARY Ascites sarcoma BP8•el , cultured in suspension in vitro were used asa 6mncrcd toxicity test system for tobacco and tobacco smoke constituents. Some 250 compounds, representative of these materials, were examined; by e,yosina cells to different concentrations of these constituents and mea- suring the inhibition of culttueero.wth, which_Hrasselated-to eoponding. effects encosnkerr.d f„r pc&i;tiyP standards. When employing the presentcell toxicity test system possible effects of factorss such aspenetration, distribu- toon and microsomal metabolism of the compounds studied, are not taken into account. The most active constituents were found to be unsaturated aldchydess and kelimes; ghm^]s and indoles. The good correlation observ- ed between functional groups and toxicity permits, within the range of fuuctionalitics studied, prediction of the toxicity for a compound of known structure. _ . INTRODUCTION The toxic effects of aa complex mixture on higher organisms can be evaluated at various levels of complexity, both with respect to the target organism and the material! to which the target is:exposed;. In the case of tobacco smoke effects on man, studies at a low level of complexity in- volving effects nf individual compounds on, variouss cells and subcellular •Supported by a research fellowship with t11e Swedish Natural Science Research Council, and.a research grant from the. Swedish'rnhneca Company. . 49
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Jnda is poten- recovery tlity and mpounds ols, ben- ldehydes '28t, most sandN- lal toxic- 4t growth ated oxi- le plasma lity in the ciliotoxic e ciliated motility '], it may is investti- 1 Kazemi- work was REFERENCES , 1 B.A. Afaeliua, The role of cilia in man, in S.V. Perry et al. (Eda.), Contractile Systems in Non-Muacle Tiuuee, El.evier/North-HoBand Biomedical Press, 1976. 2 B. Mowherg, Human tracheobronchial clearance by mucociliary transport and cough, in L Belin, B. Jirvholm., S. Larrton and G. Thiringer (Ede.), European Journal of Respiratory Diseases, Suppl. No. 107, Val.,61, 1980, p. 51. 3 W. Weiee and W. Weiea, Arch. Environ. Health, 14 (1967) 682. 4 P. Bernfeld, C-W. Nixon and F. Homburger, Toxicol. Appl. Pharmacol.,.6 (1964) 103. 5 S.P. Battiaa and C.J. Kender„Amh. Environ. Health, 20 (1970) 318. 6 T..Dillumn, Am. Rev, Respir. Dit, 93 (3) (1966) 108. 7 E:L. Wynder, H.E, Kaiser, DA. Goodman and D. Hoffmann, Cancer, 16 (1963) 1222. 9 E.L. Wynder, D.A. Goodmann and D, Hoffmann, Cancer, 18 (1965) 505: 9 C.J. Kenaler and S.P. Battiata, N.. Engl. J. Mad.,, 269 (1963) 1161. 10 R. GuBlerm, R. Badn9 and.B. Vignon, Bull. Acad. Nat. Med-, 145 (1961) 416. 11 T.R. Walker end J.E. Kiefer, Science, 153 (1966) 1248. 12 T. Delhatnn and B. Lagaratedt, Arch. OtolaryngoL, 84 (1966) 107. 23 T. Dalhuun and A. Roaengren, Arch. Otolaryngo4, 98 (1971) 496. 14 T. D.lhamn, Arch. Otolsryngol., 87 (1968) 162. 15 J.D.Charry and.D. Taylor-Robinson, Appl-Microbibl., 19 (1970) 658. 16 M.A. Sleigh, The nature and actionn of reapiratorytract cilia, in J.D. Brain (Ed.), Respiratory Defence Mechanism, Dekker, New York, 1977- 17 S.G. van den Bergh, B.B.A. Library,.7 (1966) 125. 18 LA. Timofievskaya, N.l..Lanova and E.S. Balynina, Gig. Tr. Prof-Zabol., 3 (1980) 26. 19 E. Shauenstein, H. Eaterbauer and U. ZoOner, Aldehydess in Biological Syatema, Their Natural Occurrence and Biological Activities, Pion Limited, London, 1977. 20 J. Aakby, B.M. EBiot and J. Styles, Cancer Letters, 9(1980) 21. 21 L.W. Wattenberg and W.D. Loub, Cancer Rea., 38 (1978).1410. 22 I. Schmaltz and D. Hoffmann, Chem. Rev,,. 77 (1977) 295. 23 G.M.DonneRy and H.E. McKean, Arch. Environ. Hedth,.28 (1974) 350. 24 A. PilotH, K. Ancker, E. Arrhenius and C.R. EnxeB, Toxicology, 5(1975) 49. 25 B. Petterason, M.. Curvall and C.R. EnaeQ Toxicology, 18(1980) 26 M. Thelestam, M. Curnll and C.R. Enzell, Toxicology, 15 (1980) 27 1. Florin, L Rutherg; M. Curvall and C.R. Enzell, Toxicology, 18 1. 203. (1980) 219. 55 m N 1+ !O N ~ ® O
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, nvolned ve con- r on the ie more Laed for uea was aningful t 16-d4y n•ry and a plastic HMEM), -oughout ues, and ! traehea chea was mt.1mm red to a ;assed in- ier these However, after pre• icroscopy i perspex with an md to be :ed to ex- 7-monitor v activityg luring the o tape for lete cessa• •. All com- TABLE I THE CILIOTOXICITY OF MENTHOL (1 mM) A comp.ri.onn of the tims to ci6c.GUis between oneselectad section .nd the whole Orached ring. I Selected section Whole ring 1 32 38 2 84 34 8 22 30 4 81 31 5 48 54 8 62 62 7 48 46 Me.a (S.D.) 87.9: 10.9 40.7 * 9.9 pounds were tested at the same concentration (5 mM) and examined on at least 3 different occasions, each experiment involving rings from different tracheal preparations. At the magnification used, it was not possible to display the entire c:roas- section of the tracheal ring. Instead, one part of the ring having well-exposed cilia was selected and studied. It followed, as shown in Table I, that there is a good correlation between the cessation of ciliary activity of the whole ring and that of a selected section of it. Since the compounds were added as ethanol or DMSO solutions to the medium, these solvents were tested over the period of time required for the experiment (60 min). DMSO was found to be the least toxic solvent and could be used at concentrations below 15% v/v while ethanol could not be used at concentrations exceeding 5% v/v. Both solvents were non- toxic to the cilia at the concentration used in the experiments (1.6% v/v). Compoundsexamined . Some -300 compounds, most of them abundant in tobacco and tobacco . smoke, have been esamined.'Those compounds, which were not commer- ciaRy available, were either isolated from tobacco, (e.g. the terpenoids) or synthesized from available starting compounds, using common methods of synthesis, (eg. most ethers, indoles and nicotine analoguea). All compounds were checked for purity using thin-layer chromatography, gas chromato- graphy and NMR. Compounds containing more than 3% impurity were purified using liquid chromatography, recrystallization and distillation. The structures of the compounds studied were confirmed by `H- and "C- NMR, e.g. the substitution pattern of multisubstituted aromatic substances and the branching pattern of methylsubstituted long chain alkyl derivatives. RESULTS In order to simplify the interpretation of the results, which are sum- 43 OD Q N M tD N !a
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22 3-MethYlaniaole 28 4-Methylaniaole 24 2,5-DimethYlenfaole 25 8,6•DimeRhyl.nieole 26 2,3,6-'hlmethylaniwle` 04 14 28 . 88 ~..-...._ __......-~.._ ~.._ 47 ]nduie•9+ceticadd • 48 4•Hydrozyquinaldlcacid (0.8 mM) . 49 pd'hthellcacid - ..., >60 >60 >60 ile it Dl th lb 3 Esters and .tnhydridea 84 r enaon me y 2,4- Hl lb l 4 50 Vinylaeetate >60 86 enzon e 2,6-Dinuthy t ` 51 Benzylacetate > 60 86 2,6-DlmathylbenzonitrBe ` > 60 52 Methylpropenoatea > 60 87 3,4-Dimethylben:onitrile ` 3 68 Ethyiatearate > 60 88 2,4,6-1Yimetbylberuonttrile 10 64 Benzylbenzaate° >60 89 2-EtttYlbenzonit:8e 6 55 Benzyl-8-phenyl-propenoate` >60 90 8•Ethylbenzonitrile 13 56 Coumarln . >60 91 4-Etbylb.osonitrite e 14 57 Diethylmalonate >60 92 1•Napbtbonitrlle 6 68 Diethylphthalate 8 93 2-Naphthooitrik` . 24 59 Di•n-propylphthalate° 52 94 Nicotinonitrlle >60 60 Dl-n-butylphthalatee e >60 95 Indolyl•8+cetonitrile 8 61 Di•n9ctylphthalate >60 lic acid anh dride 62 Phth > 60 Ketones y a 96 2-Propanone > 60 Amiderondnitrilee - 97 2-Butanone P > 60 >60 63 Nicotinamida > 60 98 entanone 2- 64 N-Methylnicotinamide >60 99 2-Hexanone >60 65 Ethanonitr0e . >60 100 2-Heptanone >60 66 Propanonitrile >60 101 2-Decanone 24 67 Butanonitrlle >60 102 6•Methyl-2-heptenone 10 68 PentanonitrBe >60 108 8•13uten-Sone 8 69 Hexanonitrile 16 104 8-Methyl-8-buten•2one 19 70 Heptanonitrile 12 105 1-Penfen•Sone 11 71 Oetanonitr9ee 18 106 3-Penten•2-one 38 72 Decanonitrile` > 60 107 4-Metbyl-3•penten-2bne >60 78 2-Methylpropanonitrile >60 108 6-Methyl-5-hepten•2one >60 74 3-Methylbutanonitr9e > 60 109 6•Methyl-8,6•heptadien•2-one 20 75 Propenonitrile > 60 110 2,8•Butandione 36 76 Phenylethanonitrile >60 111 2,8:Pentandlone >60 77 3-Phenylpropanonitr0e >60 112 Cyclopentanone >60 78 3-Phenylpropenonitrile 5 113 Cyclobenanone >60 79 Benzonitrile >6 0 114 2,2,6•74imethylcyclohezanone 14 80 2•Methylbenionitrile , 24 116 2-(•yclohe:enone 35 81 3-Methylbenzonitrile 6 116 Piperitone` 14 82 4•Methylbenzonitrile 26 117 Carvenone 18 83 2,3-Dimethylbenzonitrile 4 118 Carrone 6
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as TABLE I INHIBITORY EFFECT OF 320 TOBACCO SMOKE COMPONENTS ON NORADRENALINE INDUCED BESPIRATION No. Compound % inhibition at level 1mM 0.1mM Atcohola 1 Methanol 0 2 Ethanol 0 8 Hexanol 32 4 Octanol 96 21 6 Nonanol 95 43' 6 Decanul 93 45 7 Pentadecanol 15 8 Hexadeeanol 7• 9 2•Propanol 1 10 2•Hexen•1-ol 100' 19 11 Tetrehydroyeraniol 96 36 12 Genniol 96 38 13 Farneaol 91 37 14 Menthol 91 21 15 a-Terpineol 49 16 a-Ionol 55 17 p]onol 54 18 Propylene glycol 25 19 Furfurylalcohol 9 20 Benzylaicohol 32 21 2-Phenylethanol 32 Efhers 22 Anisole 23 2•Methylanisole 24 3-Methylanisole 25 4-Methylanlsole 26 2.6-Dirnethyl.ni.ole No. Compound % Inhibition a0lerel 1mM 0.1mM 27 8,6-Dimethylanbole 4 28 2,3,6•Trimethylaniaole 22 29 2,4,6•Trimethylanieole 21 30 2-Ethylaniaole 29 31 82 EthylphenyletAer Diphenylether 80 62 88 1,2,3•Trimethoxybenzene 22 84 2-Methoxynapthalene 27 35 2-Ethoxynaphthdene. 64 Acidr 36 Methanoic acid 20 37 Ethanoic acid 4 38 Hennoic acid 76ee 39 Octanoic acid 100e'• 67e• 40 Nonanolc acid 100' 87• 41 Decanolc acid 100e'• 100e** 42 Pentadennoic acid 78ie 43 Hexadecanoic acid 41** 44 3-Phenylprupenoic acid 91* 34• 45 Henzoic acid . 60•• 46 2,6•Dihydroxybenzoicacid 25 47 3,6•Dimethory•4-hydroxy 0 48 benznic acid 3-Pl•ridine cuboxylic acid 15 49 Indole-3 -acetic acid 26 50 4-Hydroxyquinaldic acid 3 51 p-Phthalicadd. 11: Esters and Anhydiides: 52Vinr-l.eetan .
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6 uarac~aI . , EFFECT ON CIIJARY MOVEMENTS OF SOME AGENTS WHICH COME IN CONTACT WITH THE RESPIRATORY TBACTs By P.K.. Des,. R.S. lGntox, P.S. StHtu Atm A.K. S/tHrxr. Department ojPltarmacoingy, College ojMedica[ Sciences, Banaras Hindu University, Varanasi - 5. iratory epithelium, on ciliary activity without using any special solvent. I ught worthwhile to study the effect of a number of agents which are liable to affect the differences in the experimental models and solvents that have been used. It was, therefore, ~(6.7,8,1o to 16). The results of different workers are,.however, variable probably because of ties, volatile oils, alcohol and cigarette smoke on ciliaryactivity'have been reported relation to health and disease of the respiratory tract. The effects of a few volatile attaes- :wholly or partly into the respiratory tract. A study of the effect of alll those agents, which are _either inhaled or excreted into the respiratory tract, on ciliary activity is of practical significance large number of abnormal gases and fumes like cigarette smoke, petrol gas, industrial fumes and gases, medicinal gases, vapours, aerosoles, etc. Bpdy also excretes a number of drugs respiratory secretions which is a distinct aid to infection. Upper respiratory tract in&ctions, bronchitis, pneumonia, asthma, pneumoconiosis, bronchiectasis, pulmonary atelectasis, eto.may result or be abetted by faulty ciliary function of the respiratory tract (I,4,5,9). No other tissue of the body with such a delicate system has to face, and~ now a days increasingly so, such a minate infectious agents and foreign particles. A defect in ciliary activity leads to stagnation of Ciliary action keeps the viscous film of mucus moving towards the pharynx, thus helpingtoeli- Ciliary activity is one of the important protective mechanisms ofthe respiratory tract. MATERtALS AND METHODS creosote, potassium iodide, oxygen, carbon dioxide, petrol gas and cigarette smoke. All Ethyl alcohol, paraldehyde, ether, chloroform, oil of peppermint, menthol, thymol, IDimts were conducted with the following agents :- ings were always interpolated between the observations with the agents studied. Experi- e were at least 3 nearly constant readings taken at intervals of 5 mirs.. A number of control trol effect (time to travel I cm) in each ease. The effect of the agent was. studied only when ce of 1 cm wascalculatelt and the effect of the agentswas expressed as per antage of the 'aHsPO,, 0.083 mM was used. In all observations time taken by poppy seeds to travel aa unit The eiliary tttotility of frog's oesophagus was studied by the method described earlier (2). ger solution of pH 7.5 containing NaCI, 111.11; KCI, 1.88; CaCIs, 1.08; NaHCO., 2.38 and
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Extension of the aromatic system of phenol to that of a- and p-naphthol (206, 207) increased the ciliotoxicity. It.is notable that some of the most abundant phenols in the weak acidic fraction of tobacco smoke, i.e. phenol, catechol and hydroquinone did not display any ciliostatic effect in this test. Hydrocarbons The acyclic hydrocarbons, saturated or unsaturated, were non-toxic. Of the cyclic, non-aromatic hydrocarbons, cyclohexane and cyclohexene showed a ciliotoxic. effect, while the three monoterpenes, limonene, a-pinene and p-pinene were inactive. These three compounds are the only terpenoids tested that did not exhibit any cillotoxic effect. Benzene itself was non- toxic, but all the alkylsubstituted derivativea (217 L28) were toxic. How- ever, no clear correlation between activity and position, number or size of the substituents on the aromatic ring could be observed. Naphthalene and its methylderivatives inhibited the ciliary function while all the other polycyclic aromatic hydrocarbons, except acenaphthylene, were inactive. Furanes and thiophenes In this group comprising only six compounds, the monocyclic furanej and thiophene were non-toxic, while the heterocyclic benzofuranes were ciliotoxic. N-Heterocycles Neither the non-aromatic (253-255) nor the aromatic (256-264, 266- 268) monocyclic N-heterocycles, inhibited the ciliary function. All the pyridines tested, including those carrying alkylsubstituents (258-263) and other functional groups (46, 94, 127, 171, 264) were non-toxic to the cilia. Consistent with this, all the nicotine analogues examined (2881L921, which also contain a pyridine moiety, were non-toxic. In contrast, the fused bicyclic representatives, quinoline (265) and the indoles (47, 95, 172, 269-284), were all ciliotoxic. As the indoles include many biologically active compounds [20,211 and as they are abundant in tobacco smoke [22], a selection of mono-, di- and trimethylsubstitated indoles were.tested. Most of them were found to be.potent.inhibitorsofthe ciliary function as they caused ciliostasis within a few minutes; despite precipitation, on addition to the medium. The two indole derivatives nor- harman and harman, also displayed a ciliotoxic effect. Aminea Practically all members of this group were non-toxic to the cilia. Of the aliphatic amines only n-decylamine displayed any ciliostatic activity. Most of the anilines were non-toxic. Introduction of alkylgroups into the aromatic ring did not affect the ciliotoxicity, while enlargement of the ring system, to that of a- and p-naphthylamine significantly increased the ciliotoxic effect. The two diaminotoluenes were non-toxic. ib ~ 52 IWA
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Go TABLE I (Continued) No. Compound % inhibition at level 1mM 0.1mM 245 Picene 0 246 Coronene 8 247 Aeenaphthene 15 248 Acenaphthylene 16 249 Fluonnthene 7 Furnnee and thiopenes 260 Tetrahydrofuran 9 261 Puran 7 252 2,6-Dimethylfuran 17 253 2,3-Benzoturan 46 254 Dibenzofuran 86 255 Thlophene 16 N•Heterocyclee 256 Pyrrolidine 257 3-Pyrtoline 258 Piperidine 259 Pyrrole 260 Pyridine 261 2-Methylpyridine 262 2,6-Dimethylpyridine 263 3,5•Dimethylpyridine 264 2,3,6•Tr(methylpyridine 265 2,4,6•Trimethylpyridine 266 3-Ethylpyridine 267 3-Hydroxypyridine 268 Quinoline t No. Compound . 269 Pyraine 270 2-Methylpynzine 271 2,6-Dimethylpyrazine . 272 Indole - 273 1-Methyllndole . 274 2 Methyllndole 275 3-Methylindole 276 6-Methylindols 271 7-Methylindols 279 1,2-Dhnathylindole 279 1,8-Dimethylindole 280 2,8-Dimethylindole 281 2,6-Dimethylindole 282 3,6 Dimethylindole 283 1,2,8•Trimethydndole 284 2,3,4-Trimethylindole 285 2;3,5•Trlmethylindole 286 2,3,6-Trimethylindole 287 2,3,7-Trimethylindole 288 Carbazole 289 9-Ethylcarbuole 290Benzimidazole291 Nornicotine 292 Nicotine . 293 8-Nicotyrine 294 Anabasine 295 Cotinine 296 Harman297 Norharman % Inhibition at level 1 CoM - 0,1 mM 12 15 12 26 47 78 62 85 65 88 17 87 69 66 14 64 90 47 75 30 76• 7 0 4 38 . 0 0 100 53 89, 21 '
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TABLB I (Continued) No. Compound % Inhibition at lenl . No. Compound % inhibitlon at tevel 1mM ' 0.1mM 1mM. " 0.1mM 128 1-Phenyl•2-butano»e 62 148 2•Hezend 86 124 1•Indanone 24 149 2,4•Hezadienal 100 20 125 9•Fluorenone 54 150 a-0yelocitral 52 126 2,3,6•1Ylmethyl-1,4• 87 151 Safranal 94 naphthoqulnone 152 Phanylethettal 83 127 (8•Pyridyl)-1•propenona - 15 153 8-Phenylpropztul 91 154 . S•Phenylpropenal 87 Aldehydn 155 Benzaldehyde 47 128 Propanal I 186 2•Metbylbenzrldehyde 81 129 Butanal 18 157 8•Methylbenzaldehyde 58 130 Pentan.i 84 158 4•Mathylbenzaldehyde 42 131 Hexanal 8o 159 2,4-Dimethylhenaldehyde 48 132 Heptanal . 92"• 160 6-Dimethylbenzaldehyde 2 38 133 Octanal 100""" 49" 161 , 2,4,6•TYimethylbenzeldehyde 28 134 Nonenal 96" 57 162 2•Methozybenzaldehyde 21 136 Decanal 100°" 61"" 163 4-Methozybenzaldehyde 29 136 Undacanei 100"" 73 164 8,4-Dimetho:ybenaeldehyde 2 137 Dodecanal 100""' 63"" 165 2-Hydroxybenzaldehyde 100 75 138 2-Methylpropanal 2 166 3•Hydrozybenzaldehyde 44 189 2-Methylbutanal 64 167 4-lfydroxybenzaldehyde 46 140 3-Methylbutanal 46 168 2,6-Dihydroxybenzaldebyde 89 141 2-Methylpentanal 22 169 3,4•Dihydrozybenzeldehyde 27 142 143 2,2•Dimethylpropanal Glyceraidehyde 33 0 170 4•Hydroxy3-methoxy benzaldebyde . 26 144 Propenal 100 80 1716•Methyl-2•furfurei 5 145 2-Methylpropenal 94 172 6•Hydroxymethyl•2-furfural 12 146 2-Butenal 61 173. 3-Pyridine earboxaldehyde 30 147 2-Methyl-2•pentena] 61 174 ' 3-Indole arboxaldehyde 66 w, .
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Aminna 309 2,3•Dimethyleniline 6 298 n-Butyl.mine 21 310 2,6-Dimethylaniline 34 299 n•Hexylemine 21 811 2,6-Dlmethylmlline 10 300 n•Decylamine 100 76 312 2,4,6•9Yimethylaniline . 8 301 2-Methylpropylunine 0 813 2-Ethylan8lne 30 302 1-Methylhutylamine 13 314 4•Ethylan0ine 48 803 2-Propenylemine 2fi 316 N-Ethylaniline 31 304 - Benaylamine . . 83 316 DiphenyWnine 58 806 Aniline 11 317 1-Nephthylamine 52 306 2-Methylaniline 17 816 2-Naphthyi.mine 67 307 3•MethylanBine 9 319 6-DLminotoluene 2 0 308 4•Methylaniline 13 820 , 8,4-Dlaminotoluene 24 Asteriska denote meterials which stimulate the buic rate of oxygen consamption and thus d.eieee6 the ma:imal nondrenaine inducable respiratory reta. . . . •60-240 nmo101min per 10' cells. - ~ •*240-i20 nmolO(min per 10' cells. <••420-600 ntnol O/min per 10' aBs. .
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Phonpfs °1~,, inh-bolt Imm a.I v.M Hydrocarbons 175 Phenol 37 210 n-Hexene 51 176 2-Methylphenol 58 211 2-Hexene 40 177 3-Methylphenol 65 212 2,4-Hezediene 36 178 4-Methylphenol 49 213 Cyclohexene 47 179 2,3-Dimethylphenol 77 214 GyFcMhexene 53 180 2,4-Dimethylphenol . 72 215 Limonene 37 181 5-Dimethylphenol 2 86 216 a-Pinene 41 182 , 2,6-Dimethylphenol 60 217 0-Pinene 42 183 8,4-Dimethylphenol 72 218 Henzene 10 184 8,5-Dimethylphenol 81 219 Methylbenzene 42 185 2,3,5-14imethylphenol 93 220 1,2-Dimetbylbenzene 81 186 2,4,5-14imethylphenol 93 221 1,3-Dimethylbenzene 40 187 2,4,6-Trimethylphenol 58 222 1,4-Dhnethylbenzene 51 188 2-Ethylphenol 100 22 223 1,8,5-'hlmethylbenzene 9 189 S-Ethylphanol 99 11 224 Ethylbeoxene 67 190 4-Ethylphenol 99 10 225 n-Pzopylbenzene 76 191 8-Ethyl-5-methylphenol 100 28 226 Iwpropylbenzene 73 192 2-lwpropyl-4-metbylphenol 100 42 227 Styrene 58 9Q 2-Methoxyphenol - 29 228 Phenyl.cetylene 60 194 3-Methoxyphenol 53 229 Indan 76 195 4-Methoxyphenol 14 280 Tetrdln 78 196 2,6-Dimethoxyphenol 9 2S1 Indene . 48 197 Eugenol 61 232 Diphenylmethme 80 198 leoeugenol 71 283 Azulene 46 199 Catechol . 6 284Nephthelene 48 200 3-Methylcztechel 75 235 1-Methylnephthelene 82 201 3-leopropylcztechol 100 18 286 2-Methylnsphthalene 48 202 Rezorcinol 1 237 1,4-Dimethylnaphthelene ' 87 203 Hydroquinone 5 28B Anthneene 18 204 Pyrogallol 75 239 Phananthrene 15 205 206 2-Hydroxyecetophenonq 8-Hydroxyacetophenone 26 26 240 Benz[a]antbrecene Chry'ene - 241 11 7 207 4-Hydroxyecetophenone 30 242 Pyrene 8 208 1-Naphthol 100 87 243 Hens[u]Pyrene . 7 . 209 2-Nxphthol - 97 21 244 Peylene 2
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:ii further testing' at lower concentraLions, the inhibiLingeffrct was still sLa- Listically significant (P < 0:001). The compounds tested were primarilyy selected: on thee basis of their abundance i-n--to ac^G-co or o ac6 cosmoke• ~cara r c. as solubilityin ethanol or dimethyl sulfoxide were lim- iting factors,, however. Included as well weree some tobacco and tobaccosmoke constituents having documented biological effects. The purity of the compounds was tested by thin-layer chromatography,, NMR or gas chro- matography. To obtain additional information on steric or substitutional effects on the biological activity, substances which are chemically related to the com- pounds selected according to the above principles were tested irrespective or their occurrence in tobacco or tobacco smoke. Most of these compounds are, however, also tobacco or tobacco-smoke constituents. RBSULTS The results are summarized in Table I. The substancess are subdivided; according to their functional groups; which does.not, however, imply thatt the functional group is responsible for the biological activity. Imspite of this, the effectsof the compounds will be discussed essentially in that order below. . A/cohols.. Most of the alcohols(1-12), were found to be non-toxic; in- Lerestingg exceptions were some of the isoprenoid alcohols. (5-8) which arr fundamental units in the biosynthesis of higher terpenoids. Since the allylic butnot the benzylic alcohols displayed activity, the. -C=C4-Ol{ unitt rather than the hydroxyl group itself appears to he the~ critical chemicali entity. Gihers The effect of anisol (13) is negligible but iiicreases on alkyl sub- stitution of the aromatic ring (15,16). Enlargement of the aromatic as well' as of the alkyl unit also seems to increase the toxicity. These effects re- quire furLherstucly. Acids. Of the acids (20-26), only indolyl-3-acetic acid (24) displayed some activity. In view of the results obtained for the unsaturated alcoholsthiss group willi later be extended to include polyunsaturated fatty acids. (PUFA). 6stersand anhydrides. A1li members of this group (27-39) which com- prise an aromatic acid exhibit toxic effect's exceptthe.anhydride (39).The. magnitude varies with the alcohol moiety; mostt remarkable is the effect of the chain length in the casee of the phthalic esters (35-38). AA similareffect has been observed for gallic acid esters [41, where maximum dis- turbance of the intact function of the microsoma] detoxication chain of livrr eells is displ:ryed by compouncls having a chainn length of 8 carbons. It is also of intr.rest that certain.esters arecapable of spontaneously forming, biologicully active clectrophilic derivatives under adequate pli conditions (51. Amrdes and nitriles. The amides and nitriles (40-51),, with the excep- 57
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so a,o-uw~cu~y~auu,na 53 Benzyl acetate 42 88 2,6-Dimethylbenzonitrile 48 64 Methyl propenoate 9 89 8,4•Dimethylbenzonitrile 65 55 Ethyl atearate 4 90 2,4,6-7rimethylbenzonitrile 43 56 Benzyl benzoate 75 91 2-Ethylbenzonitrile 58 57 Benzyl3-phenylpropanoate 88 92 3-Ethylbenzonitrile 89 58 Benzyl3-phenylpropenoate 82 93 4-Ethylbenzonitrile 80 59 Coumarin 21 94 1-Naphthonitrile 52 60 Diethyl malonate 59 95 2-Naphthonitrile 88 - 61 Diethyl phthatate 50 96 Nicotinonitrile o 62 Di-n-propyl phthalate . 29 97 ]ndolyl-3-acetonltrlle 86 63 Di-n-butyl phthaiate 9 64 Di-noctyl phthaiate 0 Xetnnn 66 Phthalic acid anhydride 19 98 2-Propanone 14 99 2-Butanone 17 Amid es and nitrilet 100 2-Pentanone . 14 66 Nicotinamide 0 101 6-Methyl-2-heptanone 68 67 N-Methylnicotinamide 81 102 3-Buten-2one 100 59 68 Ethanonitrile 2 103 3-Methyl-S-buten-2-0ne 57 - 69 Propanonitrile 2 104 1-Pentan-Sone 100 89 70 Butanonitrile 8 105 3-Penten-2one 12 71 Pentanonitrile 5 106 4-Methyl-3-penten-2ane 22 72 Hexanonitrile 19 107 6-Methyt-6-hepten-2one 89 73 Heptanonitrile 39 108 6-MethyL-8,6-heptedien-2one 98 8 74 Octanonitr8e 66 109 2,3-Butanedione 3 75 _ 2-Methylpropanonitrile 4 110 2,8-Paittmedione 11 76 3-Methylbutanonitrile 17 111 Cyclopentanone 3 77 Propenonitrile . 7 112Cyclohezanone 17 78 Phenylethanonitrile 40 113 2,2,6-Trimethylcyclohexanone 62 79 3-Phenylpropanonitrile ' 38 114 2Cyclohezenone 18 80 3-Phenylpropenonitrlle 71 115 Piperitone 41 81 Berixonitrlle 55 116 . Carvenone 73 82 2-Methylbenzonitrile 41 117 Ctrvone 42 83 8-Methylbenzonitrile 46 118 Paeudoionone 70 84 4-Methylbenzonitrile 69. 119 edonone ' 65 85 2,3-Dtmethylbenzonltrile . 67 120 p-lonone 71 86 2,4-Dimethylbenzonitrile 66 121 Acetophenone 82 87 2,5-Dimethylbenzonitrile 40 122 1-Phenyl-l-butanone 93
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tion of valeronitrile (46) and indolyl-3-acetonitrile (51), are inactive. Part of the effect found for indolyl-3-acctoniLriltb may possibly he associated with the indole ring system as indolyl-3-acetic acid (24) was thee only acid studied: (20-26) showing some activity while all other indbles examined wero found tobe toxic. '1'hese findings are somewPoatt surprising since nitriles constitute a group including many - biologically active compounds.. Thus besides the ahove- mentionedindolyl-3-acetonitrile,d which is a plant growth hormone. [6], phthalbnitrile and di- and trichioroacetonitriles havebeen used as insec- Iieides cmd malononiUrile is.uf I!sychopliarmacologiaaI intemsL [7J. More- over, over, fi-amiiiopropioniGrile is known as the Lathyrus factor in sweet't peas causing bone damage in newborn rats [18]. The toxicity of nitriles iscon-sidered Lo. he dependent on thcir mctahu)ism to cyanide ions; this type of toxicity will thus depend on where this metabolism takes place [7]. This group may thus represent aa case in which the present test system, due to lack of -deloxicatingmetabolismg in•then ascites cells,, produces inadequate results. The nitriless should therefore beexamined; in a test system involving.metiibuGcdetoxication functions. !{elones. The ketones (52-77) display - not unexpectedly in vieww of . the fairlyy laigee number of representatives - great variation in toxicity. Of the saturated acyclic monoketones (52-54), only 6,10-dimethyhmdecan-2- one (54, tetralrydrogeranylacetone), a flavour compound,, is active... In con- trast, the corresponding diketones (55,56), and Lhce unsaturated acyclicmonokctones (b7~'i3), except for3-huten-2-one (57, methylvinylketone)are toxic. The situation is much the same for the cyclic compounds (64-77):, where the presence ofaf double. bond (68-70) or an additional oxo group (76) is also a requirementfor toxicity.. The effect of the double bond is evidently larger in conjugation with the oxo group. Since this effect is a5- . sent when the double bond: is replaced by an aromatic nucleus (71,73'- 75,77)it appears that, as in the.case.of the unsaturated alt.ohols, the double bond has a considerable impacL..A possible explanation for this is.the capa- cil'ryof n,(3-unsaturated ketones to be hydrated producing molecules which, lilce the diketones should be prone to complex metals, thus directly or in- directly affecting metalfo-enrymcs (vide infra). [911. Aldefiydes. Virtually all aldehydes. (78-122) are toxic, even the saturated aliphatic (78-94). The activity of the latter are only moderately affected by thee different side-c:hainsandthe most pronounced effect, apparently associ- ated with steric hindrance,, is observed for 2,2-dimethylpropanal (94). A small dependence on chain length is also observable for the n-alkyl aldehydes (78-89); the Cs-Cya representatives (83-87)are somewhat more toxirthan the rest. InLivduction of a conjugated.doublc bond in tltcaliphatic aldehydes (97- 102) clearly increases toxicity,, but no such effect is observed on conjuga- tion with an aromatic ring. The importance of the double.bond observed in the case of the ketones thus reappear in that of Lhe.:ddehydes. Both uon- aromatic cyclic aldehydes (103,104) are toxic,, roughly to the extent ex- .58 ... . .. ,~r'._, _. _,{
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~ Volume 14 Drugs on Ciliary Movements 30t Number 4 _ ~ Direct application of ether was found to stop ciliary activity temporarily, and withdrawl of ether was followed by a short period of exaggerated activity. In the present study. however, ether solution.in Ringer was found to excite ciliary activity. This may be due to the known irritant .y~ aetionn of the anaesthetic. It is probable that long continued irritation of respiratory epithelium F for several hr during ether anaesthesia may result in an exhaustion of ciliated cells, which may be responsible for increased incidence of respiratory tract infections reported after ether anaesthesia. .~., Alcoholism being a social problem in any society, a number of workers have studied the effect of alcohoton ciliary activity (8,10,11,12,13,16). Alcohol in high concentrations (5 to 20%,) -" has been reported to retard or arrest ciliary motility of the trachea of horse, rabbit and fowf and the oesophagus of frog. There was, however, wide variations in the actual concentration .; ofakohol which depressed ciliary activity probably because of the different biological prepara- ..s tions used.. In the present study very dilute solutionss of alcoholl were used. In a very low concentration (0.02%) it was found to stimulate ciliary activity while in concentrations of 0.2 to 1.0'/ it caused significant depression. The present study thus shows that alcohol hasan effect on ciliary activity and it.is possible that the deleterious action.of alcohol on ciliary funa -tion may be one of thefactors responsible for making the alcoholics prone to respiratory~ infections. One percent solution of paraldehyde was found to be a stimulant of cilia. However, the concentrations in which it is expected to bc exaeted in the lungs in clinical practice are very much lower, and in these.concentraGons it might not have any significant action on theeilia. , Several non-specific substances including -volatile solids, oils, etc. find their place in `'verious formulations- used for respiratory tract inflammations. Oils of clove, peppermint, spearmint, cinnamon and thymol have been reported to depress ciliary activity (11,16), while ' menthol and camphor have not been found'to have any significant effect (14,18} In the present >:{`fady, oil of peppermint, guaiacol and creosote markedly stimulated cilia activity while megthot ~Tiadamild stimulant action. Thymol definitely retarded ciliary beat confirming previous '.-Ieports. Thymol, a phenol derivative, having a weak local anaesthetic and anthelmintic ;>tctions may be directly depressant to the cilia. The ciliary stimulant action of oil of peppermint, Potassium iodide often forms a constituent of various formulations used for respiratory sote and guaiacol may be responsible for theirr expectorant actions. eases: Potassium iodide in concentrations of 0.1 % was found to markedly depress ciliary ,~ .. .;IOaottlt[y. Potassium chloride was not found to have any signidcant action while the bromide was moderately depressant.The results indicate that the depressant action was not due to cation but because of the anion . It was reported earlier that the chloride ion in the 7, iger solution cannot be replaced by any other ion (3), and the present study further ahows 3the bromide and iodide ions are actuallyy depressant, the latter being more so, possibly %Cause of its irritant action.. The depressant action of potassium iodide is not of any practical : 4gGificance a.far as systemic use of the drug is concerned because its therapeutic dose is very ,;guall. However,potassium iodide should not be used in.nasal drops.
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taining 5% carbon dioxide and capped air-tight to maintain a stable plf of approximately 7.3 (phenol red maintained at pink to yellow). The cell cultures were reinoculated to a cell density of 0.1 • 10°' cells/ml'every5thday. The cell densities were calculated with an -electronic cell ` counter (Celloscope 401, Linson fnstr. AB,. SLockholm,, Sweden). For the - tests, the cell suspension was diluted with sterile medium to an initial cell - densityof 0.4 • 10" cells/mh The compounds to be testedd were dissolved in ethanol (10 pl) or di- methyPsulfoxide (]10 p) and added to this.susl,iension (3 ml), each in amounts . to givce the final conccntrationsindicated in 'I'able I. All compounds were incubated at 37° for 48 h. Solvent (10 MI) was added to the controls. After gassing and capping, the tubes,.enclosed in a gassed (air, 5% carbon dioxide) and sealed plastic box to prevent errors due to p11-changes caused by gas lL•aksinto single tubes, were incubated obliquely. 'PGSTINO AND GVALUATION-PRINCII'LF.S '1'he tests were run..in duplicutes and' the cells were allowed to grow for 'I8h. During this pe:iod the number of cells in the. controls increased ex-ponentially, i.e. dN/dt = kN where N is the cell! density and t is t'hetime, indicating that none of the. essential growth factors in the medium was limiting. The growth rate of an incubated cell culture was calculated and compared to the average value of8-1',0 controls.performed in each series..'l'he.doubling. time for control cultures was approximately 24 h~ No systematic distinction was madebetweem viable and total cell count. The effect of the tested com- pound: is given as the ratio between the growth rates of the incubated cell culture and the controls, expressed as a percentage.'1'he growth rate of a cell culture,.u; was calculated using hi(N/Nu) t. 1n2 -(Nc = celU density at the start of the experiment). Ass the growth rate may vary between the different series, eachseries also contained 3 or more internal standards in duplicate, Le. fixed concentrations of substances having documented inhibiUing effects. These vubstances were in most cases 2-aminonaphthalene,, propranal, quinoline and isopentone. Thee relative effects of these internal standards were the same for the different series and independent of growth rate. All substances were initially tested by exposing the cells.to a final concen- Lrntioiiof 1 mit! (cell medimn • solvenL . When a-tiis concent'ration a com- pot-mT nh ~bited growth rate to 50^/, or more, furtierexperimentc.a ower. conceuLraLmns were. perfornne (.1,. 0.01 mA4, etc.) until a~ concentration wass reached which produced an inhibit y 9ow 50`Yo. The fact that manycompoun s were not tested at the lower concentration does not, of course, imply that all of them were inactive. In most cases when the inhi- biting effect wiLC within the 15--50^,f region, thus eliminating. the need for 56 t (
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0 0 5 10 15 rNnutes Fig. 1. Noredrenaline (NA) stimulated oadatln met.boliaa of iwlated b.own fat cella ,.. in the preaance of eNunol (control) or harman (0.1 mM). The cell conoantntan wae approximately 10` celWmi aaaW the terinpanturo STC. The inWbitory effect of human.:' i. 51% in this experiment. . . . and tobacco amoke, have been exam+ned. Those substances, which were not . commeraially available were either isolated from tobacco (e.gz the terpenoids) or synthesized from available starting compounds, using adopted methods of . synthesis, (e.g. ethers, indoles and nicotine analogues). All compounds were checked for purity using thin-layer chromatography, gas chromatography and nuclear magnetic resonance (NMR). Compounds containing more than 3% impurity were purified using liquid chromatography, gas chromatography, recrystallization and distillation. The structures of the compounds studied : were confirmed by 'H- and "C NMR, e.g. the substitution pattern of multi- substituted aromatic substances and the branching pattern of methyl- substituted long chain alkyl derivatives. RESULTS AND DISCUSSION . The compounds tested were divided into 12 chemical classes according to functionality to simplify the interpretation of the results. The results are presented in Table I and each figure represents the mean value of at least 3 experiments, each performed on different cell preparations. Some of the compounds were found to stimulate the basal metabolism of the cells and t.hese are denoted with asterisks in the Table. Alcohols The effects of the alcohols on the noradrenaline stimulated respiratory rate varied considerably. The inhibitory effect of the primary aliphatic alcohols (1-~8) variedd with the length of thee carbon chain-and showed a:: _ maximum for the C.-C,e representatives, which were highly active. Similarly .
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Ihe hepatic concentrations of both I U DPGA (Notten and Henderson, nilarly, phenobarbital has been .ncrease the synthesis of UDPGA with isolated hepatocytes{Notten 75). In this papcr we have ul- .) probethe relationship between nobiotic metabolism and the en- the D-glucuronic acid pathway .ISe I mfa biotransformation en- 'eot induced as a consequence of istration of the xenobiotic. The s chosen for study were tliphenyF I(Dixon et n1,.,. 1977a), guaiacol 1959), 4-hydroxybiphenyl (West 56), 2-naphthol (Berebom and 51), 1-naphthylacetic acid (Dixon b), paracetamol (acetaminophen) tl., 1974),.2-phenylpropionic acid tt., 1977c), and N-propyl gallate al.,. 1959; Dacre, 1960), all of ~ been shown to be either directly and/or metabolised to products substratess for Phase II enzymeslian liver. We also investigated ,fphenylacetic acid which unlike umpounds is not conjugated with ic acid inn the rat (Dixon et a1., tes et al., 1972). A preliminary ition describing part of this work .delsewhere(Lakeetal., 1978). METHODS 1-isocilric acid, UDPOA, 4-hydroxy- opyl gallate„Tris, I-naphthyl-S-o-gluo- Ihylumbclliferyl-fl-n-glncuronidc, isoci- :cnasc.l EC 1.1.1.42,.from. pig hean6 and .r /1'.(' l...l.ll. frow hwinc liver/ were n Ihe 3ignta Chcmieal l•nrtlp:my, 1'ook, md. (inaiacol I'--mtlhnxyphcnon,. 2- pha•nYla.eli. acid wcre uhlnined from ds, Poole, DorseL England, and di- zid I-naphthylacetic acid and 2-phenyl- I from Aldrich Chemical Company, nset, England. Paracemmol (N-acetyl- • acet:uoinophen) was B.P. grade andttx were of the highest purity available. o-GLUCURONIC ACID PATHWAY REGULATION AnhuaG md Trentnrrnt 6(ale Spmgue-Dawley mts' (40-60 g) were housed io.n raoms maintained at 20 m YC with a relative lumidity of 70-70%..They were allowed free access to foodr and water. After acclimalizing the animals In.lhe exM'rirncnt:d rrKimve far 7 duys„ Ihe' Ir•al cnm pounds were adminislercdhy daily ipinieaiuus 1'ur a period of7days. Dose levels employed werea0 mglkg/ day for 2-phenylpropionic acid,. 100 mglkgldayfor diphenylacetic aeid, guaiacol, 2-naphthol, and I-naph- Ihyhceticc acid andI50 mglkg(day for 4-hydroxyhi- p&nyl, paracctamol- phenylacetic acid„and n-propyl gdlak. Control animals received corresponding quanti- liea of the com oil vehicle (5 mlJkg body wt) except fp parseetamol which was administered in 0.9% Na7 (5 mlAtg) and for n-propyl gallate where the carn oil vehicle inleclion volulne was 10 mlfkg. As Ihhere were no significant differences observed between the control animals receiving the various treatments desuibedabove, the hepatic and urinary date from all control groups was pooled to provide the normal values shown in Tables1 and 2. paper represenP those of the '-nalivi' unuelivated• enzyme. Urine Anrd.air pi,r Ibe enlleclion of 24-hr urine s:mrples, :mimal,u - were cagcd singly in all glusx "Mclalydwls.' A smnll quantity of NaF was added as a preservative tothe.urine collection flasks. Urinary creatinine was deter- mined miaed by9he methodbf Bonsnes and Taussky (1945).Thc urinary concentrations of o-gluttniclsid. 1- gulonic acid, and xylitol were determined by a gas. - chromatographic procedure previously deseribed (Lake er a/., 1976). with Ihemodifieation that the•: chromatograph column length was 5.5 m instead-of '. 2.8 m. For Ihaeslimation of total uwglucuronic acid;..' urine samples were processed and derivatised asde-'~,: scribed by Gangolli et al:, (1974)) except that 6.0 mg -_ of ol.-tanark acid was substituted for n-eicosanf:'e as the Inlernall standard,, and the chromatography , was performed as described above. The urinary t~-::-,- crction of the o-glucuronic acid melatiolites was expressed as microgram of metabolite per milligram of - creatinine instead of milligram metabolite per rat per:- Rats were killed by cervical dislocation and the livers immediately excised into ice-cold 0.154 w,. KCI containing 50 mwt '1'ris-IICI buffer, pll 7.4. All subsequentt operations were perfnrmed al 0-4'C. Whole liver homogenaoes (0.25 g fresh.lissue/ml) were prepared employing a Potter-Iypc..TeOon-glass; rllolor- driven.homogcnizer.' Homogenates were centrifuged al 10,000g., for 20 min and the poslmilochondrial supernalant fractions used for the determinations of dhylmorphine N-demethylase (Holtzman e1 ai., 1968) dphtnyl 4-hydroxylase (Creaven evaf„ 1965),.and aollline 4hydroxylase (Nakanishi et al., 1971). Portions of the 10,000g„ supernatant fractions were further centrifuged at 105,000 g,,, for 60 min to sediment the microsomal pellets which were washed by suspending In fresh homogenizing medium and again sedimented sll 10S,0g0g,,.Washed microsomal fractions were assayed for cytochrome P-450 (Omura and Sato, 1964), protein (Lowry sr oh, 1951), and UDP-glucuronyl- trmsfemse :wlivily employing 4-mclhylumhelliferonc INealc and Parke, 19731 and. I-naphthol (nrwk, 19741 asaglycoues. Aa mitnrmuu:d.fractiun.vwere nol ua'li- valldby lre:umenl with dclcrgcnts, clc., Ihc UDI'- gN¢urnnylvanxfcr,lx aclivilies referred lo, in Ihis "OLAC1976 Ltd.,. Blackthorn, Biccsler. Oxon England:.t.aboratory Animal Centre accredited. r Spratts' Laboratory Diet 1, Barking, Essex, England. A. H. Thomas Co., Philadelphia, Pa. 24-hr urine (take rr af., 1976) in order to obviate any. arors due to the incomplete collection of 24-hr urino-.samples. In the experiments described in this paper; none of Ihe testt compounds employed had any signifi-' etml effect on the daily (24 hr) excretion ofcreatinine (normal excretion 62.5 E1.4 mg/kg b,xlywU24 hr, ..: mean ± SH, rr = )6).. RESULTS Hepatic Effects None of the nine compounds adminis- tered had any significant effect on eitherihe` activities of the three mfo studied or on the microsomal content of cytochrome P-450 and protein (Table I). In contrast to the ob- served lack of stimulation of the Phase I_ enzymes of hepatic xenobiotic metabolism, some of the compounds administeredre- sullcd in increased nsicros(nnalt Ui)I'-gluc- . uronyllransferase activity. Gu.ua' I-naph- Ihylaccticacid, paracctmmol, and n-propyl-:l gallutc stimulale the rn_ny •„_ii_.. •+f~~h 4methylumbelliferone and I-naphthol, whereas diphenylacetic acid and 2-naphthol only - • icncons t.ld., Hemel Hempstead,. Hens. England. : 8'7119240
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jlt G r 11 V.SM. Bernson, P. LundberQ.and B. Pettermn, Biochim. Biophys. Acts, 687 (1979) 353. 12 B. Pettersson and I. V.W n, Eur. J.. Biochem., 62 (1976) 383. 13 JP. Skala, P. Hahn and BY RniCht, The "second mesrenger" sydem in brown adipose tissue of developing nh~ Its moleeulsr composition and mechutiLn of functian., in L. Giradier . and J. Seydon (Eds.), Effeetora of Thecmogenesis,. Birkhii4ser Verlag,.Basel und Stuttgart, 1978. 14 B. Petter.wn, Chem.$iol. Interaet., 29 (1980) 98. 15 E: Bluerlein and H. Truch, Abstract, Xlth International Congress of Biochemistry, Toronto, Canada, 1979. 16 E. Biiuerlein and H. TYa.ch, Abstract, XXIUL International Conference on the Biochemistry of Lipids, Cologne,.(3ermanY, 1979. 17 R.L, Smalley, Ch.ngn in composition andd metsbolism during adipose tissue development, in O. Lindber` (Ed.,), Brown Adipose Tissue, New York, 1970. 18 C.M. Smith and J.R. Williamson, FEBS ktt.,1S (1971) 35.. 19 E. Shauenstein,.H. Esterbauer and U. Z611ner, Aldehydes in Biological Systemc,Their Natural Occurrence and Biological Aetivitiea, Pion Limited, London, 1977. 20 J. Ashby. B.M. Elliot and J. Styles, Cancer Letters, 9(1980) 21. 21 L.W. Wattenberg andW.D. Loub,CarrcerRea, 38 (1978) 1410. 22 E.R. Suter, J. Ultrstruet. Res., 26 (1969) 216. . 15 OD N . w ~ D' C
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the acyclic terpenic alcohols (11-13) dtsplayed a strong inhibitory activity, while only one of their cyclic counterparts, menthol (14), was of corresppnding activity. While there was no pronounced difference in, inhibitory effect between the saturated and unsaturated acyclic monoter- penoids a marked difference in both basal and noradrenaline induced respiration was observed between hexanol (3) and 2-hexen-l-ol (10). The strong inhibitory effect of the Cs-C,o aliphatic alcohols and the acyclic terpenic alcohols could be due to inhibition of the proton translocation linked to the mitochondrial electron transport previously demonstrated by Bauedein and Trasch in the case of beef heart mitochondria [15,16). Ethers All members of this group, which comprises aromatic ethers, were found to be weak or moderate inhibitors of the noradrenaline stimulated respiration. In constrast to the corresponding phenols, the anisoles (22-30) . did not show any increase in activity on alkylsubstitution. Enlargement of the aromatic ring system (34, 35) did not affect the inhibitory effect, while extension of the alkyl moiety (31, 32, 35) increased the inhibitory effect. Acids Several of the aliphatic acids (36-43) augmented thee basal respiratory rate of the cells with a maximum for octanoic and decanoic acid. As demonstrated previously, free fatty acids constitute an excellent substrate for brown fat cells and are oxidized at a high rate [7). Simultaneously, there . is a loss of respiratorycontrol and a drop in energy.levels 110,121. Noradrenaline induced respiration is thus restricted both because of lack of energy in terms of ATP and because of the fact that maximal or close to maximal respiratory rate is already obtained. Surprisingly, decanoic acid was found to be a better substratee than hexadecanoic acid (palmitic acid), which is one of the major components in the triglycerides of brown fat [171. Benzoic acid markedly increased the basad respiration,-possibly due to uncoupling of mitochondrial phosphorylation,, which resulted in the impairment of noradrenaline induced respiration. The uncoupling effect as well as thee inhibition of noradrenaline induced respiration decreased on substitution by hydroxyl or methoxy groups. Esters and anhydrides All members of this group being benzyl alcohol esters (53, 56-58) were found to be potent inhibitors of the noradrenaline stimulated respiration. The ethyl esters of the diacids (60, 61) were moderate inhibitors. The activity ,. of the phthalates decreased with increasing chain length of the alcohol moiety, which, in view of the results of the corresponding alcohols, indicated that they did not suffer hydrolysis. 10
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N A fundamental feature of isolated hamster brown fat cells is their rapid catecholamine stimulated oxidative metabolism [7-9]. Under these condi- tions the mitochondrial respiration proceeds at a high rate which is indepen-' dent of ATP demand and ADP availability [10]. On the addition of 1 pM noradrenaline, .the oxygen consumption increases from a basal level of approximately 40 nmol O/min/106 cells to a respiratory rate of 600 nmol O/min/10° celxs [11]. The events behind the respiratory increase involve agonist-receptor interaction, increase in cAMP concentration [12], protein kinase mediated stimulation of the lipase [ 13] and the oxidation of free fatty acids by the mitochondria [7,12]. Hence, a maxtmalrespiratory increase after noradrensline stimulation, is only obtained if every function from the receptor at the cell membrane to the mitochondrial cytochrome oxidase is I and the effect of a test compound on cell metabolism or membrane integrity - was registered solely as an impairment of noradrenaline stimulated respi- . ration, and/or an increase in basal respiration. intact [ 14]. In this study, the cells were used for screening test purposes only MP'IHODS c . . ..__"... ....._ ._ . . . . . . . ..Fiy. l, i Isolated brown fat cells were prepared [11,12] from adult hamsters : . "' Ln~ -' ! (Mesocricetus auratus) which were kept in cages maintained at room tem- u 5 t ' perature and provided with food and water ad libitum. The oxygen con- sumption rates of the cells were measured at 37°C using a Clark-type oxygen and c i electrode fitted in a Perspex vessel of 1-m1 volume. The output of the elec- com~.' trode was amplified and continuously registered on a recorder. After addition or sy; : of a buffer solution (Krebs-Ringer phosphate buffer, 1/2 normal Ca'• con-synti.- centration [8]), the vessel was sealedd to external air except for a narrow. checic.._ channel through which subsequent additionswere made [11]. Sufficient cells and ,:. -` were introduced to give a final concentration of 10' cells/ml and the test 3%'u.:,: :. substances, dissolved in ethanol or dimethyl sulfoxide, were incubated with recry;,;._ the cells for exactly 5 min during which period the oxygen consumption was wr. re ;' registered. After this preincubation, noradrenaline was added and the oxygen sulwt:.,n consumption of theceUs was registered for a further 5 min. The noradrenaline substi: concentration was 1 pM which is approximately twice the dose known to inducemaximalrespiratoryrate[12].Excessnoradrenalinewasusedtoavoid IIksui.:inactivation of noradrenaline per se by the test substance. . Thenonadrenaline induced oxygen consumption rates in the presence of Th~ .: an ethanol solution of herman and a control (ethanol) is illustrated in Fig. 1. to fu,.. ` The toxicity of } substance is detEnninedd by comparing the noradrenaline are pr-: . induced oxygen consumption in the presence of the test compound to that 3vxp,._ observed for the control (solvent blank); the inhibition registered is expressed Sou., ' as apercentage [14]. All the substances were initially tested at a final concen- of the ,. tration of 1 mM. Compounds exhibiting an inhibitory effect at this concen- tYation close to 100%, were also tested at a final concentration of 0.1 mM. ..tlcoii,,:; Ih, Compounds examined rate v:` :; Three hundred and twenty compounds, most of them abundant in tobacco :dcuhu,. `: . . maxim_ 2
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thu acLive cumpounds using livcr micrusumal systemsare presently under way. l(ydrocarbons. The acyclic hydrocarbons (.158-164,166),, whetherun- saturated or not, are non-toxic. Of the cyclic, non-aromatic hydrocarbons (1(i5,167-1i69), the monoterpenes (165,168-169) are active. The latter are ull ydadilyauto-oxidized giving a,/3-unsaturated alcohols. and ketones [121, a fact which may serve toexplain their activity along the lines men- tioned previously. In fact, a-pinenee has long been known for its strong toxio- action on liver and kidney on oral administrationresidting imulbuminuria- in sheep [131. 13enzene (170) and toluene (171), are non-toxic,, but further substitution or increasee in the number of carbons inthe substituent yields active com- pounds (172-182). Here, as in the aforementioned aromatic compounds (145,148),. introduction of olefinic bonds (177,181) iss without effect. The toxicity'of azulene (183) and the naphthalenes (184-187)) is about the same as that oG the active hydrocarbons containing only one aromatic ring (172-181). The results for the polycyclic aromaticc hydrocarbons (188-200)6 which indicate a low activity, are less certain on account of experfinental difficul- ties. 'T11us since some of the heavier compounds (4,7,28,188,190-197,211)) did not dissolve completely in DMSO, the cells had to be counted manually with a notable decrease in accuracy and re.producibility... . I•'mrans, [hinphene: Tetrahydreofuran (201),, furan (202): and thiophene (206) are non-toxic,, while the substituted furans (203-205)) showed inhi= biLory effects. The activity of these compounds parallel tltat.of the corre- sponding N-heterocycles. . N-llereocyclcs. I:xceptfor the wcakly active piperidine (209), all mono- cyclic, aromatic as well; as non -aromatic, N-heterocycles (207,208,210,219- 226,228-230) are non-toxic. Similarly, the tobacco alkaloids (231-233),, which have two heterocyclie rings linked together bya.single bond display no or very weak activity, while tho bicyclic quinoline (227): and indoles (214-218) are toxic. As mentioned previously, the indole ring system per see seems to be toxic and this is supported by thee fact that both carbazole (211) and benzimidar.ole (213) are non-toxic. In contrast totho phenols, nciUicr pyridines nor indoles showany markedincreased in toxicity on alkylation. Indoles include niany biologically active compoundssuch as tryptophan, rescrpine, auxines and lysergic acid derivatives and are thus of great interest. 'I'heresults obtained heree must, however,, be checked against their possible effect on Lhe: level of SH auino acids in the celllculturesas Wellens [14-16] has shown that growth inhibition can occur in animals due to extensive sulphntae conjugation ofindole which in turn gives risc too a deficiency of . 511 amino acids. '1'hiss effect can be counteracted by cysteine,, eystine, uie- thiiininr or glutathiom+, compounds which are tobeo added in fuLure Lests of the toxic activity of indoles in our system. :Imicce.a. Of the aliphatic amines (235-239) only 3-aminopropene (238,,
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r aromatics showed low inhibitory effect. Neither alkylsubstitution (261-266) nor introduction of otherfunctional groups (48, 96, 127, 173, 267) increased the activity of the pyridines. In accordance with this the nicotine analogues (291-295) also containing a pyridine moiety, were inactive. As the indoles include many biblogically active compounds [20,21) a selection of mono-, di- and trimethylsubstituted indoles were examined, Of the indoles tested, all but indole itself (272) and two N-substituted indoles (279, 283) were found to reduce the noradrenaline stimulated respiratory rate. There was a great variation in activity and the only apparent correlation between the inhibitory effect and the number or positions of the methyl groups encountered, was that C-methylation increased the activity more than N-methylation. The 2 indole derivatives, harman and norharman, exhibited strong inhibitory effect. Amines Of the aliphatic amines (298-304) only decylamine (300) had a pro• nounced inhibitory effect on the noradrenaline stimulated respiration. This could be due to an inhibition of the proton translocation linked to the mitochondrial electron ttansport [16,16J as previously discussed. Most of the anilines showed no or low inhibitory effect. Aniline itself was inactiive-, and introduction of methylsubstituents into the aromatic ring was without , effect, while ethylsubstitution of the ring or alkylation at the heteroatom increased the activity somewhat. Extension of the aromatic syatem to that of 1- and 2-naphtylamine also increased the activity. CONCLUSION The brown fat. cell system has proved to function as a screening test for a large number of compounds derived from tobacco and tobacco smoke, thus providing a means of identifying groups of toxic compounds. An account of TABLE If MEAN INHIBITORY EFFECT (R. INHIBITION) OF THE 12 DIFFERENT CHEMICAL . CLASSES ON. NORADRENALINE INDUCED RESPIRATION Chemical class Mean inhibitory effect - . Number of compounds Alcohols 51 21 Ethers 34 14 Acids 47 16 Esters,.anhydrides 39 14 Amides, nitriles 40 32 Ketones 46 30 Aldehydes 56 47 -4henols 62 36 Hydrocarboaa 36 40 Furanes, thiophenes 22 6 N-Heterocycles 35 42 Amines. . 26 23
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-4 E. Arrhenius, Unpublished. 5 E.C. Miller and J.A. Miller, in Twenty fourth Annual Symposium on F'undamental Cancer Research1971, W illiams and'. W itkins, Baltimore, 1972, p. 5. 6 HIB. lienliest, E.R.H. Jones and C.F. Smith, J. Chem. Soc., (1953)'.3796. 7 R.T. Williams, 9ckncication Mechanisms, Chnpman and llall, Londoni 1959', p.:190.. 8 J.J. Clcumiunyand D,M. Angevine, Federation Proc., 15 (1976) 512. 9 J.L. Simonsen an& LX Owen, The Terpenes 1, 2nd ed., Cambridge Univ. Press, Cambridge, 1953,.p. 154. - . 10 A.L. Lchninl;er,'I'he Mi4oeliondrion, Benjnmin,.Netv York, 1964. 11 E. Arrhenius, in D..Shugar (Ed.), Biochemical Aspectsof Antimetubolites and of' Drug Hydroxylalion, FEBS Symposium. Vol. 16, Academic Prrss, New York, 1969, p. 209. 12 JIL. Sinronsen and L.N. Owen,.TheTerpenes lI, Cambridge Univ. Press, Camhridpe, 1957,, pp. 138,199. 13 .1.M. I larvry. Uuivrrsity o6 Qucenxl:md Papers, 1(1912) 23. 14 C, Wellens, Bu11...Soc. Chem. Biol., 35 (1953) 1&A. 1.5 G. Wellens, 11ull. Soc. Chent. Bihl., 35(19fi3) 1358. Ili C. Wclluns, Bull. Soc. Chem. Bio1., 36 (1954) 16fi5. 17 D.R. Cl:n•uru, Chemicai Ca ,rciuugunexis, Little,.Bruwu, Boston, 1962. 18 E. Arrhenius, Carcinogenic Aminea. Some Aspccts of their Metabolism and Inter- action with Micrusnmul Functions or Liver Cells, Dissertation, Dept. of Cell Phys- iol., WennerGren Inst'atute,.UniversltyofStockholm,Sweden, 196R. 19 I'YI'. Yipe, in L. '1'wnatisaud It. Montessano (Eds.), Chemical. Carcinogenic Essays, 1'ubl. 110. Intern. Agency Res. Cancer Lyon 1974, j),,119. '?0 E. I luhyrm;m, in 11. '1'nmalis and Et. Montesano (Eds.), Chemical Carcinogenic Essays, Publ. 10; Intern. Agency Res. Cancer Lyon 197A, p. 1.17. 21 B.N. .\mca, W.14: Duraton, E. Yamasaki and F.1). Lee, Proc. Natl. Acad. Sci. (ULSI), 70 (1973) 2281. . , 62
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l :IIAIAI:IN.  M(I'1'AGENIt:ITV:GENE MtCI'ATION til•UnIFti Anwa ussuy Salmonella IypBimurium :L.,,a. l rf.- SfaricJf:d!'IFJM Rnulr n.,v, /t.J NG R-0 RESULTS The National Instilute of Environmental Health Services obtained negative results in Salmonella muta- genicityassays. Inlmalation ahout the assay was not given. of de. lin tiit ;ed ,ht. lan t Ames assay Salmonellatyphimurium Ames assay Salmonella typhimurium TA100 platelet aggregation when induced by collagen. (R- 14) 0 2-Methoxyphenol was found to exert a hemo- lytic action on cattle blood and interfered with RNA synthesis, protein synthesis, and, to a small degree, mitochondrial respiration in rats. Fatty acid oxygen- ase in sheepp vesicular gland tissue was inhihitcd' by exposure to 2-methoxyphenol. (R-7) a The tox effects of 2-methoxyphenol weree studied in vitro to deter- mine its depigmenting action. 2-Methoxypheuul in- hibited protcin syutiLesis in latliverslices(!i mM concc gavee approximately 80y'a inhibition), RNA synthesis in Hela cell cultures (5 mM caused 50-60% inhibi- liun). atul respiratiun in Isol:Itl4I rat Ilv(-•1' IIIIWGLmIt- dria (0.5 mM caused 25-70% inhibition). This indi- catedthere was no obvious direct relationship between these effects and their relative depigmenting actions in vitro. ~R-f 15) i Ascites sarcoma 1SP8 cells, cultured in suspenston, in vitro, were used as a general tox test syslt•m lin• Inbaccoanwkt• cuuslilurut.r. (7tuiiuod wlls tested by cxposi ng the cells tu 1.0, 0. I, 0.01, and 0.11111 mM ofguaiacol and then measuring.theinhibition of' allutre I;ruwth. '1'his system of testiug did not allow fur possible effecLisuch aspeneu'ation, distributiun, and tnicrlMomal meCtlx>lism.-fhe fi)IlowingIY•stdts were obtainetl:7496 inl)ibition at 1.0 mM level and 8`yo% in- hibitiou at 0.1' mM dosage IeveL (x-20) PRARMACOSINEfICSlMETABOL[SM  2-Methoxy- phenol is largely absorbed from the digestive tract and is stored in ihe blood, kidneys, and.respiratory urg:uls. It is excreted by rabbits in combined form with snll:he (15%)) and glucuronic acid (72%).. (R-7) 0 Investiga- lions have Ixcn coud uctcd too stud y l he rcL•u iunsl) ip NG - R-17 RESULTS 2-Mmhoxyphenol was found to be nonmuta- genic. using the Ames aseay. ._ 1,000. 100•'l0• 1•.and R-84 0•1 µg/plate RESULTS At Ihese dosages of guoiarol. there was no evi- dence of mutagenlcity, using Ihe Ames assay. After dilute aqueous chlorination.. guatacol was found lo be mula- genic, OfiNl200 revertants per 0.4 mL soln minus spon- taneous revertants• between the activities of certain enzymes of Phase I and Phase 11 hepatic xenubiotic metabolism and the urinary excretion of some metabolites of the D-glu- curonic pathway.. Male Sprague-Dawleyrats were in- jected with guaiacol 100 mg/kg/D, intraperitoneally, fbr 7 1). Guaiacol did not have anyeffect on the pa- rameters ol' hepatic Phase I xenobiotic metabolism tested; namely, mixed function oxidase enzyme activ- ities, cytochrome. P-450, and the microsomal content of protein. CuaiacrA did not stinn)latl! the urinary ex- cretion of b-gluciric acid or L- ,°n'~ acid or xylitol, but it did.increase urinary excretion of tot:d (frec and conjugated) D-glucuronic acid..(R-21) EN9IA08MEHTAL IMPACT t2iVIRONMENTAL FATE: TRANSFORMATION PRO- CEsSPs-aIOnEGRADASN,ISY  3-Methoxyphenol was autocatalytically decomposed, at 14-18 C in 0.001 mula.r atlueouwsofinlions, byayualii: pl:uus.'1'hc dc- comp byaqtr.nic plants is a lirst-order neaction•.dte velocity constant is 3-5 times greater than the vedur for aumoxidatioo. Ut.•comp is due to the catalytic.:ttt- tion of the pheuol.oxidase contained in the algae;; the initial decomp products are yuinones. The reaction is inhibited by copper-containing.enzymes, KCN, and p-nitrophenoL'Che.high reaction rate and theadapt- ability of the algae to phenol concs s 5 mg/L were confirmed by tests in 90 m' ponds..The high reactivity. adaptability, and thr.tbility to develop in poor-tp«:dity water indicate that algae could be eflectively used for the purification of eflluenls containing phenolic com- pounds. (R-21i). Spectrophotomctric.tcchniquc was Iexd tu mcasore tFhee degr:ltlation true ull 2-mcthox- 8'711s25o I oat A xo,`,D1C 19M 4,
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allylamine) is Loxiv and in rent.u•kably low conccntraLion (0.005 nvlf). 'L'6is situation parallels thatencountered: for the alcohols, aldehydes and ketones and may be explained aiongthesame lines. It is also parallel in the sense that the anilines (240-255) show much less orno activity. Most active of the liitter compounds are diphenylamine (251), a- and i3-naphthylamines (252,253) :md 3i4-diaminotoluene (255)~ Of these, 3,4-diaminotoluene meets the criteria for a metalligand im having two properly spacedatoms possessing fn:c electron pairs. '1'he naphthylaminess need metaholicc activation in the nticrosomal detoxication enzyme chain in order to exert' their carcinogenic effect 1171; these detoxication systems, however,.are attsentin all celli cul- ture systems which have been cultivated for several generations. 'Phus the toxic effect observed here must bedUe either to some other functional disturbances or to a nomenzymatic conversion of the amine leading to similar reactive metabolites as those produced! enzytnatically by the detoxi- cation enzymes of liver cells [18). No effects of alkyl substitution in tho aromatic ring were observed for the anilines, but the toxicity increased somewhat with N-substitution (250,251), and in the presence of another amino group. . - 6ENERALCONCLUSIONS It should be pointed out that the test system has.limitations whichmust be considered when evaluatingg the results. Thus as mentioned above these cells dbnot possesss the detoxication enzyme systems which in many cells of Lhr in lilctani mal con vert fotrign conipounds to n on-toxic excretable produc ts or to reactive intermediates which maydhmage cell functions. The present study must therefore be complemented by studies involving metabolic sys- tems in order to identify thosee substances whose metabolic products (rather than the components themselves) are active. Such test systems, based on acontbination of the present cell culture.syst'em and hepatocytee cultures 1 19,201 or mirrnsmmaU prrparaLions 1211, arrheing tleveloped: •Phe results.S obtained here are presently utilized for further studies on subcellular nr- ;;:mrllr'systcros to cihtain further inlurmaCion regarding the functional liack:- ground of the observed toxic effec:ts. .1CILN O W LE DC IiAt1iN15 'flae authors are inde6tedi to Miss YiaNyberg for skillful technical assis. lance. REFERENCES I E. Arrhenius nud' Ic. :1ncAer, in .Il Nils.on: (Ed.), filuurcycmLL WNiteningAgentc, MVGReport _', SwedisL Nutional Seiencu Res•arc6 Council,.1973'', p. N:3. S•r.S. Rfunro and D.D. I'nrlonus,.6ri..1. IY:adiut. ninl., 21 (.IA72) 87. :1 I3. tluhn6crg,.l'e.rson:J cuumnuaicutiuu, tmdcitial inm:muscriitt. 61
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I the correlation between chemical functionality and inhibition of noradrenaline stimulated respiration is given in Table II. The most active compounds were found within the groups of aliphatic alcohols, aldehydes and acids, of a,p-unsatnrated aldehydes and ketones and of alkylated phenols and indoles. In contrast, polycyclic aromatic hydrocarbons, ethers, amines and N-heterocycles (except indoles) exhibited low inhibitory effect. These results are in good agreement with the results found by Pilotti et. al. [4] and by Thelestam et. al. [3]. Some of the n-alkyl acids which in brown fat cells function as substrates [7,12] could of course not be considered as toxicants. When evaluating the results it must be considered that the true toxic -- concentration of some of the most unpolar compounds, i.e. the polycyclic aromatic hydrocarbons, the N-heterocycles and the long chain aliphatic alcohols, aldehydes, aeids and amines is hard to estimate as the solubility of those compounds is poor in the buffer used for the cells. However, in spite of the low solubility several of these compounds displayed a high inhibitory .. ,, effect on noradrenaline induced respiration. It is likely that the results from this investigation reflect the activity of the tested compounds per se. Electron micrographs of brown adipose tissue demonstrate that there are few inclusions of endoplasmatic reticulum [22j, an observation which indicates that these cells probably lack the detoxica-. tion enzymes responsible for the conversion of foreign compounds to excretable products which are either less toxic or occasionally constitute . reactive intermediates which may damage cell functions. Redox reactions or hydrolytic cleavage of the test compound could however not be excluded. The results presented here form the basis for further studies regarding the mechanisms behind the inhibitory effect of the most active compounds on the noradrenaline induced oxidative metabolism in these cells. . ACKNOWLEDGEIAENT The authors are indebted to Ms Ylva Munterud and Ms Eeva Kazemi-Vala for skilful technical assistance. This work was fmanced by a grant from the Swedish Tobacco Company. . REFERENCES 1 CR-Greeu, Recent Adv. Tob. Sd., 3 (1977) 94. 2 RA. Heckman and F.W. Bed, Abstract, 32nd Tobecco Chemists' Research Couferenee, Montreal, Canada, 1978. 3 M. Thelertem, M. Curnll and C.R. EnseB, Toxirology,15 (1980) 203. 4 A. Pllotti, K. Ancker, E. Arrheniue .nd C:A. Enttll, Toxicology, 5 (1975) 49. . 5 B. Pettenson, M. Curv.ll and CA. EnzeLL 6 I. Floriq L. Rutberg, M. Curvdl and C:R. EnzeO, Toxicology,15 (1980) , 219. 7 SB. Pru.iner, B. Cannon and O. Lindberg, Eur. J. Biochem., 6 (1968) 16. 8 B. Petterawn, Eur. 7 Biochem., 72 (1977) 235. 9 J. Neder{aerd and O. Lindberg, Eur. J..Biochem., 95 (1979) 139. 10 D. Nieholls, Biochim. Biophys..Aets. 549 (1979).1. 14
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I Toxicology, 18 (1980) 1-18 . o EleerierlNorth-Ho(Lnd Scientific Publiiben Ltd. EFFECTS OF TOBACCO SMOKE COMPOUNDS ON THE NORADRENALINE INDUCED OXIDATIVE IdETABOLISM IN ISOLATED BROWN FAT CELLS BERTIL PETTERSSON' MARGARSTA CURVALLb and CURT R. ENZELLb •The WennerGren )netitute, Unkxsity of Stockholm, NorrtulGgntan 16, 3-113 46 . Stockholm andd bReeeoreh Depurtment, Smedid+ Tobacco Company, P.O. Box 17007, S.f04 62 Stockholm (Sweden) . (Recei.ed July 4th, 1980) - (Revision received September 18th, 1980) (Accepted September 23rd, 1980) SUMMARY The effect on cell metabolism of 320 individual smoke components have been investigated by measuring theil:inhibition of noradrenaline induced• and/or uncoupling of mitochondrial respiratory control. increased the basal respiration of the cells, probably by acting as substrates hydes and ketones. Some of the aliphatic aldehydes and acids significantly acids, of alkylated phenols and indoles and of a,8-unsaturatid aliphatic aide- inhibitors were found within the groups of aliphatic alcohols, aldehydes and tative of the gaseous and semivolatile phases of tobacco smoke. The strongest respiration in isolated hamster brown fat cells. The compounds are represen- cellular test systems. Representatives of.the gaseous and semivolatile phases of the tobacco smoke have been tested for their effects on membrane per- meability of human lung. fibroblasts [3], cell multiplication [4], ciliary about their biological implications. To clarify the interaction between tobacco smoke and vital biochemical functions, we have chosen to study the . toxic effects of individual smoke compounds in several cellular or sub- w now some 3000 compounds [1,2] have been identified, but little is known Tobacco smoke comprises a large number of chemical substances. Until activity [6] and also for their mutagenic activity in the Ames' test [6]. The ^-°° * v¢nsk daals wi+h the ihhih't .~ ff~ * f°~0 in man e com ounds on the noradren ~.nryL:e1 rtaiuf44f res iration in ividu isola rowrl "~_
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108 WOODS AND SMITH weekly to the check pouches of golden Syrian hamsters (Mesocricetus au- ralus). Biopsies were taken at 16, 38;, 40, and 45 days from the commencement of treatment. Biopsies were alsoo taken from normal untreated cheek pouches. '1'issucs were fixed in fnrmol saline for histological purpases and in 2" osmium Re:. t. An H nnd. F.prerwrntion of hamater cheek pouch trented for 4fld)rys with 4-hydiasy anisole, The epithelium shows hyperkemU)xiswith some pamkeralinirat4a) and hyperpkvo. Regions nf'lhe disthrbed hasal layer have grown down intn'the inflsmed cnnneetive ti¢aue and s huGOprvMMaliunufKUlamlmxvnwm,WdrisdeKVnnralr. X'MN)munuiliaaiun.
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conjugation was extended by an aromatic ring; a,P-Unsaturated aldehydes ", are highly reactive substances that easily combine with biochemical ` nucleophiles such as sulphydryl, amino and hydroxyl groups [19]. Short chain aldehydes lare also known to inhibit the mitochondrial oxidative meta- bolism [191. The moderate inhibitory effect of benzaldehyde was not affected by alkylsubstitution while introduction of methoxy groups seemed ` to decrease the effect. Substitution by hydroxyl groups was also without effect, except when a chelating capacity is produced (165, 168). The hetero- cyclic aldehydes,.except 3-indole carboxaldehyde (174), were weak inhibitors of the noradrenaline stimulated respiration. Phenols All the alkylated phenols (176-192) are strong inhibitors. Phenol itself showed a moderate activity which increased with increasing methyl substi, :.. tution and with the chain length of the alkylsubstituent. Thus, the most ... : active alkylphenols are the ethylphenols (188--190), 3-ethyl-6-mothylphenol (191) and 2-isopropyl-4methyiphenol (thymol) (192). Alkylphenols substi- tuted both in the 2- and 6-positions.(182, 187) did not enhance the inhibitory effect to the same extent as alkylpbenols substituted in other positions,: an effect possibly due to the shielding of the reactive hydroxyl group. . Except for pyrogallol, introduction of methoxy, hydroxyl, acyl, carboxyl ' and aldehyde groups into the aromatic ring either lowenadd the activity or . was without effect. Enlargement of the aromatic ring system to 1- and2-naphthol significantly increased the inhibition of noradrenaline inducedrespiration- - Hydrocarbons Most of the compounds in this group were found to be low or moderate- inhibitors of the noradrenaline induced respiration. All the aliphatic hydro- carbons--cyclic or acyclic, saturated orunsaturated (210 217) - - inhibited the noradrenaline stimulated respiration to about 40-50%. No significant effect of introduction of double bonds was observed. The activity of benzene (218), itself a weak inhibitor, increased on alkylation (219-231). An augmentation was observed on increasing the number of carbons of the substituent (219, 224--226; 229, 230), while the number of substituents did not affect the inhibitory effect. Substituents having a double bond con- jugated with the aromatic ring (227,: 228, 231) also increased activity but not to the same extent as the saturated analogues (224, 229). Of the polycyclic aromatic hydrocarbons only the naphthalenes exhibited some inhibitory effect. Fumnes, thiophenes Alll the members of this group comprising only 6 compounds showed low inhibitory effect. In accordance with the results for the corresponding . N-heterocycles, substitution of furan increased the activity. N-Heterocycles Of the monocyclic N-heterocycles both the aromatics and the non- 12 0D 4 N F& fC W O O~
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Nitrifes Most of the compounds in this group were found to be low or moderate inhibitors of the noradrenaline stimulated respiratory rate. The low activity of the aliphatic nitriles was foun4 to increase somewhat with increasing chain length but not to the same extent as for the corresponding alcohols. Alkylsubstitation or introduction of double bonds had no or negligible effect on the noradrenaline induced respiration. The moderate inhibitory activity of benzonitrile was not enhanced on alkylsubstitution or enlargement of the aromatic ring system, which is in contrast to the effect observed for phenols (vide infra). Ketones The ketones displayed a great variation in toxicity. The aliphatic methylketones (98-101) were weak or moderate inhibitors of the noradrenaline stimulated respiration. Introduction of a double bond con- jugated with the carbonyl increased the inhibitory effect except in the case of pentenone (105). The a,p-unsaturated ketones having a terminal unsubstituted double bond (102, 104) were found to be the strongest inhibitors. Of the cyclic non-aromatic ketones only the terpenoid ketones (113, 115-120) caused a notable inhibition. Two of the ketones having the carbonyl group conjugated with an aromatic ring were strong inhibitors (122,126). Aldehydes Of the n-alkyl aldehydes, the C,-C,i representatives (132--137) in addition to being strong inhibitors of the noradrenaline stimulated respiration, they also increased the basal respiratory rate of the cells. Experiments on isolated rat liver mitochondria showed that these aldehydes uncoupled the mitochondrial respiratory control using succinate as substrate (Bernson/Pettersson, to be published). In the intact brown adipocyte uncoupling would lead to the combustion of endogenous free fatty acids and an increase in respiratory rate [7,11). However, it could not be excluded that part of the alkyl aldehydes are . oxidized to the corresponding acids and thus act as substrate per se. n-Alkylaldehydes with an odd number of Catome (132, 184, 136) would thus after p-oxidation yield propionyl-CaA which by enzymatic carboxylation could be converted.to succinyl-CoA. Succinyl-CoA as well aa . propionyl-CoA have been shown to inhibit citrate synthetase [ 18] and thus block the entrancee to the citric acid cycle. This.s could explain the observed - difference in basal respiratory rate between n-alkyl aldebydes having odd and even numbers of carbon atoms. The introduction of allryl groups into the n-alkyl aldehydes (138-142) showed no clear effect on noradrenaline induced respiration, while introduction of a double bond conjugated with the carbonyl (144--149) . increased the inhibitory effect of the molecule. Further extension of the - conjugated systemm increased the inhibitory effect, but not when the . 11
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FIC. 3. Electianmicmgrnph of a region of epithelial-connective timue junction. A bmal epi• thelial cell (B) withh pxudopodia (P); projecting through the lamina dense of thee basement meo} hrane (() into the connective tinue (C). X2o.eoo magnification. some containing red blood cells, between the keratinized and nonkeratinized cell layers. Increase in size of epithelial cells, loss of basal cell polarity and nuclear hyperchromatism were. evident. In addition, some basal cells appeared to be proliferating into the underlying connective tissue; this produced an irregular, "saw-tooth" junction between the epithelium and connective tissue in contrast to the normal smooth junction. The corium was infiltrated to a variable extent by chronic inflammatory cells and some subepithelial collegen bundles were abnormallyirregular. Muscle degeneration was a marked feature in some areas, particularly under regions of most severe epithelial and connec•
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(SUct±a co/ A!VS_WEPd :' ANI C'Ai.i,5~( 1. ) : 1s;i.1~'iz TI Effects of phorbol myristate acetate, phorbol dibutyrate, ethanol, dimethylsulfoxide, phenol, and seven metabolities of phenol on metabolic cooperation between Chinese hamster V-79 lung fibroblasts AU Malcolm, A. Russell; Mills„ Lesley J,.g McK:enna, Edward J. CS, Environ. Fes.. Lab.,L1.S. Environ. Prnt,. Agency 1...0Narragansett, RI y USA.. SO Cell P.iol. Tox.ic.ol., 1(4), 2E,9-85 SC4-3 (Toxicology) . X ` 1 / ®1 DT .} CA~ CEs9-OE2 15 r:r742-.2i r91 PY 1985 LA Eng AB The effect of phorbol myristate aceta.te(I) f-16561--29-8.], phorbol dibutyrate C.61557-88-8], ethanol C64'~--17-57,: dimethyl sulfoxideC67-68--57, phenol C1C?8-95-2] and 7 metabolitesof phenol on metaboli.c cooperation were assessed as a function of mutant eell recov-er-y fromm populations of cocUQtiv.ztedhypoxanthine-g~~.anine phosphoribosyl transferase-deficient mutant (HGPRT--) and wild-type(HGPf+T+)Chinese hamster V-79 lung fibroblasts. Phorbol myristate acetate and phorbol dibutyrate were potent inhibitors of inetaboli~c ° cooperation. Ethanol and di--Me sulfoxide, solvents commonly used to prep. chems. for testing, weakly inhibited metabolic cooperation. t!' Phenol and' phenyl glucuroni.~.de C176i?5-G5--1] h.a.dd noeffect on metabolic cooperation. Four oxidativemet-abolites (1,4-banzoqu.inone L1n6--51-4]„ catechol C120--8U-9], hydroxyquinol C533-73-3] and quinol C123-w1-9]) inhibited metabolic cooperat_ion. Phenyl suifate _ L937--?4--8] weakly inhibited metabolic cooperation. Conversely, 2--methoxyphenal C90-05-1], aa methyl'a*~ed deri~v of catechol aome•~rA~ to ei? ance. metahol'ic coo era ic,n. fhese resurlts genera.lSU pr~~rt t:e iypm esis that.tumor premo~_nrs :ni~iba+ T~SOSi-~-~aoperation and i1 Lrstrae t e importaoc~.e_-M metabolites when testing , thishypothu-•'s_ The weak capacity r+f 5 metabolites of pfienrl~o xn ibi. met.aboli.c cooperation cor*rnl-+t•~, with the weakness.of phenol as a,tumor promoter. Interpretati.on,of thess results.is complicated because 2 metabolic cooperation--inhibiting metabol..iites (catechol and quinol): a.renonpromotimg when tested'.individmaDly im.the same assay where phenol shows promotingg acti~.vity. Su.chmetabolites may be incomplete (stage) promoter-s, and exposure to .gtoreq.2 may be re.qr.ri.redfor a promoting effect. The signif'icance of enhanced metabolic cooperation, rerquiresfurtfier investigation, particularly in r-elatinn to anti'promoting,effects. For diagram(s), see printed CA I ssue.. , _ 2 111
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Agrfc. &nl. Chem., 51 (5), 1439- 1440, 1987 Antimutagenic Effects of-Phenols on MNNG-induced Mutagenesis in Escherichia coii Atsuko KusHl and Daisuke YosHIUA Central Research lnuilute. Japan Tobacco Inc., 6-2 Umegaoka. Midori-ku, Yokohama, Kanagawa 227,.Japan. Received October 29,1996 We reported that antlmutagenic effects of a pyrulysate of albumin were observed on MNNG-induced muta- genesis in Eseherichia roli' WP2 Blf IrP-n (WP2)- We purified the active component and identified it as p-cresol. Phenol was identified in the same fraclion, but its anti- mutagenic effect was not studied. Therefore, the melhylor other substituent group of the phenol Iskeleton seems to be very important for the antimulagenesis. In this paper, the location and variation effects of the substitucntgroupsof the phenol skeleton on antimutagenesis is described. The anqmutagenic effects of 23 phenol derivatives wcre examined. The structures of the diRerent-0aivativest are shown in Table 1. All phenol samples.were of the purest grade available from commereiai sources. The antimuta- genfc elfeetswere measured by the method of Shimoi el aGo E: ryl7 WP2 B[r nn- was used as the lest strain. The bacteria were grown overnight in nutrient broth at 37°C, washed twice with I/l$er phosphate buffer(PH 7.0), and then resuspended in phosphate buffer. The cells were ne:t treated with MNNG (5 ygJml) for 30 min at 37°C, washed lwioe, and resuspended in phosphalebuffer to afrnal cell count of 3~4ic10' cellsJml. A 0:1-m1 portion of the treated suspension and a predetermincd amount of the TABIE 1. fNMR1ON OF MNNG-INDUCFD MUTAYM)N IN E. call. WP2 By PI6Ttof5 No. Sample AD.(yg/mlY o-Cresol 68 m•Cresol'. - p-Cresot' 38 2-Allylphenol - p-nc-Butylphenol - p-terr-Butylphenol - 2,6-Dimethoxyphenol 257 4-Ethylphenol - Eugenol o-MethoxyPtiol m-Melhoxyphenot pMethoxyplicnol 98 o-Phenylphenol P-Phenylphenol a-Naphthol s 10 8-Naphthol 16 2-Methyl-l-naphthol 2,bxylenol t . 2,4-Xyknol . 2,6-Xylenol 2,5-Xylenol . 3,4-Xylmol 3,5-Xylenol t Structure` a=-CH, b=-CHs c=-CHs c=-CHrCHCHi c=-CH(CH>)CHsCHs c=-C(CHs)a a=-40CHs, c=-0CHs c=-CHrCHs. a=-0CHs, c=CHsCHCHr c=-0CHs a=-r.Hs c=-r.Hs f=-0H g=-0H f=-0H, g=-CHr =CHS, be-CHs a=CHt,.c=-CHr a--CHt, e=-CHs a=-CH„d=-CH, . b--CH,, c--CH, b--CH„ d ° CHr ' Concentration of antimutagens which was corresponds to 50% mutation frequency of the controle s Decreaseof try' colonies might be due to the inhibition of Trp' expression. • c......,... 1 k C j i' - Mutagenicity initiated with MNNGwas not inhibited. n m f
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I IA::AIJ rA1(f ,'I II:M4'AIC 1  ANAI.YTICAL METHODS: ENVIRONMENTAL brne/da I'rr/xr'rrJimr Derivatizalion to 2.A-dlnnraphenyllc ether yphcuul. 11 rclicsulrotl Iltc luss uf tIV:Ibsurlwnovwhrtr dw bonzt•ur ring is ch;rved.'1'he ttxl saau t~m- t:tirred tite2-medu)rcyphenul as tlto:sulc sourcc uf i:u- Ium iu a mineral s:dt xuln..'1-he wevelength selected w:ts at 276 nm, whidl was ou ut- uc:u' dte ahsurptiun tn:tximm~t. 'I'he lostt solusweres iuoculatccl with L0 nil. of a 1.0% suspernaiun of.Niagra siltt loam. 'I'he sr:tlt•d huldc•s wcre stnrcd in tlle diurk at 2,5' C. and periexlically (np tu 64 D): aliyuoLs were rentoved( aucl analyred. 'l'he total loss of UV absorbency oecuered :u 4 I) kx' 2-methoxyphenol. (R-27) ENVIRONMENTAL FATE: FIELD STUDIES 'I'his slutly' dc:ds w.illl dctcrminiu); Ihe rate and effective.rone of persislence oi' many dissolved organits in tlre cllluent plumc from pulp nlill wastewater discharged into. Lake Superior. Rhud:unine. WT clye was used to tt'ack the pluutc., lltc milFs.efllnetlt enleretl ilno xchannel:md cmptit•d inlu a bay. 2-Mcthoxyplu:nol was of imerest because it w:ls uue of dte majur ullluenl cum1wuents, is known to arise fronl thebrc-.tkdown of lignin, and i% ane of the components known to untlerga chlori- nation in the mill process. Samplesof die plunte were taken :R various places and tintes during the stu(ly aud an:tlysi,s was perl'ormed on a gas chronmtograph. l:x:mqile OI tltrrestdts fram oue clye run: Ute canc:u lhe source was.:38Ci µglL, while at 2,000 tn frunt the source it was :3 µg/L.. The author concluded lhnt (li- lutiottas well as.b:tcterial degradation ancl volatilira- tion are responsiblce for its tlisappe'arance..(R-29) TREATABILITY 'I'hVadsot'plinn of organir eum- Iwuuds tliasulvetl ill w:uer un ar.liv;ued ctrlxm wus stutlitxl by gas-liquicl chcomalogtaphy:'I'hc.cl]iclx uf adcling );ruuµs ru plhenul w:l.rcumlxu-c(I widt Ihe ad- sorption uf phenol ou activated carlxm. The authurs statr Ihat adding OCH;t onl phenol increasesiLs :xl- stnrption. A decrease in solubility is also noted (in• 2- methuxyphcuol ifcomlxu'ecl to phenol. l.mgmuir is- ntherm consulnts were given asQ=(1.'L91i mgLtng carlx)u.:utd Il_ (1.400. (R-30) TOXICOLOCIAL DATA SKIN AND EYE IRRITATION DATA CODEN sku-rllt 501) mgF2,11i SI•:VF(:'I'OD7 Y111(Suppl),697,82 cyc-r61 5 mg MI.D FCTOD7 21/(Suppl).1i97•82 50 Lrlrrtiun ~tlNlmr!' GC-ECD (cnpillary column) MUTAGF?L.DATAnmw-am-I1N) µlJphuv TO)DC1TY DATA url-r:u 1.b511:72r) nlg/kg uu•rlt LIN.r9(Ill mg/kg „ url-nms 1.D!i11:890 mg/kg ihl-rnus I.CfilL7.!i70 mgCin' ivn-mus LD50:170 mg/kg orlrrt L-DI-o: 1,500 mg/kg skn-rbt LD50:4,60U mglkg scu-r6t L.DL.o:i,250. mg/kg .~ scu-gpg LDLo;900 mg/kg J I acu-pgn l.llla:4(R)~mg6kg UrlrrGnv l.iwil Rrf 0i01-0.09 ng R-28 CODEN I t l•:( :Tr\ I i'! -15p1L8 n CODEN '1'XAl'A9 6,978:64 FGfOU7 20(Suppl),697,82 KSG7-A3 96 .982,82 F(: fOD7 411(Suppl),697.82 PC: )'OD7 20(Suppp,697,82 FCTOD7 20(Suppl),697,82 FCTOD7 20(Suppl),697,82 Fcn'OD7 20(Suppl),697,82 FC"POD7 20(Suppp,697,82 FCTOm 20(Suppl),697,62 DATA1)FFERENCES CODEN Mutagen Skin and F.yc.lrritaut Reported in EPA TSCA Inventory, 1983 @PA Cenctic'l'nxicolnyry Progr:un, January 1984 RECENT FEDERAL REGISTER CITATIONS CITATION NUMBER 46.19.46 (46FK9ti44ff,. I-29- RI) NOTICE OF PROPOSED RULEMAKING ReviSiotl tlf 27(;FIt212, aQd 27CFK21. Uupt. ul'tln•'llrasury, Itu- rcau oC Aleuhed,'I'obacco & Fircarms: fin'nudas for clena4u'edd alcohol & rum; comments solicite(1d by 9- 30-N 1. CITATION NUMER 47.137.169 (47FR91 I80ff, 7- 16-82) FINALRLR.E AcIt140CFR300, Envirunmental Pro- tcctinn Agency: Clean Water Act, Comp- Env. Re- slxnse, CumpclLS:ttiou„ & f.iability Act; National Od & HazartlbusSubsnmces Contingency Plan. CITATION NuMBER 48.107.44 (4RFR24fi72ff, 6-2- 83) FINAL RULE Add 27(:1'R21, dtJcle 27CFR212, DepLOf (he'1'reasury, 13ut-enu of Alcuhol,'fotucco, 8'7119251 DANGEROUS PROPERDFS OF INDUSIRIAL MA7L7BAIS RD'CM
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11 Ls Ig )f f - h tl pected on the basis of the above-mentioned counteracting steric and con- jugative.effects. Substitution of an aromatic.hydrogen in benzaldehyde (108) by a methyl group (109-111) increases activity,, while substitution by hy- droxyl or methoxy groups (112-118); hass nor effect except when a chelatingg capacity is produced (112,118).. The aromat:c aldehydes frequently occur in esscntial: oils and plant extracts; they are used to a large extent in per- fumes. N-Ileteroaromntic aldehydes (121,122) appear more toxic thanO-hetero:uromatic aldehydes (119,120). Generally,, aldehydes and ketones are slowly metaholised in the mam- maliam body. The mainconvesions are oxidation and reduction to acid or alcohol,, respectively. This should be kept in mind when evaluating the tox- icity of ketones and aldehydes as the toxicity inintact animals largely de- pendk on the degree of conversion of these compounds.to the relatively non- toxic alcohols and acids. Plz no . Of the phenols examined (123-157), all but 3-hydroxyaceto- phenoue (148) are toxic." g moderate toxicity of phenol (123) is increased 0o the ihtrodue.tion of electron-donatmg, suhstitucnts such as al crT(1T14- . 14 , ry roxy . - 7 an methoxy (1~41-1461 gtouns, especially in _AUA'a and pare positions. Similarly, insertionn of electron-withdrawing sub- stituents such as acyl (147-149), aldehyde (112-114,116,117) and car- boxyi (21,22) groups either creates chelating configurations (112) or lowerss theactivity or is essentially without effect. . Incorporatiom of alkyl substituents containing olefinic bonds (145,146). hasa very minor effect, in agreement with thee aforementioned assumption that chelating propertiess are obtainable only when the allylic carbon is oxygenated. In contrast, extension of the aromatic system of phenol to that of a- and {i-netphthol (150,151) increases the activity somewhat. Phe poly- valent phenols (152,154-157) whichmay formquinones are efficient growth inhiliitors for the cell culture. Since the same effectt is observed for the non-chelating hydroquinone. (154), and its monomethyl ether (143)', it could depend on the electron donor capacity ofthese.compounds and hee due to their interference with sonre of the electron transport functions of the cell. Substituted phenols are of great interest in view of their well documented uncoupling effect on the mitochondrial oxidative phosphorylation, which may disturb several energy-consuming functions in the.cell [10]... One such disturbance of great iinportanee in tobacco toxicolbgy is ciliotoxicity; mito- chondrial respiration experiments as well as ciliotoxicity studies are pres- endly beinr' performed.on the active compottnds- .4nother, relatively newasp<x:t of the toxicity of substituted phenols is the probable disturhancee of the intact function of liver cell detoxication enzymes in in vitro systems through interference with the electron transfer bctwccn the lluvoprotein enzyme and cytochromo 1' 450 [11 J. This causes a change in the metabolism of, amangothers, aromatiaamines which may favocu the production of toxic and carcinogenic elcctrophilic.metabolitesc If this occurs in vivo,, tL••:! phenols could act as carcinogens. Such~ studies of 59
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rJ 4 I-connective tissue junction. A basal epi- the lamina.densn of the hasemet mem- iavtimt. e keratinized and nonkeratinized .s,, loss of basal cell polarity and Idition, some basal cells appeared nectivee tissue; this produced an epithelium and connective tissue The corium was infiltrated to a ;i and somesubepitheli9l collagen agenerationwas a marked feature nost severe epithelial and connec- I II1Inl1IX'YANIW ILI•: (IN IIAMB'I'14H <'lll•:ISK 1'tlll/'ll I I I live Lisvur t-hnm,m: tiulut• nl' Iht•nc nhornl.ions inrlnccrl bvA-hpdroxy:mi.olo amt be seen inhi);. I anel may he cumpnred wil.h I-he histulugyul' nurnmd chcwk pouch utucosa in I+ig.. 2. Electron micrt.vcapy Ultrastructurally, the greater part of the epithelial-connective tissue junc- tion was undamaged and did not appear to be dilferent from normal. However, .t . Fic. 4. Electronmicrograph of a "light" epithelial cell in the: stratum spinosum.. The cyto- plsm contains onlya few ribosomes,, mitochondria, and lonotilaments ITY.. Between deamoaumu. ( f), there.are regions of clear ectoplasm IE). devoid ofbrganelles. X 1?:000 magnification.
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SAl l'I'I 1 titrinn. hmuali•n IA4rxnt•rirr•lu. au- nd -15. LIaYs from the couvnencement ,m normal untreated cheek pouches. ological purposes and in 2'7osmium loou pouch treated for 40 daya with.4-hydrvq- some pamkeratiniration andhyperplaaiad into'the in0amed.connective tiasueand a ;20e magnification. IiI11lCHU\\ANI411LIti'.(INI IIAMFI'IiH ('llI'UU('ll 0 10oN -4 ,., . *1Z1 Fx.. 2. An H and E prepatati.m of normal hamster cheek pouch. X200 maRnilication. tetroxide for electron microscopy prior to embedment' in araldite resin. UI- trathin sections were stained in methanolic uranyl acetate and aqueous lead citrate. RESULTS In the treated pouches, some erythema was visible throughout' all stages of painting and after 3 or 4 weeks the normally smooth mucosal surface exhibited slight roughness. Hfstology Sections of all biopsies revealed engorged blood vessels, explaining the erythematous appearance, and other histological changes. in a variable degree. These features became more prominent as painting progressed, though the 40-day specimen contained the most severe effects (Fig. 1). The epithelium was hyperplastic,, hyperkeratotic, and covered'by flaltykeratin which some, times revealed a parakeratotic pattern. Occasionally, small bullae were found, OD a Ira N ~ w N Cn
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133 )IC coity kreec of _dnm =api: a ch. -(Sur mdit nin" He, a splitless injection delay of I min, an injee- uon, port temperature of 225°C and a detector temperature of 250°C. GS-MS was carried out on a Kratos MS-50 instrument operated at 70 eV and connected to a lUatos DS 55 computer system. The temperature programwas heldd at 60°C for 5 min after injec- tion, increased by 2°C'min' to 250"C and then held'at 250°C for 25 min: Indaction of sister-chromatid exchanges Blood from healthyy non-smoking donors was antrifuged(250 xg) to remove the erythrocytes and the supernatant was collected. The lympho- cyte fraction thus obtained was grown in Medium 199 with Farless salt. The cultures contained 25% autologous serum+ 1.25% phytohaemagglutinin (Reagentgrade, Wellcome Reagents Ltd., London, England) and 0.1 mM 5-bromodeoxyuridine. After 24 h incubation at 37'C, the fractions or com- pounds to be tested were added to the cultures..In the tests of pure compounds, the potent SCE inducer sryrene-7,8-oxide (Norppa et el., 1980) f t 8 2 21 3 4 22 23 was used as a positive control. The final con- centrations of the solvents, DMSO, EtOH or DMSO/EtOH (1 : 1) did not exceed 0.66%,. 1.0% and 0.5%, respectively. After 88-90 h, the cells were treated conseculively with colchicine(50ng/mli 2 h)) and hypotonic KCI (0.075 M, 5-10 min) and they were then fixed in methanol/acetic acid (3:1) for I h. After the culture time used, thepnrtion of cells thatt have divided more than 2 times is about 20-30%. Chromosome preparations were made by applying the cell suspension to wet cooled slides. The staining procedure was mainly according to Wolff and Perry (1974). The slides were stained with Hoechst dye(0.5pg/ml) for 12 min and then rinsed in Mcllvaines buffer (pH 7.0). They were then exposed to UV-light (Philips TUV 30W G30T8, 10 cm) for 10 min, incubated in 0.3 M NaCl/0:3 M sodium citrate for 2h at 60°C and stained with Giemsa dye (4%, pH 6.8) for 20 min. Well-spread metaphases were scored on coded slides and 25 metaphases from one culture were analysed for each concentration tested. 5 12 6 fiq, 4. Suuctures of the compoundss that have been tested for SCE a'ducrion. Numbers refer to Ta61e 1.
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~ 3 iAlAt S n. arn,IFnv:tnus:lutwidaslr,rdrnalurrdaIr.uhrd\`nnu: sorJs..llvrJ„\}ue/ur.l7kloc/,o5d'.kokha cllcrlivt• dair 7-5-8;1; cccudilit;uiuw ol tt•Ktdatiun; rruGiol:.2:7-1I. Fruni CAS:I!(1:IU~12g. ~~ dcn:unranl :nuhuritcd fnr da•muured spirits: part uf 11 I!~. t)sm,v,kii. \t. \I. I'hN. IbcicnL,¢1 .tnJ gu.ri.irul l.q.,a, -28 FurmWa C+t. :3ti-13 H8.1117,18:i), Fi)rnmla and ulin rosiu.. Gi.l'i.an !.S>urit. 29r8c,..i-.•S. Front - (48.107.14().('t(iFR964d,-I/4`RY/969)(::\:ti:lilai_'Ga. 20. 1'illuui; A., ci :d.. 1975. h:llctils ul' lobacar aud.lolsirco .80 xY 7.82 7,82 7,8°- 7,82 7,82 .)n of v, Bu- as for by 3- REFERENCES I•Lcwis,.Richard ).,ctaL(Mal.)..Jau 1981.!(rgi.alryrf•1'iaic /•f)1r•tt rf(;ltrrnirrel.SuG:slanrer Mirmliche F:ili- liuu. 1)I11IS. (NIOSI I): Ihibl. Nu: 81-Illi-I. 2. S:rx: N. I.. I!IR.I. 7)irnyrnac.7'.nlr:rlir:..n/'lnrlm,ni,d J1ntr ~ nrrh, ullrul. 1'au Nusu:md Rcinhulrl Co.,.N1`. l.t.l l.iltlc. A. I). 1970. Wnlrr (!rudilr'Crilrrin (lbtn /fmrk. vnl. ~J - ' I:mJ ll.l)t 4:mirrhrmiial luullmirru.dlirslnr.urr. t S EI4L. (ambridgc, MA.. 4. Wintlholz, M...ct al. (Ed.). 1976. TThe Mrrrk Indrx, 9dtcd. Merck and (:o.,.Inc., Rahway, NJ. 5. Hawley,. G. C. (Rev, by). 1981. The Cortdnsvd CAnnical Dirlimratx II)dt cJ. Vau NOStcmd Rciuhold (:o., NY. 6.. Versehueren, K. 1977'. Handbook of EnuirmurtprUil Data on Organic.Chemicalt. Van Nostrand Reinhold Co., NY. 7. National Academy of Sciences. 1977. DrinkingWalrrand /ira(th. 1 ~:740-741. ~HE W. 1981. Annua! Plan for Fitcai Year 1981. National f oxicology Program. Indexx and Parts I and 11. NTP 79-7. 9.. McKcc• J. E., and H. W Wolf (Ecl.). 1963. WalrrQtudit,9 (bilrrin, 2nd cd. California State Water Rc•soarccs ('.m- utd Board. DHEW. I(3. Sledur:ul,'I•. L.197(i: Slyduuurs MrrlirrJ.Didimuny. Wil- liams antl Wilkins Co.,. Baltimotc, MI).. . 11. Clctson, M• N., ct al. 1976.Clitica! Toxicology rrf Can-),a'cial PratGtc(s: 4th ed.. Williams and Wilkins Co., Sal- timore, MD• 12. Urs, N. V., ct al. 1975. Enhancement of the bactericida) activity of a peroxidase system by phenolic compounds. ('kWrslreUwingp: ('i5(fi)tfi86-R90. 13. Smythc, O: A., CL al.. 1975. UUse of :t hy/w[hctic:d re- ccptur-site model to predict novel pituit:vy harmone releasing and inhibitingagents. Hyp'odual. Hy/w/iAysi- atrofeie Harm. Proe... Conf. 1972. pp. 189-197. From CAS:83:92168. 14. Jobin, F.,.and F...Tretnblay: 1967. Plateletreactions.and immune processes. 11. Inhibition of platelet aggrega- tion hycotnplemcnt inhibitors. TkrnrnG. Diath. Harrwmr6. 22(3)14fi6-481. From CAS:73:2356l. lff, 7- ' 15. Riley, 1'. A. 1960. Hydrclxyanisole depigmenwninn: Itt tl Pro- v. Re- tal Oil viuro sutdies.. f. PaUmlogy... 97(2):193=L06. From CAS:70:1 13591. Ili. (:redlanyi, (:. V'., et al. 1980. EI'Icct ol' phcmols on lhc adapuuionn of Da/rknia mngru,:uul Dhpknio /nt/rx. Qid- rdrird. lklial Leerled. VWL Sib. 2:IOA!2(11. Ihum (lASt93:1670... :rrelli, J. J., et:d. 1977. Mutageniciiy of lienzicline and latedcompounds entployed hy tltetletcction alhetno- globin. Am: J. Clin. Pat6a!• 67(6):!i2(i-527. 18. Khudyakov, Y.. P., et aI: 1962.. Effect of phenol cnm- pdunds on dormancy anrl sprouwing.rd-rye and poutu stnokt• cun.aiwvnls nn rcll Illtlhlllhta) Wn nt VIIIb. I'rAC- irrrlirhry 5:4!1-4i2. akc, li. C., et al. 1980. 'I'he elleat of ul•anllellt wilh snnre Phase 11 std>.arates on heJxnic xcuubiolic.niclab- ulisni and Ihc. urinary'rxrrriioo rd' mt•talndiicx nf Ilbtr D-glncnruic arid Ir.idlw:ry iu Ihc rat.7ilvind..urd.4/r/diivl l'/nrrmu„dngv Bl:a':i.I-:Yi F. _. 22. ~I'aylur, J, Irf.,.ct al:. 1964. Acnuiparison of thc tuxicitv olsumr alhli prupenvL..rnrPln~rprl rouqr,nnldsiu dtr r:u. I'urmrL ruul Ap/dird /'kannmu/nX7 1i::178-:4N7. 23. Kopperman, H. l.. 1974. Ayuc•tun cldorinatiou and ozonation smdics. I. StrucYUrc-toxicitycorrelations of phenolic contpounds to Dafhnia omgtm: Cbaa.-liia(. !ii- leraclimr.v 9:24•5-251. Rapson, W. H., et al. 1980. Mutagcnit:ity peodueexl by aqucous chlorination of organic compounds. Bull. Ett- tarrm. Conlarn.. Toxico(. 24:590-596.. 25. 13urdiu,.A. K., et al. 1963. Sc.rccning ol' fumig.mi. l..r tuxicily lrn cggs:md I:uvac uf the Uricnul Fruit Fly :utd Mediterranean Fruit Fly. f. of Ecownnir Entamolagy. 56:24i 1-265. 26.Slom, D. I•, S. S. Tinlofeeva,. 1.. I. 8elykh, et a). 1978. Role of Cha(aceac algae andather aquauc plant.s iu (he dcgi;tdaliun of phatol amqxmuds. Vrnlir. 1(mta'. 4: I Ofr-I 11. Wom CAS:911:71i1189)(;. 27. Alexantltr, M., and It, K. l.ustrigmau. 1906. Effccl of dtcmiadl suvnurc un micrulii:d dcgratlatiun of subsli- tuted.lxrnxenes: f. A;(n'ii. 1•-ornlCkrm. 14(4):41(3-41:3. 28. Ichtuncn, Matti. 1980, . Gas chrontatagrrphic detcr- mination of phenols as 2,4-dinitrophenyl ethers using ydasscapillary columns and an denron~capture detec- tor. f. (krane. `102:413-421. 23). Fox, M. R. 1976. Idcutilir~alion ancl.:malysis af urganic )xdlulamx in w;u<•r. pp. 041-Ii5!I. 30. AI-13aht:mi, K. S., and R. J. Martin. 1976. AAdsorption studies usiuggas-liquid chromatography 1. Effect of molecular structure. Water Research. 10:731-736. 31. Weast. Robert C. (Ed.). 1977: Handbook af Cla•nushy arul Physics, 58th ed. CRC Press, Inc., Cleveland, OH. 32. Clayton, C. D. (Ed.), 198D. Patty'.clndssudal Hygiene and 7'nxidn/ap, 3rd cd.,.vol. 2A. John Wiley and Sons. NY. 33. I-ill:rd, D., ct at. 1975.. Aquams'. Otlar thrc•shnltls of organic pollut:ans im industria) clTluculs EPA-(ili(1/4- 75-002. 34. Shumway, D. I..,and'.J. R..Palcnsky: 1973. Impairnrcnt u1• thc Ilavor of lish by water Ixlllutants. 1{I'A-R:1-73- 0111. 87129253 1[dditiunnlRelerences CIS SOURCES OF INFORMATION (:IS, EI M:ccs.Spcctttmtctry (:IS; Carlxm-13 NMR Slxrctrnntetty: !1/1-0fi-1.Dl NIOSI I/(:IS,.I('I'NC.S: .SI. 772i4(lll
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test'compound were added to 2ml of top agar solution. T~ fl. Sensmvmrs orINDUCSU Trp' CLUNSs The mixture was shaken vigorously and poured onto a oF E. roB Bit WP2 trp- To unDere semi-enrichcd minimal. (SEM) agar plate. For rneasura nscoratxucrvn CnraDtrtarn ment of cell viability, the suspension of treated cells was diluted l0s-fokl and 0.Im1 of thix was poured onto each Mcan plate. Trp` revertants and viable cells werecounled after Concmtra- number Survival incubation for 2 days at 37°C. ThelnrtiEG9ptimutagcnie Sample tion of Trp' (~) activityofeachchcmiealw (mg/pkte) colonial the concentration at which the antimutagm red the plate muteuon fregarnry to 50% ot that of Ihe wntrol O-Cresol 0 1002 100.0 (Minakata er nf."). 1.0. 1101 109.9 The results are. shown in Table I. Among the 23 2.5 1072 107.0 compounds tested, 14 cnm unds (o<resol, p<resok 2,/r 5.0. 984 98.2 dimethoxyphmol o-methoxyphmollp-mnho.yphmol,p• p-Cmae1 0 1002 100.0 phmylphmol,, a-neph k -naphthok 2-methyl-l- 1.0 1085 108:3 - naphthol„ 2,3-xylenol, 2,4xylenoi, 2,6-xylNrol,3,4-xy 2.5 1095 109.3 knol; and 2,5-xyknol) a,Qpeared to inhibit the mutaeenie 5.0 975 97.3 - ret nf MNNG: These mmpounds were not toxa: to the 2,6-Dimethoxyphcnol 0 820 100.0 test stminn at the concentration used. To establish that the 2.5 795 97,0 results obtained were not due In the inhibitory e&cts of 5.0 844 102.9 the compounds on the expression of Trp', a tecon- 10.0841 struction experdmmt was done as described by Shimoi er n-Methnxyphenol 0 820 1000 aL°1'The Trp' coloniea grew well in the presenaot 7 1.0 777 94.8 7 compounds (p-phenylphmol, 2-methy4l-oaphthol, t - 'O+a '• i " .~,.' .,. ~..~ w.. and the five xylenols) stifled growth of the colonies, ap- PmemvaJpIKLIoE rmtl by in)abiting the expres9on of Try`. 1.0 791 96.5 ~ Y 2.5 809 98:7The ADp vsluas or the tmted samples are summariud 5.0 848 103.44 in Table 1, in which naphthols havo the most anfi- mutagenic activity (ADp; a-,.lOpglml, ft-, l6pg/ml). a-Naphthol. 0 820 100.0 0.1 756 92.2 Cresols arc moderately active and 2,6-dimethoxyphenol is 0.25 815 ' 99.4 less active. qverall ortho end 0.5 817 99.6 strongest'antim_u_tagenic elfecls on MNNGypducebmuta- a,Naphlhal 0 820 100.0 genesis, and meta_ispmerELLa_v~pQCHect. 0.25 821 100.1 37ie mahanisms of the antimutagenesis of phenols is. 05 831 101.3 noww under investigation. 0.75 825 100.6 REFERENCES 1) A. Kushi and D. Yoshida;, Agric. Bio)- Chtm., 51. 1435 (1987). 2)K.,Shimoi, Y. Nakamura. 1. Tomita and T. Kada, ' The numbers are the averages of the.2 plates. MuraationRea., 149, 17 (1985). 3) H- Minakata, H. Komura, K.. Nakanishi and T. Kada, Muenefoa Rea., 116, 317. (1983)'. Ag Nt P fi in f ac0 DN bioa of t trca sup. F~ The dmt che: N-n mn. AM N out't and the, buff MN urat thm A0. moll cxtr: onto deta cells pour revcr 37°C Te exrra etticr ter fi, the e Andt indut MNI addit
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(.r .c ..T Iuo- 152. wing zicnl. ,Ruo- igrtkn 126. ucar- "rrsd~i. )c ki- nts ill *tied tcipal )cular ench) ausetl Jtsun- From nxar- wntcr 89. Sc:uic, C.G. (Ed.). 1976. Clirrniird Cnrcitengnresi.+. A.C.S. MunngraOIr 174. Amcr. (:hem. Sc)t:: Washingwu. I)C. 41/. S:rrul:lincu.. I I., and I'. I'l:dlli. 19811. I)oxe dcpvuda-nl ncmrthcmical cffccts of 1,/,2-tt'ichlolrrl.>,2-Irilluo- tvaah:mc inluJ:uiou C'xIMITl11Y' 111 ral.c. '1'm~irrd. [.rllm li:~l'i~l!I. d 1. Ucsoillr, I1., cl vL.. 1975. Kxpcrilurul:d aludy ulsyncr- gistn- Applilaliou tu Ihc siundlaucnusnlilir:uiim of pi- Ix-tonrl hnl„side aud sunlr alildlalic. hl..rnllm,ne.:u I..uc. Arrlr. dlnl. I'nf. Mrvl: Tnrtr. .1'rnn'. .1'ru•. Gli:b.-IN. (Ihrnrhl PremP.A. 71i:151R. d'I- F:Oxtviul S.N., ce ;it. 1!Ni7. Syurrgislic laxivit,v :uld ctu- rinugcuii~il yaf"Frcnn.' andpipt•IVmyl bul uxide. Nnlnrr 21 1:521i 5214. 43. Aviadu, l).M.,:u1d U.(:: Sutidl. l'.)75.'1'nxicityufacrosol prupcllants itn thcrespitatury aud cintul:nurysystcnls. Vlll. Rrspiratiou and circulation in ptinntcs. Tuxicrd. 3 :'•1 I :r52. 44. Slrettun, R.J., et al. 1971. Tlie effect of halc~irl.)n aerusol prupellants on bacteria. J. Applied BrauriuG,,U :54:774-777. 45. I)u9'unt (./.'1'nxicity.studics wiih 1,1,2-trichloro-1,2,2- trilluonwlh;utc. PnKlun Information litdlctin No. S- Y4. Wilmiuglon.llP.. 4(i.ISUscy; W.M. 19ti7. Unpublishcd d:ua, Haskcll Rcl.nt No. I-ILO-02CiR-(i7. Hazelton lalxs. Falls (:lulrclt, VA- Slamulrrd hy I laxkcll lab„ IC-L du 1'our dc Netnutn:r and.Cu., Inc.. Wilnliuglon..l)F::.1'ubl. No. 81-123. 17. Ikn:Yky- Pr:mrnc (:, 11176. It:lskill I:J.n':Ilury krl»n-1 No- IFI~I-71t: Unpul.lisllctl tlala,.l's.l. (lu 1'urn db Nrws uurs aud (e)., luc. Wilmiugtutt,.l)@. 48: Mackisun, Frank W. (Ed.).. 1981. Omufxuimd Health Cnu/eliwsforCGaniatlHnzardc. DHHS. (NIOSH) 49. Lillian„ 1)., et al., 1975. Atmospheric fatesof h:doge- nated mtn pcmnds.Emrtrmt. Sci. TrcJmai. 9(12):11W2-1048. 50: 40 CFR 261.33. Revised as uf f uly 1, 198L 51.. 49 CFR 172.101. Revised as of Octuber l, 1981. 52. Mcllullie, R. 198 1. Pstihtation ol.octanol/watcrparti- tion coef'licients for organicc pollutants using reverse- phase HPLC. Cluuuupkne 10:73-83. AddiBonul Refrremxs CIS SOURCES OF INFORMATION' C:IS. El Mass Slnx:unutettry N IOSI I/CI S, K'I'EC:S:. K) 41H)1111110 (:IS CI'(:P, Chctn -1'oxicuingy of (:cnnntctxi:d I'nxl- IICLY: 3,}I) CiS,'1'HERMO: NUS TN-270 Series (:IS, FRSS (Federal Register Search System) CIS, GESARS NON-CIS REFERENCES F.PA, -13CA 1 n vcn tory List N li.ti/NI H, lonir.nion I'utrntial Europc:uu Econonlic Clmmumity Inventury: 345.11 F7!A, prganic.Chenlical I'rcxluuetx: 343(1 F:ROICA, Organir Prn(xctics PHS-149 C:uYinu>;cnic Activity: E0162.0224 cunwcoL NIOSIi, NOHS: 74010. , 84218 URNL, F:MIC Kl'A- Sclcctcd ( h);a n ic Ail- 1'ulhu:u I ts AI'ILYRC,'I-hcrmoAynanticc ancl Slx-aroscnl>y I'fI I,'I'oxicancl H;n,nd'.lurhtst-Cheul Safcty Manual N LM,.' I'ux ' I'ip,. NLM, CHBMI,INh::'f0\I.INh:,-1711i,'f(IxISACK f1Sl IA. Air Cuwamin:uns: %-I NFI'A N~I!IIM,Itlauual of Il:v:u'dous Chrtniral Rr- ar'tinus: HI.IM-12(i ' Aldricll (:alalug/1 laudlawk: 172H2(t (:Or uf Fedo1;J KrKulatiuus ((::IIFiMLAW). .)apau. I'riurilyList lir Asscssmcut in F.nviruumctn: 10Hti r KOl)AK I.Ihca:uury Chcmicals: 11300 Qmdensed C:hemical Dictionary  GUAIACOL ",:l~I CAS RN 90-05-1 NIOSH # SL 7525000 J See N. I. Sax. 1984. Dangerous Properties of ~r Industrial Malerials, 6th ed'. New York: Van Nostrand Reinhold. p. 1487. SYNS o-methoxy-phenol (8CI): o-guaiacol; o-hydroxyanisole; anastil: cat(lchol methyl ether: guait:ol: guaiastil; guaicolina: guajoL guasol: methylcatechol; o-methyl catechol; pyrocatechol methyl ester; pyrocatechol methpl ethei; - pyrocatechol monomethyl ether; pyroguaiac acid:1-hydroxy-2-methoxybenzene:2-hydroxyanisole; 2-methoxyphenol. COMMON USES Used therapeutically as an expectorant. (R-4) Food flavoring agent. (R-5)Used in the manufacture of perfumes and flavors (vanillin) and as a raw material for manufacturing papaverine (medicinal). (R-6) Usedd in the manufacture of catechol and guaiacol compounds. (R-6) Used as an intestinal disinfectant. (R-10). Used as a fumigant. (R-25) STRUCTURAL FORMULA OH OCHs ml CrH.o, mw 124.15
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1), 4TS of 4-Hydroxyauisole on Hanuter........................... .. 107 . The Effect of Trypan Blue on. -ride in the.Rat ........................ 115 iole of the Methylated Nucleic ............................ 129 \ M. BRYANs, AND HAROLD F.. hildren.uith Hurler's Syndrome. 141 Morphologic and Ctochcmioal Tmnsformation ....... .. .. ...... .162 :Lr•.Lnnclr:a• l'r)'N4d hlmlr.diun of ntintul :....................... 17fi~s:. Correlation nf Sc:umiug lilcc- ~ Images of Individual Cells in ............................ 186 ~-'he Hypertrophy of. Levator Ani one: An 1;lectkon Microscope ................... .............. .. 199 }xKNAGEL: Early Incorporation do into Rat Liver Particulate~ .............................. 219 v, Beltfmore, Md. 21202dor Aeademic .eweaYork, N. Y. 10003 issues eech) will be published. no: $24.00 :Id beaent to the olfiee of the publishers uRcaof I.ha,PuhliNLnrN nL lu,wl: ,otll old and uew uddruxsea. nsltimore,.Md. 21202 +, Inc., New York, N. Y: 100a$ ~- nLMtlIIMYMAI: ANI/: M/NAYnY.AII l•ATI/NAM:V 10, 1117-I14 (I!MiD) The Effects d( 4'Hydroxyanisole on Hamster Cheek Pouch D. A. WOODS 8lec7rrcxlMicrlucopy Unit, Imperial Caneer ReArorch F•und, fineoln's Inn Fields, London, W.C2. RR AND C. J. Smrrtt' 7H•IV,rlmrr,l u( lAwlrrl.Wirvrre• /fmvd f•nflrg,• u( 4rrgrv.rr, IAVrdul• W.f•.Y.. B.vveiu,vUMnhrr 15, J:NW4-Hydroxyanisole (p-methoxyphenol) is a compound used in industry and for clinical depigmentation purposes. Sea] et al. (in press) found that treatment of guinea pig ear skin with 4-hydrosyanisole produced.ultrastntctural changes in the relationship of basal keratinocytes to subepidermal connective tissue as well as depigmentation. 3-Hydroxyanisole brought about similar changes although its action was slower and 2-hvdruxiMisole had no apparent effect. Under continued applications of 4-hydroxyanisole, basal cells formed pseu- dopodia which projected into the dermis; these markedly resembled micro- invasive processes found in premalignant harlister cheek pouch epithelium which hadbeen painted with the carcinogen 7,12-dimethylbenz(a)anthra- cene (DMBA) and possible premalignant human oral mucosa (Woods and Smith, in press). However, when treatment with 4-hydroxyanisole was ter- minated, the skin npll.•Lrcntly reverted uL normal whereas the effects of DMRA were irrevocable. In addition there have been other reports of pseudopodial micro-invasion across the dermal-epidermal jundionn in spontaneous human neoplasia (Luibel et al., 1960; Ashworth et al., 1961;. Hinglais-Guillaud et al., 1961) and experimentally induced animal tumours (Frei, 1962; Fasske and Morgenroth, 1966; Tarin, 1967),.. This phenomenon has not been observed after applications of irritants such as aylene or benzene (Listgarten et al., 1963). In view of the reported ultrastructural effects due to 4-hydroxyanisole which parallel those of a known carcinogen, this chemical has been employed in an experimental site thought more likely to respond to any latent carcinogenic activity. 'Pheresults of some short-term experiments conducted with 4-hy- droxyanisole inn the hamster cheek pouch are reported. MATERIALS AND METHODS The agent was male up as a 20e,o cream in lanolin (704;% anhydrous wool fat, 29% water,, and 1f:: cetyl alcohol; Sea] et al. in press) and was applied thrice ' t'rophit L:anarer Itescarch SLUdenL. ~ 107 ~ emnudlr6) IvM, I,y. AmAbmb 14uw; Ine. . IOOOOOO i-b td W ~I,~
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Y { 7 ,. Y.':¢ , 1'S~4'i\!nOtYSap", ;: • v .1 0 0 W 0 F' \ .1 e IN~ Fm. 5. (n)Slectronmicrngmph of deKenemting muscle tdtmeing disaeminntian.nf nivolilnmenta in nn twdemn4ms wvl+ptlusm.'11te Inlin nf snrc,pLmm lu mynliber is grently incret~il fmm that in Ihc nnrtnnl cnnditimt (h). X Ye,1xxT nuwgtiliani,m: at scattered intervals, regions of the subepithelial connective tissue were in- vaded by extensions of the proximal plasma membranes of some basal keratin- ocytes through the lamina densa (Fig. 3). Theseext'ensions have been termed pseudopodia because they were empty of cytoplasmic organelles and tonofila- ments, and contained only an electron-translucent. ectoplasm. There was no
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132 TABLEI LINEAR REGRESSION ANALYSIS DATA OF RESULTS FOR THE EFFECTS ON SCE BY WEAKLY ACIDIC CO65- POUNDS Compound' CASNb.h' Purily` (%) 1. Phenol 2. 2-Methylphenol 3..3-Methylphenol 4. 4Methylphenol 5. 2-Ethylphend 6. 3-Ethylphenol 7. 4-Ethylphenol 8. 2,3-Dimethylphenol 9. 2,4Dimethylphenol 10. 2,6-1Nmethylplienol 11. 2-(.)-Propenyl)phenol ( cis: trans 1: 3). 12.2{2-Peopenyl)Phenol 13. Catechol 1 108-95-299.7 95-48+799.4 1319-77-3 991 106-44-5 99.9 90-0" 999 29471-88-398.6 123-07-9 99.0 526-75-0 99.8 105-67-9 98.6 2513{)1-4 99.7 638D-21-8. 95.04 1745-81-998.0 120-80-998.0 J$..2-Methoxyphenol (guaiacoq 90-05-1 99.8 I6. 2-Methoxy-4-(2-propenyl). phend (eugenot) 97-53-0 99.6 17.2-Methoxy-M(1-propenypphenol (isoeugenoq ( cis: trnnr 1: 9) 97-541 93.4 18. 2,6-Diinethoxyphenol 91-10-1 99.7 19.4-Hydioxy-3-methoxybenxaldehyde (vaniBin) - 121-33-5 99.6 20. 2-Hydroxyd-methyl-2-cyclopentcn- lone (ryclotene) 80-71-7. 98.6 21. 2-Hydrbury-3-ethyl-2-cyclopenten-l- une 21835-01-8 - 22. Furfuryl alcohol 98-00-0 94.5 23. 3-Hydroxy-2-methyl-l,4- pyrone (lnaltol) 118-71-8 9L9 Conccn- Regression Correlation Signif- Degrees of tralion coefficient coefficient icance freedom range (SCE/cell/ Ievel' (mM) mM) 0-2.0 0.5 0.25 ns 5 0-0.5 1.4 0.21 ns 4 0-1.0 0.8 0.27 ns 4 0-0.5 5.4 0.76 ns 4 0-0.25 -5.6 -0.80 ns 3 0-0.25 -0.2 -0.02 no- 4 0-0.25 4.1 0.2 ns 4 0-0:5 -3.4' -0.64 ns. ~ 4 0-0:1 -6.9 -0.28 ns 4 0-0.25 7.3 0.39 ns. 4 0-03 17.9 0.98 •^ 4 0-1.0 -1.9 -0.65 ns 4 0-0:25 35.0 0.96 4 0-0.25 1.3 a25 ns 5 0-0.5 6.9 0.88 6 0-0.5 -24 -0.49 ns 4 0-1.0 21 0.W • 4 • Numbe.s refer to Fig. 4. h Chemical Abstract registration number. ` Estimated by gas chromalography.. d Estimated by'H NMR. • 50% w/w ih propylene.glycol. ' n5, Mt dlglliaeanL • p < 0.05. p < 0.01. p<0.0o1. Instramentationn graph was equipped with a fused silica car inAnalytical GC was conduetedd with a Hewlett column (60 m x 0.32 mm) coated with a. <:m- Packard 5790 A instrument fitted with a capillary ically bonded polyethylene glycol polymer (. .- -)CI- injectar, a flame ionization detector and a Hewlett cowax 10) as stationary phase. The GCcono ms Packard 3388 A integrator. The gas chromato- were set to give a column flow of 1 ml mirT of He- a splirtl lion port I lemperatur GS-MS in5trument Kntos DS program w tion, lnc+M held at 250 WpdiDEl o. Blood it epttrifuged and the su cyte fractio 199 with E attologo115 ...(Reagent gt England) a ; 24 h inart POt[nds t0 ' :':-the tests< inducer ST il
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hem, rtion oxy- ~V v of uene Ig, tion OsS, L $. ~. ~ tage Dis. Interaction of BHT with other antiosidants 1195 actual metabolic activation of BHT and could serve as sort of a role of BHA in this system. So there are other considerations one needs to think about once you generate the reactive intermediate and what could happen in: terms of toxicity. IN. Simic, National Bureau ojSlandardr:Just a comment. We have measured the redox potentials of BHT and BHA, and the redox potential of BHT is higher than that of BHA. So the Bowof electrons is going to go from BHA to BHT. ,y. Trush:.Based upon that, yes. But based upon our resulls;.lhey are consistent with the data we're presenting ]n the formation, of the products we're seeing. Hf. Simia: That's interesting, but then you have to consider perhaps some different mechanisms.
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ious investigations theadminis~ inducers of hepatic xenobiotic n to the rat has been shown to ;he urinary excretion of both D- acid and other metabolites of the lie acid pathway (Aarts, 1966; '/i.., 1960;: Conney ef al., 1961;; r af., 1973; Lake et al.,. 1976; Reid, 1963; Notten and Hender- However, the present data, in with the observations of others id Chasseaud, 1976), would sug- lepatic coqjugative activity may cd withuut ;uly uvcfull stimu- he D-glucuronic acid pathway. wgh there was no increase in the .:retion of either D-glucaric acid, acid, or xylitol, moreUDPGA mably made available for the ~runyltransferase catalyzed for- glucuronides; Furthermore, the mulation of the above three D- acid metabolites correlated with effect of the Phase II compounds ad on the hepatic mfo measured. Asion, these observations on the administration of Phase II sub- I support to the validity of ineas- ctrum of certain urinary metabo- e D-glucuronic acid pathway,, Livic acid, L-gulonic acid, and~ an index of the activities of :rosumal cytochrome P-450 de- fo in the rat. However, these Juid be substantiated by studies t of chronic treatment with Phase :s on rat hepatic mfD activitiess trinaryexcrelion of o-glucuronic ,l lilcs. 'KNOWLEDGMENTS .rms pan of a research project sponsored ainistry of Agriculture, Fiaheries and n our thanks are due. The results of are Ihe. property of the Ministry of ~isheries and Food and are Crown e skilled technical assislance of Mr. iald is gralel'ully acknowledged: o-GLUCURONIC ACID PATHWAYBEGULATION 377 REFERENCES (1977d).Metabolismofarylaceticacids.4.-Pltysieo- - AARTS, E. M. (1965). Evidence foethe function of o-glucaric acid as an indicaWr for drug induced enhanced metabolism through the D-glueuronic arid pathway in man, Biarliral. Pbarmncal: 14,. 359-363. AAETs. E.. M. (1966). DitTerentiation of the barbiturate stimulation of the glucuronic acid pathway fromm dt noeo enzyme synthesis. Birxhrnr. Plmrmarul. 15, 1469-1477. AAavs,.E. M. (1971). o-Glucaric-acid excretion as a test of hepatic enzyme induction. Gmrel. 1, 659. BERENROM, M. ANI) YOUN(1, L..(1951). Blhchl'mleal rAndiex.ul'tnnic agrnls. l.'I'h1-i.wdal;m" nl'I-:u,d 2- napbthylsull'uric acid :nld I-:uW 2-naphlhylghlcuru- nide from the urine of ral.s dosed with 1- and 2- naphthol. Biacbem: J. 49, 165-169... BocK, K. W. (1974). Oxidation of barbituraless and the glucuronidation of t-naphthol in perfu.sed rat liver and mierosomes., Nanrn•n-.Sr•lunit•drbrrR'.r Anrb.PluHrnnc•nf..283, 319-3,TG. BaNSNrc, R. W., AND TAns~sKV, H.. H. (1945); On the calorimetric determinalion of crealinine by the lntfe re:mtion.J. /lie1. ('ln•nl. tEg, 581-591. BOOTH. A• N.. MASRI. M. S.. RoaalNs, D. J., EMERSON. 0. H.,. JONES, F. T., AND DE EDS, F• (1959). The metabolic fate of gallic acid and related eompounds. !. Biol. Chem. 234, 3014-3016. BURNS, 1. 1... CONNEY• A. H.. DAYTON, P. G.. EYANS, C.,.MARTIN, G. R., ANDTALLER, D. (1960) Observations on the drug-induced: synthesisof D- glucuronic,.L-gulonic and L-ascorhic acid in rats. J. Phnrmarrrl.. Exp. Tlrcr. 129,. 132-138. CONNEY. A. H., BRAY, G. A., EVANS, C., AND BURNs, J. J. 11961). Mclah(dic inler,(Clions between L- ascorbic acid and drugs. Ann. N.Y. Arnd. Sd. 92, 115- I27. CREAVEN, P•.J., PARKE, D. V., AND WILLIAMS, R. T. (1965): A fluorimeiric sludy of the hydroxylation of biphenyl in' virru by liver prepunnions of variuusspecies. BiicGnn. J. 96, g79-8g5. DACRE,.J. C. (1960). Metabolic pathways of the phenolic antinxidants. J. New Zenland lnar. Chenl. 24, I61-171.. DIXON, P.,A.F., CALDwELL, J., AND SMIIN, R..L. (1977a). Metabulism of arylacelic uciJs. 3. The metabolic fate of iliphenylacetic acid and its varialion with species and dose. Xrnafriuricn 7, 717-725. DIXON„ P. A.: F.. CALDWELL-.J., AND SMITH, R. L. (1977b). Metabolism of arylacetic acids. 1. The fato- of I-naphthylacetic acid and its variation with speciesand dose. XenubirNira.7, 695-706. DIXON, P.. A. FL, CALDWELL, J'., AND SMITH, R. L. (1977q. Metabolism of arylacetic acids..2. Thelate of I"Clhydralropic acid andils variation wilh species and dose. Xrunbinrinr7, 707-715.DIXON, P. A. F., CALOWEA.L, J., AND $MITIt. R. L. chemical, structural and biological factors iMuencing the pattern ofeonlugation. Xenubiarica 7, 727-736. GANGOLLL S. D., LONGLAND, R. C.. AND SHILLING, W. H..(1974). A gas-liquid chromatographic method for the determinalion of o-glucaric acid in urine.. Clin. Chinr. Arta 50, 237-243. H(IIT2MAN,.J. L.. GRAM,.T. E.. GIGON, P. L., AND GILLETTE, J. R.. (1966). The distribution of thee components of mixed-function oxidase between the rough andthe smoothh endoplasmic rcticulum of liver cells. Riuchem. J. 110, 407-412. -HUNTER..J., AND CHASSEAUD, L. F. (1976). Clinical aspects of microsomal enzyme induction. In Prngrexs in !)rNx M~rnMdLrm (J.. W. Hridges and L. F:. l'h:nncau,l,: cds.). Vul. 1. I'P. G9-I91. Wiley,.. London. HUNTER, J., MAXWELL, J. D., STEWART. D. A. AND WILLIAMS, R. (1973). Urinary o-glucaric acid ex- . eretion and total liver content of cytochrome P450 inguinea-pigs; Relalionshipdunngenzymeinduclion and following inhibition of protein synthesis. Bi.>- rbrm.J'hnrrnaruf..22, 743-747. IAMES,. M. 0., SMIT/i, R. L.. WILLIAM; R. T.. AND Hr:Itm.NaERn- M. (1972). The cnnjug:u'sm of phcnyl- ucelir acid in man. sub-hu(nan primmcs and some non-primate species. Prne. Rny. S.N% Srr. B. 182, 25-35. . JUULOW; D. J•, THOROEIRSSON, S. S., POTTER,. W. Z.. HASHIMOTO,. M.. AND MITCHELL, J. R. (1974). Acelaminophen-induced hepatic necrosis. V1. Me tabolir dispositionof toxic and nontoxic doses of acetaminophen. PGar.nacoingy 12. 251-271. LAKE. B. G.. LUNGLAND, R• C.. GANGOLtI. S. D.. AND LLOYD, A. G. (1976). The influence of some foreign compounds on hepatic Xenohiolic melaho- lism and Ihe urinary excretion of u-glucuronie uid metabolites in the rut. ToxicoL ApPl...Pharrrra('ul. 35, . 113-122. LAKE, B. G.,. LUNGLAND, R. C., HEROD, I. A.. COLLINS. M. A.. AND GANGOLLI, S. D. (197g). The effect of 4-hydroxybiphenyl and 2-naphlhol on hepatic xenobiotic metabolism and the urinary excretion of metabolites of the o-glucuronic acid pathway in the mt. BFwIrrnr. S/N-. Trmrs.: 6, 1246-1248. LOWRY; O. H., RosEBROUGH, N. J., FARR, A. L., AND RANDAEL, R. J. (1951). Protein measure- ment with the Folin phenol reagent. J. Binl. Cln•rn. 193, 265-275. MARSELOS. M.,.AND HANNINEN, 0. (1974). Enhanee- ment of D-glucuronolactone and acetaldehyde dehy- drogenase activities in the nt liver by inducers of drug melabolism: Biochnn. PAornlarol. 13, I457- I466. MARSH. C. A. (1963a). Metabolism of o-glucurmm- Iaclone in mammalian systems. 2. Conversion of o- glucuronulactone into u-glucaric aeid by tissue preparations.:Bint!(rnl. J. 97. , 82-90. 87119244 I
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til.rl'1'I I . '..lE* `__k~ rlu xh„wiux Airxmimniwl uI' myfJJamenls 'myl~liher'.l Krcullrincn^rsed rrnnr that inn thelial connective tissue were in- membranes of some basal keratin- tese extensions have been termed ~oplasmic organelles and tonofila- ducent ectoplasm. There was no 1.11YUROXYA\ISOl.h: Il\ IIAMRTISR CHFEK POl1CH 113 apparcntt tl:nnaga lu Ilke collngt•n lihcrro in Ihe viciniiy uf Ihtxv psrudupntliir this dill'eretl I'renr comlxtrahlc I'cnt.urcN- previously reported in t,trcinngenesis where such pseudopodia are avcociated with collagen destruction (Woods and Smith; in presv). C.ellsin the tipper sla:Ha ul' (lie epilliclium also shuwcd abcrrant( features. Intlividunl isoLnad cells in Ihe slralimt spittte;um„ churacl.erized as keralino- cyteshY Lhcir desoustnnes andd timolilantenl. amlent, pucw-x.ed cytoplasm which was k-..g ' cleclron opaquc Ih;m Ih:d ul' thcir m•ighliorsfhig: 41: 'I'hr plasma membrane ol' these "light" cells was often closely applied to: lhe ntem- branes of adjacent cells but attachment complexes were not formed at these sites and marginal prolongations containing a clearectoplasm,r appeared to be pushing between neighboring epithelial cells, in a manner which suggested loss of contact inhibition. -- In the mteuJe layer, whole muscle blocks exhibited degenerative change . (Fig. 5a); there was loss of integrity ot' muscle tibers;, and myofilaments were scattered loosely in an oedematous sarcoplasm. The overall appearance was one of prateolyticbreakdown of the muscle when compared with apparently normal adjacent.muscle blocks (Fig. 56), - DISCUSSION' Continuous exposure of the cheek pouch mucosa to 4-hydroxyanisole I'or a relatively short period produced a condition that included hyperkeratosis, epithelial hyperplasia, disorganization of basal epithelial cells, and disturb- ances at the epithelial-connective tissue junction and muscular degeneration.. Comparable epithelial-connective tissue disturhances,. at both the light mi- croscope and ultrasl:ructural levels, have been observed in developing experi- mental hamster cheek pouch carcinomas (Woods and Smith, in press), hyper- keratotic human oral mucvtsa of possible premaliKnant character (Wrwds and Smith, in press) and peripheral regions of human oral carcinoma (Woods, D. A., unpublished results). In these latter examples, basal cell pseudbpodia have been seen to attain a maximum size and number either in, or adjacent to, welLditferentiated tumors. These experimentx% have shown that 4-hydruxyanisolecnuses apparently benign epithclinl invasi+m, anvmtp+miedd by Ihodevelnpmonl, of basaJ crll pseudop<xtia. In widely differing circumstances, the latter have been incrimi- nated as associated with incipient neoplastic change but continued exposure to the chemical I'or several weeks longer than in the above oxperimentec has net substantiated or contirmed any true carcinogenic potential.... Nevertheless, a sinister lesion has been generated in our experimental tissue by employing a cosmetic material. It should be noted also that the butylated form of hy- droxyanisole is used as an anti-oxidant in many foodstuffs. The effects of prolonged exposure of the hamster cheek pouch to 4•hy- droxyanisoleare being studied but itt is difficult to estimate when long term experiments have been sufficiently exhaustive to pronounce the chemical free of suspicion.
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_ti rep- ectixel~, in frac- -4 pyro. +nent of were a tyl and but in rboxy fc _d indc- tn al6%l fdent!- phenol,- 1roXV-'•_ in con . fmctLn- oft, ils pr y1gua: 28% ~kcla. nolr FT3 F1 F10 fF5 F]g-5. Frequmciesof SCE in human lymphocytes aftef Ireat- ARE mmt with the semivolatile fraction of agarette smoke con- 5 ~ deo-vte (O)- the total weakly acidic fraction (.)„ and the _ ] aeklyy acidic subfractions indicated in the figure (0).. Dashed edor ine in the curva for F2 ihdicates.two different eaperiments.. cDr[anb~un(W/me -fi6.6. tllustntion of the rEgression caeffictienu of the fractions , F1-11, whiehhavc been normaliaed vs- the total weakly acidic focliun in each lest. Line F shows Ihe.mean of the regrrscion 13S m ,o 0 20 m 0 U y 20 a 1.0 zo ,a 2.0 0 2o 20 10 025 ni. o: On~, 02 ~ 0.5 20 i i-Ein.pne.W zo m ~-E~nyMerol za 05 O xa.D..emrim.:•a / a 0.25 0.5 a iA-D~melMynxm~ 20 m 0 025 05. 0 0.1 0,2 0 0fl- 0.2 ConcmtrD~ian ImMI Fig. 7- Frequendes of SCE in humen lymphocytes.after trwl! ment with individual compounds.from the weaklyacidicy frac- tions(9): The effect of the positive control!(styrene-7,8-oxidc) frorneach experiinent is shown (0). cis- and frans-isoeugenol. The main components of fraction F8 were phenol and the two cresols, 3- methylphenoP and 4-methylphenoi. The 2-methyl- phenol was eluted later and thus collected in frac- tion F9,, which mainly contained multialkylated phenols. Fractions F10 and F11 both comprised vinyl- and propenyl-substituted phenols. The main constituent of fraction FIO was 4-vinylphenol,. which polymerized during work.up.k coefficients obtained for the tests of the total weaklyaeidic fraction: The intercept shows the mean of thee background values (11.2 SCE/cdp from the testsaf the fractions F1-11'. Fraction F6 was signiKcant both for linear and quadratic regression and normalired values both for Ihelinear regression coefficient and for the hnear coefficient from the quadraticc regression are shown. 02 i
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o an:- m ta (' l.ambert- B.-.A. Lindhlad, M..Nordenskjbld and B. Werelius (1978) Increased frequency of sisier-chrumatid exchanges in cigarette smokers, Hereditas. 88, 147-149. I-aVoie, EJ:. SS. Hech t. D. Hoffmann and Ll, W ynder (19801 The kss harmful cigarettee and tobacco smoke flavors, in: G,B..Gori'and F.6. Bock.(Eds.),.Baobury Repor[3. A Safe Cigarette-Cold Spring Harbor. New York, pp.251-260. Liringston-G.. and R.M:.. Fineman (1983) Correlation of hu- man lymphocyte SCE frequency with smoking history, Mu- tation Res.,.119, 59-64. Nodnwto, K., S Wolff and A. Koizumi (1983).Inductfon of sister-chromatid exchanges in human lymphocytes by mi- crosomal activation of benzene melaboliles, Mutation Res.,.. 119,355-360. fJcstmann, E.R.- E.G: H. Lee. TJ. Matula, G.R. Douglas and 1.C. Mueller (1980) Mutagenizity of constituents identified in pulp and paper mill effluents using the. Salnwnella/ mammalian-microsome assay, Mutation Res., 79, 203-212. hpppa, H., and H.. Vainio (19g3) Induction of sister-ehro- matid'eachangesby styrene analogues in.cultured humam lymphocytes. Mutation Res., 116, 379-387. porppa,.H.. M. Sorsa; P. PPJflli and FL Vainio (1980) Styrene and styrene oxide induce SCEs and are metabolized in human lymphocyte cultures, CarcinogetKsis, 1. 357-361. i 139 Nbrppa. H., H.. Vainio and M. Snrsa.(1983)Metabolic activa- lion of styrene tlyerythrocyres detecled as increased sister, chromatid exchanges in cultured human lymphocvtex. Cancer Res, 43, 3579=3582: Perry, P.- and H.J. Evans (1975) Cytological detection nf mutagen-carcinogen exposure by sister chromatid ex- change, Naturc(Ltndon), 258, 121_125, Pool. B.L., and P.Z Lin (1982) Mutagenicily testing.in the Salmonella r}phinrurium assay of phenolicmmpounds and phenolic fractions obtainedGom smokehousesmoke mm densates, Fd. Chem. Toxicol.,,20- 383-391. de Raa4 W.K. (1979) Comparison of the induction by cigarette smoke condensates of sister•chromatid exchanges in Chinesa hamster cells and~of mutations in Salmanellu ryphimuriuur. Mutation Res:, 66, 253-259. Stich. H.F., M:P. Rosin, C.H. Wu and W:D. Powrie (1981)' Clastogenicity of furans found in food, Cancer Let[.13J. 89-95. Strand, L.P., and R.R. Schelina (1975) The metabolism of vsnillin and isovanillimin the rat. Xenobiotica, 5. 49-63. Wo1fG S:, and P. Perry'(1974) Differential Giemsa staining of sister ehromatidss and lhestudy of sister chromatid exchanges withoutt autoradiography: Chromosoma, 48:, 341-353.
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%Z- 3 n G, r-LZ 1; l: aa-1S i 5 . . , C f7 l o r oq -fmu,-G A-GJ 00 I Au4. ~s~~ . . . Sre-ukA'-duuslc Sn,.okt (imnAILSO~r_ f, ~ . .. 1e~,laf72 h.u.u„ 4~,-c /y~'.~- j~~d; z s 66.C~ ~tiC 9K/,, k ;4PfYLJ. , _. EG(: Tax, ~ A.,y/;~q~7•st,-s ~/h g 14 T 't, (1) f -H o S+~d.w ~.~.sa4 7S. ~SC6) TI _ To6. ~cOA5f,7'- 1$ P,di-O .~_. ~ ~ ,Grd„tn 1 -tA Ca,~al /()e r a dc Aa (U& z+krssoM1 -1 ~ ~ G'7w6-Srn,A C,aa.~cde v-rs 5sx,r--f-jb e&~J I wf .-iC.4d ~uja-AA--Ac,t~ TaX 'Dtj. Ic LG) 14 k~
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130 FI -TF3--i F31 F.-1-F5-4-Faf-Fr-I Fe--{w---4--Fn i Fig. 1. Gd chrolmtograpliie characteristica for Ihe separation of the weaklyacidic 991aterial ion Sephadex LH-20. the weakly acidic, semivolatile (WASV)subfraa tions (Ctttvall et al-, 1984). Several studiess have shown that biologicalfyaetive components; such as tumor promoters and cocarcinogens, are present in the weaklyy acidic (phenolic) fraction of cigarette smoke condensate (Hoffmann et a1.,,1983; Hecht et al., 1981). However, the data on genotoxicity of individual phenolic compounds are sparse and, to our knowledge, catechol and hydroquinone are the only ones present in tobacco smoke that are known to iuduce SCE (Morimoto et al., 1983). We there- fore decided to study the genotoxicity of the WASV fraction and its constituents by measuring their effect on SCE (Perryy and Evans, 1975) in human lymphocytes. The WASV fraetion of a cigarette smoke con- densate was separated by gel chromatography into 11 subfractions (Fl-F11). The chemical composi- tions of these were examined by gas chromatogra- phy (GC) and gas chromatography-mass speo- ;, trometry (OC-MS). The 11 subfractions and 23 individual compounds,, most of them abundant in the WASV fraction, were tested for SCE induc- tion. Fractionation of cigarette smoke condensate A cigarette smoke condensate was prepared from domestic nonfilter cigarettes of American ~~ 9oatOe wttcrta sea,olan. ..u.~el. Ir9nl M~tl[Inn ~-. .e.mt wtea.l easlt rl.au Meva /a9l.In r.ur .<Ime .aerln ts g) . Fel 9nrmtugn" FI It {l q F6 F6 F]{g'q flt 0J9g 0.329 0.021, 0:P1 0.209 0j119 0-ng 1A9g M0g U.1-. 3.a1 s.a 11.42 ).u .A1 i.rr 5.06 n:ai Iu.Ot La Fi& 2. General fractionalion schelne and yields from th, adea LH-20 gel cbromatngraphy, of' 5 g of the 9veakh . fraclion. 87119322 blend type nicotine per and the con K scribed prem tion of the using CO, different te yielded 3 fr (20A,) and fraction Wa NaNCOy, A acids (5%). .v and neutraL proccdures (Curvall et a Part of I soparated bi (8 x 150 cm, The cohmm water with a monitored (Waters Ass fractions. '11 I
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Dmemal slana$rtl~ 94 102 05 14 ~_ 26 - 30 ! 35 58 46 0 94 (inCBmmlSlantlaN) 102 119 87119357 129 4 105
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compounds were potent inducers of SCE, catechol T1 ti. (p < 0.01), 2-(1'-propenyl)phenol (p < 0.001). vanillin (p<0.01);. and that 4 of them had a ~~ - moderate effect, i.e. cyclotene (p <0:05),'guaiacol ( p<. 5), isoeugenol ( p< 0.05), and maltol ( p< r _nsorcinol and hydroquinone, resorcinol: has been ~"ihown to induce chromosomal aberrations in hu- 0.05),. Statistical data of the test results obtained for individual components are shown: in Table 1. The compounds that turned out positive in the first experiments were reexamined using two con- centrations. Resultss of the confirmativeexperi- ments.are shown in Table 3. Discussion Phenolic compoundsgenerated by pyrolysis are found in the smoke from various sources, and many of them are important flavorants of cigarette smokeand food. The weakly acidic (phenolic) fraction of cigarette smoke condensate, is known to exhibit cocarcinogenic and tumor promoting activity(Hoffmann et al., 1983; Hechtetal., 1981). Many weakly acidic compounds have been identi- fied in this fraction, but to our knowledge,, only one,.catechol, has been found cocarcinogenic, al- though the presence of other cocarcinogens have been indicated (Hecht et all, 1981). Phenol and several alkylphenols have been shown to exhibit tumor-promoting activity (Boutwell and Bosch,. 1959). There are fewpositiva dataa on in vitro geno- toxicity of weakly acidic compounds. A number of phenols have been examined for mutagenic activ- ity using the Ames test (Florin et al., 1980;. Gocke et al., 1981; Nestmann eT a1.,.1980;Pool and Lih, 1982). Mutagenic activity has been found for phe- trol on the frame-shift strain TA98 and for re- sorcinol andd hydroquinone on the base-pair strain TA1.535, and for resorcinol also on TA100 (Gocke «a)., 1981). Of the benzene metabolites,.catechol, iasan lymphocytes (Darroudi and Ngtarajan, 1983) - and catechol and hydroquinone to induce SCE in human lymphocytes. (Morimoto et al., 1983).. As hydmxyphenols are strongly acidic and water- .toluble, they are not present in the weakly acidic Traction described in this study. Catechol, thee most > abundant phenolic constituent of cigarette smoke, , was tested here and its SCE-inducing activity was amfirnred. 137 The fractions containing alkylphenols. F8 and F9, induced SCE, but when testedd as individual components, the alkylphenols did nor show any activity: In a study by Cheng and Kligerm:m(1~984). the cresols were investigated for their SCE-inducing effect and o-cresoll was found toexhibit a low activity. The commonn food additivevanillfn (Arctander, 1969b)) was found to be a potent inducer of SCE and therefore probably contributessignificantly to the activity found for fraction F7. 2-Methox- yphenol, also encountered in this fraction,. exhibited a loweffecUon SCE. These results indi- cate that the 2•methoxyphSnoL_ moieiy_.may_.bt essential for this activit,Yard that itmay,benotert tialed by an aldehyde group_ in the_ pom-posilion, t ereunimg Tor thehigh~activity of vanillin. Genotoxic effectsof vanillin have not been dem- onstrated earlier although tested for mutagenicity. (Florin et al., 1980) and for induction of chro- mosome aberrations (Kasamaki et al., 1982):.The only other result indicating that vanillin may be genotoxic is the finding that vanillin is converted to the genotoxic agent catechol by the intestinal microtlora in rat (Strand and Scheline, 1975). Isoeugenol and eugenol, two 2-methoxyphenol derivatives present in fraction F8, are used in the production of fragrances and also approved for use in food (Arctander, 1969a). When tested for SCE induction., isoeugenol (2-metltoxy-4-(1-pro- penyl)phenol) was found to induceSCE, while eugenol (2-methoxy-4-(2-propenyl)phenol) was in- active. In view of the fact that 2-(1-propenyl)phe- nol,, but not 2-(2-propenyl)phenol,, was foundto induce SCE, it seems thao a double bond con- jugated with the aromatic ring is essentialfor achieving active alkenyl substituted phenols.. Norppa et al..(1980, 1983) have shown that styrene is converted to the potent SCE inducer styrene- 7,8-oxide in' human lymphocyte cultures, a reac- tion primarilymediated by erythrocytes. It may be assumed that an analogous SCE-inducing.epoxide can be formed when a 1-propenyl substiWent is present. This is further indicated in a studythat showed trans-s-methylstyrene to have a SCE-in- ducing effect (Norppaand Vainio, 1983). Several alkylated 2-hydroxy-2-cyclopenten-l- ones,, some of which are known flavorants, were detected in aa tumor-promoting weakly acidic frac-
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134 Results Fractionation and analysis of rheweakly acidic semioola7ile malerialThe gass chromatographic profile obtained for the WASVfraction is presented in Fig. 3. Frac- tionation of the weakly acidic material by gel chromatography furnished~11 subfraclions(Figs. 1 and 2)...The individual components of thesubfrao- tions were tentatively identified by GC and MS data. The compounds that were eluted in the first 4 fractions.were either water-soluble neutral con- stituents or weakly acidic cyelio-tbmpounds of low molecular weight. Thus, fraction FS contained acyclic and cyclic alkylamides, lactames. (pyrro- lidinones and piperidinones). Also found in frac- tion F1 were some.diketones and alkyl-substituted maleic acid anhydrides. Weakly acidic y- and d- lactones were found in fraction F2,. which also contained some alkylamides. About 20% of the weakly acidic material was eluted in fraction F3,, which contained mainly alkyl-substituted 2-hy- drozy-2cyclopentenones. The two, main compo- nents were the 3-methyl-substituted (cyclotene) and TABLE 2 the 3-ethyl-substituted derivatives, which rep resented 30 and 15% of fractiom F3, respectively. This.group of compounds was also found in frac- tion F4 together with somehydroxy-alkyl-4-pyrP nes (maltol derivatives). The main component of fraction F4 was furfuryl alcohol. - The principal components of fraction F5 were a mixture of alkyl-substituted (methyl, ethyl and propenyl)) dimethoxyphenols. Also found, but in minor amounts, were long-chain alkyl carboxylic • acids. Components of fraction F6 included inde- nols, coumarin derivatives and long-chain alkyl carboxylic acids. Additional componentss identi. fied in.fraction F6were some methoxyphenols. . including the vanillin analogue 4-hydrox~.-3. . methoxyacetophenone, which was the. main corn. 1 onent . i In addition to the main component in fra:.5on i F7, vani)Gn, lvhich accounted for some 20% o; ~ilis fraction, there were other 2-methoxyphenols --e.~ ent, e.g. aiaw methylguaiacol,, and ethylg. :la- col.. Fraction F8, which comprised about 2~ of the weakly acidic material,, contained alk, -,ed p}tenols (Cl, Cz and C3) along with eugeraL nd EFFECf ON SCE BY WEAKLY ACIDIC FRACTIONS DERI VED FROM THE SEMIVOLATILE MATERIAL OFCIGi.. :TF SMOKE CONDENSATE; LINEAR REGRESSION ANALYSIS OF THE DOSE-RESPONSE DATA SHOWN IN Fig. - Fraction Concentmtionrange Regresrionmeffident Correlation Significance° Degreesoffree (µg/mq (SCE/ceBxpg-txm1x10-1) coefEcient lerd Semi.olatile Weakly acidic Fl F2 F3 F4 FS F6 0-80 12.8 0.99 2 0-80 7.5 0.91 4 0-200 4.2 0.94 5 0-300 3.1 0.97' 4 0-200 5.1 0.88 5 0-60 36.4 0:98 4 0-80 10.7. 0:g9 3 0-150 6.5 0.86 4 e F3 tti a , _Fi& S. Frequencies ...mt .+im the sar dquae (M the. I ::rApNy .ridic subh. iec in the aarve for . #
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r 136 TAOLF. 3 INDUCTION OF SCE.IN HUMAN LYMPHOCYTES aY' WEAKLY ACIDIC COMPOUNDS FROM THE SEMIVOLATILE MATERIAL OF CI(iARF.TTESMOKE CONDENSATF.; CONFIRMATIVE EXPERIMENTS Cnmp.wnds SCE/cell(control):SD' C'oncentrations(m M) SCE/ceIll(trea(ed):SD Significanecinh Iwo tailed r-tesc Degreesuf fn:cdom Vimillin 8.5; 3.6 0.75 20.1: 5.9 39 b.0 20.4: 4.1 10.0: 3.4 0.25 YR3:-4:9 - nx 48 0.5 13.6; 4.2 4s Catcchol o. 48 7,6; 3.0 0.15 15.1; 4.9 I 39 Cyclotene 10.0; 3.4 1.5 12.3; 4.0 48 i 7.6: 3.0 2.5 1I.L• 3.6 4s 2-(1-Propenyl)phenol 10.0: 3:4 0.25 13.7;.4.1 48 8.4: 2.4 (13)." 0.5 18.0;6.3 (13)' 15 Isneugennl 7:6: 3.0 0.25 10.3; 3.7 (24) ' 47' 7:6: 3.0 0.5 14.0, 5.0 39 Maltol 7:6: 3.0 0;5 t0.1;,3.7 f 48: 7.613.0 1.0 11.9: 7.9 48 j ' Number of analysed cells.whemless than 25, '1 s ns, not significann • p< 0.05: •• p< 0.01; ••• p< 0.001. `' VarianeesweresignificantlydiRerent(p<0.05)andther-teslwasp ~ erformedwithaseparalevarianceectimale(DiemandLcn:er. 1970). lSlS Induction of sisrer-clrromarid exchanges The semivolatile fraction, the WASV frac.,,n and the weakly acidic subfractions (Fl-Fl1) ::~re 0 or 0.2 z-uamo..-.- ~ a , 4acuuu.~0. LofA.R Lo 2.0 0 03 Lencenamcn (mM) tested for their SCE-inducing effect and he dose-response curves obtained are shown in _~ 5. Statistical data obtained on regression am . -.u, of these results are shown in Table 2. All frac- ~m except F11 affected the SCE in a dose-deper -_nt way(p<0.05-0:001); however, fraction Fl i .o seemedd to enhance the SCE frequency. The .- : al weakly acidic fraction was used as a refera . in each test and the effects of the.subfractionc .:rc normalized versus the reference. It is the:-- :~re possible to compare tests made at different .e:o- sions and with lymphocytes from different &-,r. (Fig. 6). Fraction F4 is by far the mostt act - of the subfractions and the onlyone that sh,.:a clear enrichment of the activity compared T - the total fraction. Fractions F2 and F8' exhib- the lowest activity. Dose-responsecurves showing theeffec! :. iin- dividual components on SCE are given in F,-.. i and 8. Regression analysis showed that 3+ ehe Fig. 8. Frequencies ofSCE.irt human lymphocytes aftc :ear menr with individual compoundsfrom the weakly aci& _ Gn.- tions(0). The effect of the positive rnntrol (styrene-7,8-, ildn from each experiment is shown (0). compounds we (p.<0.01),. 2 vanillin. (p<1 moderate effec (p.<0.05), iso. . 0.05). Statistic for individual The compount first experittter i ccntrations. R i fllraltsareshou' Disctssion . . Phenolic col .- found 'tn the ° ` many of them: smoke and tc fraction Of dg: ; i to exhibit coc activity (Hoffn Many weakly : ` ffed in this fr. one, catechol,. '~though the prt . been indicatec several alkytpt tumor-promoti 1959). ~'. Thereare f toxicity of wee _,phenols have 1 _~~`i,ty using the A 'TM ai;'1981; N 982).'Mutage _ `on the fr: -_-olandlr- 535, and f 1981).. C ol and to indc lymphoc~ - catechol z lymph heno e, they c descti t phe: .. `.,-tested her gt7119328
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C.j'a4-k ipa-u~ 7b'X • ~; C'r"J4'`G ~CI __UC~Jc'~S`~ ~?S.iAJ ea kL) f-v.<& X 7J~ ~fir Yl~ .! Y 5 c~ree~rw~ ~ ~b~ ~ l.~ns~ , r4 ~s C5 4 v-dC44 e-n -~ -. , - I-. 'o.&~ PIL~ F v l ~ ~ ~P ~ ~ C4~u,. ~ ~ 4 Jc`r~sdz ~ tl -7 cf -Q,umy~ b rb b l~.s¢- ~j4C_dt$54~i.1i `~ ~S jC-1ll`Jr4;r~ ~Yt1tryG~-~c~tS 2~~` ~ ~a ~ t---/-Z ~ t - Dr . ~ ~ y <
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,Nutarian Reeearch, 169(1986):129-139 Eluvier MTR 01038. ou."cb-p 129 In vitro studies of biological effects of cigarette smoke condensate II. Induction of sister-chromatid exchanges in human lymphocytes by weakly acidic, semivolatile constituents Tommy Jansson a, Margareta Curvall b, Annica Hedin b and Curt R. Enzell b ° pepattment o/Medhal CNI Genenieq Afedica( NobN Inrtitute, Karofinska ln.rtttutet, Box 60400, S-10d 01 Stockholm and bResearch Department. Swedish Tobacco Campany; Box 17007. S-104 67 Stockhalm (Sweden) (Received 20 June 1985) (Revlsion received 3 September 1985) (Accepted 6 September 1985). Summary Cigarette smoke condensate is known to enhance the frequency of sister-chromatid exchanges (SCE) in human lymphocytes in vitro and some of the activity has been found in the most volatile part of the particulate phase, the semivolatile fraction. In this study we have investigated the chemical composition and the SCE-inducing activity of the weakly acidic, semivolatile fraction of a cigarette smoke condensate. A number of individual weakly acidic compoundss were also tested for their SCE-inducing effects. The weakly acidic fraction was separated by preparative gel chromatography into 11 subfractions (Fl-Fll). The chemical composition was determined by gas chromatography and gas chromatogra- phy-mass spectrometry. Measurements of the effects on SCE in human lymphocytes were used to evaluate the genotoxic effects. All fractions except F11 induced SCE in a dose-dependent way. The most active fraction was F4 which contained mainly alkyl-2-hydroxy-2-cyclopenten-l-ones. The individual compounds to be tested for induction of SCE were selected on the basis of their abundance in the weakly acidic subfractions and on the basis of their occurrence in the environment. Of 23 tested compounds, most of which were alk enols, 7 ipyl;gdcCR i.e., catechol, 2-(1-propenyl)phenol, cyclotene, maltol, isoeugenol, -methox henot (guaiaco d vanillin.Many of these are important flavor components that occur not, only in tobac,co_&udloacw_moke_but_also in food, candies, beverages and . perfumes. Cigarette smokers exhibit a higher level of sis- ter-chromatid exchanges. (SCE) in peripheral lymphocytess than-do non-smokers (Lambert et al., 1978; Hopkin and Evans,. 1980; Livingston and Addtas mrreapoademx to: Prof. Cun R. FszeB:. Raearch - DRNment. Swtdish Tobaocu Company, P.O. Box 17007, . S10a 62Stockttolm (Sweden). Fineman, 1983). Cigarette smoke condensate in- duces SCE in vitro (de. Raat, 1979; Hopkin and Evans, 1979; Curvall et al., 1984) and some of the activity has been found in the volatilepart of the particulate phase, the semivolatile fraction,(Curvall et al., 1984). The effect of this material is enriched in the neutral subfraction, which has been investi- gated further (Curvall et aL, 1985). Some SCE-in- ducing activity was also found in the acidic and . 0165-1218/86/f03.50 0 1986 Plsevier Scicnce.PubGsbers B.V. (Biomedicaliniviaion)
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in mainstream and sidestream smoke of Bright tobacco for acid analysis. sloesMam smoke N O b P h ry 0 20 - a0 60 min ~J 90 110 ii0 C
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131 blend type (23 mg tar;. 11 mg CO and 1.8 mg nicotine per cigarette). The cigarettes were smoked and the condensate collectedd usingg procedures . de- scribed previously (Curvall et al., 1984). Distilla- tion of the condensatee in vacuo (0.1 mm Hg), using COz2 as carrier gas, was performedd at two different temperatures (20°C and 80'C) and yielded 3 fractions, volatiles (10%), semivolatiles (20%) and nonvolatiles (60%). The semivolatile fraction was extracted consecutively with aq. NaHCO,, NaOH and HCI to give 4 subfractions: acids (5%)., weak acids (phenols, 12%), bases (18%)) and neutrals (21 %). The distillation and extraction procedures used have been described elsewhere (Curvall et al., 1985). Fart of the weakly acidic fraction (5 g) was separated by gel chromatography using a column (8 X 150 cm) packed with Sephadex LH-20 (2 kg). The column was eluted with 75% methanol in water with a flow rate of 375 ml/h. The eluent was monitored using a differential refractometer (Waters Associates R 403) and collected in 25-mt fractions. These were combined- to give subfraa tions Fl-Fll (Fig. 1). Each subfraction was ex- tracted 4 times with twice the volume of dichloro- methane. The organic.layer was dried over NazSO4 and the solvent' removed at 30°C under reduced pressure (10 mm Hg). The absence of solvent was confirmed by 'H nuclear magnetirresonance (NMR):.Thegeneral fractionation scheme and the yieldsof the 11 subfractions in grams and in percentages of the WASV fraction are outlined in Fig, 2. All compounds used for testing were obtained commercially and their purityevaluated by GC or 'HNMR before testing (Table 1). Those contain- ing more than 5% impurity were purified by distil: lation (1-11, 15-18, 22), recrystalGration (13) or sublimation (20). Thestructures: of the compounds studied were confirmed by 'H- and "C-NMR (Fig. 4). For biological testing purposes, stock solutions were made up in dimethyl sulfoxide (DMSO), ethanol (EtOH) or DMSO/EtOH (1: 1) and stored at -80°C until used. ae FW 3. Gas chromatogapdc pro61eof thetotal, senivolatile weakly acidic fraction. Numbers in parentheses sliowin which ; tubhaaion(Fl-F11) the oumponent residea 1. 4hydroxybutanoic 1,4-lactone (2);. 2. furfuryl alcohol (4); 3. 3,5-dimethyl-2-hyd~y- +" 1`y'dopsnten-lone (3); 4. 2-hydmxy-3-methyl-2cydopenten-l-one (nyelolqm)(3); 5. 3,4dimethyl-2-hydroxy,2-cyclopenten-l-one i ,;.(3);C-methoxvnhenoli(gusfaool)(7);.7.3-ethyl-2-hydrmry-2tydopenten-lone(3); 8.26-dimcthylphenol(9); 9.2-methoxy-4mah- . :'ylphenot(7); 10..2-hydruxy-T-n-prop-yl-2ryclopentm-lcne(3);.11. phenol'(8); 12.2-mefhylphenol(o-ccesol).(9); 13:,2{(E)-1-pro- ;.pwyl}phenul (11); 141..4-ethyl-2-methoxyphenol (7); 15. 2<thylphenol (8); 16..2.5-0imethylphenol (8,9); 17. 2,4-dimethylphenol ', `.. (8-9)';18.4methylphenol( p-cxeeol) (8);.19.3-methylphmol (m-ercaol) (8); 20. ethyl-methylphenol (9); 21.2,3dimevhylphend (8, 9); ' Yt 2-hYdroxy-4metlwxybenraldehyde (-); 23. 2-(2-propenyt)phenol (9); 24. 2-methoxy-0{2-propcnyl)phenol'(eugenol) (8:9); 25. j..-4ethylphenol (8); 26. 3;5-dimethylphcnol (8, 9); 27. 3-ethylphenol (8); 28. 2-mcthoxy4vinylphcnol (9); 29. 3,4-d"unethylpheno] (8, 9); 1•;,30. 2.3,5-trimethylphcnol (10); 31 2-methoxy-4-((Z)-I-propenyl)phenol (isoeugenol) (9); 32..2.6-dimethoxyphenol (6); 33:2{(Z)-1- '_.;..prapenyl)ptienol (11);34. 2-methoxy,4{(E)-1-propenyl)phenol (isaeugenol) (9); 35. 4vinylphenol (10)1 36. 4-hydroxy-3:methoxyben- ' ~:I+Idttiyde(vanillih) (7); 37: 4-hydro.v-7}methonya¢tophenone (5. 6).
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Figure 1. Apparatus for collecting cigarette smoke. SmoRing macnine 7 M W E D INTRODUCTION Mainstream (MS) smoke has in the past been the main subject of the studies.on tobacco smoke, both from the viewpoint of aroma and taste of the tobacco, and also in respect of the effects of smoking on the health of smokers. The aroma or irritatioo-of the sidestream (SS) smoke differs from that of mainstream smoke. Problems of the effects of tobacco smoke on non-smokers, i.e. the effects of passive smoking and the pollution of the air im a room, havee increased interest in sidestream smoke. Many reports on the sidestream smoke have Uecn published recendy(1-S). Because of the difficulties in collecting and treatingsmoke foranallysis,r the number of the compounds qualitatively or quantitatively determined imsidestream smoke is still small compared with those in mainstream smoke. The acidic compounds in sidestrcamm smoke, in particular,, have been little studied and only acetic acid (9) and phenols such as phenol (5,. 9, 11, 14), cresols (I!), xylenols (Q1), ethylphenols (11) and catechol (l0), have been quantitatively determined. In the present studv, differences in the deliveries of acidic cumponents,, which involve organic aeid's andl phenols, to the sidestream~and mainstream smokes have beem determined.. t ff9 f . Vacuum pump F G A: Glass cnimney./or collecting sidoatreem awbB: Affi tray C: Ganrpnalge filter holder n: Alkak sphNOn tmp E: Mist trap F_ Charcoalitrap ' G: FIOr mster EXPERIMENTAL Cigarette Sample and Smoking The cigarettes used in this study were 70 mm, all Brights all Burley, all Turkish and all domestic (c.¢ Matsukawa): types: These cigarettes were conditioned at 20 °C and 60% relative humidity prior to smoking. Smokingwas accomplished with the apparatus showmin Fig, I and standard conditions consisted of a 35 ml puff of two seconds' duration each minute and 30 mm bun length.. Air was drawn through the apparatus by a vacuum pump at a flow rate of2 I/min, which hardly affected the puff number and the mainstream deliveries as.compared with those obtaineduttder normal smoking conditions. Phenols The mainstream and sidestream smokes of twocigarettes were separately directed, firstlyon to Cambridge filter pads and subsequently through scrubbing bottles containing aqueous 5% NaOH (40 ml for MS, SO ml for SS) to which I ml of aqueous o-acetylphenol solution (10 mg/900 nil) had previousl,vbeen added as;m itr tcrnal stand'ard (IS). Both the mainstream and sidestrcam smokes obtained 64
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O v e 7, Brunncmann, K. D.,. C. Stahnke, D. Hoffmann: Chemical studies on tobacco smoke, LXI- Volatile pyridines, Quantitative analysis in mainstream and sidestrcant smoke of cig;uettes and cigars; Anal. Lett. II (1978) 545-560:. S. ISrunnemann, K... D., W. Fink, F. Moser: Analysis of volatile N-nitrosamines in ntainstream and side- stream smoke from cigarettes by GLC-TEA; On- cology' 37 (9980). 217-222. 9. Johnson, W. R., D. H. Rowell, K. W. Hale, R. A. Kornfeld: Incorporation of atmospheric oxygen into contponents of cigarette smoke; Chem. Ind. (Lond.), 1975, 521-522. 10. Brunnemann, K. D., C. H.. Lee, D. Hoffmann.: Chemical studics on tobacco smoke,. XLVII. On the analysis of catechol and their reduction; Anal. Lett. 9 (1976) 939-955. 11. Cornell; A., W. Cartwright, V. Olender: Side- streamLmainstream (SS/MS) distribution ratios of steam-volatile phenols in cigarette and cigar smoke; 32nd Tobacco Chemists' Research Conference,. Montreal, Canada, 1978. 12.. Sakuma, H., N. Shimolima, S. Sugawara:. A•lethod for the rapid determination of formicandl acetic acids in cigarette smoke; Sci. Pap. Jpn. Tob. Salt Public Corp. Cent. Res. Inst. No. 119, 1977, 107- 109: 13... Sakuma, H., S. Matsushima, S- Munak.ita, S. Suga- wara: Pyrolysis of chlorogenic acid and rutin; Agric. Biol. Chem. 46 (1982). 1311-1317. 14. Neuraths G., and: H. Ehmke: Appamtur zur Un- tersuchung des. Nehenstromrauches; Beitr. Tabak- forsch. 2 (1964) 117-121. Authors' address: The Japan Tobacco and Salt PwbGc Corporatiom, Central Research Institute, 6-2 Umegaoka, Midori-ku, Yokohama, Kanagawa, 227, Japan. 71 / I E E
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N,.rc identified by comparison of theitGCretcntion wmes and ntass spectra with those of authentic com- M,unds: RI;SULTS AND DISCUSSION I;,,s Chromatograms of Aeie(ieFractiotrs.,,r AnalysisofPheno/t T71e chromawgrams of the TM5 derivatives of acidic c„tnpounds in the mainstnam, and sidestream smokes. ,;BrigNt tobacco are shown in Fig. 2 and the peaks Table 1.. Phenols and other acidic components Ident- 8ied in Figure 2. Peak No. 2 4 Canpound 2-Mothylvaleric acid Phenol 14 Lactic acid 17 Glycolic acid 28 2-Furoic acid; 30o-Cresol' 32m-Cresot 35 p-Cresol 38 46 3+lydroxyproplonic acid 2-Hydroxy-3-methyl-4H-pyren-4-one (cyclotene) 53 562,6-Xylenol 58 Benzoic acid 70 p•Vinylplienoll 75 PhenylAOatic acid 63 Catechol 94 o-Acetylphenol (internal standard) . 100 2,6-Dimethoxyphenol 101 Resorcinol 102 4-Methylcatechol 104 3-Methylealechol 105 Hydroquinone 110: 4-Vinylguaiacol 1155,6-Dihydro-3,5-dihydroxy2-methyl- 4H-pyran.4-one 119 4-E8lylcatechol, 120 Methylhydroquinone 123 3,5-Dihydroxy-2-methyl-4H-pyran-4-one (5-hydroxymaltol) 129' 4-Vinylcatechol 140 m-Hydroxybenzoic acid 142 p-Hydroxyhenzoic acid identified arc listed in Table 1. As:can be seen in Fi(;..2 and TableI, ntalor peaks in thcaeidic fractions of thsmainstreanl and sidestream smokes were phenols such as phenol,.catechol'andhydroquinone, and the peaks,uf organic acids were relatively.stnall. Gas Cbromatogramss of Organic Acids The chromatograms of the n-butyl esters of formic and acetic acids in themainstream smoke an& thee chro- matograms of the TMS derivatives of organic acids in the mainstream and sidestrcam smokes are shown in Figs. 3 and 4, respectively. The acids identified inFig. 4 arc listed in Table 2. Fig, 4 and Table 2 show that major peaks in the fractions of either smoke extracted with aqueous 2.5'Ye: sodium bicarbonate solution were acids such as lactic„2-furoic, benzoic and succinic acids and cyclic ketones with double bond(s)) and hydroxyl group(s) such as 2-hydroxy-3'-methyl-2-cyclopcnten-l- one, 5,6-dihydro-3,5-diliydroxy-2-methyl-4H-pyran-4- onc,, and 4-hydroxymaltolL Disrribntionof Phenols and Acids between the Mainstream and Sid'cst.eam Smokes Yields in the mainstream and sidestream smokes and sidestream to mainstream distribution ratios.(SS/MS) of Figure 3. Gas chromatogram.of n-butlyl asters of formic and acetic acids In mainstream smokee of Bright tobacco (temperature: 60°C). 9 w LL 10 min 67 i I 11, ru9
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6.14 W[R111S AND SMPfII ti1IMMAll1' Idnn+lor rbrvk Innu'hs wvn• n•gnl:,rly Irr:ded willc I-Irydrusyvmiwdr nnd hiqwir.c ,w•m ulkm al iutcrvnln fur binluhqy nud'd elcclnm mirn.+w.ry,v.. An'n illil.i:d crylbenmpnigmwively inrn•aurl msl by 4/1'd:wn rpilhelial byl.rzphwin nnrl Irvln•rkeralrnia wom evidcnl• :urnepumiirl Iry Ihr fSrnwlim of snm(I bulbw. •111repithclinl-aawarfivu liicwlc junclfnn wax hislohry;irall,v dinlurlad mv( IiwJ tnllv funned.Ir.w•udipa.lin which exlrndrnl Ibnelgh thrhunilw dnlnL Mm.mLu degenemliisratn unmrrntl. 911e uluulmm in Lhe.epithe(i,un nnd eudiepithclinl nnmrw9.ivu ti•auc wcm r.nnlumrnble wilh disturbances obacrved in developing experimental hamster cheek pnueh LarcimNnas. ACKNOWLEDGMENT3 We are.indebted to Dr.. P. A. Riley,. Uhiversity-0ollege Hoapitnl Medical Sehonl, limdnn. (a his adiria•.nnd fur rulpplyintCl.lu• prclumvl 4-bydrns,r,miwJe rnaun. RF:FER6NCRS Aanwoum, C. T:,,STSwemnse, V. A. and LomaL, F. J. (1961); A study o(.basement membrenn off normal epithelium,. carcinoma Lrn situ and invasive carcinoma of uterine cervix utilizing eko tnm m icnrwqry nnd hwuehemicvl mclhds. Arta Cv(ol• 8, :Ifd1-:pl•L. F+lvsuR, E. and MnlN:anaorn, K. (I9fif7. NJectmn micnxuxqlic studies nn the relation nf Iht basement.membmne tn thesquamnua.cell cnrcinomax inanimak. (Inrnlrygin 20,.11:1-12R. FaaI, J. V. (1964. •I1N: lilw structure ,d'thc lulwe-nlent mculbnuw in ipidenmd lemur.. J. CcY Biof: 15, 335-342. HINOLAISG1rILLMm, N., Moatoapo, R. and BERNHARD, W (1961). Uitrastructure des mnoen pavimenteux ihvasiG du cot uterin chez Is femme. Bufl. Assoc. F1vnc. Etude Canrer 48. 283- 31& Grmr~stirm, M. A., ALealnxr, J.T. and Golnxaasa• P. (1983). UltraatruRural alte®tiaes in hamster cheek poudl epithelium in reeponse to a®rcinogen. Arch. OrnL Biol. 8, 145-IB6. LutasL, F: J., S.u+p®ta,. E. and Asttwoxrrl, C. T. (1960). An electran microscopic study of m- cinoma in situ and invasive nreitmna dthe cervix uteri. CaneerRes. 20, 357-361. . Sea4 P., Ru.ev, P. A. and INUAN, D. R. (19fi8).Basal cell encraechment into the dermis causal by.applicatioru of hydroxyanisole. J. Inueat. Dermatol. (in press). Tnltu; D. (1967).Sequent.ial eleL•tron miinmcopiral study of experimental mouse skin carciro geneaie.lnt.J.Lancer2,195-211. W_. B WoOOs, D. A. and SMmt, C:J. (1966). Ultrastnicnue.of the dermal-epidermal',junctial in er- perimentally induced tumors and human eml lesinns. J. Invest. Derma(a!: (in prewc)..
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~ Hi•itrage zur Tabakforschmir International Volume 12No. ?' - June 1985 The Distribution of Cigarette Smoke Components between Mainstream and Sidestream Smoke I. Acidic Components * b,r H. Sakuma, M. Kusama, S.. Munakata, T. ObsumiandS. Sugawara. Central Research Institute; The Japan Tobacco and Salt Pu!>lic Corporation, yokohama,. Japan SUMMARY Yields in sidestream (SS) and mainstream (MS): smokes and sidestream to mainstream distribution ratios (SS/MS) of acidic components were examined for four types of cigarette made from Bright, Burley,. Turkish and do- mestic (c.v. Matsukawa) tobacco. Of the acidic com- ponents,, formic and acetic acids were analysed by GC following n-butyl esterification, and acids other than. formic and acetic acids and phcnols similarly after trimethylsilylation. In acidic fractions of sidestream and mainstream smokes. ~ of four types of cigarettes, major phenolic components were phenol, catecholl and hydroquinone, and major ,acids were formic, acetic, lactic,, glycolie and succinic. ; The following compoundsgave.SS/MS ratios > I for all ! types of cigarettes tested: phenol, cresols, xylenols, j guaiacol,. formic acid and acetic acid'. ZUSAMMENFASSUNG Bei Zigaretten aus vier verschiedenem Tabaktypen (Vir- ginia-Tabak, Burley-Tabak,. turkischer Tabak und diee einheimische japanische Tabakvarietan Matsukawa)i wurde die Menge der im Nebenstromrauch (SS) und im Hauptstromrauch (MS) enthaltenen sauren. Verbin- dungen bestimmt und die Nbbenstromrauch/Haupt- stromrauch-Verhaltniswerte (SS/MS)) dieser Bestandteile ermittelt. Ameisen- und Essigsaure wurden mittels Gas- chromatographie nachNeresterung mit n-Butyl und die anderen sauren Inhaltsstoff'e sowie die Phenole nach Trimethylsilylierung nachgewiesen. ° s.advn{, 9,h Jm,c tY1n. - a-.ymd: znh lanwn- iYSJ. t In den sauren, Fraktionen sowohll des Nebenstromrau- chess als auch des Hauptstromrauchess d'er vier Zigaretn tensorten fanden sich alk.phenolische Komponenten vor allem Phenol,. BrenzcateLhin und Hydrochinon und als Sauren haupuachlich Ameisensaure, Essigs5ure, Milch- saure, Glykolsaureund Bernsteiasaure. In allen vier Zigarettensorten lagen die Werte fiir die Nebenstrom- rauch/Hauputromrauch-Verteilungg d'er im folgenden genannten Verbindungen bei > 1: Phenoll Kresole, Xylcnole,. Guajakol,. Ameisensaure und Essigsaure. RtSUMp Les quantites d'e composes acides contenues.dans It flux secondaire (SS)) ct It flux principal (MS), ainsi que It rapport de repartition de ces. composants, flux secon- daire/Buxx principal (SS/MS), ont etcdeterminespour des cigarettess fabriquees a partic de quatre types de tabacc differents (Virginie, Burley,, turc en une variete dc de tabac japonais local: Matsukawa). On a mis ainsi en evidence la presence d'acides formiquee et acetique par chromatographie gazeuse apres esterification par n-butyle,, ainsi que d'autres composantsacides en phenoliques apres trimethylsilylation. Dans Ies fractions acides du~ fluxsecondaire aussi bien que.du flux principal des quatre types;dc cigarettes, om a decele des composants phenoliques, en partieulier phenol, pyrocatcchine et hydroquinone eu parmi Ies acides,.surtouuacide formique, acide acetique, lactiquc, glycolique; succinique. Pour tous Ies types de cigarettestestes, les. composes suivanus donnerent des rapportsSS/MS plus grand quc. 1~: phenol, cresols, xylehols, gaiaeol, acide formique et acide acetique. 63 '1
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817119361 Table 3. Yields and aidestream to mainstream distribution ratios (SS/MS) of phenols and acids for four varieties of tobacco. Bright Compound mainstream smoke (MS) sidestream smoke (SS) SS/MS mainstream smoke (MS) Burley sidestream smoke (SS) BS/MS mainstream smoke (MS) Turkish sidestream smoke (SS) SS/MS mainstream smoke (MS) Domestic (c.v. Malsukawa) sidestream smoke (SS) SS/MS Phenol Phenol 136• 241• 1.77 79• 69' 2.14 90' 148' 1.64 95' 169• 1.78 o-Cresol 19 24 1.26 14 18 1.29 15 17 1.13 13 14 1.08 m-Cresol 18 24 1.33 11 16 1.45 12 14 1,17 11 13 1.18 p-Cresol 37 46 1.24 32 37 1.16 33 35 1.06 30 30 1.00 Guaiacol 13 21 1.62 8 12 1.50 6 10 1.67 6 8 '1'.33 2,6-Xylenol 16 20 1.25 12 17 1.42 13 15 1.15 8 8 1.00 p-Vinylphenol 47 44 0.94 21 21 1.00 51 45 0.138 28 21 0.75 Catechol 362 • 292 0.81 148 138 0.93 251 168 0.67 258 180 0.70 4-Methylcatechel 80 55 0.69 29 25 0.86 42 29 0.69 47 29 0:62 3-Methylcalechol 62 47 0:76 31 25 0.81 39 28 0.72 36 24 0.67 Hydrequinone 300 285 0.95 133 126 0.95 132 95 0.72 114 91 0.80 4-Vinylguaiacol 36 32 0.89 23 24 1.04 29 27 0.93 29 24 0.83 4-Ethylcalechol 102 68 0.67 27 19 0:70 53 31 0.58 48 26 0.54 Methylhydroquinone 39 41 1.05 23 21 0.91 27 23 0.85 26 21 0.81 4-Vinylcatechol 113 40 0.35 23 0.30 69 23 0.33 65 19 0.29 Acid - Formic acid 478 665 1.39 210 341 1.62 311 493 1.59 297 435 1.46 Acetic acid 809 2187 2.70 333 1310 3.93 637 1927 3.03 655 1241 1.89 3-Methylvaleric acid 29 24 0.83 22 20 0.91 261 384 1.46 21 20 0.95 Lactic acid 174 123 0.71 63 45 0.71 163 106 0.65 163 83 0.51 Glycolic acid 126 77 0.61 37 35 0.95 60 52 0.87 111 76 0.68 Li3vulinic acid 45 43 0.96 29 25 0.86 513 49 0.88 40 30 0.75 2-Furoic acid 107 53 0.50 44 25 0.57 g0 60 0.67 90 55 0.61 3-Hydroxypropionic acid 39 29 0.75 2 1 0.50 31 25 0.81 22 12 0.55 Benzoic acid 28 23 0.82 14 12 0.86 21 20 0.95 21 14 0.67 Phenylacelic acitl 19 13 0.68 29 19 0.66 38 30 0.79 18 11 0.61 Succinic acid 129 65 0.50 137 66 0.48 112 69 0.62 163 70 0.43 Methylsuccinic acid 20 7 0•35 4 1 0.25 31 13 0.42 23 It 0.47 Glutaric acid 58 18 0.31 10 6 0.60 34 17 0.50 27 13 0.48 m-Hydroxybenzoic acid 42 14 0.34 8 3 0.38 21 12 0.57 64 15 0.23 - Ea[I, valuM 11 IM1Y nvn,a,)U pl eF,ao tlYlu~minal,ons 149~cInarBlla/.
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Table 1. Non-mutayenkiry of phenoNc emnpounds isoluted frvm Ihe PAH-free phenolic fraetion of smokr condensate No. of his' rcvertants/plate Concentration TA1535 TA1537 TAI538 TA98 TAIOO Compound (pg/plate) -5-9 +S-9 - S-9 + 8-9 -S-9 +S-9 -5-9 + S-9 -S-9 +S-9 Phenol 0 28 03 32 5 32 50 30 500 28 5000• 0 u-Cresol 0 22 5 14 50 16 500 30 50 0 0 ' 0 m-Cresol _ _ _ 0 12 Od 22 5 21 50 23 500 15 5000• 0 p-Cresol 0 12 0.5 22 5 24 50 21 500 21 5000• 26 2,4-Dimethylphenol 0 11 0-5 17 5 11 50 13 8&C6ii49 500 SINq• Is 0 11 6 7 12 29 II 9 9 13 16 II 7 4 14 20 0 0 0 1 II 9 5 II 8 IS 11 5 NT 9 22 4 6 17 11 , 20 2' 8 13 7 29 9• 8 16 7 20 2 0 0 4 4 22 5 11 16 36 31 to 11 16 30 31 3 NT 21 34 26 10 10 14 38 29 6 11 Is 37 0 0 0 7 20 12 12 9 8 20 12 5 5 10 18 15 7 12 6 20 22 3 5 7 20 17 6 4 6 19 0 n 0 11 0 26 35 100 84 21 33 107 90 22 36 100 90 0 14 98 30 16 14 91 76 17 16 NT 91 19 14 117 86 18 14 62 85 IS 17 53 97 3 2 0 32 21 23 67 76 20 30 65 90 18 31 68 81 20 31 67 87 18 29 64 100 29 9 3 12 16 41 89 76 21 12 106 65 21 20 100 82 20 20 102 85 17 14 N7 76 0 1 15 49 1' 2-1 Ff o0 19 20 71 67 pow
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Syringol Yanilline Eugenol 0-5 5 50 500 5000• 0 05 5 50 500 5000• 0 05 5 50 500 5000• Posilive controls Sodium aride 2 2-Nitrofluorene 3 9-Aminoacridine 60 2-Aminoanthracene 5 \/ 111 Ir. 1} 1/ .t 12 22 67 6 5 h 17 I1 • 17 la 25 19 20 71 67 16 16 15 17 14 23 21 22 65 65 17 20 17 17 13 26 19 23 63 69 /6 24 15 15 13 28 17 27 68 68 4 -~ 19 9 23 29 -- 63 24 9 5 14 5 To- 19 14- 67 76 21 16 7 12 13 12 20 34 66 I 94 26 13 7 16 9 11 18 36 71 93 22 19 4 13 10 11 23 14 65 81 15 16 4 9 14 9 22 16 67 I 95 25 9 0 . 13 13 10 16 - f S7 64 __fF--_ j2_. 12 9 16 6 9 41 67 76 9 16 6 9 16 10 8 22 66 88 18 16 3 10 19 5 8 25 67 77 3 12 12 8 6 16 9 11 24 71 c 71 10 14 8 12 16 10 13 25 65 77 0 0 0 0 9 4 4 10 61 79 22 12 16 14 17 18 27 41 67 76 13 19 22 14 16 22 18 39 66 78 14 - 11 15 7 17 22 20 IS _ 68 81 39 II 17 14 16 22 20 11 65 76 28 17 15 9 10 20 12 10 63 62 0 0 13 0 1 6 0 7 0 38 15 18 10 15 15 23 17 22 91 65 11 17 9 20 18 20 16 21 79 65 11 16 8 16 IB 26 15 28 77 66 12 20 9 17 16 21 15 24 77 71 14 18 9 16 16 22 6 20 100 57 0 0 0 0 0 0 0 0 0 n 526+54 - - - - - - 284 t 60 - 275 t 74 - - 810 f 95 - - - - 306 ± 60 - 269 3 47 - 1027 f 199 - 1093 ± 227 502t28 - 723 ± 159 NT = Nof tested 'These concentrations resulted in toxicity which was apparent as a thinning of the hackgroundlawn. All of the values for test compounds are means of four plates and those for negative controls are means of eight plates. In all cases the standard deviation uas within 5-10"-;, of Ihc mean. The positive control values are means ±SD from 7-I3 independent experiments. Vl.14 V i TzIS ! 05 5 50 500 3000' -~ .--a-.-.,r- I
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. I 1192 2.0 D.. C. TrtoMtSOH and M- A. Tausx increase in lung weight was 175 mg/kg. Above doses of 175 mg/kg, the lung/body weight ratio climbed rapidly, reaching a plateau around' 1.5% at 400 mg/kg: Higher doses of BHT did not increase this ratio. Thus, three doses at or near the threshold dose of BHT were chosen.n for this experiment:.100, 175 and 250mgJkg.Asshown in Fig. 1, subcutaneous injec- tiom.of 50 or 250 mg BHA/kg significantly enhanced the lung/body weight ratio compared to animals receiving BHT alone. BHA did not further enhance thee lung/body weight ratio above 1.5% if higher doses of BHTwere used. BHA had no effect on lung weight by itself at doses up to 500 mg/kg. The mode of BHA administration (i.e. subcuta- neous injection) was chosen to minimize any intes- tinal or hepatic metabolism of BHA prior to its reaching the lung. When BHA was given by intra- peritoneal injection, an enhancingg effect on BHT- induced lung injury was observed, but was much less dramatic (not shown). BHT has been shownn to increase the incidence of Ismg tumoursin mice given a single initiating dose of a carcinogen such as urethane,.dimethylhitrosamine: bcnzolajpyrene or 3-methyleholanthrene (Witschij 1985). A tumour-promoting effect of BHT can be elicited no matter whether BHT is administered by injection,.by gavage or in the diet. BHT can act as.a tuinour promoter in.at least two species;.the rat and the mouse, and in several organs such as the liver, bladder and gastro-intestinal tract, in addition to the Iung..Malkinson & Beers (1984) have compared,BHT and two analogues, 2-tert-butyl-4,6dimethylphenol (BDMP) and2-rerr-butyl-4methylphenol (BMP):j for their relative abilities to induce lung toxicity and promote urethane-induced tumorigenesis in mice. 10U 175 250 BHT dbse (m9/k91. Fig. 1. Effects of 0(O),50 (B)) and 2% (0) mg BHA(kg given rc to male CD-I mice on lung toxicity. (effect on lung/body weight ratio) induced by a subsequent ip injection of 100-250 mg BHT/kg. Asterisks indicate values differing significantly (•Pa0.01) from that recorded with BHT alone. mice are exposed to hyperbaric oxygen after adntinis- tration of BHT, pulmonary fibrosis ensues (Witschi, Haschek, K)einSmnto & Hakkinen, 1981). A preliminaty experiment was carried out to deter- mine a threshold BHT de¢e that would cause little or no lung toxicity (measured as an increase in lung/body weight ratio 4.daysafter treatment). Using 4-5-wk-old male CD-I mice,: the normal (untreated) lung/body weight ratio was always near 0.6%. The highest dose of BHT that caused no significant TaEle 4. Ability of BHT and two.anxloguev to indute moure lung.toxicity, tumour promotiou .nd quinona methide fomution Cnmpound BHT OH CHS _ OH BDMP CHs OH BMP Strutture HsC Lung Tumour Quinone methide mrlelty promotion' formalibn + ++ CHs BDMP-2-IOl-Buryl-kb-0imethylphenal BMP - 2-rerbButyl-4methylphcnol 'Detr from Malkin.on & Beera (19g/). 1. Their results are brief was observed to be mouse lung while BH promoter. These aut were cqnsistenl with metabolites are respo and tumour promot strain of mouse that , of BHT but, not to th a genetic deficiency.i form the promoting t Since BHT-quinom be the metabolite resl BHT; and since we hz ing the formation of B the ability of BHT,, quinone methide. Ch should be able to fort this was not the ca quinone methide mc only a small amount c did not form any dete spectral assay is an ac these compounds to findings suggest that t . may be related to a, methide. In summary,.we ha BHA can enhance the BHT to BHT-quinonc the in viua lung toxic phenolic compounds activation of BHT; ~ various mammalian tit can also catalyse the BHT. What, if any, releva in assessing the possilBHT?Humansinges (Gosselin, Smith & Hr enhance tumorigeneai around 35 mg/kgfday(Witschi, 1985). This r approximately 450-50 caused byhigh single although there are larg mice (Kawano, Nakao lung damage is observ(Marino & Mitdte6, " intake of BHT in hum: doses used to elicit tox in mice. However, as somal enzymes from BHT to a reactive prc that of tnicrosomes fr Therefore, while the c not cau]e overt tissue I. level of humann expoi miglit kadto more su Sinlilarly,..the chronii amounts of BHT-quin development of ttunor Bnisms so that the abil environmental stresse sponaes msy be partit intestine, a tissue Whic~
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l i as abovee were treated in the.following manner: aqueous ;% NaOH solution in the scrubbing bottle was trans-ferred to a 200 m[ separatingg funnel and about 250 mg of ascorbic aeidd added as antioxid.ant.. The Cambridge filter pad with, smoke condensate and 40~m1 of ether were also added to the funnel. The funnel was shaken tu slurry the filier pad and the aqueous.layer was urans- icrred to another 200 ntl separating funnel.. Approxi.- mately 50 mgpf ascorbic acid was added to the funnel and the alkali solution was washed with 40 ml of ether. The aqueous layer was then transferred to a. 200 m] beaker an& adjusted to pH 1.0 with 50 L HiSOa1 under ice cooling. The acidified solution wastransfcrrcd too a 200 mi separating funnel, saturated with NaCI, about 50 mg of ascorbic acid was . added and extracted twicowith 20 ml of ether. To the combined ether extract was added about 0.2 mli of pyridine and anhydrous mag- nesium sulfate. The dried ether solution was filtered and concentrated too about 0.5 ml at 50°C and an aliquot of the concentrate (ca: ]0 µl)) was put into a. 0.3 ml vial to be trimethylsilylated with 50 Id of tri, methylsilyl (TMS)) reagent, BSTFA", at 100 °C for 10min..Theresulting TMS derivatives of acidic compounds weree analyzed by gas chromatography (GC) on a 30 m X 0!28 mm inside diameter glass capillary column coated with OV-101. A Shimadzu GC-7A gas chro- matograph with an FID was employedund'er the fol- lowing conditions: temperature program, 50~to 200'C at 2"/min; injector temperature, 250 °C; helium flow, 1.l ml/min through thee cohamm with a split ratio of 1:50. Organic Aadsformicand Acetic Acids: The mainstream and sidestream smokes of three cigarettes were trapped separately by Cambridge filter pads and scrubbing bottles containing aqueous 0!25 % NaHCO3 solution (40 ml'for MS, 80 ml for SS), to whic6 I ml of aqueous n-valeric acid solution (200mg/10 ml) had previously been added as internal standard. The aqueous 0.25 9'a NaHCOs solutiom in the scrubbing bottle and the Cambridge filter pad with smoke.condensate were transferred to a 300 mi sep- arating funnel. Twenty ml of ether were added to the funnel and the funnel was vigorously shaken to slurry the Cambridge filter pad. After phase separation, the aqueous layer was washed with a further 20 ml of ether and evaporated to dryness in a 30 ml pear-shape flask at below 50 °C under reduced pressure. The following, procedure was carried out in accordance with the n- butyl esterification method (12): the dried residue wass esterified wit6 2 ml of n-butanol,. 0.25 ml of sulfurirc acid'and 2~ g of anhydrous sodium sulfate at 100 °C for15 min under an air condenser. After being cooled, the reaction mixture was: shaken with I ml of n-pentane and 5 ml of distilled water, and the. organirc layer was pipetted into a small!test tube...A small amount (1-2ml) of water was added to the test tube and the test tube ''•~',D-lhc(,dmeehyt~ilylpnnuomunamidc ss•as shaken vigorouslv. One Id of the organic lai cr vas injcctcd into thc gas chnomato;;raph. Gas cliromata, graphic conditions were as follows: apparatus, Shinmdzu GC-7A equippedl w.ith an FID; column, a. 50tn k. 0:25 mm glass capillary column coated rvith PEG 1500; column tentperamre,. 50 `C(isothermal); injector tcm- purature, 120 °C: nitrogen flow, 0:8 ml /min through the column with a split ratio of 60:1. Calibrationcun=es for formic and acetic acid Nscrc• ob- tained bv the follbwing procedure: to the mainstreant or sidestreamtsmoke of I cigarette, obtained in the samc way as described above, known quantities of sodium formate and acetate were added and the mii:turr was esterified as above. The peak height for the added formate or acetate was obtained by dl•ductingeach peak height for formic or acetic acid contained in the main- stream or sidestreamn smokes from the peak heightt for formic or acetic acid of thc mixture. The ratios of the peak heights (added formate or acetate./Snternal stan- dard) wereplotted against' the added amount of formic or acetic acid. Organic Acids other than Formic and'Ace[ic Acids: Thee mainstreamm and sidestream smokes of two cigaretrtes, were trapped separatelly by Cambridge filter pads and aqueous.2.5 % NaHCOs solutions containing 0.2 mg of m-toluic acid as an internal standard in scrubbing bottles (40 ml for MS, 80 ml for SS). The mainstream and side-stream smokes obtained as above were treated inthe same manner, as follows: The aqueous 2.5 °lo NaHCOs solution in the scrubbing bottle and the Cambridge filter pad with~ smoke con- densate were transferred to.a 300 mi separating funnel. Twenty ml of ether were added to the funnel and the funnel was vigorously shaken to slurry the Cambridgee filter pad. After phasee separation, the aqueous layen wass washed with an additional:20 ml of ether, transferred to a 100 ml short neck Kjeldahl type flask and evaporated almosn to dryness at below 50 °C under reduced press- ure.. The dried residue was acidified to pH 1.0 with 50% Ht.SOa under ice cooling and then continuously extracted with ether for 3 hoursusinga 30 ml'receptor containing about 0.5 ml of pynidine. The ether extracn was treated with anhydrous magnesium sulfate, filkered and concentrated to approximatel,v0.2ml at below- 50 °C under atmospheric pressure. A pan of the con- centrate (about 10 lll) was placed into a 0.3 ml vial. After addition of: 50 µli of BSTFA, the vial was heated at 100 °C for 10min before analysis. Gas chromatographic eonditionss were as follows: ap- paratus,. Shimadzu GC-7A equipped:h with an FID; col- umn, a 30 m~ x 0.28 mm glass capillary column coated with OV-101; column temperature, 5VC - 200 "C (2°/min); injector temperature, 250 °C;, helium flow, 1.0 ml/min through the column with a split ratioo of50: 1. Identification of Phenols and Acids Phenols, organic acids and other compounds in the acidic fractiom of the mainstream or sidestream smokes 65
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'o.. IOf•- (Alviu the ta Dieler) C{iamo nat vrere tine and ,us and samplt Penirk tLs were n to use. were ad• njeotiua. :e agents g taku- the oils ;anut dF ril alone, specificd :d under ng exlii- I'. to con- in indivi- Suppk- : surgny es, raleN ays after r on day wed and te organs The liver tted, was LNng the xof646 autopsy ore than ,ht were im orin- liver in- jected to 10 days. tl als aaf 3•14•• 2.34. 2-74• 329'• t•19 1-69 1 74 1 44 Liver regeneratiommnd essential oils175 Table I.-tmuiitued Oil or other teal material d d l~ N Body weight(g). an ose (mgRat/day; ovaall)) o., of mts fnitialj TerminaljLiver increment (g) kries 6E-conrinued - Panley-seed (100;283D) 5 242 ± I9-0 251 ± 17fi b746 ± 0-243 3-60" Okoresin cdery (100; 29401' 9 225 t.98 251 ± 11-3 2.402 t0107 3-23" Snics 7E ' Control 12 244 ± 39 265 f 53 3171 ± 0-093 Anise (100;.2735) - 11 256 ± 43 276 t 6-0 2593 ± PI12 1r91•• Cumin (100; 2825) 10 242 ± 45 253 f 66 2490 ± OfyM 2,38• Eugmol (50; 1765) 11 249 ± 4-5 262 t 5,5 1997 ± 0066 150 44Allylanisole (50;.1335) 12 252 ± 4.1 271 t 64 2592 ± 0-143 2d6• Saies 8E Control 12 289 t 15-6 296 t 15•2 2267 f 0126 Tarmgon (100; 240D) 5 288 t 139 296 t 7-7 2-938 ± 0241 2-71' Guaiac wood (100; 2475) 12 288 ± 144 277 f 144 2268 t 6114 001 Suies 9E Control 15 344 t72 339 t 7-7 2-148 t 1M97 Nutmeg (I00; 2065) 13 332 £ 7-0 346 t 5-7 2,623 ±(H197 3,42'• Sniu 10E-kmalwControl 10 203 3 26 213 t 43 1•607 ± 0073 Tarragon (100; 3450) 7 202t 29 205 t 8-3 2097 ± 0158 - 3-10" 4•Allyknisok (50; 1675), 12 203 ± 2.4, 215 t 50 1.914 ± 0094 250• Grrot-seed (75; 2480) 10 208 ± 2 5 1221 t 2 7 1833 ±0144 1-40 Parsleyaeed (50; 1640) 8 209 ± 2-4 216 t 30 2•130±& 103 4-25" Saies HE Cont7ol 12 302t 123 299 t 13-7 6773 t 0 115 Sassafras (50; 1130) Camphor sassafrassy. (50; 1240) 6 6 316 f 23,4 282~ t 26-6 313 t 26-1 283 # 30-3 3095 t 0164 2-187 t 0-159 6b1•• 2U9 Snies 12E Contml 10 264 t 108 292 t 7,4 2-127 t 0095 tPropenylanisole (100; 2450) 12 269 ± 8.9 295 t 7~6 2-613 ± U08K 3-77•' Isoeugenol (100..2640). Isossfrok (15; 375) 8 11 2523 7.3 272t 10-3 276 t 7•1 290 t 94 2.248 ±P120 2423 3 0087 PB0 2-31 • ieier13E Control 13 213 ± 2.5. 264 t 3+9 2.206 t 0-115 Ssafrok (10; 295) 13 2l0 ± 3.4 266 t 66 2-565 t 0-078 2-58• uirs 14E Conlrol 13 283 t 88' 292 t 96 1-891 3 0'108 Dihydrosafrole (IS; 360) 12 282~ t 7-1 299 t 11-7 2-3% t o-155 2-70• in es ISE Coatrol 12 285 t 64 313 ~' 6-6 2168 t D105 3-Allylanisoie (50; 1160) . 15 285 ± 5.8 311 31 5-3 2-585 t 0088 307•• .Propylanisole(ISQ 3440). 13 280 ± 6'1 323 ~' 6r5 2-555 t 0-140 219• rPropylbenzene (200; 4670) I I 288 ± 57 310 31 SI 2-481 ±.tM88 b27• Tarragon (50; 1760) 12 287± 50 314 ~' 6-0 2-543 ± 0-112 *45• inirs 16E Control 1I 249 ± 4-5 271 ~' 88 1985 f 0098 AByI phenyl etMx (50; 1320) 12 256 f 43 275 ~' 46 2-487 3 0106 3-46•• Pipetine (25; 660) 10 250 144 277 ~' 91 2481 t 0071 403•• aia 17E Control I I 301 t 64 355 ~ 90 2.503 ± D091 AGyl6cnunc (100; 2330). 12 289 t 9-8 319 ± 8A 2893 t 0087 322° fMethylstyrene (100; 2330) 14 291 f 56 320 $69 2-665 3 0,126 1.05 AByl phenyl ether (100; 2260) IS 302 3 80' 321 ± 65 3-273 ± 01 18 547•' itnes ISE Conuol 16 282t 72 291 ± 89 1-892 ± 0088 rltopropylBenrrldehydc (50;.1250): 15 282t 79 279 ± 66 2-278 ± OU74. 336•• 3-ppMethoxyphenylpropunol. 1150';3750) II 282± 85 278 t'84 2-076 ± 0082 P46 itries 19E Comml 11 324 f 8-3 335 ± 71 2-256±DI1S Guaiene (100; 2135) 13 325 f 7-7 331 ± 7-5 2-402 ± u1J0 1-40 Guaiazulene (50; 1065) 12 322t 7.3 336 ± 80 2697 ± tH362 345•• Yerivert, Haiti (150; 3330) 13 320 t 7-1 310 tB,6 2-383 ± 0-121 D75 fepverol 000; 21701 . 12 318.3 76 327 ± 68 2d87 ± fH 12 206 CM1amomik,.H-1(50;IIU0) 12 32138'8313t78 2548±UI18 Ih77 tyected on daysl-7 after surgcryin a daily volume of peanut oil ranging up to O50ml,.the eonlrols rcceiving ponut oil alone. Except for series 10E, males wcre used'throughoul lvuall.weights were recorded prior to surgery and terminaliweights at autopsy 10 days later. fsulls are means ± SEM for the numbers of rats stated. Significant t values are marked with asterisks:. •P <UUS; ••P < 00.1.
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fd C.tem. Toxir. Vul!.24. No. 1011. pp, 1189-1195. 1986 027a-6915186 f3.00+0.00 pnntcd in Greal Britain [hr8amonJournak Lid THE TOXICOLOGICAL IMPLICATIONS OF THE INTERACTION OF BUTYLATED HYDROXYTOLUENE WITH OTHER ANTIOXIDANTS AND PHENOLIC CHEMICALS D. C. TttoatPSoN and M. A.. TxusB Department of Environmental Health, Sciences, Division of Experimental Pathology and Toxicology, The Johns Hopkins University, Schooll of Hygiene and Public Health, Baltimore. MD 21205, USA Abstract-Butyl%ed hydroxyanisale (BHA) enhanced both the. !n vitro o' m Imt binduutnf butvlated hydroxytoluenelBHTJ to m:+ aom-L---te.n an t ormation of BHT-auinone methide. Eugeno , met y para ivanillin. gyaia~P ll ferulic acid and several other phenolic compounds commonly used in food and tosmetic producls also enhanced the metabolic activation of BHT. BHA was the most d&ctive compouruftested: Microaomes from lung; bladder,.kidney medulla and small intestine of various animal species, including man, were also able to support this interaction of BHA andBHT using either hydrogen peroxide or arachidonic acid aa the substrate.. These !n vitro observations were extended to an in ohn moure lung model. Subcutaneous injections of BHA significandy enhanesdthed lung/bady weight ratio ofmice given intraperitoneal injections of subthreshold doses of BHT. The toxicological implications of the interactions of BHT with other antioxidants and phenolic chemlealsand their potential relevance to human risk are discussed: Like many xenobiotic compounds, the toxic eBects of . BHT are thought to be caused by metabolites rather than the parent compound. From a toxicological standpoint, one of the most interesting metabolites of BHT is BHT-quinone methide (2,6-di-tert-butyl- 4-methyleno-Y,S-cyclohexadienone).. BHT-quihone methide is a relatively recently described hepatic metabolite of BHT in mts (Tajima,. Yamamoto & Mixutani, 1981; Takahashi' & Hiraga„ 1979). The formation of BHT-quinone: methidc has also been . observed in vitro in mouse liver microsomes and in vfoo in mouse liver and lung (Mizutani, Yamamoto & . Tajima„ 1983). BHT-quinone methide has been suggested to be the ultimate toxic metabolite of BHT responsibhfor causing lung damage in mice (Mizutani, Isbida, Yamamoto & Tajima, 1982; Miiumni et aL 1983) It has also been shown to interfere with vitamin K-dependent protein carboxyl- ation ation (Takahashi & Hiraga, 1981) and thus may play ;~ a role in BHT-induced haemorrhagic death in rats (Takahashi & Hiraga, 1978). - BHT-quinone methide is a reactive compound ` capable of reactin:g with various cellular nuclco- phiks, including glutathione (Tajima, Yamamoto & Mizumni, 1983 & 1985). Administration of high doses of BHT has been shown to depleteJevelsof glutathione in rat liver and mouse lur,g, (Mizutani; Nomura, Yamamoto & Tajima,. 1984; Nakagawa, - Tayama, Nakao & Hihaga, 1984). Depletionn of glu- e tathione levels with diethyl maleate or buthionine sulphoximine enhanced the toxicity of BHT to mouse lung, whcreas dietary administration of cysteiae A3brrviarions: BDMP=2-rrro-butyl-4.r+dimetAylphenol; BHA = butylated hydroxyanisole; BHT-butylatcdhy- droxytoluene: BMP =2-rerr-bu1y144-methylphenol. protected mice from BHT-iaduced lung toxicity (Mizutani et a!. 1984; Nakagawa,. Sup & Hiraga, 1984). Because of the high reactivity of quinone methides. (Turner, 1964),.metabolicreactions that result in the formation of these compounds in mammaliansys- tems are of signifiwnt interest. BHT-quinone methide is thought to arise fronu the biotransformation of BHT byy a cytochrome P-050-relatedenryme (Kehrer & Witschi, 1981; Tajimaet af. 1985). Wee have recently demonstrated that peroxidase enzymes cancatalysc the in vitro formation of BHT-quinonee me- thide (Thompson, Cha & Trush, 1986). This report describes the in vitro formation of BHT-quinonc methide by peroxidase enzymes, the inAuence of various phenolic compounds on this reaction, and the possible invivo.significance of theseperoxidasereactions:. . Exper®ental Materials. (RingU-1eC) BHT (20mCi/mmol) was purchased from Amersham Corp. (Arlington Heights, IL). Arachidonic acid was obtained from Nu Chek Prep (Elysian, MN). BHT, BHA and other test compounds were obtained from Sigma Chemical Co. (St Louis, MO): or from Aldrich ChemicaU Co. (Milwaukee, WI). Horseradish peroxidase (type 11) was obtained from Sigma and prostaglandin H synthase was prepared from ram seminal vesicles obtained from Dr L. Marnett, Wayne State Univer- sity: Microsomes from ramseminal vesicles and other tissues were prepared as 25% homogenates in 0.15 bt-KCI adjusted to pH 7.8 with dibasic potas- sium phosphate. The homogenates were centrifuged at 9000 g for 20 min, filtered; through cheesecloth and then centrifuged at 100,000 g for I hr. The final micro- somal pellets were resuspended in 0:15 st-buffer at
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Liver regeneration and cssential',oils 177 from ca» dubomiletea(Id). !ion, r4u / 7es[ malttial BodyweigAt (g) kg a:.~ ' rnddidary levdt No.. tQO10p ' trSr ol rarss rmtwlF /ermmalj Liver increment Igl r c cinme. mioLh.-i~ }rmat ._ ' rrMo.L v^' 1 tulene ar ~~ limelhu} C.~ae (040) mg k; ~ Sos "!E 5,InelyaaT :~35E eq 4c1~4 4uvo1 Ouoomile oi4 Engluh.(035) .nZ){ jjW Ouitomik oil H-4 (0-35) t.erh~ 0 aamomuCUmtrl-Jtvtu) •. ollf r•R •~•• I~ r~ [b-ilrole (025) _. Ln _8E 's sP°~' Caa6d s. eannp.. Yanen oiI•.Bourbon (035) vdac~naL h.u 29E ~ the ~(omd ~hti pp . fanaq oil• Haili (050). smlmn NaV) nn.n- ~M i 11°c rv+vr •~aa.i 1~ry'.VI1111Fr1(tlpKRyYC IVJJI. 4rka PriK fruol • POMM fioluulene (020) sge. t~r'' iba 3?E~emakc mC waQ C<btrd ;mimak a• J>~e 1025) ;= yd ~=!e3_E-females indiomill. f wnw rtsidaa~'F ~: • wcvr `eR mp~ 4uuM ~ ~ Y 3.meg oil (0601. rdb•P•~yrauil(U60) rlteu~4tae/. f6m-snd oil (060) CYoresin tumeric (060). ru-mr nmlvwl `n-se.ed oil (060) 39E fud I u +Enn oB (050) ,4iEund .Imi, j,;amegdccomntedand Fomd (61)) s ou r/iFIDQS1E. (aod Laa allspice. ground; (6-0) uE ftrol r ] iaraom•ground 150). fabsxd.ground(601'. s6E-lemales Satol it6ole10251 fumosafmle.(0251. Tablb 2.,'ominurd ___ _- . ...._. ... • ~ 1 v ~ .rvc .i .. . ...rv~ 1 ..•. . : ~ 14 11 . 14 12 II u 332't 7-8 320t I1-1 239 7•5 t 235 ± 7•5 235 ± 74 JAa?trr 349 t 7-4 332 3 11-2 278 3 7-8 263 t 9'9 276 t 8-3 trt2trn x.vJ 2540 t 0115 2958 ± 01113 253• 2'034 ± 0074 2•234 ± OI 12 1.52 2•431 ± DII? 7t)3•• lvFJ=vinn Jvb^ n. cvo. S _I 9 254 t 107 256 t 120 2,636 ~, U 119 4•49•• 17 230 t 11.3 259 t 108 2135 * 0'066 14 243 f 142 257 t 134 2•166 ±0073032 16 250 t 3}P 287 t 45 2.178 } 0(170. 12 258 3 38 297 ± 47 2-715 t 0077 5-J I'• 1] 256 f 5'8 291 ± IUI 3775 ± UI_'6. 4O9'• 11 aJ•r 2 sv nr S b-r = ru * vru> nov__ rt tJJ S J'O cbb $ J'b [•J[b Y wn J'N 12 245. ± 5,4'. 264 t 54 2-005 ± 0.118 Ill 243 ± 7J 251 t'TI 2-814 t O18fi }80`• 16 182f 2•9' 192 t 3-8 U071 1630 t 12 182t3J 186±50 . 1.911±(t10214+• 9 172t 1,0 ' 183 ± 4-9 2218 ± 0068 544° U IYt 3=! L.!3 t SV !w! i UU63 ~J 1>J S J'b uY 2~> - Low S vVY~ J•]o~_ 16 245± 24 260 t 31 1'897 ± 0062 I I 243 t 29 260 f 5.5 2•438 ± 0 140 39?'• 12 242 ± 73 255 t 9.8 2iI92 ± 0081 8 247±'44265tI1•9 b170±U224 038 8 245 t 119 260t 9-2 2•420 ± 0-093 '_62• 8 245 ± 101 258 ± 12-0 2083 t 0143006 rJ 1JV I rvu JJ> = rv rant Y vrma'. rcn I I 335 t 149 307 ± 15-6 3-212 ± 0166 5•92•• 15 272 t 5-0 298 t 63 2.294 t 0056. 12 271 t 5'5 282 t 7,4 2240 t 0tr114 036 I I 264 t 7.6 270 t 66 1-783 ± OU76. . 13 274 :1, 8-6 279 ± 7r0 3•555 ± 01679.09a. 13 275 t 9-9 281 ± 84 2083 t 0611S 13 275 t 13'8 273 ± 122 2037 ± OI 17028 14. ' 276 t 11.8 284 f 12-4 1911) ± (Y087 10. 279 ± 14-4 289 ± 12.6 2035 t 022 086 9 290t15-5 283±12fi 2011±0110 051 - 13 223 ± 90 235 t 79 1998 ±(IUM[ 15 218±80 213 8 1 'I 2945±UUJ3 ?67• 13 222 ± 9-3 233 ± 99 2655 ± 0123 429•• Sns wbreAxPmf f76. for 10 days folbwing surgtty. Males were used in each scries except xiurt indicated 21T +b.ise /serits 32E:.33E..35Eand 46E). .eip)tts were ranrd~d prior to surgery and'tcrminal wcights at auropsyron day. 10. rre means ~ SEM for the numbers of rats nated Signifiunr I values are murkedwith asterisks: •Y < 005; 31P' ~ 7<OUI.
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CHROMOSOME FRMiHHRTXPION Pi~. . .{1liurn Cr/.n• Peui;;cn cmr•nrsnl rnul allrunrosoutr,. i~t rL r> rlinurrf~. niol. .faiucA iU ap'dal ciulrlr. ~- n--t, p:.:vrapllexr. f, lr: Irrl.• aunlthasr. ir.lobrplr.ur•, rt. rl. r.-.rr,r.rxtrr.mr r.h:rnt;>s. m:rnv nunanic.liurr•.auJ I:~sgPtrrrl.-_ 1. _Nvn em:dl Ir:r;;nu~uln; In~r uurl nne u/qn'ItrJl lrs.nunl.: /: :Y. rraptncril.: r{: Irnn.?or:dir~n Lridge• rrv dlr:rti.r n( .ia.rr rruui..u; le: Iragmrnl.: /. It.rr ndrranurlbl ;nd ou...l:ugo. Irapnrrnt -- .7111i11.
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in the 'dbefg,a acalyp r er the Effccl of essential oils nn.drug mcl:dadi.nl I TABLE 3. No. rats Treatment Interwal between treatment and pentobarbital (hr) Sleeping rats (%) Sleeping line (min } S.E,) controls 80 43•3 .!, 3 Eucalyptol 7 100 368-t:I, „ 18 667 31,6 } St „ 36 60 30-3 i 3' 48 60 44•0 .L P Pentobarbital was given at the dose of 30 mg/kg i.p, a0er euculyptol (500 mg/kg s:c.) 'P <0-01 vs. controls }P <0-05 TABt E 4. 20N3 No. rats Treatment Inlerval between troatmentand pentobarbital (hr) Slecping lime (min ;- S.E.) Pentobarbital concentration (g/brain) 20 controls - 50 ;~ 3 16 a- 0~6 8 Eucalyptol I8 21. ::6" 11.9_=1•3' 6 36 29 ~'- 6' 130 i 0-7' 9 Oil of Pinx.r prmdfio IN Sfi !~ 6 1T5 +- 1.9 5 ., „ „ ,,. 36 66. ~: 6 17,7 -I- 0~9 Drugs were giJensubcuraneously (500 mglkg) beforc pentobarbital. &ain concentration was measured 90 min a0er penrobarbital (30 mg/kg i.p.) injeetion, 'P <001 vsaontrols. TABLE 5. No. rats Treatment Aerosol. min of admin. in 4 days Interval between treatment and pentobarbitaladmin. (hr) Sleeping time (min -;- S.E.) 25 controls 62 2 8 Eucalyptol '90 18 §24'. E._ 6 9 „ 90 72 57-F- 8 S T3o 18 §33 - 3 9 controls 52 5 9 Oil of Pinnr pmnBio'90 18 . 48 ~+ 4 8 controls 42 ~ 8 5 o-pinene ;6U 24 33 -I- 4 5 $-pinene ;60 - 24 35 ? 5 Pentobarbital levels in bmin (nglg + S.E.) 165 ~-07._- §7-2 -i: 1'5 171 1-2 §12-4 i 0-7 15.3 -i-1 14.4 -I- 1 12~7 I09 11.1 !- I-5 10-4 } 0-3 Rats were submitted to aerosol daily for 4 days during pcritds.nf • IS, 15. 30..30 min respectively t 5. 5,.10. 10 min respectlvely ; 15. 15, 15, IS min respeclively §P<0-001 in respeot to the controls Pentoliarbital (30 mg/kg i.p.) was given at tlte reporterf huurs afrer the liut acrosol and was mcasured in brain 90 min after the injection.
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1. Frrr,rlmrn6dirw pererfdngr in diffrrrnl roolx of /nur lrenhmml. Ini1L O,noo: molJf fronrenfrn- 11 13 12 16 17 5 63 1 ?ri ? 31 liurl N~ pi1rogrd(o(. SumLrr af cv ll+ wilh Ilngm enlx \lenn nuluLer 1 :4 1 s r nf Irngn.ents IrM cnlI 4 6 II 4 4 -_ 1} 8 7 1 3 I 1 I,5 A 11. f 1 6 4 2,s /: 2 1 3 :4 I,a 4 . ~ 1 3 1 1 2,s :51/ 29 16 12 15 9 I,s I:1 9 3 1 1:1 11 ! 2a 11) 7 r 24 211 / 5 N :III 14 9 711 Y /i 'fnlul aJ:~ lUl QS 6.f Il,x 6.a Erngmrn- lalinn Tolol cells unallxed 61,x :47 69,s 33 66.11 47 54.1 37 75,n 20 64,s 174 66,0 -. 6s 31.1 45 00,a :13 i3s 36 n1,r 52 36;e 7N 31s 43 lu,l 140 24 v 0:1 . _-__. . ____. ... ....:._. _ lA,r 515 :Ha 66 IUS I, aa,a IiA 30" 23 :ts,a GO 0, 23,5 34 0y 31a 43 _ ~-~ --~~-- 0 •s 30,2 341 O,v 16,a 60 0,9 1 ],s 63 0,4 20,s 34 0•t I,s Y1 0•x 19,6 56 - 0.1 364 U,n 20,. 303
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388 IA L. PooL and P. Z Lru Table 2. Mutayenirity, testiny of phenol using SaI monell a.typhimurium TA9g8 and S-9 from indured andmtinduced Spragut•Dawley rats , Concentration No: of his" revertants/plate (pmol/platel. (pg/pdate) With S-9Trom Aroclor induced rats With S-9 from untreated mts 0 0 22 31 10 940 19 31 20 1880 28 35 30 2820. 33 26 40 3760. 30 29 So 4700. 30 33 60 5640' 32 IB 70 6580 28 23 80 7520. 22 20 90 8460 16 16 100 9400. 17 18 •This concentration resulted in toxicity which was apparent as a ttlinning.of the back- ground lawn. Values are means of four plates except for the control valuesi which are means of eight plates. In al0casesstandardAeviation was within 5-t0°o of the mean. Table 3. Muragevicity^of pherrol towards Salmonellatyphimurium TA9N in rhr presencr nj caryingg amounts of Arocfor-inducedS-9 mi.c. Preirrvufwtian was rarried'out prior to. platiny (aee'experfnrenraf), Phenol concentration No. of his' revertants, plate S-9 protein pmol/plate) (p8/plate). (mg/10m15-9 miz)..- 20 60' 100 0 0 36 32 31 41 10 940 39 39 34 30 20 1880 35 43 37. 19 30' 2820 0 0, 34 0 40 3760 0 0 0 0 'This.concentration resulted in toxicityy whicliwas apparent as a thinning.of the back- groumflawn. Valuesare means of three plates except for the control values which are means of eight platcs. In all cases the standard deviation was within 5-104; of the mean. Table 4. Efrurr.f norharman (-'Mlyty/plme) on the muroyenirif)"q( phenof in Salmonella typhimurium, TA9N'8 after preincuhation u•iNi S-99 mic, tonrainirryp diQereur prora•iir ['unecnllanrms No- of his"' revertants, plate S-9 protein (mg/IOmIS-9 mix)..,.. 20 40 60 Concentration ' (pg/plate) Norharman - + - + - + 0 '_0 20 21 21' '-0 20 m o-5 17 16 . 16 18 ?-1 1g 5 ~ 16 18 19 10 22 19 N 50 ' 17 ' IS 20 19. 23 17 N 500 . 15 15 20 17 I% 17 5W0+ 15 9 17 10 15 ~ % 'Thisconcentration resulled In toxicity which was apparent ac a thin ning o f the ~ background Ihwo-. Values are means ofthrec.platesf rfta ta.1,
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2082 A. lonr. A. Ilunuurrn and1'. Li. NuM nen p-aminophenol, 4-amihoantipyrine and p-nitrophenol rt.spectively, formed inthe incubation medium after 30 min, were analysed according to Gilbert and Goldberg.$ The liver weight and the liver proteins were not affected by the treatment with eucalyp• tol aerosol. . RESULTS Table 1 shows the results obtained with pcntobarbital,. 18 and 36 hr after the drug administration: In the case of eucalyptol, there is.a signi6cant.decrease in pentobarbital effect. The sleeping time of l8-hr-pretreated rats is approximately 50 per cent Icss than the control group. This c6ect is dose-dependent (Table 2), develops several hours al)er administration of eucalyptol and is long.laaing (Table 3). TAUt.e 1. No. Treatment Imerval between mts ucalment and pentobarbilal (nr) Sleeping rms Slceping (%)~~ time (min :~i $.E.). 41 . coNrolc 805 36.8 +. 2 30 EucalYprol I8 66.7 '196. L.1 36 t00 '22-2 yl ataco TS ttA)-"----_-34'U .!: 7 6 36 66 42.0 -'v t ---~-- Mrn o IT 100 ~9~~"'=f`3 6 36 83 400 _ 2 6. Oil of Pimu prunil7n. Is 100 42-0 -~4 6 .. 36 100 38-0 _ 3. 6 6 .-pinenc Is 36 100 g3 36~8 _ 3-3 324 !- 4-9 6 S-pinene IS' 83 34•3 ± 4 6 „ 36 100 31-9 L 08 Drugs were given s:c: i300 mg/kg) before pentobarhital I25mglkg i.p.). • P c0.01 vs. aontrols: TAOLC 2. No-. rats Treatment mg/kgs.c. Skepingrals (%) Sleeping time (min-S.E.I 41 controls 80 36~8 '- ? 10 EttcatyptWl l25 30. 3).7. : J. 10 ,. 250 40 •26-0 :- 1 30 ,. 50o 66-7 •19-6 ~, 1 Pentobarbital was given at the dose of 25 mg/kg i!p. 18 hr after Fucalyptnl. • P !001 vs. controls. Table 4 reports that the levels of pentobarbital in eucalyptol treated rats showing a decreased sleeping time, is significantly lower than in control rats, when it ismeasured in brain 90 min after pentobarbital adminis(ration. The results reported in Table 5 show that the inhalation of aerosolized eucalyptol can reduce brain level and the narcotic effect of iqjectrd pcntobarbiLtl in rats:.Agreeing with the data obtained by subcutaneous (s.e.)) administrati.An. the effect induced by the ataroh als.• Sigtprescs. '2 hr a('irr tha l::;t inha!ar.l*n. ~ No. rats 20 8 6 9 5 Drugs Brain cc •p <00 No. rats 25 8 9 5 9 9 g 5 5 co Et rA, Oi cor p 2-F Ra:> •:er: 5 • 15, 15. 30, 3 0i t5 5 10, 10 1 ~ p5t0 001 In ... PentobarbtaVt in brain 90 mi- Ell
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, ~• Eryect of Tobacco Lea/ on Smoke Catechol Form®tbn J. Agrb. Food Chem., Vol. 30, No. 2, 1982 373 Teble LPyrolyzate Ostechol Yields and Leaf Polyphenol Contents in Bright and Burley Tobacco Varieties tromato- re pyro- :entative orted tn tg lignin ,6)., were ne their varietir< tcco Re- ce plot.,. t of uni- vred a> ured, aJ; Here rr- rial wa, vrolysi-. noiatun g a trac or 48 li. pass „ r high of leat cid, mll ntly the temical, ellulosa (Grad, ical C '. ..G. A- d lignii: erimeotien•ir( . >'usini.i7).andl 79) prr, ctcd ir. istille'i- ,oncen ')to:,•r) ntmt.-: i. AI: with .: but.i catechol %tot'al polyphenolqe chlorogenie variety" type , mg/g pyr colorimetricbHPLC° acid isomers, HPLC° White Mammoth Bright 1.L7 3.89 47 2. 1 66 • Speight4G-10 Bright 1.04 3.93 2.05 1.46 North Carolina 95 Bright 1.03' 4.26 283 209 Canadel BrighL 1.02 3.81 2.66 1.78 North Carolina 2326 Bright 1.01 4.17 2.41 1.68 Virginia 115 Bright 1.01 4.01 247 1.72 Speight0•28. Bright 0.98 3.68 2.79 1.83 401 Cherry Red Free. Bright 0.91 4.17 2.28 1.60 Coker 319 Bright 0.88 3.72 2.46 1.60 Kentucky 34 Burley 0.82' 261 1.47 0.97 Gr 46 Burley 0.68 1.88 0.97 0.48 Warner Burley 0.60 1.22 0.48 0.20 Kentucky 67 Burley 0.53 1.61 0.59 0.23 H 47 Burley 0.46 1.74 0.89 0.38 average for Bright 1.00 8.96 2.49 1.71 average for Burley 0.62 1.81 0.81 0.46 a All values obtained for mean of three replicate plot samplea b Determined by colorimetric method of Williamson '(1975). ° Determinedby high-pressure liquid chromatography (HPLC) method of Snook and Chortyk (1981). HPLC method more specific for polyphenolic components than traditional colorimetrie methoda 3. ~~ ' 1>•ble II. Major Pyrolysis Products of Selected Tobacco Leaf Components compd pyrolyzedAfr chlorogenic acid 354 quercetin(dihydrate) 338 rutin (trihydrate) 601 caffeic acid 180 lignih undefined cellulose 80-600 000 sucrose 342 fructose 180 yield (mg/g of compd mol % product pyrolyzed) converted catechol43,313.9 4-ethylcatechol 40.2 10.3 phenol 13.9 5.2 6(hydroxymethyl)furfural 13.2 3.7 catechol 19.6 6.0 4-methylcatechol 4.6 1.2 catechol 7.8 4.3 4-methylcatechol 5.5 2.7 4cthylcatechol 4.6 2.0 4-propylcatechol 1.2 0.5 catechol 3.6 0.6 c hol 9.5 a guaiaco 5.2 a methylcatechol 4.3 a phenol 23 a isoeugenol 1.1 a furfural 5.4 a levoglucosan. 2.9 a furfural 27:7 9.8 6{hydreaymethyl)furfural 19.5 5.8furfural 38.6 6.9 5{liydroxymethyl)furfural 19.5 3.5 , in a reaction vial. Excess BS'1`FA reagent[N,0- (trimethyleilyUtrifluoroacetamide] (Pierce Chemical ) was added, and the vial waeaealed and placed in a ting block at 75 °Cfor 30 min: The silylated eamplea directly analyzed by GC-2 on a Hewlett-Packard 20A gas chromatograph equipped with flame ionivation and converted to utilizea 25-m S1i54 capillalry umn which had been dynamically coated (Arrendale et 1980). The detector temperature was 300 °C, the in- on port was 280 °C, and the oven temperature was ed from 50 to 260 °C at 2°C/min- The cerrier (heliumm) linear flow velocity was 25 ®/s, the epGt flow 100 ml/min, and the helium makeup was 30 mL/min. hder these conditions, the trimethylailyl derivative of 01 eluted at 15.9 (t2%) min. Quantitative data, ed with a Hewlett-Packard 3351B automation sys- were based on reeponae and retention values of au- tic catechol. High-PresaureLiquid Chromatography (HPLC). itationof leaf chlorogenic acid contents was obtained the method of Snook and Chortyk (1981). Tobacco samples were subjected to an ultteaonification extraction with water and were directly analyzed by reverse-phase HPLC. This method has been shown to be reproducible with a relative standard deviation of less than 10%. RESULTS AND DISCUSSION. Pyrolytic yields of catechol(TableI) showed that flue- cured Brighttobacoo varieties produced significantly higher levels of this compound than for flue-cured Burley varieties. The Bright varieties exhibited a higher range of total polyphenolic leaf constituents as determined both colorimetrically and by HPLC.. Chlorogenic acid data, obtained by HPLC, indicated that chlorogenic acid isomers (oldorogenic, neochlorogenic, and 4-O-caff'eoylquinic acids) constitute a greater portion of the total polyphenols in "higb" polyphenol varieties than in "low" polyphenol to- baccos. The average catechol yield for the Bright varieties was 61% greater than thatdetermined for the Burley varieties. Average total polyphenol and total chlorogenic acid levels ranged from 118 to 271% greater for Bright tobaawe. These data indicate that although polyphenol L El'
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1194 D. C..Teosteson and M. A. Tnusn Mamelt L. 1. & Eling.T. E. (1983). Cooxidation during prostaglandin biosynthesis: A pathway'for the metabolic activation of xenobiatics. In Revieas in Bioch'rmical.. To.rimlogy 5. Edited byE. Hodgson. J...R. Bend & R. M. Philpot. p..135. Elsevier. New York. Mizutani T.. IshidaI., Yamamoto K. & Tajima K. (1982).. Pulmonary toxicity of butylated hydroxytoluene and! re- lated, alkylphenols: Structural requirements.for toxicity potency in mice. Toxic. app6.Pharmac. 62, 273. Mizutani T.- Nomura H.. Yamamoto K. & Tajima K. (1984). Modification of butylated hydroxytoltiene- induced pulmonary toxicityy in mice by. diethyl maleate,, buthionine sulfoximine, and cysteine: Toxicology Lerr. 23, 327. Mizutani T.. Yamamoto K. & Tajima K. (1983): Isotope effects on the metabolism and pulmonary toxicityy of butylated hydroxytoluene in mice by deuteration of the 4-methyl group. Toxre. appl:.Pharaurc. 69, 283. Nakagawa Y., SugaT. & Hiraga K. (1984): Preven6veeffecl of cysteine on bulylaled hydroxytoluene-induced pul- monary toxicity'in miee. Biochem. Pharnaa:33, 502:, Nakagawa Y., Tayama K., Nakao T. & Hinga K. (1984); On the mechanism of butylated hydroxyloluene-induadhepatic toxicity inrats. Binchem. Pharmac. 33, 2669. Shlian D. M. & Goldstone J..(1986)::Toxieity of butylated hytlroxytoluene. New hngf. J.Med. 314. 648. Smith:L. J. (1983). The effect of inethylprednisolone onJung injury in mice.,J. lab. din. Med. 101, 629. Subrabmanyam V: V. & O'Brien P. l. (1985). Phenol oxidatiomproduct(s), formed by a peroxidaseteeaction, that bind to DNA. Xeno6rntfca 15, 873.. Tajims. K., Yamamoto K. &Mlzutani: T. (1981). Bio- transformation of butylated hydroxytoluene (BHT) to BHT-quinone methide in rats. Chem. pharm. BulL, Tokyo 29, 3738. Tajima K.,, Yamamoto. K. & Mizulini T: (1983). Idenli- fication and: determination of glutathione conjugate formed from, butylated hydroxytoluene in rats. Chem. phurnc.Bull., Tokvn 31, 3671. Tajima K.. Yamamoto K. & Mizutani T: (1985): Formation of a glutathione conjugate from butylated hydroxy toluene by rat liver microsomes. BiocNem. Pharmac. 34, 2109. Takahashi 0. & Hiraga K. (1978). Dose-response study of hemorrhagic dcalh by dietary butylated hydroxytoluene (BHT) in male rats: Toxic. appL Pharmac. 43, 399. TakahashiO. & Hiraea K. (1979): 2.6-di-tert-butyl-4- methylene-2,5-cyclohexadienonc: A hepatic metabolite of butylated hydroxytoluene in rats. Fd Cosmer. ToxicoL 17, 451. TakahashiO. & Hiiaga K. (1981). Inhibition of phyllo- quinone cpoxide-dependent cazboxylation of microsomal proteinsirom rat liver by 2,6-di-rerr-bmyl-4-methylen, 2,5<yclohexadienone. FdCosmcl. ToxiroF..19, 701. Thompson D. C:, Cha Y. N. & Trush M. A. (1986). Thc peroxidative activation of butylated hydroxytoluene in BHT-quinone methide and stilbenequinone. In Biologtca!' Renctice Jntermedfates, IIa. Edited by1. J. Kocsis, D. J. Jo11ow,.C. M. Witmer, I. D. Nelson &R. Snyder..p. 301. Plenum Press:. New York. Tumer A. B. (1964)'. Quinone methides. Q. Reo. 18, 347: Winchi H. P. (1985). Enhancement of lung tumor formation in mice. In Carcinogenes"u, Vol. g. Edited by M. l. Moss, D. G. Kaufman. J. M. Siegfried, V. E. Steele & S. Nesnow. p. 147. Raven Press, New York. Witschi H. P., Hasehek W. M.. Kkin-Szanto A. J..P_& Hakkinen P. l.:(1981). Alteration of diffuse lung damage by oxygen: Determining.variables. Am. Rev. resp. D"u, 123, 98. Witschi H. &. Saheb W. (1974). Stimulation of DNA synthesis in mousc lung following intraperitoneal injec- tion of butylated hydroxytoluene. Proc. Soc. exp: biof. Med. 147, 690. QUPSTIDNB AND ANSWERS lohn Danief, Life Sciences Research, Chelmsford: I would just like to make a statemenl' and perhaps you would react to it. In 1966, 1 reported the formation of the carlwxylic acid from BHT in the rats which was based upon,.l presume,.the formation of a quinone methide, which then reacted non-enrytnaticallywith glutathione or cystine. I also reported an that time, the failure to observe such a metabolite in man and that the major route to metabolism in man, as Dr Conning showed this morning, wasthe formation of a dicarboxylic acid. Would you like to put your comments into perspective now for human risk assessment? M. Trush: All I can say is that in our studies we can actually demonstrate a direct interaction between a qpinanc methidc and glulalhione. It would be interesting to know if humans can, in fact, metabolizeBHT to quinonc melhide and how well they can do it. In going back into the literature it's questionable how well even some of the animal tissues can metaboGze it to its quinone methide in the absence of one of these activator compounds. And you can see that where we had absence of some activator compound, there was actually very little metabolism and covalent binding of BHT in and of itself. But inn the presence of these other compounds, there is.a dramatic increase in that. A:. Sevanian, USC Institute of Toxicology: I think your proposal here of the redox coupling between BHA and BHT and quinones is an interesting and, fascinating one. I wonder if I could prod you into a little more speculation. Could it be possiblelhat compounds such as BHA or BHT, when they undergo redox cycling, could set up a set of redox cascades with endogenous compounds of suitable redox poise relative tothemo such as catechols, ascorbic ad& and, other things, which could them precipitate a whole series of cascade reactions? Therefore,. BHA andBHT may not necessarily have to be the agents that deplete the glutathione pool? M. Trush: Right. That's possible. We've also done some reading about the quinone methides„which prompted thoughtt about alternative targets, such as NADPH oxidation. We looked for that and didn't find it. We even looked for the possible intercovalent binding of quinone methide.to NADPH because of the possibility it could be binding.to phosphate groups...lt's verypossibley you could seft up a redox cycle. Another possibilitgwe've also thought about and haven't tested is that there may be endogenous compounds whickcould drive the actual'metabolic activa other considerations or happen in terms of tox. bb Sfmic, National Burr and BHA, and the red, go from BHA to BHT U. Trush: Based upon I in'the formation of the M. Sunic: That's inten I
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Interaction of BHT with other antioxidants absolute amount of any one antioxidant present in the food product itself. The eBectiveness of certain individual antioxidants and some combinations of antioxidants has been attributed to the ability of the antioxidant to regenerate itself (Chen & Shaw, 1974; Kurechi et al. 1980). In a study on the synergistic effect of a combinationn of BHA and BHT on hydro- gendonation to the stable free radical 2,2-diphenyl- 1-picrylhydraryl, Kurechi etal: (1980) attributed the mechanism of increased hydrogen: donation to the regeneration of the parent BHA molecule. This re- generation of BHA was acsomplished,, however,, at the expense of BHT, which was converted into BHT- quinonee methide. The oxidation and regeneration of BHA at the expense of BHT might also explain our . results in peroxidase systems. The crucial point is the direct interaction of an oxidized metabolite of . BHA with BHT. If true, this would represent a novel mechanism of activation for BHT in biological . systems. `) We investigated the.ability of a number of other phenolic compounds to stimulate the metabolic acti- vation of BHT. This.was assessed by measuring each compound's ability to enhance or inhibit the covalent binding of BHT. Compounds that stimulated BHT binding are shown in Table 2. Althoughh all the simple phenolic compounds tested weree able to stimulate BHT binding, the most effective compound was BHA. The compounds selected for this experiment were chosen because of their presence in or addition o food and aoimetic products or medicinal prepara- li ons. In many instances one or more of these com- 1191 pounds is present in the samee product with BHT: The abilities of peroxidase enzymes from variouss mammalian tissues to catalyse the interaction of BHA with. BHT are presented in Table 3. Usingg arachidonic acid as a substrate,. BHA stimulated the covalent binding of BHT in microsomes from sheep, guinea-pig, dog and human lung, as well as dog bladder and rabbit kidneymedulla. Using hydrogen peroxide as a substrate, microsomes from rat, mouse and human lung;.as well as.rat intestine,.catalysed a similar enhancement of BHT binding by BHA. These results indicate that peroxidase enzymes from a vari- ety of mammalian tissues,.ihcluding human lung, can peroxidatively metabolize BHT to a reactive inter- mediate, which covalently binds to protein, and demonstrate that thisbindings can be enhanced by BHA, presumably through the.increasedtormation of BHT-quinone methide. In a related experiment,, guaiacol stimulated the formation of BHT-quinone methide from BHTusing myidoperoxidase isolated from human neutrophils (not shown). In order to assess whether the interaction of phenolic compounds with BHT to fovn BHT- quinone methide might have any in ufi:o significance, we measured theefiect of various doses of BHA on BHT-induced mouse lung toxicity (Fig.. 1). BHTeauses the destruction of type I alveolar cells(Marino & Mitchell, 1972; Witschi & Saheb, 1974) and pul- monary rndothelial cells (Smith, 1983). This toxic response is generally reversible (in 6-10 days) unless a second stress impedes the proliferatiomand repair process byy the type H pneumocytes. For example, if Table 2: Abilityof various phenolic etiendrals to enhanm peroxidaseolnlysed eovalent Nndingof BHT to protgn Compound Use Relallveenhanacmrnt• BHA Anaoxidant +++ Methylparaben Pmervatlve in bererages: foodr and eosmetics + + EVgenol In potfumc, manufacture or vaniuin,..dmW uWgeaic + + Vanilbn Flavouring .gem ++ Diahylailbudtrol Uestrogen + + Phenol Disin&..1.nt; andmicrobiallagent + + Guaiiwul Fspexonnt, + Aceeamieophen Analgesic, antipyretie + Ferulis atid Natural plant cunstituent + oesuadial Destrogen + •(+) repnsrnn 1-2-totd eNUneemem of BHT covalent binding; (++) rrytercnb 2-3-fold eManoement; (+++) represents >3-foH enhano:mmt. Table 3. Peroxids.dryetdent rovaknt binding of BHT lo proteih.by mismsumes from various animal asues Covalent binding (mnol'.gHTboundJmg proteinjlOmin')in insvbaons contalning Tuue BHT BHT+BHA Sheep lung Andido.k adOdegeptenr 0.18t0.02' 1.63+0.06 Guilw-pig lung. Dog lung 0.15 t0A2' ND 0.37±0.10 0.68¢0.0g Human lung 0.10 } 0.01 0.35; 0.05 Dog bladder R bbi kid d ll 0.44; 0.03. 7.90 t 0.65 a tt ney me u a ND Hydrogen peoxldaMpe.ds.t 0.69 f 0.23 Rat lung 0.0630.02 4 0.32 0,04 Mou.e lung 0.13t0.01. 0.80}0.03 Human lung 1.03 ±0.01 6.14 ±0.03 Rat intestine 4.33 }0.51 27.66.t4.15 ND - Not detectabk •Except (or hydrogen peroxide-0ependent binding by human lung (3Umin incubation).
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J phenols and: acids. of four varieties. of tobacco (Bright,. Burley,. Turkish and d'omestic)are given in Table 3. In the acidic fractions of either smoke.of four varieties of; tobacco tested, major phenols were phenol, catechol nnd hydroquinone; major acids were formic, acetic, lactic, glycolic and succinic acids, witb thee exception of a large amount of 3-methylvaleric aci& in the smoke of Turkish tobacco.. The compounds for which SS/MS distribution ratios were larger than 1.0, were phenol, cresols, xylenols and guaiacol among the phenolk, and acetic and formic acid among the acidks for all tobacco varieties tested.In. addition to thesee compounds,. the ratio of3-methyl- caleric. acid for Turkish tobacco smoke was 1.46. The SS/MS distribution ratios of dihydric phenols.such as catechol, hydroquinone and their alkyl derivatives and acids other than formic and acetic acidss were less than lA. Those of 4-vinylcatechol,, which is mainly derived from chlorogenic acid (13), m-hydroxybenzoic acid, and dibasic acids such as succinic, methylsuccinic. Table 2. Acids and other acidic components Identified in _ Figure 4. Peak~No.. 1, Compound: 2 14 3-Methylvaleric acidLachc. acid. 17 Glycolic acid 27 Levulinic acid 28 2-Furoic acid 38 3-Hydroxypropionic acid 41 3-Hydroxybutyric acid 46 2-Hydroxy-3-methyl-2-cyclopenten-l-one (cyclotene) 58 Benzoic acid 67 5-Metttyl-2-turoic acid; 75 Phenylacetic acid 80 Maleic acid 84 Succinic acid 86 Methylsuccinic acid 87 m-Toiulc acid (internal standard) 91 Itaconic acid 92 p-Toluic acid 93 Fumaric acid 106 Glutadc acid 115 5,6-Dihydro-3,5-dihydroxy-2-methyl- 4H-pyran-4-one 123 3,5-Dihydroxy-2-methyl-4H-pyran.4-one (5-hydroxymahol): 128 o-Hydroxybenzolc acid (salicylic acid) 140 m-Hydroxybenzoic acid 142 p-Hydroxybenzcic acid and glutaric acids were especially low. It is known, (I) that small moleculhs arc preferentially delivered to the sidestrcamsmokc: The SS/MS distribution ratioss of phenols and acids.in Table 3 show a similar tendency in that the larger the molecules,.the lower the ratios. There have been some reports on the SS/MS distribution ratios of phenols and aeids.. Cartwright et al. (11) ex- amined the 55/MS distribution ratios of steam-volatile phenols.in some cigarette and cigar smokes and showed that the SS/MS ratios of unfiltered cigarettes or small cigars were 3-5 for phenol and 2-3 for cresols, ethyl- phenols and xylenols, while the ratiosaf filter cigarettes or medium to large cigars were 9-16 for phenol and 3-18 for other volhtile phenols. Harris et al. (5) have reported the SS/MS distribution ratios of phenols to be 1.7-20 for various filter cigarettes. Brunnemannet al. (10).found the SS/MS ratio of non-volatile catechol to be 0.78 for plain cigarette and 0.65 for filter cigarette. fohnsonn et al. (9) have shown that the SS/MS ratio of acetic acid was0.67for the85 mm Kentucky reference cigarette. Of the ratios reported by these authors,, only that of "catechol(10).is in approximate agreement with the value obtained'in this study and given in Table 3. Direct com- parison of the SS/MS distribution ratios obtained in this study with those obtained by other authors, however, is difficult because of the differences in methods of sampling, an& analysis. REFERENCES 1.. Johnson, W. R., R. W. Hale,. J. W. Nedlock, H. J. Grubbs,. D. H. Powell: The distribution of prod- ucts between, mainstream and sidestream smoke; Tob. Sci. 17 (1973):141-144. 2.. Yoshida, D.:.Detertnination of nicotine in the side- stream of cigarette smoke; Sci. Pap. Jpn. Tob. Salt Public Corp. Cent. Res. Inst. No. 118,. 1976, 203-206. 3. Jermini, C., A. Weber,.E. Grandjean: Quantitative Bestimmung verschiedener Gaspliasenbestandteile des Nebenstromrauches von Zigaretten in der Raum- luft als Beitrag zum Problem des Passivrauchens; Int.. Arch, Occup.. Environ. Health. 36 (1976) 169-181. 4. Cornell, A., W. Cartwright, V. Olender: Side- stream/mainstream distribution ratios of ammonia and volatile pyridines in cigar smoke, Influence of cigar components and smoking regime; 31st Tobacco Chemists' Research Conference, Greensboro, North Carolina, 1977. 5. Harris, J. L., L. E. Hayes: A 20.port sidestream smoke collection system; 32nd Tobacco Chemists' Research Conference, Montreal,, Canada, 1978: 6. Dong, M., I. Schmeltz, E. Jacobs, D. Hoffmann: Aza-arenes in tobacco smoke; J. Anal. Toxicol. 2 (1978) 21-25. 69 II
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ov; V.I., and Yu.V. Korogodina, Institute of Zical Genetics, the U.S.S.R. Academy o( Medi- .j Sciences, Moscow (LLS.S.R.) Lgenic and teratirgenic effects of thiophos- #rmide in CBA, 101 / H, and TPS mouse.embryos ~~b estimate the mutagenic and teratogenic ac- ,pp of thiophosphamide, a trifunctional alkylating - , pregnant females of strains CBA, 101/H TPS were treated withh the substance in- ~eritoneally (5 mg/kg body weight) at the 12th ;~ of gestation. To evaluate the rate of mutagenic ~y chromosomal aberrations were scored in ryonic liver cells.. Embryotoxic and teratogenic were measured in terms of postimplanta- lethality; body length and weight of 19th-20th embryos, and the rate and spectnm.of skeletal alies (deformation, reduction or absence of bones in limbs, spine, ribs and skull). . fraction of aberrant metaphases in em- nic liver cells induced by the treatment was to be about 62% in CBA embryos.and about in 101/H and TPS. Thiophosphamide-in- increase in postimplantation lethality was onlyin TPS strain (about 35%). Body length weight studied in 101/H and CBA strains by about 35% and 45rR,.respeclively, in former but only 20% and 35% in the latter veto untreated~mice. Overall rate of skeletal es was found to be about 90% in 101/H about 60% in CBA embryos. us, interstrain differences were found in the of mutagenic and teratogonic effects of hosphamide namely, TPS and 101/H em- were much moresensitive than CBA ones in ;respects. n', T., M. Curvall'• and L. Zech •. • De- ent of Medical Cell Genetics, Medical Nbbel te, Karolinska Institutet, Stockholm and ch. Department, Swedish Tobacco Com- Stockholm (Sweden) of sister-chromatid exchanges in human es by neutral and phenolic constituents 2N from the semi-volatile material of cigarette smoke condensate A cigarette smoke condensate was separated by distillation into a semi-volatile and a non-volatile fractioo, The.fractions were tested for induction of sister-chromatid exchanges(SCE). in human lymphocytes. The activity of the semi-volatile frac- tion was higher than the activity of the non-vola- tile. The semi-volatile fraction was. further sep- arated into acids,phenals, bases and neutrals by solvent partition. The acidic and the neutral frac- tions showed high SCE-inducing activity. The.pBe-- nolic fraction had a low effecc while the basic fraction, which contained mainly nicotine, did not affect SCE. The neutral and the phenolic fractions were further investigated. Flash chromatography on silica gel with gradients of hexane/ethyl acetate as eluents of the neutral fraction furnished 7 neutral subfractions,.2 hydrocarbon fractions and5 more polar fractions. All the neutral subfractions af- fected SCE and the effect of S increased with increasing polarity. Major pone ts of the neu- tral subfractions we iden ied by g chromatog- raphy- ass spectr etry: One of the hydro- carbon ctions~s co tained a larg amount of neophy ene, wliich origin tes fr chlorophyll. This p und was 'sola . tobacco and tested, ut dt not en ance the ftequency of SCE. The heno fra ion was separated by gel chromat raphy on ephadex LH-20 into 11 sub- fractions, which were tested for SCE induetion. All except one showed effects on SCE determined by regression analysis:.. Major components of the phe- nol subfractions were identified by gas chromatog- raphy-mass spectrometry. 66 of these compounds (phenol, ua'a eugenol, cycloten, furfurylalco- hol, 2,6-dimethoxyphenol) were tested, and twoo of them (guajacol, cycloten) were found_ to induce SCE. Tests for SCE induction of other major components from the phenolic subfractions are in progress. 43 Jantunen,. MJ., A. Lumatainen„T. Ramdahl * and A. Ilkonen, University of Kuopio (Finland),
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tl P:u / Mutapcnicily.of'i.phcnols from smokehouse smoke Lanovmrr.>,n,ku fao,lucur SSnuhC cOm IU•nsate ' 0,).. epue ' [Ijni eil1, ctnl'1anelaLiv, ItI i A4uco - phasr I el. (:.) tt'N. rar fCactl4na (Jq: rt.n. aa•, .~,. Irzcr.luu. . ~ .r.inarlo exrranuoi, elv~cu,yl aecrau Iltnanie pha- -------- I I evaporarimc [o drym.e> PhenuliC extract I IMS-slliea gel epluun eprifird pLenolic fraccinn' (FAa-free) (mono. and ntfunCtlnnal phennls) I paititioninc vith ethyl acetate ~i Yenvfunecivnal aifanccional pbenols' ptmnols' Fig..2. Preparation of fractions containing mono and bifuncli6nal phenols from smokehouse. smoke condensale. Fractions indicated by asterisks were tested for mutagenicity: plied by Dr.. B. N. Ames, California. The test for their reversion to histidine prototrophywasperformedming the plate incorporation assay as previously de- Ieribed (Ames,. McCann & Yamasaki, 1975). Fach~ compound or smoke condensate fraction was dis- , mlved in DMSO and tested im five concentrations in ~ quedmplicate in the presence of buffer pH 7.4 or S-9mix with cofactors (60 mg S-9 protein/10 ml S-9 mis)) hom, Arodor-pretreated Sprague-Dawleyy rats. The S-9 protein content was determined using.tNe biuret- method-kit nethod-kit of Boehringer, Mannheim. The sensitivity I of the S-9 and of the indicator strains was monitored ( routinely using positive control compounds during E: och experiment. Table 2 summarims the combined ' poutive control data obtained ib the individual ex- ~ pcriments, Significance levels for positive dose-res- Pnme effects were obtained with the Jonchecre test ~ IFIolPander & Wolfe,.1973)• l Special proceduress in addition to the standard i eethod were used tostudy the mutagenicity of the 11. asrent compound{ phenol. These included incorpor- ~A umn of S-9 from untreated Sprague-Dawley rats, anncubation of the compound, (8dded in I00pli H:ll) :with the bacteriaa and 5-9 mix prior to plating. Wllion of$-9 mix with various protein contents,. and 385 inclusion of the comutagen norharman during the tes1.. RESOLTS. The mutagenicityof the ten phenolic compounds detected, in smokecondensates were investigated in the Ames Test (Table 1): Each compound was tested in quadruplicate at five concentrations in each of the five bacterial strains;.both in the presence and absence of the 5-9 mix. The number of hislidine revertants scored in the presence of the test compounds phenol, o-, m-, p-cresol. 2,4-dimethylphenol, bmnaLatechine, uuuiunl, syringol, vanilline and eugenol,.never more than slightly exceeded that number of spontaneously arising revertants. For all of the compounds the only dose-related effect was toxicity at the highest tesle.d concentration (5000pg). This was apparent as a thin- ning of the bacterial background lawn and as a reduced.number of spontaneous revertants. Phenol was further tested using a variety of ezperi- menlallmodifications with S. ryphimurium TA98 as in- dibator organism. The test was carried out using.5-9 from untreated Spraguc Dawley rats. to determine whether Aroclor-induced detoxifying enzymes played B:
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6Er~iNf w(. i~'r alogbemeul Phermacoloi,. Vol. Is, oP'-atll•'UPl. 14rpamon Prcs 1069. PrinmJ in Grem Hrnaln 113ti.5 EFFECT OF ESSENTIAL OILS ON DRUG METABOLISM A. JORI, A.. BIANCIIETTI and P. EF. PRESTINI Islitulo di'Ricerche Farnmcologiche "5tario NeSW°. \'ia Grilrea.62:'-u157 Milano;,llaly. (Rerek+rd 31 Jununrr 1969: uceepred15 .IPer( 1969) Abstraet-A number of componcnts nf essential nilsk sudi as nsnlhnl. n and a pincne. guaiacvfand eucalyplol and the oil of PLuu pmuiLo, ere sludicd t,+ calahlishuif Iliey affect the metabolism of other drugs in ial.e. Eucalyptol,adminislered subcutaneously or by aerosol +cas found to increase utc in uirro (9000g) liver metabolism off aminopyrinc. P-nitro-anisol and anilincc and in citro the metabolism of pentobarbital. RECENTLY se.veral papers have been puhlished,ahout controllithle faclors.that modifydrug action arrd toxicityin experimental animals..Of the various environmental factorss studied,, the inductive activity of DDT,' chlordane- and'~ related pesticides on micro- somal enzymesystems, which is responsible for instance for the modified responsive- ness of animals to barbiturates, was most dramatic. The aim of the present study wass to establish if constituents of essential oils, widely employed in disinfectant sprays andairfreshenesaswell as in pharmaceutical preparations, afleceothedrug-metabol- izing activity and ponsequently modify the pharmacological responses to drugs in rats. MATERIAL AND METHODS Female Sprague-Dawley rats weighing 150gji_ 10 were used. Drugs were dilutetl in arachid oil_andinjected subcutaneously(s.c.) or undiluted _nebuliz by mcans of pressurized air in aerosol treatments. The inhalation of aerosolized drugss was repeated for 4 days consecutively but fnr ' different lengths of time, as raporled in the tables. The drugs were nebulizedd at the L ratee of 50 mg/min: Two rats were placed in aa round cage (4 (m+') with solid sidcss for the period of inhalation.. 2 Drugs used were: eucalyptol (cineole); uaiacol (o-methoxy-phenol); menthol (hexahydrothymol).(obtained from Faravelli- dan): oil of Piuus pumilio (containing pinene, phellandrene, dipentene, sylvestrene and 5 0,/of bornyl acetate) (Esperis, M ilan ): a-pinene and P-pr'nene (nopinene).(C.Erba, Milml). All products are pure according to the Italian Pharmacopeia (F.U: - 7th edition). Pentobarbital effect (sleeping timc)) was ev:dualcdl as the limc elapsed between loss and regaining of righting reflex.. Pentobarbital concentration wus determined by the method of Brodie et al.a with minor modilications. Enzymatic activity was measured imrilroon 90IXYg supernatant fraction of a liver homogenate of treated and control rats,, according to KatoandTakanaka.' The following enzymatic reactionswere estimated: ring-hydroxylanion.(aniline),.N-deme- thylatiom (amiRopyrine)) and O-demcthylation (p-nitroanisul). The mctabolites, 2081 .d¢,p7a1*
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138 tion of cigarette smoke condensate (Hecht et al., 1975). Cyclotene, 2-hydroxy-3-methyl-2-cyclopen- ten-lLone,, which is the major component of that fraction did nou show any'tumor-promoting activ- ity whentested on mouse. skin(LaVoie. et ali,. 1980). It has recently been demonstrated that cyclotene iss inactive in the Ames test (Bjeldanes and Chew, 1979). Cyclotene was the major com stituent of fraction F3 and makes up 28'R, of this fraction. Although the effect of cyclotene on SCE' I was low ittill contribt th , may sue significantly toeeffect of thissubfraction. The most active fraction, F4, contained alkyl-2•hydroxy-2-eyclopenten-l- ones and also alkyl-substituted 3-hydroxy-1,4-py- rones. Maltol (3-hydroxy-2-methyl-l,4-pyrone); which is known to bea base-pair mutagen as determined by the Ames test (Bjeldanes and Chew,. 1979) showed a low effect on SCE in this study. The major component of this fraction, furfuryl alcohol,did,.not induce SCE, but this compound as well as'some other furan derivatives are.known to be dastogenic(Stich et aL, 1981). The compounds found to be active in this study can only in part explain the total activity of the WASV fractions of the cigarette smoke con- densate. However, the present findings c)earlyshowthat many weakly acidic compounds are inducers's of SCE. Since these compounds are water-soluble. and may be readily taken up by different tissues, they are probably distributed in most tissues as is catechol in mice exposed to cigarette smoke (Hwang,et al., 1982). They may therefore.contrib- ute significantly to the in vivo effect on SCE seen in cigarette smokers. Since some of these com- pounds are found in food, extensive studies for other genotoxic effects are required. Acknowledgement Thiswork was supported by grants from the Swedish Tobacco Company and one of us, T: Jansson, is also grateful to the Funds of the Karo- linska Institute for a grant. We wish to express our sincere gratitude to Dr. L. Zech for valuablee ad- vice and criticism of this work. We are grateful to. C. Bjarklund.for skilful technical assistance, Dr. T. Nishida for recording the NMR spectra,.and to Dr. J. Bielawski and Dr. O. Dahlman for record- ing the mass spectra. ,. ! References Lamben, (1978). Arctander; S. (1969a) Perfume and Flavor Chemicals. Vnl. 1, eigare Montclair, New York. . uyoie, F Arctander, S. (1969b) Perfume and Flavor: Chemicals, Vul. 2. The le~; Montclair, New York. G-R. ( E~ Bjeldanes L.F., and H. Chew (1979) Mutagenicityy of 1.2-di: .s Ggare r. carbonyl oommpounds:.Mahol. kojicaeid, diaextyl and re- /.ivingslo r lated suhstances, Mutation Res., 67, 367-371. man 1, BoutwelC. R.K.- and D.K. Bosch (1959)~ Cancer. Re..-.19; : ution 413-424. ~ Morimolo.. ®' Cheng, M., and A.D. Kligerman (198•1) Evaluation of the _ suler-. ~, <notoxicity of cresols usin siiterchmmatid ex<h i g g anpr t (SCE). Mutation Re.c.,,137, 51-55. ! 119, 3: 'i. Curvall,: M.. T. Janssnn, B. Penersson; M. Thelestam anJ C_ ~ Nestrtrann Enrell (1984) Evaluation of the biological aclivirt. of( JC. M cigareue smoke condensate fractions using six in vitro .iinre " m pull term,tests, 1. Toxicol. Environ..Health, 14, 163-180. mamm Curvall, M., T.. Jansson, B. Petterssom A. Hedin and f'.R. ~ Norpp; I Enull (1985) In vitro studies of biobeical effects of cie:,-eu.. {-"m.dd smoke condensate,,1. Genotoxic and cytotoxic effec.nf neutral, semivolalile constituents, Mutation Res.. .57. 169-180. Darroudi, F.. and A.T..Natarajan (1983) Cytogenehc ar.....,,. I . ,..human a.: of human peripheral blood lymphoeytes (in vitro) ud with resominol, Mutation Res.- 124, 179-189. Diem, K., andC. I.enmer (Eds.) . (1970) Dooumenta i,_ Scientific Tables. Geigy; Basle, 173 pp. Florin, I:, L Rutberg. M. Curvall and C.R. Ervell . Screening of tobacco smoke constituents for mutag using the Ames test. Toxicology. 15, 219-232. Gocke, E, M.T. Kind, K. Eckhardt and D. Wild (19B1'• ,n genicily of cosmetus ingredients licenced by the Er ,an Communities, Mutation Res.,.90, 91-109. Hcchl. S.S., R.L Thome, R.R. Maronpot and D. Hc nn (1975) A studyof tobacco carcinogwesis, XIII. Tuc moling.subfractionsof the weakly acidic fraction. Cancer Inst., 55, 1329-1336. Hecht;:S.S., S. Caramella, H. Moriand D. Hoffmann (' I .1 study of tobacco cardnogenesis- XX•: Role of catev, , major cncarcinogen in the weakly andic fraction o =lc condensate, J. Natl. Cancer Inst.. 66, 163-169. Hoffmann,. D., S.S Hecht and EL Wynder (1983): nwr promoters and', cocarcrtwgens in tobacco oarahr., ..a,. Environ. Health Perspect., 50, 247-257. Hopkin, J.M., and HJ. Evans (1979) Ciga¢tte -inke condensate damage DNA in human lymphocytei. .mrr (London), 279, 241-24L. Hopkin, J.M., andH.J. Evans (1980) Cigarette smokeiccd DNA damage and lung cancer risks, Nature (I.ondc: I&). 388-390. Hwang, K.IL, 0. Sonko, D.R. Dansie, R.E Kouri a' C.1 Henry (1982) Studies on thc deposition and distnu- •n:o/ catecho/ from whole cigarette smoke in BC3F1/e: rn~.e•: Toxicol. AppL. Pharmacol.,, 64, 405-414, Kasamaki, A., H.,Takahashi, N. Tsumura, J. Niwa, "i .ujN and', S. Urasawa (1982) Genotoxicityy of navouring :~penn. Mutation Res. 105;,387-392.
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a « 306 as '!EL . t..am. rudiw- vua I S ea rn-I sI . Yttsy.e rtaa 1977, PM40 ~n ce<o a,ilu,n REGENERATION OF RAT LIVER IN THE PRESENCE OF ESSENTIAL OILS- AND THEIR COMPONENTS L L QgasnnerN Northwest fnsriture fo, Medical Reseacli, 5656 West' Addison Street. Chicago. Illinois 60634. USA (Receiaed'1 OcroAer 1976), Abstnct-Irver regeneration wasiolbwed over a 10-dayy period in paaially hepatenomized?ats given se injections of high levels of essential oils: terpenes or aromatireompounds daily for the first 7 daysor fed ad fi6: for 10 days on diets supplemented with oils. spices and bounicalsThe liver inerement (the amount of tissue rcgenerated) was increased signifieantly' by sc injection of oils of anise, fenncl, larragon, parsley seed, celery seed and olcoresin, nutmeg, mace-cumin and sassafras and of the aromatic principles- 4-allylunisole, 4propenylanisok, p-isopropylbenarldehyde,safmle and isosafrole. Most of Ihe essential oils were ineffective in total doses up to 3ll(10 mg/kg because [hey contained a high percent- age of terpenes, which proved inest Many'of the agents'effeetive by the sc route were also activc when added4o the die1, although a possbk exception was tarragon oil (050%) Azulenes(e.g. Marricaria chanorniRa L oils and guaiazulene) stimulated regeneration whengiven by injection but Haitian vetirert oil, the akoholmikture vetiverol and guaiene, in massive doses- did not. However, these last three agents, as well as the blue chamomile oils and ground' Marricarla flowers, stimulated liver regeneration when given in the diet Several.of the test agents also increased the wet and dry liver weight expressed relative to body woght,.in intacL rats,, whether given by'the se or aral rnute. To throw further light on the rests with the active principle; certain aromatic or phenolic compounds, including monosubsti- ruu:d derivatives-.were screened in operated animals by the sc route INTRODUCTION' ksseruial oils and flavourings are widely used and must figure prominently in nutritional surveys as possible factors in cancer and some other diseases Sdrole (4allyl-l,2-methylenedioxybenzcne);k an im- ponant, componentt of a number of oils, has been rtwwn to be.a hepatic carcinogen (Borchert. Miller, ]GI)cr & Shire& 1973a; Epstein, Fujii. Andrea & 1lantel, 1970; Hagan, Jenner; Joner, Fitzhugh, Long, Brouwer & Webb, 1965; Homburger,. Kelley,, Baker & Russfield, 1962; Long, Nelson, FitzLugh & Hansen, 1963) with I'-hydroxysafrolc constituting a strongly hcpatocarcinogenic metabolite (Borchert er aG.1973a; gorchert, Wislocki, Miller&Miller, 1979b; Wislocki, gomhert-. Miller & Miller, 1976~ In a short-term h'igh-dosagetoxicologicale study, gross pathological changes were reported as extensive with ssd'rok,.isosa- frofc, dihydrosafroBe and aliphatic allylcrompounds, moderate to minimal with the 4allyl-, 4-propenyl- and 4n-propylanisoles and essenliaBynegative with benzene- allylbenzcnc and anisole (Taylor, Jenner & lottes- 1964) Reports are also available on. safrole and other principles in relation to accidental poisoning rCraig,. 1953kto hypothermic activity (C:mjolle & ylcynier,J960[and to differences in gastro-inleslinal +bsorplion (Fritsch, de SainlBkrnquat & Derache, 1975) The effecl of nutmeg.or myristiet powder, as xell as of its. principle, myristicin (4-allyl-6-methoxy. 1?-methylenedioxybenzene)t .has been determined on the nervous system. (Weiss, 1960; Wesley-Hadzija & Bohinc, 1956) and an explanation of the psychotropic activity has been attempted on the basis of a sug- gened relationship with the structure of sympathomi- metic amines. (Seto & Keup, 1969; Shulgih, 1966; ~ Shulgin & Sargent,. 1967} Yet another principle of several oils, namely. asaronc or 1;2;4trimethoxy-5- propcnylbenzene,. has been studied from the stand- point of physiological and psychotropic properties (Dandiya & Sharma, 1962; Seto & Keup, 1969; Sharma & Dandiya, 1962) Recrnt biological data on safrole have been reviewed by Opdyke (1974kL and for more genemP information on essential oils and flavourings, the reader is referred to the latter work as well as to several treatises, especiallyy that of Guenther (194B-)952). The general problem of enzyme induction has been investigated in relation to safrole but the lindings have varied with the criteria employed. Liver micro- somal-enzyme activityy hasbeen reported as decreased (Friedman- Arnold, Bishop & Epstein, 1971; Fujii, Jaffe, Bishop, Arnold- Mackintosh & Epstein, 1970; Seto & Keup, 1969), as increased (Gray, Parke.. Grasso&Crampton, 1972;Lakc& Parkc, 1972;.Lot- likar & Wasserman, 1972; Parke &Rabman, 1970) and as essentially unchanged (Burchcrt rr ul.- 1973b; Lollikar & Wasserman, 1972). The formationn of a safrole metabolite-cytochrome P-050 complex which can be cleaved by safroleand other agents has beenn advanrxdrecently (Elcomhe, Bridges, Gray, Nimmo- Smith & Ncttcr, 1975). The present investigation. was undertaken with a vie<v to screening the eBecn of a number of essential uils, Oavuurings,s spice.s and plant materials on dre extent of liver regenerationn in partiallyhepalecto- mizr.d r.us. Where an oil displayed a definite acrivity,. a correlation was attempted on the basis of the action of terpenic and aromatic components tested in a simi- lar manner. In this connexion, previously published findings with a variety of aromatic compounds engen- 173 /5, 1'Y3 : /g/
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INDUCTION OF CHROMOSOME FRAG- MENTATION BY PHENOLS svALIIERT hF.l'.4.\•. .%su. JOE 1ilA! TJIO CYTO.cK%l:nC LAlfux.CruaY, lc.;.ifP niMdl blood I. GENERAL OBSERVATIONS. SINCE our recont discovery ILIiVAN nnd I'JI(). 1948) '. that phenols hneea fragmenling effect on allium Cr•pn chromosomes we have Ie.ted snme forty phenols, phenol dcriv:divcs and related compounds nn. Ilteir fragmenting activitv. Allhuugh the majority of them exhibited sume acliVily.most of them were I'imnd lo bc but very little active. In many cases only a systematic search through many slides gave a positive result. This may be one reason for Ihe non-deteclion of this reaction earlier in spite uf its .ridee dislribulinn wilhin this type of substances. Another reason is that the reaction muy he well-defined and distinct only in fnvournble cytological III:Iterinls. Thus, hARMENTIER and htts•rtN ( 1!l48) .evidently dcal with Ihe same ruaclion in fhe mouse a17ar hydroyuinnne Irealmenl. They mentiun,., in fact, a chromosome frag- mentationn as one amon„ four ultcrnatire explanalions of their findings, bnt they considcr il as more likvly th:d .ume change in the cenlromerew induced hv the treatment iss r(•sponsible I'or the charncterixtic cytolog- ical picture at hand. On each side of'tlte ntelaphasc plale little groups off chromosomes migrate towards the poles. These chromosomes arc usuallyIhey smallest onesR. and Ldi•r: -,4n inlrinsic cause of the ab- nnnnal chromosome migralion appears Ib Ire the most probable. It is suggested that au impeded division of the cenlromens of the longest chromosmnes gives the besl.explanalion of(he peculiar nlitotic figures. (t. c.. p. 527). In the more 1'avrrurable material of .4!lium it is casily secn 1h;d the ~smallest, c.hronwsumes miFratinR to the poles ure lhe 16 centromeres logelher wilh so much of their chromosomearms as has been left• while the remuindc•r ofIls chromosome malter lags. ut llee equator region as free fragmeols.. Our standard melLod of testing Ihe suhxlances ml Iheir fragptentin.- acticilyy is the same as LEVAN and coll:dxuators have employed since 1!138 for analysing the c-nrilr.licc aclivily of ch(•mical'suhnhmces. A HEREDITAS 34 (1948) pps. 453-484
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aaffected.italation, aered by ;stration, , aificamtly ris with ophcnol . - S.E.1 z 5 16 16 16 10 _ 10 uss werc -nsolized ;ures are atration. 4 aniline uch as of the ITerent rity of zy.mes ration drugs ¢yme :"allce Uusly tivity ngs.a vood :5tilr ctice Elfeenof cssenlial oils on drug.m<tabolism 20a5 Our results confirm this.possibility and show rhat relatively low amounts of com- pounds such as eucatyptol, may aflect the microsomal activity when inhaled by rats.. These data should be stressed inn view of the large usc of cucalyptol aerosol I'or therapeutic purpose and in view of the possibility that sprays of other compoundss iii the environment may contribute to change Ihe drug metabolizing.activity in animals. AcknonJedgernrar-Thisworkwasfinanciallysupported byContract DHEWrI'HS. NIH/PH43-67-N3. The technical assistance of Mr. Luciano Guarmeci was panicularty appreciated. REFERGNCES 1. L. G. HART and 1. R. Fours, Pror. Sor. e.rp:.BioL Jrrd. 114,.388 (1963). I. L. G. HeaT,.R. W. SnuLrtcE and J. R. Fours, Toxic appl, PGmvuar. 5„371 (J963). 3, 0. 0. BrtontE; J. J. BURNS, C..MARK, P. A. LEter, E. BeRNxreta and F:: M. 1'ArreR, J. PLaruurr. r.tp.. TAer. 109,.26 (1953).. 4. R. Kero and A. TAKANAKA, Jap. J. PGnrmar: 17, 208 (1967). 3„D. GILBENT and L. GOLD9ERG: Pdl CoLaer. TOSir. 3,417 '(I J65): 6. H. C. FEaarrsoN, J. Phann, Sri. 55, 1142 (1966). L E. S. YrSELL, SNentt 157, 1057 (1967)... 8. E. S. VeseLL, PtwramratagY (8ax!) 1,.81 (1968). g. A. E. WADE, J. E, HoLb, C. C. HILLIARD, E..Muuns and F. E. GRe[w•.. Pharamrnlqrr(&+srp i, 317(1968).
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Mutagenicitg ofphenals from smokehouse smoke Table 5, .Ilmupenirire levlinq n1 crude undpuriliid .vmukr rimdensureqrurfinns rvrurriitinU phruulr 389 Nuof hii' rev<ftantsplare Concentration TA1535 TA1537 TA153R TA9% TAIOO Fraclion Ipg'plLtcl -5-9 + 5-9'- S:9+S-9. -S-9 + S-9 - S-9' . S-9 -5-9 + S-9 Smo4cmndcnsuu l to 16 U'. 11 Ig 22 16 I_ 76 113 ILYr BnCtinn 05 20 21 11 13 IR 25 19' 36 66 96. F¢s I nnd 21 5 20 21 13. 23 20 34 20. 36 69 102 511 19 24 13'. 18 17 28 16. 3N 69 107. 5at 17 15 13 . 23 14 30 19' 0: 65 103 5(ItIM 0 12 0 17 0 29 0 0. 0 96 Smoke cnndens:nc o Ifi 18 13. 12 18 20 16. 29 76 108 Wqocuas fracliim: (}5 16 19 13 I5 IlS 25 IS _'S 64 117i FigsI and!'-) 5 16 19 1.5 11 16 26 17 21 63 98 50: 21 20 II 13 16 23 14 27 69 104 Sac 17 19 14 lo' 14 20 14 n 77 104 SncKlr a 21 0 . 13 0 18 0 0. 0 108 gaxtztmct 0 19 22 6. 9 14 27 33 47 145 Illl Ifrom.lar fraction: 05 25 16 7' 14': 20 21 34 48 121 125 Figs.I and 21 5 24 20 9 II li 30 '27 46 146 78 su 211 24 5 12 14 31 23 32 123 113 500 24 16 8. 7 20 26 24 43 154 136 50Uf)f 0 13 . 0 9 0 0 0, 21 0 90 gaw-exlrae0 0 21 23 14 22 23 33 19 43 127 163 drom aqueous frmtion:. 0.5 ?9 16 14 25 22 70 IS 38 109 153 Fies I and 21 5 27. 26 14 20: 19 34 20 36 121 157. 50: 29 26 20~ 21 18 32 21 39 131 134 5(10. 27 19 18 24: 27 32 ?5 311 144 142 5001 0 IS 0 13' 0 26 0 30 17 152 ScutruI Icompotmds. (Fig. 1) 0 0.5 9 7 10 12 9 8 6 9 8 5 13 15 19 13 Ib 13 163 176 97 103 5 8 9 4 7 2 15 II 14 161 101 501 6 7 4 7 3 16 8 19 165 101 500' 3 7 8 7 3 15 9 14 189 94 5000t 4 7. 0 10'. 0 19 0 5" 229' 108 1lonofunctional 0 7 13 9 6 8 14 19 20 174, 120 phenols (Fig. 1 f. 05 7 9 6 II 6 11 17 19 151 112 5 9 10 4 6 7 18 19 14 157 117 50' 7 13 3 7 8 13 17 20 172 100 500 6 9 2 10 10 12 24 14 177' 109 5000t 0 6 0 6 1 11 0 16 108: 120 Purified phenolic 0 17 13 9 10. 14 18 17 171 178' traction (Fig. 2) . 0.5 21 13 8 7 16 20 19 23 152 143. 5 20 IS 8 5 16 22 20 24 168'. 149 50, 19 12 8 5 IS 20 21 23 175. 150 500 50U0t 18 4 16 15 6 4 5. 6 14 6 17 12 7 0 23 17 209' 0 152 150 Nonutunctional 0 7 13 9 6. 8 14 19 20 177' 103 pkenols (Fig. 21. 0.5 7 9 6 II 6 11 17 17 178'. 104. 5 9 10 4 6 7 18 19 14 176 101 50 7 13 3 7 8 13 17 20 184 99 500 6 9 2 10'. 10 12 24 14 204 101 5000t 0 6 0 6. 1 11 0 16 236' 86 &functional 0 9 10 9 6. 8 13 19 18 163 97 fie ot (Fi 2 0.5 p n s g. 1. 5 8 7 11 10 II 8 6 5 7 7 12 12 11 111 19 17 152 157 99 86 50 8 9 6 6. 9 11 9 15 162' 75 500 10 6 7 6. 8 12 7. 15 167' 71 50l)Ot 7 5 5 5 1 12 0 13 194' 74'. •Ihese.values resulted in toxicitv which was apparent as a thinning ofthe background lawn. tdues aremuns of four plates except the control values which are means of eight plates. In all rases.the standard deviation was within 5-10%;.of the mean. Values marked with an asterisk show a significant dose-related increase rluncheere tesq in th'e.number of revertants tP <0011. ~ role in masking a mutagenic eNect of phenol. Table in the presence of S-9 mix with various protein con- : shows that for the concentration range shown centratibns no phenol-induced effect was observed, 'rW00pg/plate);nomutagenicactivitywasdetected, other than ihereased toxicity at much lower doses N'hen the procedure.incfuded preincubation of phe, than was apparent from the plate incorporation assay " •ith the bacteria for 30 mfn at room temperature (Table 31. Table 4 shows thavevemaftcr including the
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2084 A. hwi. A. InA,urui i u mrd. I'. I'. Pwsu~.r Other constituents of essential oils, such as guaiacol,, menihol,.oil of Piirus plvnilio, a- and fl-pinene, are inactive on thepentobar6ital sleeoing time (Table.l). Tables 4 and 5 show. that bratn pentobarbital concentrations are also unatfected when some of these compounds ane given subcutaneouslyor by aerosol inhaVation. The results reported in Table 6 indicate that when eucalyptol was administered by either subcutaneous injection or acrosol inhalation, 24 hr after the last administration, the activity of liver enzymes present in 9000 g supennatant fraetion is significantly increased. On the contrary oil of Pinass pwnilio,, thao was inactive in Ihe cxperimentswith pentobarbital (Table 5), was also iitetTective in rirru (Table 6). TARLL 6. Treatment Way of adminislralion p-Nilroph¢nol (mpMolc S.[.) 4-Antino- +m(ipyrinc fmµMolc I Sli.) p-Aminnphenol ImpFlole ~S.Ej. controls s:c., 225 Y0 190''-0 68 -5 Eucalyplol s.c.. •5(M1. 59: •J73 : 7,. '227 Ifr controls :Ieroull 242 34' 280 . 40 148 - 16 Eucalyptol aerosol '1?0 -- 34 *534 - 46. '2'0 -'- 16 controls aerosol '-65~~48 341 77 83 r: 10 Oil'.ofplmrsPnnriliu aerosol 278 i38 257 31 70 10 Eucalyptol was given subculancously 1500 ntg/Ggldailv for 4 days. The aerosolircd drugs werc given daily for 4 days during paiods ofl5- 154 30, 30 0 nun respectivety. The amounts aerosolized were 50 mg/min. Controll rats received arachid oil subcutantt+usly or acrosolizcd. The ligures are the aversge of.siit determinal ions: Determinations wcre.pcrformcd l24 hr ancr thc Iaut adminislratiom, • r <Or.ol vs. controls. The figures mean.the mctabnlltes formed (mpmole/g/hr) frum p-Nilroanisol. aminopyrine, aniline respectively, used assubstrates by 9000, superrrarxnt of rat liver:, inducers of microsomal enzymesviu the inlralalion route of administralion. hydi'ocarbmu constituent of cedarwood. sucfi as cedrol and cedrene, are elTectire may be responsible for the induction. More recently Wade cr al,n showed that volatile It was suggested that ingestion or inhalation of compoundseontained in softwood reported by Ferguson." may be dependent on aln increased microsomal cnzyme activity displayed by these animals as compared'to animals kept on hardwood beddings.8 to barbiturates shown by mice or rats housed on softwood beddings, previously by mucous membranes of Ihe respiratory tract. The possibilityahat terpenes are enzyme inducers had,bcelt suspectedby'Vesscll%llc dentonslrcucd lliat incrcased tolerance is particularly used in practical medicine because af the high absorption of these drugs when ilis given byaerosol route.It'shouli3 be.notcdth:u (tikkind ofadministratiom The results reportedd here, shcm also that eucalyptol induces the microsomal enzymes pentobarbital: enzymatic reactions and iu riro on the metabolismand pharmacological activity of microsomal enzyme systems. This effect has been demonstrated itrrirrn on different terpenes,.alcoholsand phenols, tuealyptol wasfound to increase the.activiiy of the In a number of components of essential oils, showing.differcnt structures such as DISCUSSION Our results pounds such a These data therapeutic pt: theenvironme Acknou•ledgerncn . The technital as I. L. G. HART ar ' 2.L.G.HART,R 3. B- B. BROOIC;: 7her. 109, 26/ 4. R. KAro and . 5- D_GaasRr an 6. H. C. Frncust 7. E. S. Vrs[a, : 8. E. S. Vrsrl.c, l 9. A. F.,WAnF,J ]I7(1968).
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Citn(U1oSOMEFn3Galilr.c'rtuY -t1i5 in Ihe series. (: 7011-200 cells Nvere :malsscd in ench ufcunf•.enlndions }-I:S, only in 4-7 have single fr:f.rmcfns l,rf•n rucowdurc.d: Cunavnlrnlii,n A'u. : .l r ./ . T1 (1 hours 1 ~ I. 2 r. l I day . . ........ .. _/4 I 4. Hgdroquinone. - rf: 1-ili. b: Ruuts ~att: 1--'S 14 hoursl,, 3(? Jnyst, 4-fJ (3 davs)'. 10 (5 dafcsI. v: C'-Imu<arrs in 7---l/i. d: Lren in I nol inslaut:fnrous killing,,liul strnn;; al,l;lulinadifrn in, 1--d'. r:.C-mitosie in I--S, c-tcudcnf•ics.flown to 8. In 2 L1LC r•-fnitosis iv nl' hall-I.ypc. in 3 oC bnw-I;'pc. I;nncenlm- Rumber nf frn;;monls prr rell Pr J .............. :IO I) IO li.. .............. 7 75 73 17 Ii 4 i 2 ' I I: - -- 1 25 •r7 ................ 8 .............. SI ................. li 76 7 1.1 l B 5 { 1 :1 y .. . 1 -- 1' - . _ I {7 21 ill ................ S:i 8 J - - = :1 17' 11 ...... ..-.._... 4-4 °-~ 11 .. _ 1 li I, 12 12 ................ /Ni 2. 1' 1 4 1:1 ................ 1110 _ . 11. 3. PI/rr.,f(nllol. -- rY. Concf•nlr:f]i::nisorir,: 1---,LL h: Iluuls .ul'I: I--1 I~-t huursl: o It, dn)•I, li 12 Ja}'ti1.. c: t:~luulnurs xrallcrrdin :F-Id. d: 1--'2 show slronti,:l;;nlulinntinn, y: li:r. iunltlcd chrnnrosntnrx. :4-5 haveIhr peculinriiy Ihat oslremcl}' Il•w nn9,iplrawa :ind nnnphascs While mam' prophaxes.:vrs presrnt. •1'lii.. maV Ix• ranscdbc nu ur+c crlls ;;piuo, into mulnplf:uo dtnin~ lln• Irr.dmr•nL .cllilc all nId np.'fulru•. SumL...rvmr Ir .nl. l:.~r,rif .mu Su- II I _ .. 1 .~ h.._..--..._.... :II o. 7 2 .IY. : ...... .... ..... 33 13 10 4 ' I 1 - - '}Y il _-. . . . . -. . . ... . 63 .i/) 29 lii 12 I./ a IIU :I ................ 63 17 7 :i 1 I 1 :I7 10 .......... .... lla 5 1 1-- 7 I I 38 3 I' 22 1~~ 74. I? ...... . ._ 200 - . 11
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; thetusuel ' Or in water e the ability liscibleco- lerating in ng are ap- -ogen (and selectivity,1) required nin to le9s t guaiacol etic meth- 1971) but t yields of tual lignin 179; Jegenr, f aromatic y fission of d methane !r phenyl- 3recursors id Ceylan, :r, present -ing lignin d Gavalas, night react Id thus be energetics lechaniatio ~xplain the ticetic ob- cin e n terms of e 8a. The phenoxy- which can I catechol, colradical idical that 3ortionate 3a to form 1 of Figure aiacol (G) ,and char i for neat If water is 0 occur in ; solvated ldrolyzed ecessarily ndergoing tccesaible nebulous e solvated chol and oa 0.1 0.9 I .. IMecYon T - te mn 'IIICr@~ p~p'P;'. o ro ~ ~ r 02 aee°wrr -s0mw a, I I ~ , . . ,1 0 u.A ae u w 2 2.4 Figure 7. The dependence of gueiacol eonveaion on reduced water density for Pyrolyslis for 15, 60, and 90 miu at 383 eC. methanol.. Pseudo-steady-state solution of theaetwork shown in Figure 8b was accomplished by: (1) considering the salvation to be rapid at reaction conditions, and thus allowing guaiacol (G) and solvated guaiacol (G') to be in virtual equilibrium; and (2) recognizing that gas chroma- Wgraphic analysis at ambient conditions would yield only the sum of guaiacol and solvated guaiacol (G + G•) as the observable guaiacol species (G°): in kinetic analyses. Rate equations for observable guaiacol (G') disappear-mce,. and methanol (ME), catechol, o-cresol, and char appearance that explicitly account for the effects of water are shown as eq 6-10. -d(G°)/dt = ((3k1 + kHKWh/(1 + KW) + ks [kt/ke(1 +KW)G°jt/Z)G° (6) d(CA)/dt = [(kHK{Sn + (1 + a)kt)/(KW + 1)1Q°~ (7) d(ME)/dt = kHKHnG°/(1 + KW) (8). REACTION ELEMENTARY STEPS OR ~o~ OR ~OCN3~~ V• + •CN3 421 •[Ht. + nQ'aN _y ~yaN+ rH.. OCN~ l:•nCHq' 17/ a0H + °~00CM, - V-OCHt+ G(OH (,) IOI aN+ 1i100CH3- a~Hr~ ~~t5) v. (YaN - ~'0H + Ca.+ Nr lJ.ocNr v Ind. Eng. Chem. Fundam„ Vol. 24,.No. 2,. 1985 207 d(0C)/dt = OkIG°/(1 + KW) (9) d(char)/dt = yktG°/(1 + KW) (10) Note that methanol and catechol rates both increase with water concentration and that o-cresol and char rates both decrease. It is interesting that therate expresaion for guaiacol disappearance can be in agreement with the conversion data in Figure 7. Recall that these data show that guaiacol conversion decreased with increasing water concentration at low levels of water concentration, reached a minimum value at intermediate levels of water concen- tration, and eventually increased at high values of water concentration. Note also that this behavior is like that expected from the formal superposition of a fission-con- trolled free-radical reaction pathway andd a bimolecltlar hydrolysis pathway. The rate conatantfor the former pathway would have a.positive partial molar volume of activation (Eckert, 1972) and would thusdecroase with added water (increasing pressure). The rate of the latter pathway would always increase with water concentration becauae its partial molar activation volume would be negative and its rate expression would be positive order in water concentration. In brief, the candidate mech°„ia..s of Figure 8 and the implied rate eq 2-10 are in qualitative agreement with the present experimental findings. Conclusions We conclude that two competing reactions occ(:r during guaiacolreaction in water. The first is neat pyro:ysis that leads to catechol and high molecular weight material as major products. The second is hydrolysis of guaiacol to catechol and methanol, the selectivity to which is a con- tinuous and increasing function of water density. A superposition of free-radical and hydrolysis reaction mechanisms, the latter invoking an ill-defined solvated and caged guaiacol species, allowed formulation of rate ex- pressions that were qualitatively consistent with the tem- poral variations of observed productyields. These results suggest that hydrolysis reactivns may contribute to reduced yields of char and high yields of single-ring products during the "extraction" of volatiles N //~y OXI NO~, OR ~ON (6) 2~ -'p1 101 f~ e o I +QIOI +rNYWM aCNr• v~OCXr CNr-O^/ J 0 ~/.N (a) Elementary steps for neat quoiocoi thermolysis aN a nNqa ocHr ~ _ L H sN. ox a + CH OR CHtON T -~ Ns°1 eu°iacal T1,ermalr,is, °e I° (°1, ae°.e (b) Superposition of neat quaiacol thermolysis wi4hh quaiacol solvotion and hydrolysis Figure S. Elementary steps for gueiacol pyrolysis neat and in water at 383 eC.
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it pyrolysPe m guaiacol :h) in a pu- i,conatant ,ctora were :inga. The wne small ntative ez- ,e internal amount of ictor. The - ath a 1-µL nL syringe. ; confirmed sealed and nd, after a ly removed vater. The rature and .or was op 4y4mLof : and iden- s collected ued. A 50 qillary col- :Y nNed for L i.d. boro- n 100-12D of all com- xbing cat- ed column rt was used imple with malyaea of matic ring on 1. (1) at 383 °C, iatic rings action was ation and ,). 201, cresol, mounts of of methyl s product -e 1. The Nith reac- nate of a ai 0 e M ee 1e M W ee. lo p reOG e..oion fY.w ~+. riaNm 1. Temporal verietiau of the yielde of major producfa frem eeatauai.col pyrolysis at389 •C. " Il.,oem ni,r. nin Fk~ure 2. Temporel vuiatione of tbe yielda of ine)or paodecte fmm rustiacol Pyroiyeie in water et8&9 •Cand reduced density of 1.6.peeudo-fira4order rate constant, k= 0.0198 min r. The principal primary productwae catechol, the molar yield (mol of i/initial mol of guaiacol) of which increased with reaction time to em asymptotic level of about.0.90 at a residence time of 90 min. Yields of the leaeer of the major products, ocreaol and phenol, also inaeesed with pyrolyais time. Phenol showed no signs of an asymptote at 90 min. Methanol was observed in only trace amounta. The influence of water on gueiacol pyrolysis is illustrated in Figure 2, where the changes of the product spectrum with reaction time are shown for gueiacol pyrolysis in water at a reduced density, p,~„ of 1.& The continuous decrease in guaiaool concentratton corresponds to a leaet-equaree estimate of a new pseudo-first-order rate constant of k= a024z min t. and methanol were primary produefa. with theasymp ttc molaryte of atech m ' e 2 being increased over those in Figure 1 by approximately 0.25. The yield of methanol in Figure 2 is approximately 0.4. It is interesting that while the aeymptotic yielde of inethanol and catechol inczeaead grea~tly-wltli f t]-e sldl~n-Fo water, the te~eeol redchar.t!_ecreeaed311d-th o guatacol and phenol wer sentl~-oec[~an e~ fie-i:oTal`ylefd'of emg~rmg phenolica m~ igure 2 ia approzimately 0.70 for a reaction time of 90 min. Thisia a substantial increase fromm the Ind. Eng. Chem. Furtlam., Vol. 24, No. 2. 1985 205 . . n..rw. n... w,.: Pigere ]. Ttie inlluenoe of weter dendty oo the temporel verietinm of atechol yield from pyrolysis of `uaiacol at 383 •C. . 02 aa u w ai e b al: m0 m M A 0 W N em .,.rw.~,,:. Figere 4. 7Te intluence of waterdeneity m the tempural varietions of inetlund yield from pyrolysis of'ueiacol at 383 •C. e.eew.nr.+r. Figare b. The inlluence of weter density on the temporal verietinu of ~creeol yield from pyrolysis of guaieml.at 383 •Ca yield of 0.35 illustrated in Figure 1 at the same reaction time. 8'7119366 fi p;; l;
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0 rrrn.~~ rv1 . ., nr_ .1. ws2 nm,A m< x n.^m. nu m: ..n ° Grt4-cc-v-<~ ~, 143 (41 /YOSa,.pqgll,Unn'. l'op,r n P~V, 1asqm P.. Litl. V/' MUTAGENICITY TESTING IN THE SALMONELLA TYPHIMURIUM ASSAY OF PHENOLIC COMPOUNDS AND PHENOLIC FRACTIONS OBTAINED FROM SMOKEHOUSE SMOKE CONDENSATES B. L. Ptxtt and P. Z. LIN' Insli/mrul liaaiinlmlrnmf (7rrnxrlhrrupll Grrnrtn CnnrrrRrstxrrch (-r+nrr- 6n Ntxrenhntirter Feld:.'/n; M19ta1 Heidrlber4- Federal, Rrpnhliu u/ Grrmanc t Rmrired.. 6 Orrorhrr 19M) . Abstract Smokchouse smoke, which is used for Oavouring mcat products,, was investigated for its mutageni< aelivityy in the Salmmelhi rt•phinnvium assay. We were chiefly ooncernedwith, theiractions free of padycydic aromatic hydroeartbns.hxt rontaining phenol eompounds. which are responsiMefor the preservative and aromalizingpropertie.c of the smoke. The mosrt abundantlyy occurring phenol compounds tphenol. cresols. L4-dimethylphenoll brenzcstechine, syringol. eugenol, vani0ihe and uma- coq gave n rauve results.when the were tested for muta enicit 'a five concentrations un to 511/ Tte, wn and wilhaur 5-9 mi_• usi: _nm --ium. Even when phenol wass further invesu gat rn avariety~of tesl condtuons.no induction of his' revertants was observedi Whemsmoke- house smokee was condensed and fraclionated the majority of the various phenolic fractions also gave negative results when tested at fiveconeentrations using five strains or 5. rephimuritmc However there was a slight increase in the numberof revertants in a:few cases. The presence ih the phenolic fractions of very small amounts of mutagenic impuritias, the.nature of which needs furtker, investigation, cannoo be excluded. These results support tNefurther development of non-hazardous smoke-aroma preparations, based on the phenolic components of smokehouse smoke. INTRODUCTIO\ 1n the Federal Republic of Germany, a relatively large number of foodstuffs are subjected to smoking for preservative and',aromatization purposes.:The type of wood used to generatelhe smokee is mainly depen- dent onn the aroma required. The only restriction is that the wood should remain natural and not be treated'with glues..paints, impregnating or other prep- arations. During the process of smoking,.the woods arefirsfignited'and then, allowed to.smoulder at tem- peratures of 400-500-C, although higher temperatures up to 800~C may also be used (Toth; 1980). The -«sdtingsmoke contains at least 300 different com- pounds, mainly phenols;, carbonyls, acids„ furans, alcohols, esters, lactones as well as PAHs (Hamm, 1977; Mohler, 1978). The proportions of the iionstitu- ~eatsdepends on the type of wood used, andd the pro- -ress of smoke production. The PAHs are, however, well known for their mutagenic and carcinogenic properties (reviewed in: Brookes, 1977;Eiscnbrand & Wiessler, 1981). Therefore, much effort is beingdi- ncted towards the elimination of PAHs from the smoke aromaslo avoid unnecessarycontaminationy of food products:.Since the preservative and aromatizing properties of the smoke are rnainlgattributable to the _ phenol-containing,fractions, the enrichmentt of this fraction is one of the first goals in the development of aun-hazardous,smoke aroma preparations. 'Originally from Peking.Peoples Republic of China, holder of Alexander Humboldt Foundation Scholarship. .A6breuiatiam- PAH = Polycyclicaromatic hydrocarbon The genetic toxicology data on phenols, derivatives of phenol or even on the phenolic fractions fromm the smoke condensates are sparse or unavailable (Dean, 1978): Some phenols have been shown to exert; car- cihogenic or cocaminogeniu effects in mice (Boutwell & Bosch..1959).and rats (Boyland. Busby, Dukes rt af. 1964) after topical application. The most elaborate mutagenicity study was performed by Levan & Tjio (1948a,b) who observed that 40 different phenols and related compounds were weakinduacers of chromo- some fragmentation. Only benzene, which is thought to be metabolized tophenol, (Cited in Dean, 1978)J rescorcinol. phenol, cresols,.2,4-dimethylphenol and eugenol (Florin, Rutberg, Curval & Enzell, 1980;.Gil- bert, Rondelet, Poncelet & Mercier, 1980; Gocke,. King, Eckhardt &Wild, 1980; McCann, Choi, Yama- saki & Ames, 1975; Shahin, Bugaut. Gilard & Kalo- pissis;. 1980) have been tested in the standard Ames system using histidine-auxotrophic Salmonella ryphi- muriurn strains, with negative results (except in the study-by Gocke er aL (1980) who found phenol to be mutagenic towards S. typhimurium TA98). No other data on the genetic toxicology of the phe- nol compounds found in the smoke or in the smoke condensates is available. We therefore investigatedphenolic smoke fractions and isolated phenols for their potential mutagenicityin the Ames S.typhimur- fum assay. Thistest is well knownn for its sensitive detection of mutagens and therefore possible carcino- gens (McCann & Ames, 19761. The aim was to pro- vide preliminary information on the possible biologi- cal activity of thisclass of compounds.and toaid,their evaluation. (PooI,J980), rcr.p//~ 383~. Gz„&l-a><
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176 L. L..Grasonnrv Thcy were killed as described above and the percent- Other cum rur s eliCilinj~_little cli:mgr_ from,, - ages of wet and dry, liver were determincd- BFSLL75 Srrhnnancnux injection oJ ayrrrrs impurtial7yn hrparerro- sni.rd'rats ' Positive findings in the liver following sc injection off very highI dosess of oils, flavouringsor principles on the first 7 days after surgery appear in Table L Of the oils, nutmeg (series 2E and 9E), mace, fennel: celery-seed, anise,. cumin, parsley-seed and tarragon (series 3E, 5E,.6E, 1E and 8E).stimulated liver regen- eralionn in male mts. The principles 4allylanisole, 4propenylanisoleand p-isopropylbenmldehyde, screened at overall dosages of 1335,. 2450 and 1250.mgikg• respectively;.increased the extent of liver regeneration; as did safrnle, isosafrole and dihydrosaf- rule injected daily in a dosr of 10-15 mg(rat (overall dosage 295-375 mglkg): piperine (460 mg/kg);I sassa- fras oil (1130mg/kg) and a high safrole-containing fraction of camphor oil (1240mg/kg}The incremental increase with the last agent lacked signifuanoe at the 5% level of probability: The activity of tarragon oil was compared with that of several aromatic compounds (series 15E1.. 4-Allyl- anisole was very eMective at. 1160rng/kg (P < 001) but the increases in increment with tarragon oil at a comparable level, n-propylanisole (3440 mg/kg) and n-propylbenzene (4670 mg/kg) ) were significant at the 5;a level of probability. Allylbenzenee was' a more marked stimulant thann-propylbenzene, whereas J{-methylstyrene (2330 mg/kg; series 11Ej, and com- pamble levels of cugenol, isoeugenol, 3-allylguaiacol and 2-allylphenol were inactive. Allyl phenyl ether proved, an eBective compound at an overall dosage of 2260mg?kg. When tested in adult female rats (series 10EG 4-allylanisole (1675mg/kg1; tarragon oil (3450mg/kg)'I and parsley-seed oil (1640mg/kg) elic- ited significant rises inn liver inr.'rement and, again in linee with observations in males, carrot-seed oil (2460mg/kg) was without significant acflom. Of the azuleneconmining products (rable I; series 19E), the increases in. liver increment with guaiene (2l'35mglkgk Haitian vetivert oil (3330mgJkg)I veti- vcrol (2170mg(kg) and one blue chamomile oil sample (H-I ;. 1100 mg(kg) lacked, significance. How- ever,, in several preliminary runs, two other chamo- mile oils (H-0 and. H-5) and guaiazulene, at similar levels stimuhrted,the regenerative process. rrgenerutiom trol vn ucs rn the extent •_.reguwtation._IdL , given rn an ovem dosage of 24_ 5(Y_'950tt1gr6g eta-•: whcr- a mrLcatcrl otherwise. - wcrc the laryit., ,. eadinenc, camphene, evrvone, caryophyllerw- cinr+e - citral,. citronelloL. dipentenc, famesol. germliul. ku- t. roxycitronellol, linalool. trnnsp-menthanm. . L-m.r, . thonc,, myrcene, x-phellundrenr. x-pinene, rhoJina- x-terpinene• y-terpinene, x-terpincrol, terpiholenejcc wood turpentine,. as well as 4-allyl-2,6-dimahm'•- ~ phenol, allyl isothiucyanate (g0and 120mgke'ez - latter being.rather poorly tokrated), anisole-J-cumi ' pheno ucol,.gttairne, 2-hydroxy-4:and 5-m.tihae•- t benzalde y es I200mg/kgk 4-nrelhoxybcnzyl ak; hol. 3-)rmethoxyphenylprop•anol (3750 mgkg} ptir:: . lole and piperonyl alcohol. The following oils a.;. similarly without significant effect when injectI , ~ an overall dosage of 2400-2950 mg/kgor as spe Abies sibirica, bay, bergamot, cade, wjapuL canaep ` .: cardamomseed,m cassia (1100mg/kg), ccdarwlI eitronella; chenopodium (25 and 125 mg,kg4 the L:. being rather poorly tolerated);• clove• copnih.. ::r- . iander (oleoresin); cubeb, davana dill, eunl% ptu;•. L~ anium, ginger, guaiae wood, juniper. kmon• rl~ joCUn,m neroli, opoponax, origmrum, paprika ta: resinh patchouly, pennyroyal (1300mg.kgt ).i'!r • (bluck), peppermint. perilla, petitgrain. pirts: essence berries;.pimento leaf• roscmary. vee. u:.',:--' ~ wood,. spearmint, thuja (14f0 mg/kg). thyme. ~.:- '. wood (1200.mg/kg) and ylang ylang. Body-weight losses were fairly high in mlinu:, ;; jected with parsley-seed., tarragon, sassafras ar:d'-; Sr • umphor sassafrassy oils at the levels inU;:.- 3(1-50%of the rats being excluded from con$A•rsws : ~ Several of the terpenes, oils andphenuls were rqc- sibk for extensive skin necrosis, which nti~--{ ,a m5, the shifting of the site of injection from .u% Admrnisfration of diets suppfentrnred tritL,o::.. *.-( ciples and plant tnateriafs' to puriiull r irrryr.,:. rats The feeding of diets containing chumonuld..,r: ~p ~ safras-bark teasat both 1-5 and 7-0°v blue Hcc_-.: chamomile oi1s,. H-I (&20",01: H-4, l0'20%), guaiazulcnc (420,'a/. ptaoprop3lbrnt_:,::~. - Y (0-35%), celery-serxf and parsley-m.d oils. :...= . 660% and ground nutmeg (6-0.".y)) to ncrli ..-~ increases in the liver increments over a days(fa61e 2} The Egyptian. Ronwinc ami E:_.i chamomile oils (each at 0d54o), ground .vr:_ _. Tablh I detion of dirts's sapplennrntrd with essentiaf oils rmJ principles midcLarmmrlle and sussurr,rs ;m,I ,, :- 1c Tcst material , and'dietary levdt 1";.) Body'weighr (g). No. of rats Initialj Terminalj. liver iiwra-mant iet 9eries.`U[ Conuul IS127f5d 266±72'--IV4±UUMt Ch:mwmilctea.(I•5) tir..dr,u h~.rrk tca (}0) Sriu.. ,IL. r,n,n„I .un...diel•ntl.dll 1 .,.m. i nt~ , 13 232±5~5 271±4~4 2-l+dl+UII' 10 230±6-1 250 ± 6-7. '~743±UIIc 17. ~xtl' 1 6-b 311.i F\ll. 2-1_6 ? tltlaJ It '1+ I tr.1 11111'7'1 2'4r4±rr113 n I r•..I "I I\ t 'u"I 11 L Iu1U t rY.tll `r1 1 uorn S7119345
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i 178 L. L Gcxsnneuv (6y%) and Bourbon vetivert oill(0•35%),were inactive. as was tarragon oil(050%): However, Huitian vetivertt oil (035and05(1"y), vctiverol (o50y,;1 and guaiene (0-40:and 050%) increased tlieextent of regeneration (Table 2). Other agents having no.significant.eHect when fed at the dietary levels (",;,) indicated in: paren- theses were allspice, Jamaica (6-Ok alfalfa seed and leaf tuts (7-0 eachk cardamom, decorticated (6~0), car- damom-seed oil (0,60), cassia oil (060), chia-seed tea (70), chieory'root, greenand roasted (7~0O each); einna- mon(t2•0k doves (5-0), fcnnel tea (7:0), fenugreek-seedtea (7U); garlic, instant powder (43), mustard seed(3,5k onion, instant powder (45), poppyseed, Dutch (7•0), sesame seed (9q), saffron, .. Spanish whole (1-5), sarsaparilla, ethanol''extract (2-7), tea,.black (7-0), tur- meric, oleoresih (060J; and yarrow flowers, whole (6.0). Diets showw to be active in themalewere also checked in the femaic rata Thus, chamomile oil. H-I (020°-„), nutmeg oil (0-60'/) and safrole, isosafrole and dihydrosafrole (each at 025%) stimulated the regener- ating liver (series 32E, 33E, 35E and 46E; Table 2). Effect of agents on tlre fnracr liver of rats of either sex The se injection of parsley-seed and tarragon oils, 4-allylanisole and p-isopropylbenzaldehyde at overall dosages of 1300, 2550, 1280 and 1215 mg/kg, respeet- ively, to intact male rats caused marked increases in the ratios of both wet and dryy liver weights to body weight (series 48E and 49E; Table 3) Liver enlarge- ment was also noted in females injected with oils'of nutmeg (3200'.mg/kg), cumin (3100 mg/kg) and parsley seed (1400mg/kg) and withh the blue chamomile oil H•I (3100 mg/kgk but even in'simihrrly massive doi-. I eugenol was' inelfective in. rats of either sex. Tz~: : 3 also presents data for immature males fed ch.tm:-' mile tea and safrole 1025°06 Only'the lam: i diet caused rises in the liver weights. Microscopic examination of liver sections remm:: i from treated groupss of operated and intact tr revealed few notable diKercnces-from, the corresponi- :~ ing controls, ; 1)18CUS91ON Most of the essential oils, flavourings and pbc , materials included in the present study were comp.' cuous for their lack of any'signifioant effect on d: regenerating.rat liver. These.Bndings are understa.d able when it is considered that the maib matrix nm, prises terpenes and that such agents, when tested ind> vidnaBy in massive repeated doses, were found c• have little inNuence on the regenerative processIe variably, when a stimulatory response was eviden: the behaviour was due to an aromatic principle sua as 4-aBylanisole,.4-propenylunisole. safrole and isos;- frole, among others, although components such i eugenol and isoeugenol had no such effect. Of the oils producing an effect on the regeneralir( liver, tarragon oil contains 4-allylanisole as the rtuc aromatic principle, and significant amounts of Ihi, ~ compound occur also in fennel-seed and anise 66 4-Propenyhtnisolc is present in fenneNsecd oil at; levelrof 50fiU°ioand at even higher levels IS1F94r in oil of anise (Bouchardut & Tardy, 1896) Oilsti nutmeg and mace are very similar in mako-up en.; Table 3. Bodp mtdlicer-weight findings for intuct rotr gioen duily ., ' injectionsoJ' essa+uiul oils for 7dnys or Ji-d'dns • supplemented.with chamondle tea or sufrole for /0 days Oil or other test Liver weight material and dose R ll Body weight (g). (g/(00g bodyweight) g overa ) (mg(tatlday:mg/ No. or dietary level (%) of rats Initialt Terminalt Wet t Dry t Series 48E (x)-males Control Parsley-seed (50: I3001 4-Allylanisole (50:.1280) Eugenol (IW: 2610). Tarragon (1011; 2550) Series 49E (scy-toales Control rrlsopropylbenrnldehyde (50; 1215) . Series 49E(set - females Control Cumin (100:.310DI. Eugcnol (1(lli; 3200) Nutmeg (IOU: 3200) ChamomileH-1I100;.3100) 12 12 12 11 11 12 12 12' 12 12 12 12 245 ± 64 245 ± 4-5 254 ± 5A 252±51 255 ± 5-7 264381 271 t 7-S 206 ± 3-1 214 ± 2-8 212 ± 29 208 ± 27 210±34 295 } 5•8 286 * 6d 296 t 69 294 t 118' 291 ~ 6-I 297~112' 307 t 101 234 ± 3g 231 ± 3-5 226 ± 27. 223.t 27. 239t20 4343 ± 0,103 4754 ± 00'l2 329" 4694 ± 0-0s2 268- 4461 t(y085087' 4931 f 00g84.29^ 3931±0058' 5278 i,' 0903'. 1142" 4207 ± 0096 5371 ± 007g9-46" 4i925 ±0069 1 IM) 4752 ± 0096' 4(Yl" 4'5'_7±0073 274' 1248 t 0026 • 6391 t 00]9 3G" • 1u39-i t 0034 340" II324 ± 0U29 i9! 1'442 ± 0-018 606" I138±0-0P0 • 1538 ± 0024 I'LR' 1 216 aJ 0032 1552 ± OU3n' 78+" 1221 ± 0026 01: 1314 t W33'1 . ' Seriia SOE (sc)-rcmaies Control Parsley'seed 150; 140(I). Series SIE (dietf--males C'ontrol' Chamomile tea (15i )' Sufrole (035';;,I 8 8 12~ 12 12 240 ± 59 240 ±28 97 ± 18 98 ± 19 100 + 1.5 252± 6U 261 t 34 157t 3'8 145 t 4 0 147 ± 37' 3-844 ± OtN4 4371 s 0(160 4-71" 4500 ± 0056 4 660 ± 0122 119 5,222 ± 0109 54)"' 1/95.± 0032 1319± 001'_i'I" IQ08t(HI'-0 , 1-296 ± tyWB Itd 1-479 ± 004g 5;1" - tlnitial weights were.recorded prior to surgery and terminal weights at autopsy om day IU .. Results are means + SEM for the numbers of rats stated. Significant r values aremarkcd with asterisks: 'P Qn: ••P c001.. ' S'711934'7 vntain ! myristici to wide be prese: dimerho; I-alfyl-2,, and my agents r.. stimulac individu contains or cumi study, t stimulab buted in fms bar' derivali• on the I low lev 13671) m~ Ther uleoresit ating lit the terf which s Table 4. Compot Type: +allvla 4•Prope 4Fn-Pro• 3-f.Met '--All)l(• 4-CumS 4-Allvl-. 4-Prop. 4-Meth. bHydr, 2-H.dn Anisole ,.Meth '-Isopr AIhlhc: jt~Meth n-Prop' AIly1 p: Type: S+frol: Iws::fr. Dilndr Piperin Piperot Prperot ~ 'IDe d
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31.ItP.i1'1' LD:\'AN .1\Il J1)E rllY 9•JIU dilulion series of IItc ,ulrvlauce In be tested is prepared,, consisliug of IS--14 con_c_enlralinns rurorinq, the rc;;inlt 11,1or, in some r.tses,.. I--OIYxI'dNl'1 I1101/I. SI11:dI .!{lIIIIn r:r•nn llulbs. ol' the "/.ilt:mrr vuriety,, vrbicb havce bt-en ;;rne'unn Inp-11'ali•r until their runls arc nbonl 1 ctm in leugth,, nrc Ihen wurrd rcccr lu these snlttiinus. Thce nlols arc ob-. scrved ntneroscupicalh' fur drnnrnslraliun ul' tnzicity; in Ihc highest c.uoecJltrntiuns Iher .rflen In,r lhcirIa1V~or. After one dan• in Ihe snl- Idiuns Ille c-lunnlurs Il.lscAs. I!IaRIi bc;;in It, appear. Cytological fis_. atiunsarc made nn Lcno ocrasiuns during the trenlmeuf, viz. aflen un rcposurc nl 4 :uu1 24 hours. 'I'brse fisatiuus supplynl:tli•ri:d for the anah'sis oF c-mitnsfs :utd clu•uulnsurnc fra};mrntalivu. In the presrnl paper few tlr[:lifc are 1-livrn ol' Ihc mnrpUolttgyy of Ihcsc reactinns..l!le.aim being lul rrrard Ihruceurrencc ul' frngnlentinR activily amonl-, tho clwulic:d.Icslyd.llu.ccvcrk sonlc general cylnlogical feadures characteristic of thcse trr:rllnenlv will be Izrieflyy described inn this Fil•slt choplcr: In Ihe slrnugrsl conccntrutimis toxicity is usually high, and thc4issues.arc inxl:ultlpIcilli•tl Ly the Ireatolent.Cnnsequenlly, in this first eancenlr:dinn zune Ibrlus• called zone 11. . noo c-nlitoses and no anaphase bridges are Ibinld. A cerlain uliililnanr of cilal processes is nor.t_ssan• il' Illeac reaclions are tu br m:mifesled.. '['he dring off oftbe iissue is often very ehuraclrriaic inn its t•.ylological type. Varinnss pllrnuntenn Couuected e'illi Ihe sn-r.Jled.:r;ghdination of tbe citronlalie nlaleriod prcdntuinule in zone IL, bui nlar also rcach Jue.n f:v belolc.. Ihis Icthnl.zunr. "I'he nlalrix uf Ihrclu'nmusomcvv is liquefied and has :t. ' Icutlt•ncv, to Iluwuul the bnrdrrs ul'plnsnla vacunlrs ur to be Iwupcd togcllrer inlir unr or uunrt• bumuticncnua.. chrmntuuurc cluslcrs. 'I'hcrc plmunnlrna nrc ullrn rufrt'rvd !., as p1'cnusis, and according to Ihe cm•rt•nl cclu-clicnlicall r.irw Ihty :Ire. Iluc to depolcmerizn!ion of nucll•.ic acids. 1C1Lilr Ibo pccnu!ic Irpc of rluln'1' cs inulliwiti•d cbrunlosunlt•• nlurpl4ulo/;ieulintcre,L ulher ch:ut~es uccurrin;; in cnnsequcnce o@ tlle liqurfacliun of thc cltt'nlllulic ul:dcl•iai are of greater interest. Often Ille .Ihiu:lblr subs!nncr i. ~cnllrWa.hrd uu[ Pnnn Ibe c6ronmsonres. recc:d- iil;; :r delicate intcrnal slrueturo: :Cllhuu;;ll ILe chronwsomes.arc dcld. ILcse slrtlclures mae be uf significance fur Ibo- interpretation of1'ilal ..Iruclures. Some fcalures ul Ihr cliromoanmc nlorpholu;;}•y mav 1!e up= prnacbable only a.l'It•r the cllrulnusunles Im.ve becn killed under Nvell- dofined cundiliins.. Tbna. in sumc ul' uur Ircututeuls the subdivision of tbt chrurnusumcs iuloo hulf-eln'nm:didss is cen' clcarh' strn- Anolhcr Iv.piral rc:relion lvhicb ul:lr nppt•ars :dler quYtc particular IrtminlcnlsOD _4 lli N ~ W OD C!1
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162 1611HIt'i' LI:V.\N AKII Illli III(\ 'r1111 them in stt f:lr a, Ihre iulcrftrrc vvilh nur I'raglnenling reacliun. The pseudochi>ISm.d:r mnc gircrise lu IYaSnlt•nls Ilvouyh Ilhee purely mechan- ical lensiuu rd 1114. :tnaplluw•tnuormrnl. These fr:t~mcuh are of n ch:lraclerislic appearnnce, hmcrrl•r. :md muc lic readily di.tinquishetl from our Ir:y;melds. vrhich, a, monlioned. arc nlrcady free before the anaphasr It•oainn hcts, ,IartrJ. Ili crrtnin of the cxpcrimcntal series de.crilxd' bclutc. r. ~. :tliz:rcinnntl nuphlhoresurcinnL pneuduchiasuull:t :rre of I'roquenl occarrenrr. Ili nwnl .ubsldtnctrs thry utay be found .catlcredwilhin uur zunca. "I'llr rrlatinn of nua- I'rn;;nu•nls to pacudu- chinstnnta is in untv':tc cnu.uutl. Ili pyro;;ullol.. 1'or inslance,, which is nur of lhe attLel:mcrs ..'ilh wu,l fren fr:t;;mculs. onlj• one.siugle aue.nf psrurbctliasma ha, hevu 11mud autuni, Idto Iltous:mds of ccllss that have IKen scrulinireJ'.Jln'ing Ihr cnur,r uf tdte in.rsligation. Nevertheless it is somcwh:dhnrJ nul lu Iliink thatl ttmre is n cnvnectiou tmlvreen those rractions. In subsl:mee. Nrhtl•.e I'rerfrm;menis are very rare it may hupprn that u p,rudochi:um:r is fuund. When Ihnl is. Ihee case, mu- ccprricnt•c i,Ih:d it nut• h:rv palicucr 1., studycnuughcclls unn cau alworvs u.lsu./'ind :dl:u'Lt•d and irrr• 1"rctr;lucnlc. Wt• h:trc :tclunllv uusl psrudbchiasmala.nc Irad-fm,il, • I~tr our rr:tcliun.. . Thcque,linu as In 11u.V luu„a tniuimunrlimc rdcspusure is,uocrsssn' fur Iltc• Ilvl-mrnls, tn starl. uppt•arinl., li:o Lrt•n ludibd iit a r•nulAc of 'C.1fif.hC. l~rnrynrrnlidinnF pur' crnl n/lrr r/i/J,rrrnl r.rpasrrrc liiilcc in rrr•r.u~s. rutd Irnurrnlrlrliim N, pprnqnlluF. 'llime r.l'trrnr~ I. qnr.qi 17. ILLIY tl. .\prfl L. HItV nrt in r r •dI'iu. nu nbdlou \'umLri ..I' Prvprnou:alinn L•mrv rrir. m •edrx. , 1 11111 1) ._- I II)1) ( / ^01 11.; 100 ll ~ - 2 :ilill '?.': 11LI 'S:n ?.-, _11 6:. al 131; :1:. Illli a.v I i'1 12 .: - . 4 :SJ 1 :uL: alla :r 1 1111; 7. 147 IS.I 1116 I n 8!i ?Il.; 7, 114 1;: ltl 7.:, 5' iR 3:fr a~t. :'.Lr :q) Ii1l,: :tll 0 12 :'1 -1.< 20 It
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(:nnOMOSOUI: PIUGMENTA'1'ION 455 isting of is the differential staining of Ihe helcrocLromulin dm•ineg contraction te cases, stages. Especially the proximal or pruchromosomic helerochromalinl curiely, btdl in sonre cases also Ihe dislaL end he(cruchronmlin retains Ihe slain of I cut . stubbornly. Such slainiugs us were drscribrd :d'tet- killing Ihr•e roots in are ob- mereuricnitrale jLevsN; 1945, l<J3 4i are ullun I'nundin the present hipltcsl Ircalmenls withinn zone I. (:rrsuls,. I'or instanet•. show these differential the sol- prochromosomic stainings reguL•irly and Ihrough Ihc entire meristem, ;ical fix- a•hile mercuric nilrnte only had Ihis rl'ftr.t in tile superficial layers. In after an extreme effects I1F these lylxts Ihe entirt• mctaphasr chronmsnmes with for the Ihe exception of the centromeric rrlgiim mnv ht^ diswlved. .I'rcv!> pro- chroatosotnesremainings on thc rqualnri:dl plate. •I'his effect described ology of by STe[NFOCSU and LsveN I1<Jf7h) I'ur chlurufor.oo is found, for in- ;menling 'slancc, in naphlltol Ireulmrnt.: lologicnl Often a more irregular nmltlinri of Ihe chronmsunnes is brought :ribcd in about, stainless und slained reRinns altcrn:diug.' All types occur from usually chromosomes with only :ra fewunslnined .pols mtd such with a more !quenlly, equal mullling of lhe culire chromosotne complement to those which <"t'` tttt<l - are without colour except I'ur one or Itvoo deepiy stained spots. The "uocoses `, transilion front slained' In unstained area is sharp bul the distribution tr off of of each area may be quite irrrgulhr. ilmo}• involvethe .vhole chromo- 1'ariuus solncC hreadth or nulv a section <:f it. I•:vrn if Ihe stuinlr,.s substance is hruur•dic quile Irnosl)arctrt it cun lu• evrn Ih:d Ihr <•hromosomuuutliuc iz ml- :n beli.w dantawcd all around Ihce rhrnutn.omc. Ont staining with Fctd4(•n Iltc ~ml.h:tsa .clainless.;lreas also ucquire :t red cnlour, though nfthn palrr thun the or lo br rest of Ihr chromusomr. •1•his rlTrrl is brrr:dlcr refcrrod Ib ns clustcrs. •mul1liu;,,• . unding '. lo 13clotv zuue IIht•rr conir, n cuncrntraliun ru. 7 ionn where c-milo.is. is zation of prcvailinfg pzonr Y)... This rcactiim muc te complctc in one or c%n conccntratians, :Ibocc- and beluw. .rhich Ihrrc occur mixed c-mitusrs ~unnsome- anduurmal miloses. In. Ihe fnrmvr case Ihoher rnncenlr.rliunsi !he cr idlhc prrscncr nf nurui:rl d.irisions is dm• luInck of ciabilil}-, which kct•ps t)flrn the ~n<trmnt d.icisicus Prrscrtrd 17onn briLrethr Ircettnrnl to Ilhr momrnt ut s. reveal- Iix:Jiun. E)nlc o. few ntilnsn nrrcinLle <-nuol"h (n :asutnc 19tc c-Irpc. :n-v di+ud. In the I:dU•rcuse I lutvrr cunr.rulr:diuus) Ifhe prrsence <if normal utiln.es a ul' vital <Mpeu<L• nn Ihr proximity of Ihc Lntrr 17veshold of c-mitn.is: The cnn- .a}- Ite uP- centralinu is not high rnouhh tm git-v c-milo+ra in all dlvldillt;crlls. tJl•c well- %one l moy pnrlly ocrrlnp r<tnv ?., in Xchiehh case we may get e-mituvrs livision of irilhi ntutllydd chrutmts<+nu•.. .lnuther •1-he: urxt cmtcenltaliun rr;{ion. or zone a, iv.s characterized bv rrulmcnlss nul'maI milnscs :u:dd it i.. hrrc Iltal chramosumc fragmeul;ilicmn arc
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Liver regeneratiun and.essenYal oils 179 massive doiK her sex. Tabi les fedd chamo .)nly the laitQ ngs and phnt rwere conspi- ~ etfect on the re understand- n matrix cote- .en tested in6- >ere found b eprooess la- :. was evideot principle such ole and isoss- tents such as fect. _ regencratmg e as the main ounts of this nd anise oils seed oil at a vels (80-90".,) 1896)_ Oilsd make-up and uin small amounts of safrole in addition to high isticin contents, but their compositions are subject wide variation. Among the compounds shown to prarnt in oil of parsley are apiole or I-allyl-2S- thoxy-33-methylenedioxyhenzene (Thoms, 19Q3), ,Jlvl-2,3,4,5-tetramethoxybermene(fhoms;. 1908) myristicin-(Ciamician & Silber, 189d). These ts may contribute to the efficacy of the oil in ulating liver regeneration, and a studyof tlRir vidual action would be timely. Cumin-seed oil aains up to about 60% p-isopropylbenzaldehyde cuminaldehyde, an agem shown, in the present , to heighten the regenerative proaas. The ulatory action of sa&ole,.a principle widely distri- in nature, is reftected inn the findings with sassa- bark tea.or the oil, among others. The propenyl vative, isosafrole, simulated the action of safrole nthe liver and presumably occurred at a relatively level in the ylang ylang oil sampletested d0mg/kg,.Table 1). Ihe observed activity of celery-seed oil and the in (Series 3E and 6E, Table I) on the regener- Gr is of interesL These mixtures are high in nnes sedanolide andsedanonicd anhydride, are responsible for the characteristic odour, and also contain small amounts ofaronuttic com- ponents. Although terpenes tested at high sc levels were invariably without effect on liver regeneration. it might be useful to fractionate the above agents and aromatic moieties with the aim of pin-poinlingg the one or moraactive principles. Representative agents were also administered by the dietary route with resultswhieli, with a few excep- tions, followed the sc findings. In contrast to nutmeg; ground cumin seed at 6ff/was ineRective (fable2), presumably because of its low level of pisopropyl6en- raldehyde. As tarragon oil added to the diet at. P50%was inactive, it appears than effects may vary with the route as well ass with, dosage, since a positive effect followed sc injection of 2400mg/kg (Table 1). The configuration of phenols and other aromaticc agents and their efficacy in stimulating liver regener- ation when given by. the sc route are summariud in Table 4. In marked contrast to n-propylanisols,. the corresponding alcohol, 3-p-methoxyphenylpropanok did not alter the increment from the range seen in. controls Inactive phenolic agents included guaiacol, the two phenolic aldehydes, 4allyl- and 4-propenyl-2- methoxyphenols (eugenol and isceugenol, respect- 4:..Smruary of lloer/'rndinpa in.partiaffy hepatectotnizedmis given seoen daily ac injectious yt onrious aromati[. .. cnmpounds Wlyhmisok (estragole) +hoprnylanisole (anethole). ~.Propylanisok ~rMethoxyphenytpropanol :-AHylphenol sCumylphenol aAHyl-2-methoxyphcnol (eugenol) sPropenyl-2-methoxyphenol (isocugenop &Methoxybenzylalcohol '.-Hydroxy-4nsethoxyben>alrkhyde :-Hydroxy-5-methotybenrnldehyde Isisale .lfethoxyphenplJgygigfdt)pro~so'hyde (cuminaldehyde) t0ylhcnzene 1•Nethylstyrene .Propylbcnzene 1hl phenyl ether idrde axafrole Oihydrosafrok - fyetine (l-piperoylpiperiAine) hperonar' Pynanyl'akohol -0HyCH:CHx -CH:CHCHs -0HiCHiCHs_. -CHyCHrCHrOH -CHyCH:CHr -C(CHshCsHs -0H -OH -CH,OH -CHO -OH -OCHs -OH -CHO -CHyCH:CHt -CH:CHCH, -CHyCHyCHs -O,CHrCH:CHt. Group B Group C. Egecr. -H -OCHs + -H -0CHs + -H -0CHs + - -H -OC'H, --0H -H - -H -0H - -OCHt -CH, CH:CHr - -0CHs --CH:CH-CHs -H --0CHs -OH -0CHs -CHO -OCHs -H -H O -0CHs--H -H -CH{CHs)i + -H -H + -H -H - -H -H + -H -H + -LHyCH:CH~ -0H:CH CHs -0Hr-CHyCHr-0H:CHCH:CHC":) -CHO -CH,-OH + - Significant inereaso-in the liver increment - - Little effect over the conlrols'Ihe dosages appear in Tables 1 and 2. Both 3-allylguaiacol and 4-allyl-2,6-dimethoxyphenol proved ineffective.
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484 A7.IIF.77'r I.P.S'AN .c]t) JOE lu8 TJIO 11i. m.Crexal ............... ............................... iGl1 17_ p.f.rrtiul ....... ..........................._.................._......... 4Mf. IN. a.ie.%plunul ........... ............................ 469'. 19. 1lrYrlml ................. ...................................... . 4 7111 : '?II. LJilhVipgr,.ralrrh.d ... ........._ ................................ .1711 21. llrciunl ................... ................... 1711 .. ~... '1'hcmuhy.lr.~qui n..u:~ ...... ... ._ ....... ....................... 1,'/l 4:. \Irlh.mrpllrunlv ....... .. . ... ........ ....... 170 _L. liu:uaa'ol !u.mrlLu~.y6.••~~~1.~ ............................_._..... IAI ~ •al. IL'dru(iuinnu.. Immulnvlhpl rll:rr Iy.m.~I6nsryLrn~d1 ........ ..... 471 2:r: \Scihus)'h5.b'o:p:iuonr ............................................. 4iAl 11, . :\miuuLrnxrn.•r :md awiu:,phrnul.. He ...... ... ......._._.. ... ... 171: •.11: Amlihr ..... ... 4711 ": .............. . .......... 471 2ti. m-Phrirvlrnrdinmiun ... . .............. ..._ ... -............ ..... 472 LHJ p-1'hrnylrn.dinminr ....... ..............._........_.... 472 :111, p-Aminnphrnui Irotlinall ................. !78 :11. 2<I.l)i:nninuphrnnldihcdruohluridr lawidull' . .................._..., 173 :12. p-\/rlhl'Inminnirllrnulaulph..tc(mrlhnll . . . .......... ~ ......... 471 :S:L p-11VArnscph.•nrhlurinr ..... ...... ............._-..........._. 47:1 1'.. \IhnhLrlall. .............................. ..................... .... . I/A :I{.. o.yilrnphenul ......................... .......................... 174 :I:" , w~\ilruphrwd ...... ........_.._...... ....... ... 411 :Ill p-\iiruph.•nnl ............................ .._..........---- .. 1il :C. i'icrir nri<I ....................... ........__.....,...,........... I7.1 P. Ilidu~rn plirnolr rlr . ........ 67,5. :1%. ^_.li-hichlunrphrunl ...... .........._..-. ......... .~. ........ 1751 eg. 'rri6romuphruul .. . ...... .................._.-... 175 11/. u:Jliv :Wi.l ._._ .... ... .. ....... .... ... ..............._... !]] II. I'urpurnp:dlim• ... .. .. ............_.. :I:i 1?: l;et'Inhrxnnul ................ ..... - Kli 111. l:i:nrLl.i~n:n ..... ............ . .. .......... . ... .... .. .. ..... ... . IGIi G:un:n::rc . .. ... .... .. ... . . . . ._. .. ..... .. . ......___,..... -/N'! I•il~.rnltm~ rilc.i ... . .... .............. ....... ....................... IM2 a t
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172 .•.I,inai"r I.fiC,, i?11 Hni nIS T:nrl Itrr diltllion sltril•a. )C.ImI•i;dlv ~nl !hl• s:•cnud dtlt• nl Ircnllnl•nl such cclls :u•c uul' iutirllurnlly Liunrl. /: Clin~.•ntrnllau ]u:. .. 1 ._ e C e n W II CI L., o, ~I Iwurs •, . . . ... . U I) ?. °_I O (1 I r „ 11 da~•.......... III -~- II. ll F 2 ll 0 Tlnn. uuly few c:lsr•. I,I nud/.ublc1l'. Ir:rgwrnl, wrre eeclorded. 6ut n prunonucrd slickinc.v, rslnmi.. ;dl !hruuglt ILc_ sorirs. ldiscnriug Ihe diamnu.is nf G:II•mcnl.. srcrrnl rrnlrrnir,lliuns hcld'p.onrlnchiawlnla :Ind altnc.hed 15:r;mrnl;. 28. nr-Phrfqyh'nr•dirrulinr. rc: il 13. L: .\II' nrrrLs 1c19•c lnrq- esrent :Iflcr onc dnr• a-I:S at6•t' 2 rtacs. tmllr.rlunalleli- I'?~ werenot uSx•rvl•d llrnger II¢ul (,ur da, %'. lellrn :111 muls were lised. t-: (i-61 Ilave Iendencila lu c•Itmlum•.. d aud r:'. Snlnrc-nlilnse~ ucc.ur io I, miclxl willt unrmal milVnes. AIIt•n I'I duy 2 al,o shows c-nliHdir. c.haract- erislios. mainl) ' aunr-,, rhr:nu4).omr r•nntraeliirn.. Wlnugh :dl nnallhnse, nre nurmui. /':. ,.uu•,Wr:qlull Su. _ . I li . >: 1 II11 1_ 111, I'It honl•5..... i/ Ifi a Q_ IL 1 IP.. •t 11 11 U )•1 ~`I 1 ll:l1'.,..._ 11 li _ )U. ir I) U'. 11 11 l1 (1 29. p-!'lu7lgh~nrdi~+nrirrr. n. 1' ia, li:: iti,••Is vdt: I I~i hwnrtif, C; rl dasl. ~1 12 duysL. 16tn nu.nr I:rcir Illnn thr• n-:md m-Inrm. r': : I:a, dl•Cidt•dl1•luul'r drvrli~prd c-IuInuun. Ilctn Ihr pmrnJinq t.cn ,ulr.l~mrc.. J: Slrull~ µnk iu 1. r: Iu.Y Illrrc :ur lirunll rcrr slicky r-Inilnse... wllich nl''Jrr 1~I:In~ .,il luno Lnli Iepr•. l:. (luer/l Ihr uwsl :Irlice. sub.lnnee~. :nul! Ilr.•1!ef~~nr• Ihr ilrr! Ir:lglnrnl;diorn calw•sare ~icrn lu•rc: Ft~,.~n.Y •.r 4 li O. . I •14, 111 1. :I I I 0I! _ I q G:: fL fl_ I t`!>1: la 1.. rit En. Ib - I _ ;U 1; Z 2 1 F3 19 1 2 i I•J. Ill .• 1~- tl 1:, 2 2 1 1/1 Ali 1 1- _. 14 rP wl . t I IS 1 - 2 111/:1 I 1 19 . 1 _ 1_ tuu u Inu u I:t IWI 'r ilnL n
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,Lltall'r Lli\,5. .,,b J014 IIIR 'r,(U tatul<I. It is ouhy ilr 1•sct•plilm;d t~la.vs IhcllIl'utinu•nts :Irc I'bunrt in c-miluscs. In cuneonlrnlivons ;;iviug luizcd c-miiusvs and nurlnut mitnscs. li1 e. Ihr Iu,.or pnrl ul' znnt• 7! I'nl,nu•nls m:n' Irn 17) 111111 ' hul aImust cxclllsi,rh•. within tllc uurluul miln,rc TIlis is uul unl,° duc tno thc ~nc:rlrr ciil'ficullp. ul idanlilciu;; fragnu•n1. t,•ilhin n r-Inilosis, where no rltrutuusume murt•mrnls r.ccur. 1'+rr rlr•:Ir frnrulenls Ilnct• :Ilsn Geen rlli.orcecl in nurnlld mtd:lph:rsrs 1Vi_. 1-: II is cvitlcul Ihnl thr frag- 1'iC. I. alliurn l:rrer. Pe.nlprn .rnaxn ni nr~•1 rLromo.unwr.. •n mrlnpbv.rin pulnr .iv.c. in Iwaro rhrumuaomrc. L:rru:rr.Iuli~m. h::cr n, urrvIl rin..- In. Ihrir .~nds.L.. ~•: m.i:q:Lanrs in.i.lv ri.•..r; L: •m:~ Irn.mrul1~..tldJr Illr ylat•., Ill ::dJilinm, ...u nllar.llyd Iraymo.ul:.r_ un••Iar;;r elI:.rIr•~dI Irnpm•ul: rl: L.~u .w:dl ::Ilnrhrd 1'rap. nn~uI.,: I•: iu uur vltrumaxomr I.ullr .i.l.:r. rlu'•:vrdiil. nrr :dl::rkrd, onr i. .n:rpirlrir. .u. rrcd. Ih.- ~d6or uuv La. •.nlh dr..:lbpedla .~uusl:lrlinu. ,..'_'//llll. mt•nt.•liion nrcJ. :I Irrt:liu nuen.urr :I ~ ~iabiliiv.: Ihr Ibc:lucucr curvc M' I. r:q{m/•Irlaliun in dil'frrrul e.Mucl•nlralirnns u/ Ilic .mm• subsl:mcc• :J,cnyn .Iw." . :m inhlimuur I:Illiu~; , hulll I."eanls hil-llet• :uldloterr cnnrrn~ h•:Iliuus. Gi•nr.r:rll}~ slrr:&in;;. Ihr r.-utilulic nnd Ihr I7u~mrnlat~innrr. at'liims, t'.cc•.ludu 1•:Irli tlllrr.. llt urJrr Ir, ulinin to :ut rslitn:diun .rf'Ihc 1'ra1`mlentin"' arlicifY nl' rac•.h c•uurrntr:rliun of :1 sulxslmlcr Icard ,rl• haer studicd a nnntbcr of anapiul.rs :and drlrnuinld Ill:• uurnLr•r uf Ira„lucuk in rac•h coli. Th: ft'ryucucc nl crlls ,cilih /.r:ri'otr.nl, rspmssrrl :rs.prl'ceul:llge./l'lilocuunledccllv ~rnllt•d", !r':, a piclla'e rd Iht• uclieih'. 111 is at;drul Ihal c•:Iclr
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Interactionof BHT with other antioxidants Their results.are briefly summarized in Table 4. BMP was observed too be the most toxic.compound to mouse lung while BHT was the most effective tumour promoter. These authors suggested that their data were consistent with the hypothesis that different metabolites are responsible.for causing lung.toxicity and tumour promotion. They further reported a strain of mouse that was sensitive to the.loxic eBects, of BHT but not toahe promotion effects, suggesting a genetic deficiency in the.abilityof these mice to form the promoting metabolite. Since BHT-quinonemethide has.been suggestedto be the metabolite responsible for the toxic effects of BHT, and since we havea simple method for observ- ing the formation of BHT-quinone methide,.we tested the ability of BHT, BDMP and BMP to form a quinone methide. Chemically, all three compounds should'be ablelo form a quinone methide. However, this was noo the case. BHT was converted into quinone methide most efSciently;. BDMP formed only a small amount oGquinone methide, while BMP did not form any detectable quinone methide..If this spectral assay is an accurate measure of the ability of these compounds to form quinone methide, our findings suggest that the pulmonary toxicity of BMP may be related toa metabolite other than qtdnone methide. In summary, we have presented evidence that: (1) BHA canenhance the inmitrometabolic.activation of BHT to BHT-quinone methide; (2). BHA can enhance the: in piuo lung toxicity of BHT in mice; (3) other phenolic compounds also enhance the peroxidative activation of BHT; (4) peroxidase enzymes from variouss mammalian tissues, including human tissue, can also catalyse the interaction between BHA and BHT. What, if any; relevancedo these observations have in assessing the possible human risk of exposure to BHT? Humans ingest up to 0.5 mg BHT/kgJday (Gosselint. Smith &. Hodge, 1984)..Doses of BHT that enhance tumorigenesis have been reportcd, to be around 35 mg(kg/day for a period of 2wk in mice (Witschi;,1985). This represents a cumulative doseof approximately 450-500 mgJkg, Overt lung toxicity is caused by high single dosesof BHT (y400mg/kg), although there aix large differences between stminsof rnice.(Kawano, Nakao & Hiraga, 1981). Microscopic lung damage is observed at doses as low as 40 mg/kg (Marino & Mitchell, 1972). Thus,, the average daily intake of BHT in humans is generally far less than the doses used to elicit toxicity or promote tumorigenesis in mice.. However,, as we have demonstrated, micro- somal enzymes from human tissues can metabolize BHT to a reactive product(s)) in a manner similar to that of microsomes from susceptible animal tissues. Therefore, while the doses that humans ingest may not cause overt tissue toxicity, it is possible that at the level of human exposure the metabolism of BHT might lead to more subtle forms of cellular toxicity. Similarly, the chronic formation of non-cytotoxic amounts of BHT-quinone methide could enhance the development of tumours or alter cell defence mech- anisms so that the ability of a cell to respond to other environmental stresses is compromised. Such re- sponses.maybe particularly important in the small intestine, a tissue which is very active in metabolizing FC.T. M 1611'-M 1193 BHT (Table 3): and would probably be exposed to thee highest concentrations o0 BHA and BHT. Finally;.while food andcosmetic.products are the major source of exposure of humans to BHT, there have.been recenereports of persons ingesting up to 6 g BHT/day for the relief of herpes virus symptoms (Shlian & Goldstone, 1986). In a 70-kg (150-Ib)) human this would be thee equivalent toa dose of 86mg(kg: BHT capsules (usually sold as 250mgBHT/capsule in a gelatin coating) are nowavailablu in many health-food stores. More signs of human BHT toxicity will probably appear if this practice continues. Arknowledgements-We gratefullyy acknowledge the finandal support of the National lnstitutee of Occupational Safety and Health OH01833-02,. National Inslitutea of Health E307141 and Johns Hopkins CAAT. We also thank Marietta Regner for typing the manuscript. REFERENCES Bauer R. H. & Coppinger G. M... (1963). Chemistry of hindered phenols. Terrahedron 19. 1201. Becker H.. D. (1965).. Quinone dehydrogenation. I. The oxidation of monohydric phenols. J. org. Chem. 30, 982. Chcn C. & ShawY. S. (1974). Cyclic metabolic pathway of a butylated hydroxyloluene by rat. liver microsomal fraetions. Biockem. J. 144, 497 . Cook C. D.. Nash N. G. & Flanagan H. R. (1955). Oxidation of hindered phenols. IRI. The rearrangement of the 2,6-di'aert-butyl-4-methylphenoxy radical! J. Am. ckem., Soc. 77. 1783. Filar L. J. & Winstein S. (1960). Preparation and behavior of simple quinone methides. Tetrahedron 25, 9. Kawano S:, Nakao T. & Hiraga K. (1981). Strain differences in butylated hydroxytoluene-inducad deaths in male mice. Tosic: appl. Pharmae. 61, 475. Kehrer J. P. & Witschi H. (1980). Bffectsof drug nwubo- Gsm inhibitors on butylated hydroxytoluene-induced pul- monary toxicity in mice. Toxic. appl. Pkarmac. 53, 333. Kharasch M. S. & Joshi B. S. (1957). Reactions of hindered, phenols. II. Basecatalysed oxidations of hinderedd phanols. J. org. Chem. 22, 1439. Kurechi T. & Kato T. (1982). Studies on the andoxidants. XV... Combination eHects of butylated hydroxyanisole,, butylated hydraxytolucno and their analogs omhydrogen donation to 2,2-diphenyl-l-picrylhydraryl. Chem. Pharm. eull:, Tokyo 29, 3012. Kurcchi T. & Kato T. (1982): Studies omthe antioxidants. XVI. Synergistic reaction between butylated hydroxy- anisole and butylated hydroxyloluenee in hydrogen donation to 2,2-diphcnyl-l-picrylhydraryi. Chem. Pharm. Bu!!., Tokyo 30, 2964. KurechiT.,Kikugawa K. & Kato T. (1980). Studies on the antioxidants. XIII. Hydrogen donating capability of anti- oxidants to 2,2-diphenyl-l-picrylhydrazyl. Chem. Pharm. Bu!!., Tokyo 28, 2089... Kurechi'T., Kikugawa K., Kato T. & Numasato T. (1980). Studies on antioxidants. XIi. Combination effects of antioxidants on the reduction of ferric ions. Chem. Pharm. JJulf., Tokyo 28, 2228. Lowry. 0. H., Rosebrough N: J., Farr A.. L. & Randall R- 1. (1951)i.Protein measurement with the Folia phenol reagent. J. bioL Chem. 193, 265. Malkinson A. M. & BccrsD. S. (1984)'. Pharmacologic and genetic studies on the modulatory etfects of butylated hydroxyloluene on mousee lung adenoma formation. J. natn. Cancer liul. 73, 925. Marino A. A. & Mitchell d. T. (1972): Lung damage in mice following intmperitoneal injection of butylated hydroxy- toluene. Proc. Soc. Exp. biaL. Med. 140. 122:.
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F 1190 D, C: THOUpsou and M. A. TRUSH protein concentrations of approximately 10mg/ml and were: then rapidlyy frozen in a methanol/dry-ice bath and stored at -80°C. Covalent binding of BNT meinbolile(s) to miero- soma! protein. Reactions were initiated by the addi- tion of 330 pbt-arachidonicacid or 0.9 mld-hydrogen peroxide and were allowed to proceed for 10min at 37°C. Reactions contained I mg microsomal protein, 100)<xf-BHT (0.25pCiltube), and IOONSt-BHA (when specified) in a total of I ml 0.1 m-phosphate buffer„pH 7.5. Incubationswith ram seminalvesicle microsomes were carried out at room temperature with 200Nxr-BHT (0.25pCi/tubc) and 100pst-BHA (when specified) in I ml 0.1 xt-Tris buffer, pH 8.0. Reactions were stopped with 4m1 methanol,.and, the protein pellets were extracted repeatedly with 2ml methanoll or methanol-elher (3:1) until. no further mdioactivityy could be extracted (generally 12-15 washes). Pellets were dissolved in I ml I N-NaOH, and the radioactivity of an aliquot was counted in 10 ml of scintillation fluid. Protein was determihed using the method of Lowry, Rosebrough, Farr & Randall (1951). In vivo mouse lung toxiciry:. Male CD-I mice (4-5 wk old) were given subcutaneous injections of BHA 30 min prior to intraperitoneal injectionr of BHT. Corn oil was used as the vehicle (:£0.2mlJmouse ip,. <0.1 ml/mouse sc). Animals were allowed food (Purina 5001) and water ad lib. throughout the experiment, were housed on Alpha-Dry. bedding, and kept on a 12-hr light/dark cycle. Lung (wet): and body weights were recorded onn day 4 after injection. Toxicity was measured as an increase in lung/body weight ratio (%). In vitro measurement of BNT-quinonev methide and stiibenequinone Reactions contained 15 U horseradish peroxidase (0.1 mg), 0.9 mtd-hydrogen peroxide, 100 pxr-BHA and 2l)0 µxt-BHT (or analogues) in I m10.0CM-phos- phatebuffer;.. pH 7.0. The rate of formation of BHT-quinonc methide was measured spectrophoto- metrically at 300 nm using an extinction coefficient of 27,000/M/cm (Becker, 1965). BHT-quinone methide has an absorption maximum of 285 nm (Bauer & Coppinger, 1963; Becker, 1965;. Filar & Winstein, 1960) in organic solvents such as isooctane. We offer the following observations as proof that thee product from our reaction (with an absorption maximum at 300nm) was really BHT-quinone methide: (1) when the product formed in aqueous solution wasexlrac- ted with hexane, the absorbance shifted back to 285 nm1. (2). the product in aqueous solution had similar spectral properties toBHT-quinonemethide synthesized by the method of Becker (1965); (3) upon standing, thee product formed 1,2-bis-(3;5-di-tert- butyl-4-hydroxyphenylkthane and 3,5,Y%5'-tetra- terf-butylstilbene-4-4'-quinone (stilbenequinone), twoo compounds known to be formed byihe dimerization. of BHT-quinonemethide; (4) deuterated'BHT, syn-thesized by the method of Mizutaniet al. (1983), in which the three hydrogens on the C-4 methyl groupp were replaced bydeuterium; dramaticallyy slowed the rate oC formation of this product; (5) glutathione blocked the formationn of the product in this reaction and caused the rapid disappearanceof previously formed product immediately upon addition too the reaction mixture, leading to the formation of a BHT-glutathione conjugate. Similarly, we observed a spectral shift iit the ab- sorbance maximum for stitbenequinone from. 445 rtm (hexane)) to 460 nm in aqueous solution. The rate of formation of stilbenequinone was measured at 460 nm using an extinctionn coefficient of 72,000/M/cm (Kharasch & Joshi, 1957). Authentic stilbenequinonewassynthesized (Cook, Nash & Flanagan, 1955) and compared, (UV-visible spectra. NMR) with the reaction product at 460 nm, to confirm the identity of this spectral peak. Results and Discussion Prostaglandin H synthase and horseradish per, oxidase have oftenn been used ass model enzyme systems for the study of the peroxidative activation of xenobiotic chemicals (Marnett & Eling, 1983;. Subrahmanyam & O'Brien, 1985);. The ability of BHA to enhance the metabolism of BHT to a reactive species in these two model enzyme systems is shown in Table 1. Three endpoints were measured: covalent binding of BHT to microsomal protein and the rates of formation of BHT-quinone methide and stilbenc- quinone. Inn the presenceof BHA the prostaglandifr. H synthase-catalysed covalent binding of BHT was greatly enhanced (3.9-fold). Similady,, the horse- mdishperoxidasecatalysed formation of both BHT- quinone methide and stilbertequinone were greatly enhanced'in the presence of BHA. In the absence of BHA, no BHTquinone methide or stilbenequinonc were detected. Prostaglandin H synthase also cata- lysed the formation of BHT-quinone methide in the presence of BHA, but no stilbenequinone was formed in either the presence or absence of BHA (non shown). The lack oOYormationf of stilbcnequinone might be due to the presence of microsomal protein which would bind to the BHT-quinone methidc before it could dimerize to form the stilbenequinone. Synergistic eAects from various combinations of antioxidants have long been noted (Kurechi and Kato, 1981 & 1982;. Kurechi,. Kikugawa & Kato- 1980; Kurechi; Kikugawa, Kato & Numasato, 1980) . For example,, enhanced protective effects in food prodtrctscan be achieved by using a combination ol antioxidants, with the added benefit of reducing the Table 1. Elfect of BHA on the N vlrra pvoxidatiwc metabolism of BHT Covalent BHT-quinone meWide SUlbeneyufnone binding fomution fomution (amol BHT-bumWf (nmotiformed/min/I5 U (nmo/ formed/min( Rwctsntr maproteie/IOmin) HRPase) 15UHRPess). BHT 1.9f0.3 ND ND BHT + BHA 27.6€1.9 43.5;22 7.7±03 absolute amoun the food, proditc individual antio anlioxidants haF antioxidant to rt. Kurechi et al. I effect of a com6b gen donation to I-picrylhydrazyl mechanism of i: regeneration of generation of B the expense of B quinone methidc BHAA al the exp, results in peroxit direct interactic BHA with. BH-' novel mechanisn systems. We investigatr phenolic compot vation of BHT. - compound's abil: binding of BHT binding aR show phenolic compo- BHT binding, [ BHA. The comr were.chosen bea to food and cosn uons-. In many it i p. HRPa1e-Honemdirh peroxidsse ND=Not d,aectaHle
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184 B. L. Poa and! P. Z. Lw~ LaWraoevY' gnnuke Prnnucer I oliM and H20- onaenaanon t f ar W¢ioti (a) Smoke <onUensa[e , (b)aquanus Inac[ians eztracti'un vith etnyl acetate.,.pry ] I aqueous phaze __1 O_GaniC phlse I evaporavtom tn nrYness Nav excxacc (a7 rcon or fracciona ~ (b) froo zqueuus iractiona a1 H-Napn exrracrion ulrn penrane/erAer ~ Organic pEase exrracGibn with I N-Inl 1 Aqueous Oreanic. pnase p)lase evapo[acion cv aryness I 9queous phase fSaccionauinn ui[h 1110j ooLU+n Neucrat Puri(ied phenolic wnWUnnss fkacrinu (PdH-frco(from aand b)nnnofwstionat phenols from a antl t) Fig. 1. Preparation uf fractions containing neutral compounds and monofunctional phenols from smokehouse smoke condensate. Fractions indicated by asterisks were tencted fbr mutagenicity: EXPERIMENTAL Preparation of test samples. The PAH-free phenolic fractions to be tested for mutagenicity wereprepareJ and kindly supplied by'L..Toth, Kulmbach, according to the schemes in Figs I & 2 (Toth, 1980). Smoke- house smoke was produced by slowly burning 200g Hirchwood spans at 600'C in'a laboratory smoke pro- ducer. Fromthis, two smoke condensates, a tar fmc- tion laland an aqueous fmctiom(b). were obtained through condensation by waterr scrubbing. Further extraction of the condensates with ethyl acetate or through perforation yieldedtwo raw cxtracts (from (a) and0l.respectively)iwhich were ultimately purified as showmin Figs I & 2. Extraction with NaOH yields organic and'aqueous phases (Fig, 1) which are further purified to yield neutral compounds and monofunc- tional phenols, respectively. This methodAestroys.the bifunctional' phenolic compounds, therefore an ad- ditional purification,scheme was employed. For isola- tion. of the mono- and bifunctional phcnollc com-pounds: IFig. 2) the raw extracts were puritied by aausous.phasa pH 3-5, percolation Orp,aniCpTase. i'henoLO extaaet steam distillation at. 170`C and partition chrottt+' tography. on silanized'silirr gel. This phenolic cxtratlwas subjceted to capillary gas' chromatography after which more than 100 different phenolic compoimds were fbund and',partlyy identified:.. The ten most abundant constituents of the purified phenolic fraclion of birchwood smoke are phenol (1•95n;). n-cresol (0g6%);, m-cresol (047%L p-cesol IP53%;k 2.6.dimethylphenol (l}32';;k uaiawl brenzualechine (I85%). syringol (942%): eugen (131%) and vanillihe (0-42%), Phenol, m-cresol, Pcresol and vanilline were supplied by Schucherdf- Munich. o-cresol and eugenol came from Merck.. Darmstadti 2.4-dimethylphenol.guaiacol and brenua- tcehine from Fluka..Buchs. CH and the syringol from" EGA-Chemie- Steinbach. Theyy were all either 98e8 pure or of analytical grade except for 2;4-dimelh)'1'phenol which was 90";,. pure and contained 5'7a dimelhylacsol as welll as.tracesof cresols. ,SArtuyenicitvrest6q/. The histidine auxotrophic in- dicator strains Sidrwmeflu tpphimurium TA1535' TA1537, TA1538-TA98 and TAI00 were kindly suP ® i _• Icd 15% ~trslot m2 th -rthed +k•ed in . .u;[drup - r.^.Ir wltl. " A ` '-9 prov ~: ~aethod -dithe S ~anine:- =ach c> CD cositin ~ ; cnme, M :'o,se 10-111 Holltn . ~ Spec i r_e Inctho, ~arent j; ~tlon ~i preinc I 14.01 ulditii /i~
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458 .\LHI•:II'f I.FR`AV ANII JOIL nIN 'I'Alo Ikterminulion of fr ':, is.liablrs to cuosidereJJle error. In order to judgf; tharcliabilily uf lhc indiciJunl fr "; value \cc mndcsomc coulrol counts uP slidesfroln difl'erenl Irenlmenls .vilh thc salme clotteenlrntiou of one xubslancc aud from dil'll•rrul rouls.th;N had undergoue the nanm lreal- ment. •Is expected. \v.r fuunJ a conciclerablc rnrialion. :lnvhmr, ne- curdiugg to our opinion, the nclk.ilp of :1 substance is ruthcr .velL dcscrilxd il° af Icua lolf cells ol' cach cunceull+aliolJ within Ihe series Ilave bxru :tualvsrd... I)Plen. h... rrrvr.. v:lrihus circumslnnres. as for instance lacl:ine numher of JicisionsN h;rre cul shnrt Iltr couJding ul. n lower nwnber ot' cells. l:specialll- atter 24 hours tJte loxic effect is usually high evcn in zuur :1 anJ eouslilules n+ourcc nff error. (:ounls a/'ler 4 hours, have Iherel'ure mninlc IJeI•u used for judging Ihe :rrticily of a certain subalance. 'Phe resulls ul' the longer treutmenls furnisli unly :t complelncnl lo Ihis.Unly 1.'hen really new char,lcteristicss have In\m revealed bythc longer Irealmonlzs have Ihese also becu laken in/o cunsidcr:rtion. L<u:lflyv the chnngc in the picture from4 hours. lu 24 huursis as follows. IliriLar concrnlr:rtion, of zone 3 tunyy rithec rii\ra complete lack of ruito,c..nr a,nl:dl. nntnberol' mitoses .vilhhigh frag- mcutnlion frequency. Lower c.uncenlralions, \vhcre toxicity is lo\r: usually ~ivc: a decreasetl fr "d. :u comporrd \\.ifh -I Luurs' trenlmeul. As :ur instance of' Ihe v:vialion in ir ";, Table I is alLmilteJ. It cout,•lins lllr resulls of fr:rtimenl cuwds on fottr diffcrenl nccnsions w'ilhiH the utost :Ic•tire ulalnltcr. In rac•h Slirlc the indicidual'ruols \\'crc recordecl srpnr:tl/•Ij-. 'I'hc cnrinlinn Lel.rc•en various lrratmeuts is ,rc:d• fruttl ?t (u. (i:i~5.. ll'nHl1- nar ruul had Lc/mcuuutcd un ruCll occasiim it would lulce been pos%iblv to: gI•1 :Is tliffcrent c:tiuvs ns 4.1 :wd i'S•o\Pil]lin rach firlliuu lcss \srialiun is liound, Ihe limils briug denoted by t11e r:liio. I: 3. We duo nni know n'hulis Ille cause of the variation. hul even ;m:Jl variations in Ihe external condiliun, mnv presunlaLly cnu,rcunsideraldcslil'fcrencrs in Iberce+ction: Since many of Iltrw comlmunJsemplnevd are easily oxidizablc and uudergu rapid chanrciu l:lltnvalrr. ae eousidcrud Iha4 Ihc cuudllinns vnuld bc. krpt untlbr bcllcr ruHlnd if \ri• usrd distilled \calrr 1'ur c,ur diluliun sarics. allhou;;h we are a\c;u•r Ihat diailled a':der. e,lrcciallr Jurin;; luui. h'ralntcuM, acls poinum":uly un Ihr uniutu rools. \Ye m:lde. huq'ever., srct•r:d comp:n;llirr scrir,.:ts. Lelw:ncu hip-waler and'dislilled a'alrr and fouml thal IIJe 17npmrnlation aiso occurred in Iop-\calrr. 16ou;;h It•ss rv,"Itlat•ly. 'fhis prold\•In \rill lie Ltlcru up IaUCr. csprcicdly axs rc,1ards: the c-Imuour reacliou. \ehiahisnutchh nlrq•1• cn.il\• brrlial( :Ibuul iudiaillyd \rrlur Ihan iu I'np-wnlvr. ~ ~ N 1 11- 1 `1 rb Gj
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ALI36:11'r I.IL1'A\ A]Il nIK Ill\ TAI(1 Phenol . . . . . . . . . . .... ....... . . . . ....... Q. Xr., . I'A rnc:dcchul . . . . . ... . .. . ...... . ...... lit•snrciuol . ................ ILuwO. ! Ilcdrnquintmc .......................... O.nu P c rug:tl l nl ............. . . . . . ... . . . . . . ... . l Wrv' r licdrrnsncydruquiuuuc ... ....... ....... <11x. 1'hlurug lurinul1.. .. ....... .~ ... . .. . O.,n:. One mrthcl group inir.~ducrd intu phrnul.lur.,ltul ch:w{;t• lhc lh. a., LoIG 1!tt tl4roslwlil :md I!ir snfuldlih~ Iwiug. hrnadle spt•:Ikiug. tilt! rc:lme: a-1'.i', sul ._ . . .. . .... . . ... . ... . 0 .,xn m- ..... ._ . . .. ~ ........ . . . ... .. Il.nnt U- .......................... O.wn II';.on Ihc other h:tuJ. ltru In,lhvl ur uon mc111eInuJ our iauprnpyl group :Irc.ntldcde u+ in <y.lrunl omd Iltvrnul rrcprclier!a', thv W:dt•r sn)tlllililp is ln,cerctl and ul.r. IILtr Ihrcsltulcl calun., Scltich resulk in a llt, a.. of O:mi im xy9t•nrd1 c[nd (1!r- iil Ihc.u:ul. 9-hr di-pht•unll hurnnbt;;ucs cihrl hyro- calucltnL. nrcinr.Il aud thrrnnUydlvquiuuur h:lvc adtuul IItP samcwulcr sulubililv n'ith a t~qleNoh:lll liiglx•r tln'pahuld. which t."it'cs n Ill. a. nf Il~~_-(Lt... Ifcre.ul is~'umltasr,l i%ilh.urciitol and Ilhynrol Willt Ihvma- hy.droqttinunc. Ilte Ih.:l. i, iut•ra:uedd in ISulh casc. .itnull:lnrous!n' ns UII is :Idded. ALuv. tht•re il nu vimltlev cunnrc6iun heim,!cn lihr Oll cuulrnl nr :I ntnirculv aud its cLnmical :INfiru. Thr thrcr InethuXephcuul; ,IuJi(ld. ^oaj•ICUI. It-wulhuc)•pLenol :uul methuxyhrdruquinunc I:u:r:t suinLilils of :dluul t.:tud n Itlrt•shuld :II a--G. Their Ih. a. Ncill 6r Il.!. 0.16 :md 4.a,. rrsprctibclv. So ill Iltir casc the inlrudtrctinn ut n1wdhcr Ull int'rr:lscs Itlc chrmic:tl acliou. '1'uc :tllrre,r dul••t dn ttnl ailn al :tnylhinti liu9 n dn:cnipliou ui Iftr -;Illlrrod ntalt•ri:d. /7>ciu:; tn !lu• !4Irr:li suurcrs uf rrrnr frum whirll..:te wc src wall ax'arc. nur cspe.rlwruls uffi.r •,cr• cunviciar ilt neccssary ill rrpr:rl thr t•xlmrilncnl+ 211d :u lnl.ciiuu usurr, dr•hli!,d ;ICCUUnt ,tuclt Inclurs.:ts Ihr pnrlly afllLV rlirutical.. Ihc pruprPtivy „IItlc wdcr usrd. Iln• plll'.nttlu• rtiffrrcul nlnlfium. Ithe pcrnlin'r.,:uid,n nn, ltcfnrv Ncc Vrntura to cou'iitrr the rr>ulls :IS 6:tciru;; rnouyh slu'rn;:th (ur ihc Luild- ing (1t nul)~ 1Lt'pulhrsi. ua Iln'ul. '1'Itr Illtliu uiljrt'.t u(IIw Isrt•vnl worllia !L~• ludc nf Iltu inlu.l'r,tinri, nrtc frt;mtcttlin, rflrrl. T!icdivlriLuliim dIliv !Y';,. ill t;ilTcrctll cun- rrulrnliunI n( 111, ,::n"r•-rdi,larrrr i, .Irifiin;_.1 it iiirrc:ncs !eum l.uth..
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1/ lI U ihe ;Ja [:uaouusnsie. rnararn.rcTnTtnN ki ', :10. p-Antinophenol (rudinal l.. --- rr. 'S- l;i.. b.: Bnals lurgnscenl in all cuncenlraliitns. c: li-t3, d amd c: U.niv in ?wrre.c-mitnses foundal'lcr 4 hours. Clear tendencies lu c-ntitn.i.,were seen aflrr I dnr in une rnut Ireated wilh enucenlrcdiuu :i. No dicisinnx occurred in adjacent cuncentrations, so tLe finding, cnuid nul lie verified. 1: Fra;;mcnls com- mon, their Li•equencydislribulion resemLles.lhat of p-pTienylenedi:tmine: f.mucutrmimi Nu. 4 .1 fi ] 4 {r In It 12 13 1'r ~, 11 hours .. 8 1 L 8 22 ~ 7 3 2 2 1 L dav .... -- -- - - - - --- 8 @8 J7 1 0 31. P,4-DiuminopherloGlihyrlrueht(irirlr Iaroidnt). -- n: I-Ul (2 serics a nnJ b). L: liouts'sufl: 1--F (1 hour), 7- Y 14 hoursl'„9 (I dny). c: 10-13 (a series), P2-16 (P seriesl. d: 9'he eulire.upper region. 1-ti. shows pronounced toxicity with insl:ml breaking of thcvital procrsses. imli isles with cital stractures and fiucchromosnme structures beriu to appear. e: No concentration gives Irleicel c-mitonis, i--Nshonwenk c-tendencies together with n ccrtain nmount af mottlin„ [: Fra;;ntents rnrely accur in 8-13. dllhough in :i.cries 100 cells in cnch of the cnncetttralinns 14-16 act•rr :mnly,cd. nu. fr:r',mrnl tcas srrv in Iltcsc cnncenlrstliotts. ronrrntrnnau \a~, r i m I2 hours b 5 ;' :1 14 • n ... . . ... I /~0 I;,r.. o~ i 4 „ b ... - ..... . -_ F 4 t ? ^ !:I da Y rt ....... .... It 1). ll , ~I r...... ... lt :R'. p-.lltlltghutliirofJlrurdsull~hrdi• 1 tuulhul,': __ rt: (:--1:3. 6: liaul< .nfA'.. C -Y /1 lwltrs).. ~1--~S I l d;irh r.:. 9--Ia. J;md e:. C:utd I. trrre prunnunct•rl pcemtsis. "...:r zhots (rndrncir. In c.tuilusi.: LuI nrtrr vrrr .Irnur,. 1': Fr'''} hours .. 1 G n ' 2 II D II "I I (ay. .... --- ~ - 40 . 0 /) 1) U 1 y r :13; p-llrlaraxyldir'ulllrllyrinv: --- n:, ;. 13. L: IPnol.k snft: :3:-_It nl d:tc);. d und c: A'fter 1tomu'ti :: .iL,srso nu pni.uurtl'rrt..;iFler 1 i~ ; a::rt•:tll vii:d aruclnres ara aqnptctrlr In~,l.rn dutc:r in
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(:IIR(Ifd0S6\IE t'RAn\IIiNTAT1ON 475 r in were encountered only in 12 allrr 1 hanrs, when 2 cnlls.nat uf 200 had eacti 1 fragment,, while I cell had 2 IraFnlenls.. P. HALOGEN PHENOLS, ETC. 38. 1di-Dichlorophcnol. ---n: I--l:f. b: Rutds soft: 1-4 4 hmlrs),, 5-8 11 day),. e: 9-12. d and c: luI-T;dl divisiuns fi/rnl deeply stained p.-cnntir lumps. 'I'hiss pas.ec iil 8 direclly /rverr inlu a more typical c-milosis, also0 has sc,^Ilerl•d c-miluscs. but mostly uormal Iqlt'.s. /f - CunCCntntinn Bu. !r !1l II 12 13 Yi• % .13 htulrs ....... 0 I 0 Il 0 1.1 day'......... - 3 0 - 0 3i1. TilLromophenol.-n: 10-C1. Only 16ese fourcnncentrations were Icsted, and the delerminntion uf Ihe cotlccntralions is rathcr ap- proaimalii•c, h: Rools.sol•1: 10 (4 hours)~ II il day). c: 12-13. rl':Ind u: C-mito-cis in all concenlralion.s,a completc in 10 aud 11, mixed with . normal miloscs in. 12,, onlyc-lendcrlcics in 1;3- 1: fu 12 and 13 100 cells were analc:stwl; no fragments were frnmd. -ltl. Cnllicrrcirl.-n::1--13. N: linols suft: I---2 1:'40 min./.:t--f (it hourL il-li (4 hours): c: 7--8: d:..5lronh toxicity in 1-4. fn :3 -1 the uulcr l:q'rrss uf the roots conl:riu 1-,rud cliromoaonrce sII'uclut'cs lvilh Irelerchromatic dil'ferenliations. c:. a--ii „ier sanlc smnll IenUCncics ln c-miluais: f:'Pflr concentrtdionsl-Ia were anatr"rl balh al'tcr ~I Ftours n1111 1<Illy.. Olllyy in Ullln (•,rtlcelltrlltiull IVi allfVl' f IIOIII'S ;Pe!-d':nly frPn Ra;;rnrntss met with. viz. •+ne c!-II l%illu 1 Ir;lentrnl and ntlr erll :cilh 2 fra_lor•nls. Allachrd Prn^mcnl, :oui Ixrudnclii^slnala urrr }nu!c- linlrv srvn. 4 1. Prrrpurrrgnllinc•.--rc 1$is ,!Ilr,l:wcr In!acnndenxd ii, andrrnc 7•rin„ around which uuc krtrr :,nd fuur itvdruxx-,roups :!rcpi,lcud'. iu tl:r pn.itiilnsI. 2. i• 8 aud'J. It i, unln- .pmlinrlc suln0lc iil lr. Irr. I+1 mg did nut dissolve on Lniling in IS r.c. Lul this suinlu-m wrrsnse,l l'/rr :r dilution scrirs• which apprrrcinulmlc corn•sponds too ourc•rn- crntrntionv.:i--11. .. Prunt 7 tlulrmw:rrds Ihtr sl•lutilm. mcl-cv c.lr:rr. b: 13unls vnflL.S. 7 11 d:ICL.M i2 da}•s!. t^ /4 rl :nrd ,~: Il:dl c~inilrrscs nccurrrd in 5. -7. Iln- nr-sl h:!d nurmal nliluses. •I'his ,ulrrl:utcr. Ihus. .1in.cs. nu ,1lvcilic o-urilulir aclik:ilc.. tiuli.lanct•s lehic4: Illa' cnlchicinr, cont.•lin 7•rins:s. are. ufspcci;ll inlervst in respect ol c-alilusm j: fn 7 and 8 .rerral p,r•tldllchiaamllil and ilrirl'lir. Icerc+1on.. Frrc i7aolmcuts nncumrod in 16c I'ullnw:iu.g. I'rrqurnov:
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374 J. Agrlc. Food Chem., Vol: 30. No. 2. 1982 . /r Table III. Effect of Agronomic Variables on Pyrolyzate Catechol Levels° pyrolyzate catechol, % change fi'omrecommended field nitrogen application Pale Yellow-10Kentucky 34 recommended nitrogen low nitrogen +51 +137 high nit:ogen: +6 ` -50 plant stalk pyrolyzate catechol, %change from bottom position position Pale Yellow-10 Special Yellow-A Coker 139 bottom middle +1,6 -2 +7 top +90 +23 +74 °As determined by relative response vs. the p{sec butyl)phenol internal standard. content of tobacco leaf correlates to a high statistical probability with catechol levels (probability = 0.0002 for total polyphenol vs. catechol, with 0.01 highly eignificant), additional precursors (a.g., lignin) as previously suggested (Schlotzhauer and Chortyk, 1981) contribute to total catechol content of smoke. Previous data (Schlotzhauer and Chortyk, 1981) had shown that 50% of the catechol-content of the pyrolriste was attributable to the polyphenol-containing ethanol- solubie leaf extract and the remainder to the sugars (methenol and water extracts) and the lignin-cellukrseresidue. Accordingly,, individual leaf constituents were pyrolyzed under smoke simulation conditions (Table II). Chlorogenic acid was shown to produce the highest yields of catechol and 4-ethylcatechoL Significant amounts of 5-(ttydrorymethyl)furfurel, a thermal degradation product of the quinic acid moiety of this caffetannin and previously reported in pyrolyzates of the ethanol-soluble leaf ertead, were produced. Quercetin and rutin, generally presentt in tobacco leaf in lesser quantities than those reported for chlorogenic acid, also produced significant amounts of catechol. Interestingly, rutin produced a series of 4-al- kylcatechols, which have also been reported inn cigarette smoke (Snnmemann et al., 1976; Schkrtzhauer et a1.,1978). In the latter study;, the presence of vinylcatechols were reported in a biologically active fraction of cigarette smoke condensate. 4-Vinylcatechol has been reported.to be a major cigarette smoke compound by Ishiguro et aL, (1976). The-source of this catechd,derivative iseasdy rationelized fromm the thermal breakdown of the caffeic acid moietyaf chlorogenic acid. Under our smoke simulation conditions, only catechol was found as a major pyrolysis ptoduct. When we performed a destructive distillation of caffeic acid, at 190 °C (oil bath) and under nitrogen, and directly reacted the distillate with BSTFA, we obtained as the major product a compound with mass spectral charac- teristics of trimethylailylvinylcatechol. If formed under our pyrolysis conditions,, apparently thiacompound was not stable under the standard workup conditions in our analyses. Of the sugar and leaf.fiber constituents, only lignin produced signihcant.amounts of phenolic products, in- cluding catechol and guaiaeols: These data suggest that lignin is a significant contributor to catechol levels inn to- bacco smoke. The polysaccharide cellulose, the di- saccharide sucrose, and the monosaccharide fructose pro- ducedfurfural and aubstituted1urfurals as principal py- rolytic products. However, recent smoke studies (Carmella et al.,. •98l) performed with 14C-labeled cellulose added to Schbtzhauer et al. -: cigarettes suggest that cellulose may be a more important precursor of smoke catechol than mightbe.deducedfrom pyrolytic data alone. Although lignin content shows relatively slight variation among tobacco varieties, polyphenol levels vary consid- erably amount type, cultivation, stalk position, and curing parameters (Tso, 1972). Effects.of some agronomic vari- ables on pyrolyzate catechol levels for selected tobacco varieties are shown in TableIlI.. The apparent inverse relationship of pyrolyzate catechol with nitrogen applies- tion closely correlates with the observations of Andersen et al. (1970) that leaf polyphenol content is lowered with increased nitrogen availability. The trend in pyrolyzate catechol yields for leaf samples obtained from the upper, middle, and lower stalk positions is reasonable,, since leaf tannins increase dramatically from lower to middle stalk positions and reach maximum concentrations in the upper stalk positions (Tao, 1972). These data reinforce the previous conclusions as to thee importance of polyphenol content of leaf in determining catedtol content of tobacco smoke. ACKNOWLEDGMENT The technical assistance of K. Childers and D. Finco is gratefully acknowledged. LITERATURE'CITED Andersen, R. A.; Chaplin, J. F.; Currin, R. E.; Ford, Z. T. Agron, J. 1970, 62,415. Arrendale, R. F.; Smith, L. B.; Rogers, L. B. HRC CC; J. High ResolutChromatogr. Chromatogr. Commun. 1980, 3, 115. Be1L J. H.; Saunders, A. P.; Speats, A. W. Tob. Sei: I966,10,138. Bock, P. G.; Swain, A. P.; Stedman, R. L Cancer Res. 1969, 29, 584. Bock, F. G:;Swain, A P.; Stedman, R L JNCI, J. Natl. Cancer Inst..1971, 47,429. Brunnemami, K. D.; Lee, H.-C.; Hoffmann, D. Anat. Lett. 1976, 9,935. Carmella, S.;Hecht, S. S.; }lotffmann, D., 35th Tobacco (2temista' Research Conference, Winston-Salem, NC, Oct 8, 1981. Cermelle, S.; Hecht, S. S.; Hoffmann, D.; Tsa, T. C., 34thTobaoro Chemista' Research Conference, Richmond, VA, Oct.27,1980. Clark, 1. T. J. Gas Chromatogr. I968, 6, 53. Hecht. S. S:; Carmelle,.S,; Mori, H.;. Hofflnann, D. JNCI, J. NatL Cancer Inst. 1981, 66,163. Higmaq E. B.; Severson, R. F.; Arrendale, R. F.; Chortyk, 0. T. J. Agric.,Food Chem. 1977, 25, 1201. Ishigum, S; Sets; S.; Sugawera, S.; Kaburald, Y. Agric. Bio1:Chem. 1976, 40,977. Jones, T. C.; Schmeltz, 1. Chem.Ind. (London) 1968, 1480.. Schlotzhauer, W: S.; Chortyk, O. T. Tob. Sci. 1981, 25, 6. . Schlotmtlauer, W. S.; Chortyk, 0. T.; Sevenon, R. F. Tob. Sci.1979, 23,103. Schlotzhauer, W. S.; Walters, D. B.; Snook,.M. K; Higman, S C. J: Agric. Food Chem.1978; 28,1277. Smith„W. T., Jr.; Chen. S. P.; Patterson, J! M. Tob. Sci. 197L, Consir economi mental I number, of herbic et al., 1 knowled: herbicidr their usr (1978)). for the (2,4,5-T) 1978). 'I informat The ra ' persister " 1970; Lat . ' mititee,.1 withnor harmful ertheless ~ ~ eztentof conditior ; junction ' -' evaluatio ' Variousc Techni ' 19, 50. . . . Snook ,. M E:;-0hortyk ,. 0 T Tnb. Sci . 1981; in press . Tso, T: C. "Physiology andBiochemietry of Tobacco Plants"; ~ Dowden,.Hutchinson, and Rose: Stroudsburg, PA, 1972 ~ Van Duuren, F. L.; Katz, C.; Goldschmidt, B. M. JNCI, J: Nat(: ~ Cnncer Inst. 1973, 51„703. Williamson, R. &., 29th Tobacco Chemists' Research Conference, W College Park, MD, Oct 8;1975. Wynder, E. L.; Hoffmann, D! Adv. Cancer Res. 1964, 8, 2~ Zane, A.;. Wender, S. H. Tob. Sei. 1963, 7, 21. Received for review August 12, 1981. Accepted November 18- 1981. Reference to a company or product name does not imPl,yapproval or recommendation by the U.S. Department of AI~' culture.. w measure , Wolfe, 1 ' measurec Lavy et a beenshor during a f et al., 197 Since it it al.,.1977). the dose, (Kolmod: Depart Universit (T.L.L., I ment, On (J.D. W J.
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390 B. L. IYxrrL and P., Z. LIN Table6. Mumyenioifrreccriny.oJhighconrenuvrionsrfrhreesmnkeeondertsarefrnctlonsin.Salmon- ella typhimurium No. of his' revertants/plate mixlStrain... Concentration TA100(without5-9mix) TA98 Iwith S-9mix) offraclion (pg(plalel Fmction... Neutral Monofunctional compoundss phenols Bifunctional phenols Neutrali compounds 0 IOg 101 108 29' 500 117 114 102. 47 1000 120 130 100 49 2500 132 145 100 62 5000 166 156 109 76 7500 183 147• Ito. 84 10000 160• O1 124 96 20000 Ot 0 147' 106 tThese concentrations resulledih toxicity which was apparent as a thinning of the background lawn. Values arc means for three plates except for that indicaled by a double dagger which is the mean of six plates. Those valuermarked with an asteriskshowk a significant dos-related increase poncheeretestlin the number of revertants IP< 00p. comumgen norharmam.using different amounts of 5-9 mix,.nomutagenic effect of phenol is observable.. The possible mutagenic activity of the crude and purified: phenof•containingfractions described im Figs1& 2'was investigated Table 5gives the resulis of testing one fraction o0 each type.. Results with all of the fractions isolated were confirmed in at least a second experiment. In almost alll of the tests the numbers of induced his' revertants did not reach a doubling of the numbers of spontaneous his' rever- tants. The crudest smoke condensates were also tested an 1000pg/plate in the presence of S-9because of their high loxicityat 5000pg.bul this also:produced nega- tive results. A suspicion of possible: mutagenie activity was observed im.TAItlO, without S-9 for some of the more purified fractionscontaining neural and phe- nolic compounds. Further experiments. (Table 6) at higherconcentrations.showed: a slight increase in the numtierof his' revertantss ina dosc-related manner up to 5000-20-000 pg/plate. These increases were sig- nificunn (P <001), as determined byy thenonptra- metrie Jonchcere test. a multisample procedure to test against ordered alternatives (increasing doses: Hol- lander & Wolfe- 1973). For Ihe neutral compounds a further distinct mulagcnic effect was.observed with S-9 using TA98 (Tablh 6). nrscus5ro. AND CONCLUSIONS The ten most abundunt phenolic compounds found imthe phenolic fractions that maybaused in smoke flavouringsfor meat products.did not show any mula- genicily in the Ames assay under standard conditions. Even when testing. Ihe parent compnund phenol, using a variety of modifications of the standard sys- tcm: no enhancemen0 of the number of his' rever- tants was observed. These modilications. including the use of S-9 ftom unindhced rats,, performing a preincubation prior to plating or varying the protein content of the S-9 mix (Ames et al. 1975: Bartsch, Malaveille. Camus er u6. 1980) are often, necessary to detect otherwise weaklyactive mutagens by offering more sensitive test conditions. The comutagenic ac- tivityof norharman has also been, described (Nagao. Yahagi, Kawachi er aL.. 1977)) and its use has been proposed to aid the detection of some mutagenic compounds evading detection under standard con, ditions. However ils use did not enhance the acfivih of phenol. These results are in agreement with those of Shahihn et nf. 11980) and Gilbert rr at.(t980) who also failedd to detect any mutagenic activity oh phenol in the S. rvphimariumm system, but are in contradiction to results published by Gocke er rd. (d980):. These authors used a different agar for cxpression of his- revertants and counted c. 34 TA98-liis' revertants at 50pmol phenol compared with 14 fon the control- Whether this low'activitv' which isbased.on the luu'% number of spontaneous revertantss in this strain is a sign of actual genetic activity of the compound remains to be clarified and could be further evaluated using a more sensitive indicator organism and teet system. The smoke condensates and their separate ftactioos alsoproduced Ihrgclynegative nesidts:.Smoke conden- sate.does.ihclude mutagenic polycydic aromatic h)- drocarbons. which mav, however evadedetectiun because of their low concenlr:uions in the crude 00 extract. Aslighl enhancement of mutagenic event,. %I was recorded in S. ra-phinumirmr TA100 for the neutral. ~A compounds Ipurilicd'according,tn Fig. 11, as w'ell a± Mi for mono and bifunctional phcnolic compounds of thr (p phenol extract. (purified according to Fig. _').This. (Q slight enhancemcnt of his' revcrtnnt yield IMh¢h N never reached a doubling of the number of spon" taneous revertants- but did increase in a dose-rdatd' manner) was confirmedwhen the experiment "a' repeated! usinghigher concentrations of the b^t samples. W hether or not tte are dealing with traces t'I true mumgcnir compounds. in thee fractions: whi are inducing reversion of the tester bacteria. musl tx ..
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r:II1tOMtlSU\tEPItA ,MHCNTXrIf1N' •f8a ,.. IIuu4>rnn;,0..1). IIIJA. Handhonk of chemislrc. aud}rhysirn. - Clivrlaud, Uhiir. 1. LEt~iN, A. I:1:Ri. Theeffecl o0solchieiur• un roul milosis iir:lfllurrr. - Ilnredilus XKIY: 4s1-a+o. il.. --- 1915. Cylolopieal reacliuns ludu.rd bc inurgauir :dl srrhdions. -- S:durc I:A1:T.Al IAJIi:.. HetvrochromWl•in r•hroRllnolneI dilriol;Ihrir ~~onlr1letion ph:lsu.. -- Ilcredilus 7:A\II: 4,0-eW;. t. Ltn'aa, 3. and Tno, J. II. H1{y, l:hrnuroxnwclrn-Wmralalino iudurcd hc Phenul.. - IlcrcJitas \XSI1': ses-:3:.' A. IA:ea9, a. alul OsTE1toREx: G:. 11H8. Thr merJh:minln of r-¢lilolii+ :uliilu. (1h- serrnlions mr lhe naphlli;dene seriuc, -- Itrrodil:u.NN/S`.:=il~-4ia. u. 1'eunteyrlER, R. and fh'sTls. Juu.. P. Illlx. h:iulr e1fecls of hmdrnquinouco0 milusis. ~- Nature 10. SiEINEG4EA, E. 1947. flelvrol)IoidiCrvl'Hlleho.uu .lr[nripllNrrru. -- YWII'm:rc. Ada H clc. 22: ra•;-yaJ.. - Il. S'fRl\F'.GCER, E. alld LEVA], A. IUI: n. (AnslilulioL :md .•-milolic aclirily af iso-colchieinr. - Hrr.dilas SXSIII::zO- ;m. 12- - and - 1917 L. The cvlolugir.dt effcel of' chlolrofunn and colcllieiue o0 13. dfrfunl. -- llcredilua X\Xlll:als- :mJ -- 15HN. TFe l'-I:/IIQIir[ /tflaht::•5 nl rulr. Ininrrllf)'I erdchicinie acid and two pheuanllrrrue dr'rivaliroa. -Ilrrediln I\\IF: er;---_+c. 1{. C/DTEIIr1RF.N,.Ii. 1914.. (:oIC111C111C nrilosi9, rlLrmm.. FOrnc CnnUacliuu, uarlrAis and prolcin elalin foiding -- Heredilns YSSrx_r-~~:~. CONTENTS. L f•r•nor:d ulinrrc:JU.u. .........__.... .-....._ ..... ............... ... II. >I/erial oL.r,rraliuo. .\. I'Ilrrod. auul .1)riuooo. II !'h~~unl . _ ................ .. .. -. I'v.rorarorLuf ........._.. ..... .~. Ci.urrivul ,.._... .~ ...... .. a'. Ifodrriqpinnur ................ . fr. 1's: ru_:di ~ i I ........ .. ... .... . . . G: Ilydrnscharh'uipriwmr~ ..._...... l. i'ldurr~:;lurinal ..,.........~... .. ri. P.Rrua,~•piinuae . ........_ . .... ..... .. ... .. . .... Iqf !1- ltuinhcdrune... ........_ IriP rt.\:Iphllv.l ........................ ..._........... .....,.,........ IUu 11 ;f.]npWhul ................ .... ...... ......................... {Gi 12. \uphlh,n~rsor<~ilr,l ....... ._........... 16i I:1; .lliz:u'io .......................... ........._............... ....... If7 11.. I•hruaullrrrurquli1Whrr . .. .............._._.. ................ IIiM It. .Vh_chdPd phrnuls .... ..... ......... .... .. . ....... ............. 116Y 1.1:. u-/lrr.ui .........._ ............. . ........__..- .........._...... .4II9
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REPRINT 87119416 ry, ie~9go s9 ~9_ 1Pk ' n ~ o /NMh-HO nsk WOIWUw Ltd. SCREENING OF TOBACCO SMOKE CONSTITUENTS FOR MUTAGENICITY USING THE A61BS• TFBT c-+1iI WTCt INGRR FIARIN•, LARB RU99RRAe, MARGARETA CURVALV .nd CURT A BNZRLLb efHpe.finexl ufEec4rWery, KerobYU4clne/i/u/e, gJ0101 StocAkolm, and k&ernrch OeyaMment 8wd4h To6atto Compmir. P.0. Bo~ l f0p1, SIO! 64 3mNMhn (Swden) (ReeeiwdYen>•n 10th, 1980) (R.rilion,.e.ind AprJ 101h,1i80) (AeceOted April 12eh, 1980) SUMMARY To clarify the mutegenic activity of individual smoke components, 239 compounds, representative of the gaseous and semivolatlle phases of tobacco smoke, were assayed for mutagenicity towards 4 hictidine-requiring mutants of Sadmanclls typhtmurium (TA 98, TA 100, TA 1586 and TA 1537). All compounds were tested qualitatively both with and without metabolic activation using a liver fraction (8-9) from Aroclor 1254 or methylchol- anthmne Induced uts. Without 8-9, only 2,3-dimethylindole and 2,8,6- ttimethytindole showed mutagenic activity that was not enhanced by the metabolic activation system. 2,6-D3ominntoluene end coranene, which like the above compounds un nGt documented carcinogens were found to be mutagenic for stmin TA 98 with S-9. Mutagenic activity was also observed for the previously known mutagens benz(u]pyrene, eh[ytene, bens(a]- anthracene, perylene and P-oaphthylamine, on expmum to strains TA 98 uid/or TA 100 with S-9. INTRODUCRON Strong epldemiologicsl evidence for a close co.relatioa between tobacco smoking and lung cancer In man ik available [1], ee ere animsl experimenta showing that tobacco ta le eareinogenie (2): In view of the facts that weh studies are laborious uid that there Is good correlation between mutagenicity Addre. all coneayundenee to: Prurevor Lns RutbaN. Depsrtnenl of Bseuriology, Rerulimka ImYtute, g-10101 gtockholm, 6wedeu. .aY IiptIC6: 7i1.= c..'arlat InaY he prot~4Gt4 py copy(19rt taw. (11ue 17 U•~ ~~ 219
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ant At.nFarr t.tcV<tN AN'D. AUh. 111N T.leu Most af'Ihef eu6st:tnccs Ilncc Lrnn buul"ltl 11'om lic6u; Slockholnl, un Scnncn, Lund.. tiumt° rarvsuVirLtnvc, n'rrc, kiudlp prucidrd hy Pru- fr,sur H. L•'nD'tat.t.N: Slurkhulm. nuJ. Pmfrssur 1.. Sa71TU, l.tl.nd• for tt'hich .ce Icndrr nur.inrrrc Ih:mks. rrnirumrritt runstrirtiun.. j.: 1°racoumis ucrucrrdin :he follulc.in," lir- .unte cnds tt'Crrt err.n. Ilt _ nunl7!in"' urruL 1. t': C-tnitusis in 2_._li- in 3 In'unuunrcu t'-mibt~is lv,illi .Ihnnq cltruuw.umt• ruulirtc.uiuu und Innrkt•d xumos. Alsu IIilTcrt•ntia .I:liuiu"s ul rl•nlt,+nucrirrcgiuns :wd chrnutn- uLsnrct•d. rl: l:-_y haI r Il'nuq loxirilc lc.itll.I a_plulinniliun ul' nViroma- Iwnrsii 1--13.(?I) huursl. i- Ill il dnn;. c: No rrntniu c-luuwurv w•rrc 2. Pyrnrvtlrrhn7. -- u: 1- Ia. 6: Ilnols suf'I: 14 IJ 6nursl. 7F (IIIi :Ind 10, pscuJurlli:rsm:dita und IrtnJrncivs lu I'ratimunl:diunc Wrre sr.•n. ,d' Illcuncchrunlowme tcu, . cut. ~-If. In.. +nmr Inarr concentrations, 9) unr CI'agtnt•nl Nuts srcn tr•ilhin :rn :Inaphnzicc c-pair. itl which unr pitsc . ?li. iila nlwusl li, : 100 por ornL 1: Pr:,(nu•ntnliun is vrrvv rare. lit :1 .1. PHENOLS AND QUINONES. 1. Phrftuf. -_ n:. li_.Ilt. li: Ifilut6 .uil: It la wiaL. ('.-_1 115 min.), 2 14 hnursL.:i (I ducl, a---L I I.a dacsl: r! 1:-tunuours uccuc from cun- cl•nlraliun a dute.n Ihrou;;L Ihr .crics. Itl Ili. Thrv rcactiun ib, kulvcrcr„ prescnt uuh' inphrnrzl - dislillyd wnlrr, lit runtruls willl phvnol -!- t:rp- tr-:dcr ltunuurs worcGmud unV\ ' in,A-e. d<.li- Y qiec slrnng toxic c11Act Wilh n;;aluliitaled. xc:JdkslniurJ ('6ruwo.snmcs. r: Gmitusio occursims qurucic in,eunt•rnlralii,n. 1- Ia: c.ni•.,~ntia. .. . l ., ~ 1, ti•.n N0. I ...... ........ !li Cr . _ ... . , . . . 1)7 .. li ............... N!I.. 7 -11 H ..... ......... r; Y !1 ...... .. ..... . . 1~(11). 1 ~.. I1/ ....... . . . . . .. 97 ^_I I -- - 1 I---~1:b .......... . 100 h"tr. mrinl/huso.- ll.t:c\N. 1!11L[ 't'rnJ, nric.<,.d c-mi6u..ic 1,.L for do.vn .li'nur, puiann t•/IltK-i: 1... -. r: /.unr L. .• :r. lit a cvrr ulcu'ar.lcri.lic. i) a. Rrznr(innL -- n.: I--lli, h:' Hauls sull_• 1'?.:• huur.s). ?.-b l; Imnrs7. r: (.-t11m!tUrv rnwlnp'u in Ilir nrgiun 7- Ili. •l: %unc I wilh
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{fi$' 'J.IiImJfr 1.1(\'.tN FNU InP:nll 'NlU slickitlcss IllpIglUlinaiiire ~ti umlrisl mar lc:n1d lo I7cts frnglncrnls of ehrumartic sub>tancv. 'I-hr.c ::nu r:lsily distinguished froln our frnGments, Ituwmcr. Fnrpmcnls ratncd lie Ilsvuduchiasmnlu alsu occur :Is .cell :Is Il'oRlllrflla whirll mrem 1rr 6v!r:rrncJ iudeprudr,nllyIrvstickiness and pst•ndnc6iusmul:l. UsunllV reueIrpe nf m:lciinn In'erluminales mithill 1111'SIIIIIC 1'ool, w•hile tlifl'rrt•nl rnnlc rll 114• ~amr In':dmrul ut:n• rrlttesenl differt•nt Iypt•s of rcnctiuns. .1s :lu in>lauvr ~:I' Ihis.. thcrrcnrdsnf :dl rools in 4 hours' Irc:dnu•nt in s :nr ;!i.rn .rpar:dnh•: nonr Su.n b.: .~r ., a., • ~:oc rnmph,h•Ic Sn. I........ . 17 1 n;;gIo111mL~d 3 Pr c,. h 2 . ........ ... 9 1 - - 2 5 25 3 ........• . . . . 3 41. 100 4 ........ ., . 1 . 1 . .. 34 11111 5 ......... ll 1 I !1 l; 1 2 8 75 Io rnots~Nos. 1. 3.nnd:r r:n,.in„ I~rrqucucV 'uf I~r,hmrnthliun is prescut. v hile in 3 and I mna dlrisirmu :Irr :I,~mInlinalt•dd m:my bridgt•s• pscndn- chin.mnta :ultl lnrtcr Ir:lgnlrtds bting, l:rrsent: Lclwccn Ihc :ln:lphnsr i;rnups. 14. Y7lrmrnf7lrrrlrguiur1nrr1 n. Irnnn's:llur:rlcd..:rlldian.lvil1rr[crs. uf mnli4sulct•d >v6SLrne:• duo~n, I:: lls• ,a«ralyd, %,Ilieh c<lar5ch`corrr.- pnntls tu n- !I. h: liuuls ~.n1U.5 - li: i I rlnci. c-j: Sa c-Iwnnurs.:rntl'nu e-mitusus 1wrr srcu..nor :Inc rrn~l:lin I l;lyn~ruls. I lr 5Ihcrc wrrc r:Lu•n'rd .r•ct•ralrnav. ul.Il.uud:,:'hiasnr,lla, lt. ALIt_YLATEU PHENOLS. Ia. o-/:•e~vol~. n: i. ..13. 7r Ilr:ute,.nlt- b. 2 1'5 miu.J. ; 4 Gnnrsi• -1 !i. !l dayL C: !I-- -1p •im;lcclunlnnrs.. rl: 1--?immcdiulrl}- killing, nuclcoli JeepipsL•tim•d., 2 Ilo. .rrv clrur I+rnc3trmmosnmic diffl•r- r•ntialiuns in mclap6:t"r--:lunpllasr. AII aal;)•> ufdissnlution of the rltranlosnmr w-nls- :.: ;P br,itrv lo .huw r..milu/ic lcndi•nc.ice,.; luls full r-milosisteittt mullleJ rhruminsnnlc.. SItns iulcrminy;lcd c-miinses :utJ m"rma lailos<•s. j: I:unrener::ltan ]c• ~~ k 9 W II Ytl1' u u rr":....-...... z s n 11 0 1
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CnROMC/SOME PRM: M8ST1\TION sides lo a muximum- Since llle lcft part nl' thiss curve iirs probublv u fuuctinn of two counleractiul; tendencies, uur fnvourin{l tra{~ntentuGiolt and one checking it through Snme form ol' toxic.action, unly the riqhl part of the curve. fronl the mnsimuln down lo the.lowesl cnncentration, camn be considered to give a si;;nil'icunt trl2resentatfon nf the activity of thcsubstance in question. The cur.cs have Ibc following appe:lr:mce in aumc of thee most active subslanccs: Cimcentmlion Nn. / .• fl 7 F 9 1n 11, . L"1:1 1. Ryrogallol .............. .- -~ :18. 47 f:i :t7 7 11 lY 0 ^. Itydruquinone ........ - 10 25 27 47 21 17 12 ~/ 11 3. HydFoxyhydroquinone .-- - 7, n. 1/1 14 2 0 0 1 4.. Ilenzoquinonc ........ - ---- -- I G 8 12 ?' 0 5. p-Phenylenediumine ... 1 9 13 36 14 IR 1 1 0 11 6. p-Aminophennt ....... e I a N 7? . 3 2 _ I. While Nos.. 1-3 and 5-6 are in fuir a;,7ermeul. No.. 1, benxoquinone,, deviates somewhal. This deviation,, however.. mny be. dut• only to the toxicity of higher concenlrationsIxinri more pronounced iu this sub- stance. The values.of the concentrnliunn 171-1:1 :Ire in good al,•reement with these values in tlie other subst:tnces: The dislribulion of values in the descendiu;; p:u'I of these curves is of interl•stl There iss a fniFlv good prnporliun:dily between cnncenlraliuns :nld fr 4;: hrnol . L..I,t ,RCh I[nua nmlI'. 1U^, Uhnl. 101, J•II Ins', Su JII ~ Pc rogallol lound ...: i = 66 3 : 7 12 0 expeclcd . 69 "_fi 14 7 :1 tiy drolluinun c lfound ~17. ?1 17 t? esPecled , . --- liil •~ :b /;t. li 3 I ( l'uund -... :ui It1 10 ; I {I•1 'houclrnv/l iaminc;I acpectcd .. aG Ili : 1 _ 1 t 1. I p~- :lminuphcu nl i....._..L..I .t/ .. I l:\Cn It Inc :I',rf'f'lltllOl IL LLnt \-lvt'":IIIC~.Inu III Lut lvrNl. tl/ltt~ :> n,. uVUl1C Illllt tnc .'3LLmi ul VCIICt uao U wlccl plVpuluL'nwln LVI inst:mcr, into :nt rxponontial prolaorliun:dily. .:1.... r• .... .......:.:..L. ;.,d:.•..L.~.~ u••,./ rl,i. r:.•,..... •,,...•..,•1:..•. t::IC :r\I:uuSluul•rrnl Ifuul ulin..n IuP.~nmrlr\ nlu:Onc fY'acunn5. ~uru I Ilnlh .. -
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174 L. f1. Grxsnurm dering stlmulatory effects on the regenerating rat liver were applied to. the current experiments (Gershbein, 1958,.1966, 1973 & 1975). For several agents, changes in liver-to-bodyy weight ratios were followed in intact rats. EXPERIMENTAL. . Test materials Of the essential oils bay, eucalyptus, peppermint and spearmint originated from Gentry Corp., Fairlawn, NJ, cadinene, caryophyllene, myr- cene, rhodinol, cajaput, carrot-seed, opoponax natural G• origanum Spanish, panley-seed, petit-grain and black pepper from Givaudan Corp.,.New York, cade rectified, cananga, cassia USP Chinese, eedarwood,, celery-seed, citronella, copaiba NF IX geranium Bourbon,.lemon, patchouly and ylang ylang from J. Manheimer Inc, Long Island City, NY; dtral, ger- aniol, isosafrole, safrole and piperonal from Matheson Coleman.& Bell Co., Cincinnati, Ohio, vetivert.(Bour- bon and, Haiti).and veliverol from Florosynth.Ine.,. Bronx, NY, oleoresins from Stange Co.,. Chicago, III., except for oleoresin celery, which was from Fritzsche Dodge & Olcott Inc., New' York, carvone, cineole, f.isopropylbet®]dehyde,,Umenthone and isoeugenol from Eastman Kodak Co., Rochester, NY, 4allylani- sole (estragole or methylchavicol), 4allyl-2,6-dimeth- oxyphenol, 3-allylguaiat.•ol, 2-allylphenok allyl phenyl ether, anisole, 4-cumylphenok, dihydrosafrole, famo- sol, umaco guaiazulene, guaiene, 2-hydroxy4-metM oxy hyde, 2-hydroxy-5-metlioxybenmldehyde, 4-methoxybenzyl alcohol, 3-p-meBroxyphenylpro- pr4nol, (t-methylstyrene, piperine, piperonyl akohol. 4-propenylanisole (anethole);.4n-propylanisok, allyl- benzene and.n-propylllenttne from Aldrich Chemical Co., Milwaukee. Wise:, and camphene,, dipentene, rrans-p-menthane, x-pinme, a- and y-terpinenes, ter- pinolene and wood turpentine from Hercules Inc, Wilmington, Del. The remaining oils and principles were furnished byFritzsche Dodge & Olcott Inc 5: B..Penick & Co., Ntrw York, supplied Spanish whole safron, sarsaparilla ethanol extmct and yarrow flowers The garlic and onion instant powders, ground nutmeg, decorticated cardamom and Jamaican allspice were commercial products (McCormick & Co.. lu. ' Baltimore• Md),s as were the herbal teas (Ah:; Products Co., Huntington Beach. Cal.) and thq ::. from Matricaria channmilla L. (Seelect DicuG , Products, Inc, Huntington Park, Cal.): Of the cham;- mile oils, J..Manheimer Inc. supplied four that wn: ; essentially free of blue colour (Egypt,. Romaine at: two Engluh, batches) and two very viscous lo: intensely blue-coloured oils (H-44 and H-5). A smpl• (H-1) similar to the latter originated from S B. Pen:i & Co. The herbs and other plant materials wa; ground in a Waring blender andsievedd prior to uY ' Animalsand treatment. The test materials weteaa . ministered to rats either in the diet or by sc injeclwa The diets were prepared by admixture of the agew° with Rockland rat meal, all percentages being akrr lated on a weight basis. For sc injection of the as and principles, solutionr were prepared in peanut 4: and the corresponding controls received the oil akue • in daily volumes ranging up to 050 ml. Prior to treatment, Charles River rats of spxif.i ' weight and sex were partially hepatectomized unA: ether anaesthesia, twothirdsof the or n5 bein xv- paled (Higgins & nderson,.l 31) and~ ned toaw- siant.weighrat 100°C.The rats were housed in inJiii- . dual cagesand given diet and water ad fih. Suppl!~ mented diets were fed for 10 days following surgu.• • and the ratswere then killed. Inthen se series, inlu;, tions were carried out dailyy for the first 7 daysaiic surzery and the rats were killed with etheron 10, at which time the entire livers were removed ar. dried as described above. Small sections of the oress were retained for microscopic examination. The bra increment, or the amount of Ussue regenerataL ay calculated from the dry hver weight by aakfng Iht , product of the weight at surgery and a factor of.0:r and subtractingg this from the weight at autopy (Gershbein & labow. 1953} Rats losing more tlu: about 12-15% of their initial bodyw•eight wa: excluded from the calculations, since starvatlon or s. anition cun cause extensive decreases in the liver in• ' crement. - In another series, intact rats were subjected k several! of the treatments over a period of 10 d!}s , Table I. Body weight and liuer increment findings Iar Pariiulll' hePOterromized rats injecredd sawith rssrntiul a0c compounds Oil or other test materiaU d N Body weight (gl and oset (mg/rat(day: oveiall) o. of rats Inhialj Terminalt Liver increment (g) I Series 2E Control IS 260 t 69 289 ± 7,5 2•244 ± 0094 Nutmeg (100;.2535) 12 261 t 8-1 ?91 ± 9.5 2-665 ± 0093 3-14" Series 3E Control 292 ± 99 301t 93 2-034 ±0092 Celery-seed(100;.2375) 9 290 ± 7.7 300 ± 11 2 2-472 ± Pn5 :R1 . Series SE Control 12 269 ± 87 29.3 f8g 2024 t 0o-103 25W). . Mace (100; Op 279 t100 2g0 ± 90 2421 t 0-102 274' . Fennef (Iql; 2415) I!1' 2g2 ± 11.6 299 ±10.9 2629 t 0157 3"9• Ytang ylang (I50; 3670) I I. 274 ± tI•1 298 t.7,5 '240 t U152 119. Series 6E Control 9 247 f.163 254 ± 12-0 1-879 ± 0-121 Cardamom seed (Ilq; 2705) Coriander seed (100; 27901 10 8' 249 ± 11•3 243 t.12-1 21i9±12-8 260 t 13-1 2•184±06131 313e ± 0076 Ia9' 191 • Carrot-seed (100;.2725) 8 250 3 98 264 t 128 '2?0. t 0,'-11 144 t Series Pan Oleu Seriei - / Coni E:;' Anis Cun• Eugc a-A1: Seriies Caa' Tarr Gua Series ' Con Nuu Series Con Tart 4-Al Carr Pan Series Con Saa: Can' Series Cor 4-PT Isoc Iws Series Cor Safr Series Cot DiL• Series Co: 4-A n-P n-P I- i; Tar. Srnr> Cu: All PiF hne~ Co All P-3 All Seric! Co Ia 3•( t 5<de C. Ga Gt Ce Ct Hnc pe ;lna Rew
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i4 hours)., stnined to slainini;: anilosis in ire. in Ittc Ihc lallow- ta I) 0 (4 honrs), rlion. r: t;- esin3and 11, c-milosis "erc 1113110,t s with the . frn;;mcnts Is sol'I: 1-2 : 1- 22 tiviWh me.l intt•rnal ,:t•utrnlion a p:lirs. su:nc- i clu0.%tustfisn•: r•nnaAtr:n°rA,rtns 4"i1 Coneentrnti0n No. I'. n n i n'll In. 11 12 13 Fr'+~14 hours •. 0 1 0 4 2 1 0 0 0 0 11 day.... - - - 32 - - - 24. llJdroquinonr mnnomelhllf r/Lcr Ip-melhuxvplrenol)..-n: 1- 13. b: Roots soft: 1-2 14 huursl.:i-1 {1 doysl, u(2 duys)'- c:7-13. d: 1Iws nhnust conlplctci * ~ lii.csnh-cd cllromusomes. 2 has strong pycnosis. e: C-mitolfetendencics in 2. I'tdlc-mitosis.in a-dwith strong r.hromosome conlraclion, in 3 lvndeneies lu moltlinR. -ahns mixed c- :Ind norntnl miloses.. l: .. I:nnrndrnlIu4 No. .., f- ] y 'l IU Fr ~' J4 hours ....... I U 8 8 5 2 ~11 day......... --- - - 33 1) 2 II IL 19 0 2 0 0 0 0 25. Blefhoxr,lhgdrnquinunc. - rr: I---1:i. 6: hools soft: 1-2 (4 huurs). 3-4 f t day), 5--7. '2 dtlysl. c: U-Cl. d: Zone 1: 1-2. Gun- mosume dissolution dreidedl}' less Ihan in Illc precedin;;t.co substances. c: (: mitosis. in 3-5, in 6 we;dc Irndrncics, inno cnnrrutration. however, completoe c-mitosix. f: ll.,ncrn1r:rtlun So. . •, ' N tC lu tt. r^. 1-1 Itours ....... 4 1 ; ? I. 4 II -t Irr '~: (l dkty ..... - t) Il -- n (1 (I 0 D. AMINOBENZENESANll AMINOPHENOLS, ETC. _II. aniliitr. -- u: ('.-1 fi. b: sut1~:(I !imlcutllv?! 1 Liolrsi. 2 11 dnyl', r:. a--4;. d': 7•onr I: c'„-1. r•: C-rnilosis in 2-1, completc in thr furmcr'. f: Surpri.iaqlr at•livr :ttilra;;mculaliun-induciilg a~cnt: n-nn~,~nlmmi .n s,.. a~ I, . u~ : n p~ In n n I:t 1'r 14 hotlrs •....... :1 1 1 I 0 0 1 2 5 1 0 _ I',I doy.....•,... -.- Il /). u 0.- 0 D 27. o-PltrnyCe-uedirtminr. -.: 1--1:5•, 1,: li o ol,ool, Galt-lirrgl'scent: 1-2 11 dayl. C.: •rt•ndeneir,v lu c`~Ilumnurs ill. 17-13.. d-e; In 1 a ntick}-lype nf o-milu,is uccure inirtnlliu;;lc•u wilh norncd amtphnscs with Drid',;r,. 2has.c-ntitulic Iendrncir., bul wm,l mi6ssets arr uorntnl. ,11'tvr I d:iv Lulll Iht•s<• concenlr:dinus Il:lir Iull c-tailr.sis inn iLr Irwdicisious. thcn ucnurrin:q. In silnilaritc la it, i,mnrry ;his .nlianncr is rlV:tract- rrizcd b.~ shu>ciuti~„iilar}-insiSl,ln•r Irpic:d r.,,tiI„sis lar duarn in H
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GIIII/I\fUtiUt11< F'H \(i\Ili\'.I!:1'1'lll\ 177 ducs nul .-1o (urlhci' dmcu helno' ihr w:dcr ~ulabiliia, thamn tIurt thv ncinliou l)ohc.ecn thexcC t.vo cnncoulr.diuu.. i,. t; 10. In inCt. Illis !ltrrsltnlrYrnnrrntrrdirzr7 t Inatrr.cnfubilily rrlnli~~n Yin•s un.iudic.nlinn u( . the nludr .d aCtion ul:t suLst:mea. If thi.ss ml:dinn I nxl;;idc mprest•uliu°il tltrrnmdyu;unic activity, as iillrndltcrd inlu Ih(- cirlcliiriut+ \•~urk Uc Gav+cnav. and cu-\rurkcrs). is less Ihan it is lo 6u: iuax•cir•d th:d 16r• wcrhanisnr ot nctiun is: nut pwclc pli~'.ic;ll. Lul Ihsll rr•rlain c!ICmlr.:d rcurGons are alsn.inruh'cd: Thus. in :I!lirrrn runic culthic.inc. Itas :I vrm• low t!vc}!wld r;[lur :tnd its lllcrmndynnmic :lClivifc !Ili'tv:lllrr nb- 6rr\'i:dcd. Ih. a.: )l is U,1.wr:l: STI•:In~F:or,tac :Ind''. LrVeN / 11117 :.. I!+IS/ uml STl I,1947) LIWCV shrl\\•n Ilt:d. lhr•.I,•nct rlr.uq;r iv Ihr• riu-, , C„( thc cnleLicinc mulecldfr r•limin;dcs its chcmiCUl :Iclivilr. lzn-cu!chiciuv :md cr>IcIlicilinc hact• lhs n.'sul' ll; -'a.nd tI rra!oclivt•Ic,. tilill, il" cidcllic6uris chvr"rd into trimrlhrt-crolchiciuic acid Ly Ihr reuiar:ll uf'nn ;rcrtcl:md nncu.•rnilh ul :In aminu !;rnnp, t'crnh chnmic:d :irIirilc i,. aequin•d. . 7"hr ulrst:mcos \corl:ed.vilh. in ihis+fudc. Ihr [i6r•nul>. ., \rcll ;r,. Ihc :Imimr... mae bt• czpt•ck•A lo hacc cternti¢nl :rclicit}': Phcnr,I \\:1, lr<Irdli)'tL\v.\cn.aN. Duur:.andPncsan. , I'.Ilal.:rnd hlrn,•rl ran Iu h;m• a Ih..:l. ut 11,,•,a. The santc aWhorsulso rvrnrd aniliur :IV cul iuO4ncr• „l' cLrutic•:dlv ;leti\-c suh.tmtccs Ilh..a. D:na.;'. f.lc['as :mrl (>sTb;netua (ayq:4. 1'i_. _, :Ind ilsrenr.ntt~-, ~ 1!!14 . I:.I'., +~lo:ar dS-:nrn up, rurrrl:niirn di:hr:mis ul'. c•milnlic Iln'r.hnLds :uld ~:~nlrr >rrluLililc in the Url¢cur nnd IthC nnlrillhrnlrmr• ~rntq}s. S riis;inrl .-urroLliiun\eal l.;un[t hel\crrn (rlllinl-, Ihrraxdd. :mJ liillin;; \c:.lrr vd- u6ililr:. It :r ,ituilar dia;;rmt is u•I u!l !hr Iht• prl lui +Uli~larcr... it i., fbuurf Sh.J Ihcn• is u\. cr>' rlr•:Ir rnrr,•!nlir~~n prc,rnl. :ind. il '!-r .,Jurs. 1Con1 ILr c:u'lice pnprrs:IrcinevYlyd inD' Ihr c:lmu din;:rau;. id i< .rrn Ill;d fhW uo\c suL..t:mrrs nrc 11,1nnlq,rd :dbUL: !lu• awr r''r:'rla{imu fi:lr. T!x• x!rrrcnling ;vnund tllc corrcialiun,.ilneii. Imn,- rri, ~. pru+rnun<'i;h. w Ilu• e,lrrtdali„n nt Iltu prrcrnl cu6~1'rmer, is: r,unid,rablr !.n.srr !io:ul thnt ol' Ilie eorlivr nuo.. F.lrccinllA- mlm, c:lrin!i•s urc ptl!,.:I ill,\, u- wnrd< iu Ilu• ri,hlpurt ul tLo ili:u;r;tm. \cliieh mr:lnv.lirlltli.. nrlicil••Iht'r:hidd i, Inir la\c iu rul:dium Io !lt.r olrn6ilik . i. i, il]r!il. n. i.: Lnc wllirtl, imGc:ilca.chrntic:d ar'lion. il i, qi'.itr ,!h'i!.iug that Cvliaiit •~cuups•.t..nbrlanrr. Icr\v ~rbwul !lir, n•.iu~dlcs. r.I ilir ,alu6ill 'fhn:c Ihr phrnnh hncr Ihrr.h~d~i. ISIIIII e:nie• :~iunidrr.illl\' ,c.i+idr 16,ir -„iubiliCnI:ICrordiuti ~!livv :dl bmr :1 lule Ih. ,i : ~ ~ (J ~ ~ ~ m
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{7u SI.It1:11'I' I.b.C.\N' \]IV ltlPl /eIN '1'.IIU llan•~eulr:dinu ]m. %. ~. IU el I~ ~ .~. ~1 luoult. .... .... 2 _ 7: 1/ 11 1 ° II Jav....._...... '" .. II I 7L'. f:;/t•lohr.r[n!uL :r. 1 13. L:. Rwuls.tnll; I 13.; hutu•sI..-/ !1 d:tcia r: ali-. d aud r: Iu I mid.•! prnunmtro:l sliekinc,s is prescul. lu ?:t cerc characleriltii• pieltero r.-adl.: .\II annts uf Ihc separniinr.:uta- plYasc rhromusuntr, h:nr Ihii•k lrrrnin:,l pnrlie:n,. Wleich cs•idenllvcunsist of'ari;lulinalcd mall'ix vuhstaucr Ilualiiig :tunl lo.vartls the cliru-c wusutnc cntl... Iu I:md? Ihrrt• ar.- linlnd crull'uBtcric dilPrrt•nlialinus.. 'i•hrsc cunerulr;tlirms :Jw. h:lirt'Irun~ r-milusr.. nllhnu."h ~:nlv. p:trlial. f: l:ccluhe~nnul is. c.rn' inurlico. Imb, url~:rllu•Irs. more wul:mbled I"roe I'r:li-mcnls wcrc. I'uuud llmn.l in plttnol. Cnu.. nntratLm .o. r :. s. .~ In. n r-- u~ ..'~k hnttrs~ ........ , U~. , I u !1 ;I !1 I! (f , I r .. II [{nl' ........... ll. II !L 11 it . _ . 1 ._ _ .. 4. Ili. CONCLUSIONS. 'I'hc subslauce, t•utPlnvrdl in Ilti, itlvrsli;(:Ili:lu ure uC :: suutt•what tliflrrrul h'pc Ieum Ih-• inditlrmnl [. mit::Iie•ub>I::nrr. 'lltev nll cuul;till reactive groups. ltcel!'ow. i:rlo .,r nneinm, 'Phe :~rcemblauce iu Ihvil• eflecl on wnik•l' is .Irir.ia;;.. In hi;;h cuiiecuirntiirnx !hrr Lnw• all imtttrdiair killiug :uii[:n.. T'bi. zouc I p:ts.rss uccr iuR,, tlice r-milimis zotne 2. and lurlhrr In•Imc, Ihix:unt- Iln• I'r:rqu:[niiu, tauorn- 1'r:ltiuus slarl. 7.but• 1. rslrudi Irum ,:dur:di•d cnnt'rnlrulinn duNtn lo, cuncculraliuns.:tl tehii•h Ibc r.dluIar drulh is rrlardi•:I I,:n-, ruuu;h In. :illutc Ihc c-milusis lu In• uctttd'esl. [I. Ane rh."'• an;il~sis rl~ turc-milnsis lut•t t...itli hcrt•lalls tr.ulsiJr Ihr uf Ih,, pn•,t•ul paprr. unlc ntt•ty I:IrI,N rluvaclct'i.,lic ui lltvr~uiik:•i- iu ucl: sult,lnures as Ihosc hrre cnlplosrd! will ht• discuz.rd. :1, iswPlt kuaacn. c-tnii~:<k i, :1 hpivai Ihrrslwld n^!rtinn; it d,Ks nul uccuel bclmr:t rcrt:!in eum~ruli:uinu. 'f1!c Ihreshuld is uul uhw:t!'.s ,hUl'h. IInnvCFP.r. ItIAI in sume nl Iler .nbslnuces cnncrrnrel in lhis studk, .+iu,lr ornilns'es :tud c-tentleucics in:n' br I'nuud lnr helmc !hc cuuct-n- IT:Hion selticll luuvt Itr. ruu.idrrrd Iltr Iln~t•?ltmld.t•:mu,•ult'ediun. In ivust ul Ihr::t'diuarc r-InilUlir suh.lant'~r. ull Ihr n:!rruiics Ispr ali i!dnrrstity; r!;rl'rlalibn has ticru t'mu[t b:•hc+•,-n Iltcir ::v.licik Iltrosiiolds and crt;lain pilenic:.114rultrrlirz.:rl'ruet:dl Ihrie w;it"rl~.wdle Ih,• acli.:ilv dneg 1 rdalio thnzA the nl, the th LdYhl Inecllg rcadi, lowp hreria STntCI lhe C, and c, is cha and , acquil T the ai tested a Ih.: of clir lL havet solubi correl: ubilih found ft'otn /hat t! Tlte sl wtlhc Ifuti r w!trds litrrah whtch Ii 5!tale /itrech 19-LJI
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8'7119419 TABt.E I (Centlnued) TABLE I (Conlinned) Campuund tbmpnund Comment Ctreenone Carvon Antophenune 1-Phenyl-2-bulanune 1-Phenyl-l-LUtenon. Tqted 'qdvntitrtledy With TA ]00 eet.d quentiuU.Ny With TA 100 up to 3.d-DlhydrouybentddeEyde V.nllOm Ventzumeldehyde 6-htethYlturfunl 6-HydrouymethylrurNral 3 I d l h A Predpit4n -I d 8 rnwl/plate • n o eeai axalde yde 9-Pyridineeeebcieldebydr mone n 4Fluurenanv '$0,8-TrimethY41,4naDhthoqulnone (YPyddYl}t-propenone AldeM1yde. Propand Sutmel Precipilatee Piecipit.tee PSmo4 Phenol ncreed m-l4ezol P;SeW 2 8-Dl lh l h l eted quanHtetiedy 30 pmd/pl.te th TA 98; tasic at PentmJ d H • me eno y p 2,4Dimethylphenol Teeted quaetltaHwly With TA 100; tmdc at e.en Heqan.l at.w NonanLL DedenJ nd.c.uY 1-bt th l J Ned qu.ntltAwly with TA 98 md TA 100; toxlL.t S 3yntal/p4N 2.6-Dimethylphenol 2•B-DimetbYlphena 8,4Dlnuthylyhewl 8;f-Dlerethylphend ,BS-Trenethylpheltol 30 pmol/pla. ted qeuUtatleely With TA 98; toxlc at 30 Mmd/plate a y pmym 2-Helbylbutand 2-HetAylbutanal ented quentitafivvly with TA 98, TA 100 ind 2•a,STrimethylphend -Td th l h d 2 d Te.Gd quuttiteHeely with TA 98; tuzic at 90 rmnHplate d Ht l T R li ith TA 98 t i TA 1535; pmeipiteta Y 30 .mul/pl.te me y y en , , e quan y w . e n ; ox c vt 30 ol/ i l 2-Hetbylpsntend 2,Y-DimethylVropenal Propsnel - Preciplbtu pnt ted: 0.3 rmd/plct. (highly tbxie at 2-Bthylphrnel 9-BtLylphend d-Bth l henol p e e Mm 3 pmol/plale) y p TAymcl y-hLthylpiopenal Teetvd qhanlibtinly with TA 100; toxic at FS~meN i hand 9~Eth R-Hmeml 3pMmo1/plete y - et oayphen 2-Hexend 2.bHex.diend 2-HethYhR-pentenal eeNd quentitzHMy With TA 98 and TA 100 u to s md/ late oay .e I-YethosYPhand 2,3-Dimelhauyphend 2,&Dhnvthdzyyhvnul rtM quenHteHevly wfN TA 100 Tested yuanYtitieely With TA 98 and TA 100; 8 f d p r p preelplYtee at 00.md/pLte e nn Phenybthanel 8-Phenylprop.nJ 3-Phenylpropend Bemeldehyde 2-Nethylbenxaldehyde 3.bkthyltienzJdehyde t-tdethylbenxaldehyde. 2Hydroxybenvldehyde 3-Hydroxyhenuldehyde .Hydroiybcnzddehyde i ld h d Preeipitatm Prtelpitelee Preeipitetee Budeml Lomyend 2•Hydroxyeeelophenon_e eB-HYdrory.eetoyhenane d-HYdroxyteetnPhenone a-Nephtd T•Nepbld Catschol Bnuminal Hydroyuinom S-Netho.yhydroqulnone .rt.d quentitaU.ety, dWith TA 98 and TA 100 Highly toxic In epot teet Highly tozie In zpot tett Teeled quentitatieely with TA 98 Tetted quantitatlerly with TA 100 T.tted quentitatieety with TA 100; toy~e at An e y w e 3 umd/plete 224 226
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482 al.lnilcr f.l:~-ts .\sn Jux InN rnu und I)usR9tv !ILflffj havo IwwnIllal hvdr..quiuuttre enu I)e injected iulo. Inicc intrapcriiunnall}: nr ,ahrul:,:acnusl~•. iu duses stranr onnug3t tao give a ccll rr;tcliun. rcidrullt' id.•nlii•:11 tcitlr nur I1ra;;Ptcntitliun reaction. :\ Ibslilrg qumirr rl .Mulr 111 wul,, vtn.t aclirr adtsl:lncc.e is Idannvd. SUiNIlV[ARY. .Wnut furts >ldtsltwcos ruulaiuiu;; pllrnulic r)li ur iandl `E[s have becnIt•sted trilli Ihr .ll!(urn-lr,f as I-o Ih.•ir aclirilv in iitdncinqchrmnu- sotue frngnlcnlntiou. It was i,mudlihnl ;dlnust :dl nl' them had snmuc activilp.:dlhuu;;L grurrally a ratllrr lutrnnr..'Illcfalloaving su6slancrs tvore musl.urlicn: locrugpllud. iintrnqpinunr. p-pilrnplenedintrtine..pt•ru- calcchol• p-tuninopllruol. 6vdruNNhcdrnquiuanr: Ilenzoqui'nuuc. In. Ult+. tuasl aclivesubslanr•cr.munu tb:ln til).';; of thc In•:dcd cclli.mavs conlaiit .. fru~lnents. T6e.fragmertis tcerr uf kcr, lcpes l:unlpleleh '. Il'ee frngments, avltirh were lcf4 in Iltc cqna6rr rr•,iuu :d auaplmsq nud att:lc-Ited feaements.. which were Inutglnr Ibnrlher ~,ilh the rest nfIhe cltrmuosnme Lv a thin thread. In addition, thrrc uccurrvtl 1'rnl-mrntx elue lbo sliekiness. these prubaGly ..beingnnlaullrntnnlin.rdrulc,g and Irnglnuuls.dun to tht• hreak- in;f uffnf Iltr. clirnmosulnc Ihrr:ul Ircltvrrn a cruLrnnr.rc autd a-pscudu. cltiasma". lu n1'i•w . r:tsr, sivlrr rlrrnlualirl rrtmiim was dcutnnctrnlyd, hul, un lltc whrdr. Iranslur•tli„u.. tcrrv c~arrntelvrarc. •1•ht• rhrumrrswurlr:~~nlrulnli,rn ucrurrwl illl a ttuncuulr:diou. zune. slarling nl O.na-11.i¢mul I irt ,umr suisl:utcrs nlurl lower iu ulher suL- sl:lnct•,, rr:rrhin;" dumn I', •!1.:,.xm,t nrnljl. \Irnvc Ihis zn1lC IhrrC. nccurrt•d inallnusl :dl ,ul~,laun~, smmr c,mcrulnllirrus.shnwiug c-nlilusis and furliu•r :ibnvv ihis z.mc iilm•r •,A vnrrrrl -.U-un~l luxic ~•I'fCclstrith c;trinus ur~;;ltlliltntinu Pitrnurmuil. CLr c-ntiiriir threshold tc;r1 ofteu siluated 111rtltet' dincn Ih:r,l tL.+ nf ihrxcalrr olubililv cuncenlralinq, trltic.h indiealcs thnl iunac ,rl !lu•,t. ,il6dunrr, imluce ranila<is liv a speeif'ic clmmical :tclinn. LITERATURE CITED. I. I1,tuLrXr:TnN. (:..U. auJ liuu•P:o. P. C. I:t!J, Tlm chnt,ical,llrt•akupe ul Yhramn. .umcs. - Ilor-.lih_' I, _. r:evArnts.. I'.. f)ul,r. \I. .nd P"n',..nr.. /L I:IIB- I.'imporl:uwnr Ir la unli•w d'aai,dL~ IlLCrurudcnamiqur rn Insil„Ib::ir cI cmph:vinarudpnnntir. -- H". Yrat'. Stall d-fas. 131 I... _...
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cu¢oMostaNle rn.tc.vcv'rvrlun 4(il I oau.rd liv luck ill repnodurliun. \I'e pualrnnr Iht• elosrdrecripliou o3' Ilir psuw6~rltia,iuala lo a IoG•n acr:tsimn, Itcin; ILrne onFc runcrrnrdl,,il'i p:vt of Ilte'e chromosomes varies rerp mue.lt; ill some casesouly tha xmalle.t prochromosotnic part of the anaph:tsc c6routosomc is stru It, pruct•.•d lo lhe pole (Fig. ?tL.al 7 u"clock). Those +muli chrontosontrs :u'c utlen nhead ot• the lnr;;e c6romuvoumes duriu.g Ihc polee w:mdt•rinl;. If Ilte pull on the eenlromere is Ihe sante iu :tll chromosomes,. Ihose which have lbst tfit•ir :u•ms should IurnisG Ir'.s rr<islnnrc lu Ih[• plastn than the tmdama{;etl chromosomes. Yi;;. j qives some instanctw of :dt:ICIScYl and free 1'rngments at=onapirasr-li•lupliase. Wltilr°rr contains ooh• crodrd chromosomes•, It and c huer srt•er:tl clearly t7eo 17agmeutx. ,'. _t d aod r nre more esthcmu nrutitrslalion, od" this rcaclioul in r, pracfcallc nll cltromosontes Lei[t', pulverized inlo >nroe liltpft•a{;menls. 1'i;;. t!'r. repre+ents :t rare cusu ul° clo~unwsuutr Lridpc. trhich has nuI Lcen caused by merc.stickincss. Ilt i., prub;thh• a trrmi4rccrtion bridge. %thic6 is cin exception:d occurrenryv in.Ihcsr• trentnteuts.. .Vter nmstard ;; t. trrnlumut ol' Ilte samc maleri:d suc6 LriJress nss indicalr vSstvr rhrom:tiid reunion nre uur ol' Iheemmuuouca rharnrt•. mcl .r.ilh [S-rxtul?cr•[in and LEVAN, nnpoLlisht•di. Pi;; ?~f nuJ !t arc lab• nna- phases aud I is a tclotrltase w'hert• two Il'aptornl. hotv,• iurmed tuicro- nuclt•i :md onc. Inrticr fra;;ntenl is loilerin- I':trutt`in Ihr Itrripfu•rir-plasm. Par:dlrllp ill the IrilgIttetll:tllollb :III(1 tll[" Iolilmtliun of censtriclions Iltrrr occurss nnulher churnctcriair phruum~•uun. wltic6 w'• u,nall)' nrcord Imtirr Ilhe Jr.it'lno/iuWut pstudurhiasmnlo ,. II cuu.inl+' Il/ Ihr hnn:.iu;;li~l"etltel• :d unc poinl ni' Iltchcn .vp:orniin„ ~isier :umpltnse cltrnmosutne<• wIVich ofl'cch a pulliu.l., utd. Ilc tltr covlrunn'nrn lrl' Ilw purliooy pruzimnl lu thn psuttduehi:tswy ioln thin 16I'~•;td,. Whilr Ihc I%cochroutu.utuc. pcu'Ic dist:J lu tlii.v I*uiul: tm"w :t pairIII lrauscr[5[' o ruta.. This Ibrm:tliou li:n':r arikin" rrstanLi:mcr to ;i ruJ bfc:dent rritli u nom-Icrtninnlirrd cLlnsina. lu :t I't•a' ranr, unr pacud6rhi^ctno ba. i,rt•n sreu ou cnclu ul thetIcu ann>s ill n vltar:tlhinr :uctllhaw chrumo:- .onur pair: u6ich Ihus cw.tuncs Iltr appearanrr IlI n[iir;. 1•iwilrnt. rrtnainiu;;, ill thr rqualtn• Ion" :Jlcr the rr.l nl' Iiho clirrnuusome., h:tnrrarhed Ihr pules,..l'sruducliiastotnl:nlt:nr liern dohislvd In- IT•nu.lse:r[7s cind {~UIJ.PII: i IU{7.I. whu uriLv Idtr a.vuntpliun th:d16rsc i'orn:alion. :trr ['uu.vd. 6% : lack, ill repruducliun ul sneill pnrl. af Ihr cheutm•zumes. ]'1•[clsp:oun>n nnd: I.er.t~ Iunpublishodl I;ncr ,rrn Ilwm in iual'r-rial ,nal.•d Ic.itlt utu.ltod:% g;ts.aud l.r:vnN ~ ItutyuLli.6ril9hn, rru lht-m in' Irr:dmrnt< .illi n~•.~•ral iyury{meic Salls, Sinrr p.rtudnrhiu.mn[n utn\.idrra•Ir occor nl'.lin :i athnrl Irr:dine.nl- %rv du mol a',lt'v Ilcit. Ihctam•
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474 AI.}3EIPr t.1:V.1)\ AND .H)1: HIN TJIU l:nllCraltl':IIIDII Ne. J Ur ^j' 1 Ilolln .. Il •: ~1 11 .{ I 0 11 n i) 1(laY.... -- II' 12 0 9 17 O. 3 0 ;Vtnchcd Il•ai;mcnlk trcrc recordcd in ll)..I I and 12. pacnJuchiaslnnl:( in 1. 5 and 13. N.. NTTROPHENOLS. :id. n-Nllrophcrml• n: I--~la L: liuols sol'1: 1-2 (1 hnurl.:1 i-4 huurs), {~-:r 41 do(.N;, fi- 7 12 d:l.~s). c~: 8-1'L. /i, Strong toxicity in (-_.2. in 2 cit:ll strnrlum. Nri1h hrteruc•Irruulnlc of l6l• cunlrunlcric region, tltis Lcing t•sprriadlgl6slinr/ on Illcseruntl daty. I!-5.h:trc.pnl- ntulnced slickincss. r: I:-mi(usis ilr'L-e, in u c•nmplrle c.-tnilosis (If bow- IVpc. (: Rragmcut:rlion nnly in isolatod. cases: Cunocu~rmiuu Sn. e 7 R. !, m Iu 1d. IA 'r 4 hour> 0 d I) I 1) I 0 1) I tlay . - /) 0 I.r+, li: O 35. nrNifroplhrnol. - rr, : I--1:3. h: Iiools sail: 1--2 !instnnllyr, 3(&) min.). d(-l 1lnurs), ia (;l dacl. fi--7 (2 clapsl. c: ill-I i..J: 7.oue J: t--3, slrung.slickinosv I--F. hi 1--9 di.sulutlon of chrmuo,uules wilh dislincl prucllrulnns:nuic /iilfvrl-nIi:ISians. r: C-Initusis in -F--7..IraceS ul (.•mil(isis also in 8, G hns l;lirit• conlpirle r•: nlilusisul buw-1Ype f: Goll.'rntrelrm: NL. ., 9 lu 11 12 1J Fr 14 huun.. ...... 11 1) u I 1) 9 ( 1 tlnc ... . ... . . . 2 _-- 9 11 --11 :1li.p-Ni)rnphrunl: -- :r., 1--Ia. ir:: RnnL; soCl: 1-2 (liusnutllrl. ;t I:1Q min.l. ~1 I-k LaursI. 5. 11 dap/- G-N (2dacs)2 c: J--L•'. d: Strnn;; lo\icitc: I--3. slirkinrs.: t---R. In i, Illoc6rmnosnine- arr ollun urolltk~J. in (i.slainctl l•hrnmalin tlrnps aro IIxmrd frum 1Bc nutlines of Iht` chro• n(usulncs: e:. C-mitosrs in :I- U. ecorywLerc mixed witlr nonnnI nnen. J- I'Fa;mrnl sccu onh• iu unr rnsc (II?i :intl' hc•rc }mssibh' nlG•IChrd. :li. Picrie recid. - r¢ 1---I:1, !+: linols soft: 1-7 (1 hour), S 4 h(ursl. !)i l dayi: c: lll--7P? /;: Stri-nqfnsicilr.: 1--a, sliclainess nccnrs ulmusl 111run~llntll fhc~~~~Itolr scnii•x (I---911 r: 1:-milo<i.a is luund in .I- .li.., cuutplrli• in I--.'.. Slrmuil '1•nnng;h: 1rclow this 1•.c;;inn in 1i--8 Ihrrcisgrrat dr-!Irucliun uflllr. eilnl sIrurimres. In (i IILC axlorn:ll Invrnz ul Ihr rnnis.hinr rlrar Ilrurhrnnlna/luir Iliilrt'rnli:dinuv.. /: I'rap;(nruh ~ ~ Illa N to ~ Q riT
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. 206 Ind. Eng. Chem. FunOam... Vot 24, No. 2,.1985 Fhrure 6. The dependence of methanol yield on reduced denrityfor pyrolyeie of gueiacalYor 15 and 90 min at 383 •C. The influence of water concentration on guaiacol pyro- lysis is further illustrated in Figures 3, 4, and 5, where, respectively, the yields of catechol, methanol, and o-cresol are plotted vs. reaction time for parametric values of re- duced water density ranging from p, = 0.0 to 1.6. Figures 3 and 4 show not only the general increase in catechol and methanol yields with reaction time and water density, but also that the influence of water is continuous and evident at values of p,.,, both greater and less than unity; similarly, the decrease in the yield of o-cresol shown in Figure 5 is a continuous function of water concentration. The influ- ence of water concentration on guaiacol pyrolysis is further illustrated in Figure 6, where, separately for pyrolysiss to 15 and 90 min holding time, the yield of methanol is plotted vs. reduced water density p,.,,- Methanol yieldsin Figure 6 appear directly and linearlydependent upon the initial concentration of water. As indicated in the above,.the conversion of guaiacol was rather insensitive to water density over the range 0< p,,,, < 1.6. More careful scrutiny of the dependence of gusiacol conversion on water density, at a given reaction time, suggests that at low values of p,,~ (0 < p,,,, < 0.10-0.15), the addition of water actually decreased guaiacol conver- sion relative to neat pyrolysis; guaiacol conversion reached a minimum with continued addition of water and, even- tually, increased with water density. Relevant data are shown in Figure 7, where the yield of guaiacol(N(:u/Ncu.o) is plotted vs. water content for theparametric.valuesof reaction time of 15, 60, and 90 min. Inspection of Figure 7 reveals that, a4the highest water density investigated (pr.. = 2.1), the conversion of guaiacol was greater than that observed for neat pyrolysis at the identical reaction time of 90 min. Discussion The present results indicate that at 383 °C water reacts with guaiacol in a manner formally equivalent to hydrol- ysis. This hydrolysis reaction provides the hydrogen (and oxygen) required to redirect the selectivity of guaiacol pyrolysis away from char and methane (Ceylan and Bre- denberg, 1982; Bredenberg and.Ceylan, 1983; Vuori and Bredenberg, 1984) to catechol and methanol. It is inter- esting that the hydrolysis appears rather conventional-. That is, methanol and catechol yields were continuous functions of water density with no apparent discontinuity aC p,,,, = 1. Hence it appears that the primary motivation for operating at supercritical conditions is that water is both hot and dense, the latter attribute having the usual accelerating effect on a rate that is positive order in water concentration. However, itshould beewted that the ability of the SCF solvent to dissolve a normally immiscible co- reactant could be a profound motivation for operating in the dense region.. At least two implications to lignin processing are ap- parent. First, just as water provided the hydrogen (end oxygen) required to improve guaiacol pyrolysis selectivity, water could provide the hydrogen (and oxygen) required to process a carbon-rich resource such as lignin to lese carbon-rich products. Second, although neat gusiacol pyrolysis yieldedlittle methanol, lignin's aromatic meth- oxyl groups are appealing (Allan and-Matilla, 1971) but chemically unreasonable origins of the modest yields of methanol that arise from the pyrolysis of actual lignia (Allen and MatiOa, 1971; Iatridis and Gavales,.1979; Jegers, 1982). During neat pyrolysis the reaction of aromatic methoxyl groupss would occur predominantly by fission of the woukor phonoxy-mothylbonds to phenol mul nwthane precursors rather than fission of the stronger phenyl- methoxyl bond to benzene and methanol precursont (Ceylan and Bredenberg, 1982; Bredenberg and Ceylan, 1983; Vuori and Bredenberg, 1984). However, present resultsehow that the water that evolves during lignin pyrolysis (Allan and Matilla, 1971; Iatridis and Gavalas, 1979; Jegers,1982) in a typical yield of 20 wt % might read' with the aromatiemethoxyl groups. These could thus be eources of inethanol even in the light of the bond energetics suggested above. Finally, the present results invite consistent mechanistic speculation. A set of elenientary reactions that explain the observed product spectra and also qualitative kinetic oG sen•ations is presented in.Figure 8. Neat gueiacol pyrolysis can be interpreted in terms of the set of elementary reactions shown in Figure 8a. The first of these nonchain steps is fission of the phenoxy- methyl bond to catechol and methyl radicals, which cann in turn abstract hydrogen from guaiacol to yield catechol; methane, and two guaiacol radicals. The guaiacol radical can undergo either A-scission, to yield a phenol radical that can abstract hydrogen from guaiacol, or disproportionate through steps such as those outlined in Figure 8a to form cresol, catechol,.and char products. Pseudo-steady-state solution of themeahanism of Figun: 8a yields eq 2, 3,.4, and 5 for the rates of guaiacol (G)) decomposition and catechol (CA), o-cresol(OC), and char appearance, respectively. -d(G)/dt = 3k,G + kb (k,G/kap/s (2) d(CA)/dt - [1 + a]k,G (3) d(OC)/dt = dk,G (4) d(char)/dt - yk,G (5) Note that the mechanism requires a correlation of o-crerd and char yields. Our extension of this free-radical mechanism for nat guaiacol pyrolysis to account for the influence of water'a shown in Figure 8b. We model neatt pyrolysis to ocma in parallel with a hydrolysis pathway that involves solveltd guaiacol (G'). This soWated or partially hydrolyzed guaiacol (Gf).would exist only at dense(not necessar9yy supercritical) conditions and is prevented from undergoilg the free-radical reactions (Figure Ba) that are accessible to the unsolvated guaiacol (G)) by admittedly nebulow "cage eftects";;subsequent addition of water to the solvated guaiacol (G') effects the hydrolysis to catechol and 8'711936'7 Fh dei mi sh, th all vir to* the ob: I am apl are -d d(C
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TABLE II LIST OF HYDROCARBONS NOT FOUND MUTAGENIC A8 hydroorbom .en IerYd s demibed in Table L In adElYon, they wan tested quantihtl.ely ud.q TA 98 and TA 100 with and without 8-9 from 8-methylrholaathnnw indund nta: The ronnntntlone med ren 0.03. 0.3. 3 and 30 p mollplete anlese otheF .Ye indicated in lhe Table. Aa Indinted In the Table mmt of the hydrocarbouz oe tede to the Eacteria. Cununent Toztedwee •mnkz/plate n-Eieusane Tested up to 15 pmd/plale precipitates at 16 ynndl/plat. Limoneee P,ecipltateratJ0amo1/plau > 9 Cydnhez.e. Praipitater at 30 umol/plele a-Pinene Redpitetn at 30 ymol/plae > 8 p Pine > J Benzen. Toluene s0 m xylene 30 P%ylene 8o Ethylbenwne 30 v-Propylbenzene > 8 4opropYlhenaan. S J Stynn 80 Phenylretylev. 80 IndN 30 ntnun > J Indene 9o LfVhenylmethur Aaulen. Naphthalene > J >.J ~ 8 1-MNAylnapt6zleae > 8 2Mmhglnapthaltaa I,bDimet6ylnaphthJen. > J s 8 •Aauvacen. sPh.nanthnne pynM •Pinne eAnnaphth,ine •Aeenaphlhg4w eMuoranthene Teeted up tu 8 umW/plzte Tnl.d up to J Mmd/plab Tested up to 3 umal/pYte TeWduptnoAeamol/Wte Pne/pit.lae et 30 pmol(plate Predpitatez .e 30 pmol/PIaY Tested up tc 8 umd/plate •Diewlved in dimelhybulfoxidea carbons in the ayar, and that the hydrocarbons constituted a limited number of substances, they were also tested quantitatively u9ing S9M. The 9 substances found mutagenlc in thls study are listed lu Table III. Five of these are polycyc8c aromaYNc hydrocarbotlp, 2 9xe methylindoles and 2 e® eminee. Of the polycyclic aromatic hydrocarbons examined only 6 ete mutaganlq i.e. benz[u)anthf9cene and benJ[a]pyrene towa.ds TA 98 and TA 100, i 87119421 TABLE DI BUebTANCEB FOUND MUTAGENIC The mutz9enle activity of the aubzt.nme zhown /n this table waz either MeaOfled In spot tezY or lo quantitative te.Y. In the Table en indiuted (a) the hyh.et daee tested, (b)1he number of teverbote obt.ined at the opumal dom (the number of apntneow ntvtenv /. auhtracled), (e) the typ, of 8-9 und (&9A- 8-91'rom Azeclorlndueed raY, &9. - 8-9 fmm S-methgldlo6nthnna-Induced ratt). All eduez ue calculated averages from the remlts of at knt 2 empedmentz Ni9heat tnted doee l / l ( t ) 69 OpimYdote (Ymoleafplate) Number of nnnan4(pLte nmo ra p a e TA98 TA104 Hydroearbov •Benz/o/anthncene 3 B9tt 0.100 36 326 eChrynnf 1.5 89, 0.005 0e 196 •Benz/alpymne J 89, 0.003 233 255 •P.ryls` 0.99 89y 0.096 91 0r •Cnronena ' 0.6 99N 0:090 82 Oe N.hen,veyeke B,J=Dimethyllndak J0 = 2.6 99 a.t.6 x,JS-Tzis.ethylindda 30 - 1.U 918 355 ANinr CNaphthglamine 80 89p 1.0 n.ta 886 2,6-Dieminntoluem JO 89A 1.5 1088 127 e DWalwd in dlmetbyLWfnzlde. ` Not elpnifiean0y above bach9rouvd. a Nattatedt e Wahly mvtyenic alru without 8N. a O 0 03 Ot Pmule./plpte O 0 I 2 1 t+naet /plate Fig. 1. Dun taponm to wronene of TA 98 eHer metsbolic activation. Fig. 2. Dnre nzpnn to 2,IdimeiAytindoN Ic TA 98. No met.boNe aetl.alion, 228 229
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' TABLEI LIBP OP VARIOUS OROANIC COMPOUNDB NOT lOUNO 10 86 rAtrr.nnurr! The aubrtencee wen tested In rpot tete uany the ctrdne TA 95. TA 100, TA 1636, and TA 1637 with and wiWout E-p fram arneler-Indueed ntn Bech eubetutea se teeted at S mmoyphte unkw otherviro Indir.ted In lheTable.ln ahrehn of aback6round Lwn of baNeia as the platea (indka0nt todelty) the tut wes repeated with a la.er canmotra- t'wt of the subtutea: BubrYna. ylrinB an unnerldn m.ult in the apnt teetl wem teded quan8tatirdy at 4 condrntretbn keeb (0.03. 0.3. a uid 30 pmollplate unleu other.he indic.ted In the Table) with the YNnU) shown in the Table. 8omw subbnoec pmclGl- taud on Ote plaba (eae Tabk), ,eeulte obtained .Ith thae whetane., ue difOcult to aealuate. Several of the leatad aubetanoee ate toxic to tb ba-b.Ctark and their mate- penidty eun only be t..ted below a mrlain dae (ne Table). Most wbelanma wen d"utolved in ethanol, Ihoee marked e were dlnnlvd Iv d'uuelhy4ulfaxlde. Cempound Comment Aleohoh Methehol Ethanol 2-ProPanol 1-0ctidec.nol Pndpitale. .-Terpinecl - Cennld yamewl Pr.elpitatea Propylene glycol Purfurylalcohd Benryleleohol 2-PAenylethand E/ken Miwle Dlphenylethei a-Metbylanleole CMe/hylaNwle 2-MethnqneThthelene 3-EthaxynephtAdena I,E,aTdmethoiybensene TeWd4uantltWrdy.ItbTAl00 Adde 3,bBiAydAZyben.ae add t8,6-I1 i mAhdxy-4-hyJ msy W oac eNd •Nkolink.cid TutedqumOtatt.elydtbTAlOo Indoly4a-¢.Lk acid •Kynunnk eeid Taaad quantitati.dy vkh TA 88 up to 8 pmnU plat. apot t.a 0.8 nmallplata •TeiephMelk edd 222 TABLE I (Confbwd) Compound 8(fan •nd auhydrida Dethyl mdooeb Bthyl elevate Vinyl aeetaR Methyl crylate Bensylaeetete Ben.ylberuaate Bensylelnnunate Coum.dn DlethylphlhYate DI•n-plopytphtAelale Dibutylphtlulat. INOCtylphthalaM Phthdk add anhyddde AmMea end nfl.dee Nicotinunide NifelhylnkotinamMe Acetonlldle PropionitNe 9utyraaedh Lobutyronlh8. Vaeronllrlle IeoNeronilrile AcrylonitNe Benaylqanide Nkotlnonitr0e Indulyl-}satonluile ]tetonn 2-Roputone 4Butanorre 2.8-Enlan.dioM 2,8-Peneanedlone a-8uleo-2-nne a-P•nten-2oue • 8-M.Nyl.6-hapNn-2vne 8-Methyl-8,6hep4dkn-2one Pcwdeienene /-lun Cyelopenlanon• 4ydohes 2~STrimethYlcyrsbh.x.npoe Pipedtone 87119418 Coouvent T..Ied quriNlatiedy with TA 08; toxlc at > a pmol/plale; preeipitatea at 30 .moN pLM PreclplYlea Preciptatm hxipitatee Tested qusntiLNvely with TA 98; pndpilhta inepotteu Te.ted quantitatirely with TA 100 up b a umoUplate Terted Vuantlt.tirely wlth TA 08 T..ted quantitatiwly wlth TA 93 T.aed quaotitatlwly, wwith TA U T.etad quantlthti.ely w1Ul TA 100 uy fo a umoUplate Teated quentitatiedy with TA 100 Tested quantitatively with TA p8 and TA 100 Tested quenObtiedy with TA 28 and TA 100 223
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Liver regcneration und ssx7nial oils I81 nsccigila. :ujiir K, Jaffe, H., Bishop. Y.. Arnold. E.. Mackintosh, Long, E. L_ Nclson, A. A., FitzhuglL O. G. & Hunsen. :aet rats D. & Epstein..S. S- (19701: Structure-au:tivity relationsW: H. (1963C Liver tumors produced in rats by feeding aomiYOi ar methylcnedioxyphenyl and related compounds on safrole: A.M.A. Arrhs Peth. 75, 595, o no scs hepatic mierosomal enzyme function, as measured by 1 otlikac. P. D. & Wasserman. M. B:. (197:1 ElTrtss of lisecmid prokngationofhexobarbital nareoslaand zoxazolamine ssfroleand isosafrole pretreatment on A'- and nng-hyd- s under- Faralysis in mice To.nic. appl. P/oonnac 16. 462. roxylation of 2-acetamidullunrene by Ihc ml and ham• ~mburga t'°rshbein,.LL(1958). EOeuofcarcinogenicandnonear- ster.Biocdirnr..J.129-937: - tsia lht onogcnic hydrocarbons andd hepatocarcinogens an nt Opdyke, D. L. 1. (1974). Monographss on fFagrance raw .vere ItA lisxr, regeneration- J. nani. Cmrcrr Insr. 21.'95• ~ materials- Sufrole. Fd Covner. To.eiroL 12. 985. Cmshbein,. L. L. (1966). ERect of various agents onn liver Parke. D. V. & Rahmaq H. (19701; The induction of hepa- rageneration and Walker tumor gmwth in partially tic mierosomal enzymes by safrol6. Bicrherrr: J. 119. 53P.. h:pateclomizcd rats. Carcrr Rn': 26. 1905. Ruhcrnann„S. u: Lewy. K- (19'-7L Uherr das blaue 01 des Oenhbein, L. L. (1973)t ' Psycholropic drugs and liver re- Bmunkohlenl"iencratorteerss und uine Bexicbun@ zu the eor. -erneration. Res Conunun• rhem. Path. Phanrate 6..1005.. dem Azukn und den Sesquderpenen desKamillenoles: md Ptt.t Gersh&in, L L. (19751 Liver rcgeneration as. influenced Brr di. rhe,n. Ges. 60. 2459. 1. Eiiak bl thc structure of aromatic and hererocycGc com- Seto-T. A. & Keup. W:.(1969k ERcts of alkylmctho.ryben- anh.tiina pwnds Rrs Cormnun..chrm- ParhPlmnnuc. Il. 445. zene and alkyimeth)lcnedioxytxntene essential oils on' of S. B Grrshbein, L. L• & Labow. 1. A. (19531 ERect, of various pcntobarbiuland ethanol sleeping tima .-Irdrs irn. Phur- shon, w0urcompounds on rat liver regeneration. Am J. Phy- nuarodJm Dnb. 180. 232.. .of. 173. 55. - Sharma. J. D. & Dandiya,.P. C.tIB6?I. Studies in Aconr.s tinil..T. 1. B.,. Parke, D. V., Grasso, P. & CSnmpton, R.. rolumus Part Vf. Pharmacological actions of asarone F. 119721 Biochemical and pathological diBerenca in ' and /Lasarone on cardiovascular system and smooth hepatic responselo chronic feedingof safrok and butyl- muscles. 6rdian J. med Rra. 50. 61. sud hydroxytolhene to rats.Biochem J. 130. 91 P. Shulgin. A. T. 119664 Possible implication of myristicin G•rmthrr. E(/94 8-1 9 5 2 1 The Essrnriaf Oils. Vols I-VL asa.psychotropk substance. A'arurr. fand: 210, 380, res, T, C B Van Nostrand, Ina, Princeton, Newlersey. Shulgin, A. T. & Sargent, T. (19671. Psydrotropic phenyi- rnno8= it.r.a: E C., Jenner, P.. M., Jones, W. L., Filzhugh, O. isopropylamines derived from apiole and dillapiole. uu•er Rrs 0_ Long, E. L., Brouwer, J. G. & Webb. W. K. (1965) lGamre: 1-and. 215. 1494. ToJcproperties of compounds related to safrole. Taxic Taybr, J- M.. lenner,.P. M. & Jones, W- I. 119641 A com-\1iBer. E :ppL Pharnmc. 7. 18. parisorr.of the toxicity of some ullyl. propcnyl- and pro- occurrm{ BqPrrs G. M. & Anderson. R. H. (19311 Experimental pyl compounds in the rat. Tacic. uppl. Phurmuc..6. 378. e and tk pulabgy of the liver. 1. Restoration of the liver of the Thoms, H. (1903} Studien Ober die Phcnol3ther. Ber. dr:. u. cn Re, shitesat following partial surgical removaL Arrks Path. . chrus Ges. 36. 1714. II186 Thoms. H. (190g). Uber franzSsis:lte> Pelersilieniil und d-anu & &-cyurger..F., Kelley. T., Jr.,.Baker: T. R. & Russfidd, einen darin entd.xktcmi ncua Pficnolathcr,. eln 624. t R/1962A SexeReet on hepatic pathology from deb- I-Allyl-2.3.4.5-tenamethoxy3knxnl. Ber dr. dirm Gr.r. c h)Pow :au diet and safrole in rats. Archs Path. 73. 118. 41. 2753. ole. Aim. ;nner, P. M.. Hagan. E. C. Taylor, J- M, Cook, E. L Weisx G. (19ht)1. Hallucinogenic and narcotic-liLr rlfesas k Fnrhugh, O. G..(1964). Food flavourings and cwm of powdered Myristicu Inutmcgl Prnldw. Q. 34- 346. 1zt0ani, prmds off related structure. L Acute oraCd toxicity. Fd Wealcy,Hadzija,B. et Bohinc P. (1956). L'in0ucncu de sfatmo+ C.m Taaicof: 2. 327- qudqucs essences sur Ic systeme nurveuxcentral des jaianobdr lau K. /1942} Untersuchungen Bber den Azulengehalt poissons. {nnls p/nrm. fr. li, 283. hi 19. ?- •x Fbres Chamomillae und der deraus hergestellten Wislocki, P. G.. Borchert, P.. Miller. J. A. & Millen B. F, 1195YS ,6rmazeu0schrn Zubereitungen. Arch.. Phorrn., Btrl. C.(1976E The metaboliractivatlon of the carcinogen ~tatnearm llg. 424. 1'-hydrox)saGole in ufw and;in ritro and the ele<trophi- rac6u~ i+icB.G.&Parke,D.V.(I972).Inductiomofarylhydro- lioreactiviliesofpossibleultimarecarcinogpns Cancer ~. 14,155s wbunhydrosylaseihvarioustissuesoftheratbymethyl- Rrs.36. 1686, mstiluua yrJroxyphenyl compounds Biuchrm- J-.130- tGiP. Gea 2). R. 1197: t ,nisok- m :drok It
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SLRY.tt'I' l.h.l':\S C1'i1 .IOE IIIA"t'•Ilr). as c-milosi._ c.tunwurs, e•:im phm--rr•nclinn ill ycast, etc. In iltce fr:tg- ntenl:tiion il isnnl Ilre (taraii)nul :t llu'esitulrl rvactiuu Lrd ul':I reacliotu m:ulifcainr il.rll' in Ire~~I,nrliuu:Jile luIhc nnuthcr af tnulrctdcsol' the nt•li\r suhal!nce•prrseiil. '1-Irr :Irlin~r zuuc nlnll ~edistnnces strctchcs ai,uuF equally !'ar do\cu. \lusl ~tf thr sulislances hncc a crrq louw urlirilv. \rltirh"dnrs ool rnron Iha tlrrl'r Ihrnainld is high, ILOt th;ll Ih:•ir Ilo '1, i. low iuIhr eulirc ir.ur. CvrIVdlrzaltul4 fnr in.l:tncr. \chich i•. uf rrl•N ' Imv cfl'icicucc. c~hiUils im lhrannly,ed zunc. a-La. 1 I•rngnlcnt in c:rehu ill thc concenlruliun+ I. !f nnJ ;fl, Iliis ondouhlcdlv sho\rs sumc ronvmhlanec to 111c vlruu_,,s! Icpe ul' c-milmisN uccurrin;; in sotncnf Ihc .u6.lancrs nindi,d. r. g. ill u-I,hrn)'I:audi;uuinr•, Mherc c-tnilnsis is prcxcn/ pxr!I' N' iqlhr lliShrsl tnnt-t ~ulrutions, I muf Y•':nd p:vll}' in.snrnc cvr\~ sulitarc case•s in Ilx. !o\cr,l runccnlnllirnta. 9 aud 11. In u.nilrn- phenul, where Ilic Ihn•ain!d is ihrulr-d :d; h, Ihc lrn•srncr• iss rrcnrded nl' pcrfecllr h•pical <•~nlto.,•s ;u. liu• du\cu ss ill l I. Thc cen- worphnlrl.t;c ill 1!lc lra:;uuulnliirnx.indii:nlrs Ih:tl nu :dt:tck is madr towardsccrlaiu puiius e,f Illr shrmm~wmr•.. Tltis c.an.ta.iilrollg rr<nll iu dillcrcnt ultinl:th•rlfucls, irertrnrioleols ur seeuodnrv con- slricliuns. \ChclLcr 111t• p.rudurhiu.nr•da :ur :rlsn lo-.bc includet!'.in tGis cuntplrs ol' rcaclinns. or \vhwIhrr Ihrc cuu.tiltrtr.quitc onothcr effect, fs nut' Ic.nown• Thc tilgure. uf ToLl.- I n<.,rell :es sontu d:tl;l ;;ivrrt in the sl,rriitl chapter give in'tonrv•s nf Ihrt truqunucc rut frec... rragpleots urcurriu, pcr cell, This Ih:d \\•hi!c a-lh \rilih 11 fr:lglncnl% arc cunnnuurst Ihcir frcqu,-ncc Ih116 frum 1- .-i 1ri.•mrnl. pcr cell. Art illlcrcainS nh>rrcaliun mhiclt dnc~. nul nppror ill Iht~sc datn is the Ivr.e•ncc nf siu"'It•. crtls aith n:rc nmm• Ir:taurnlS. :11/-a6. Io 1•^.rt. Iho rnlirechl•uutn.umr aclt c:ut Lr romp!etulpy i4o,cu up IViri: 2 rll .\ siutil;u' cunJiliuu i. alsn .':Ilid ill Ihe rnsr nflllc iodrrctl cun.,iricliuns !Pil-. : rr~. ISoside•. I1LC nul'm;tllccxpcclcef ruosirirlioms, "mr• In :' I~•m prr ce!I, thrrn will he found cerlnin ccll+ in ., hicSt alt rhrumosume, tire r!rerheapt•d n•ith runsirirtiun:...lltrchrrmrnsmltrs :Ipprv'iu_ likc ,Irin;;s oil Lrad.. li Ihv,io!tlalra4mcnts:InJ rnnatrirlinus mar6c cc~g:ucicd:rs pninGeff,•cls in \chich each d:utm~c conreSpunda lit ooe Incaliscd rltemical :dlark, IheW IruicerizeJ ur ct'wdhd rrllz uulst be cunsicler.d :Ins duee to. anmu rdhcr cbfccl. It scrnts :1+Ihnurih the cnlire ccilul:lr phrsioln~g iuILesc <•:ISCS has bcen s\cilchrd occrintonn nl'1-raacl. IJu Irpiit", lu.malrc acump:n•ivun nOlhc te;lyd sub.lanrrsacc,jrdiit;l lo Ihcir fr'xglncnhrtian actieile onr c:,n uunsidcr 01 i Ihc nonthcr nf cuncunlnatiaus \cith 1T, rmcnis. 12 pl;lr ruasim:rl' 1'r ':, 01 carh .uitstallcc. This fact sysle R'hice orlln dccid ihese the l sirinif livint at Ih, Ihis I t-rn,w ho\vr srrm muct tn 11 sprea firok c:tsc: lislkv large root fragn hich \cithr hirihl
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(32) e) (33) 1, indi- ' HNOr leoualy !nt was ure. (N'= ) (34) , (35) a) (36) he mo- en cor- iing 31 he mo- del was model. (eq 24) oncen- :aea the rrected ucially, md the u have ), from esses is eactant +ary for the al- vn, but : water : water ;ofthe I water ~enta in ta weU, ysterrrs ised for -d used process udying dcohol, Icohols. )ritten- -iety of Sfound /nd. Eng. Chem. f.ndam. 1985, 21, 203-208 for data sets 5-8. For butanol, the value for Ao wass taken from Marcus (1977)) and a good'overall fit was obtained. The established additivity of Fr and Fb calculated from appropriate data is an indication, though not proof,.of, the validity of the proposed modeL A few additional conclusions concerning the dissociation of the different acids in the.alcohol cen also be drawn from the model: HCl is fully dissociated to ions. H2S01 ia dissociated to H' and HSO: , and also possibly slightly to SOt~. However,.diseociation to S0' eannot bee signifrcant; in view of the accuracy of the correlation that did not take this effect into consideration. HNO3 appears to partly dissociate reversibly to N20b This dissociation is a function of the amounts of total acids present. H3POlis not significantly dissociated to ions below concentrations of about I g-mol/kg of alcohoL Acknowledgment We are indebted to IMI Institute for Research and Development for permission to use its proprietary data in this study. Subscripts aq= aqueous i = species NH = unreacted HNOs S= solvent W = water Ptr'°( .=AN 3 4 'S$B Registry No. Water, 7732-18-5. Literature Cited BroMey, L A. AICJIEJ. 1973. 19.313. Cepa,, J. D.Sc. Therh. TetA4dHVW Inselule. of Terhmb9y. 1059. CMlstlnl.E. D.;,HL,°n, N. A. InQ. Eng. Chem. 1939, 16, 266. EYrore. K. L: 1mt9el0, J. 0.; D°m, R. L J. Pny. Chem. 1965,.39, 3520. Eneerby, J. E.; Sapar, S. In "[onlcilpWr", Imwn, D.;.lcverYp, 0.0., Ea.; Pbnuln Reu: New York. 19e I; p 7. FreOentYa, A.; CmeliL J.::ILurvnren,.P. "V.pw Lipa7 Ep,bYan U.MO UIIFAC-: tleevb/:. AmqerNn4 1977. Gd. J. F. In ^P'°Ce°b,ps ol 1SEQ 197T; Luoe, 6. fL;,Rreal, G M.; SmMt, H. w..:Ed.:ISEC. 1977; pp 316-322. GrY,W°m, B. D.Sc. TMtY, TeWlarlrnN IataFAe of TerL,docy, 1983 pn /bEre.l. Hebert K. L ACM Tn,a. Aeri:Seaear/ 1111111. 7. 1.. IMI ImtlWte fa Rerwch an7 tMvsbpwa. InMnW tlapats. 1969-1963. IMSL Inc ^IMSL Ibrry Itefwence MenW", 9m aa;Hoaaam, TX. 1982. Ku.k, C. L: Melerrwr, H. P. AICriE S)T,p. Su. 1976. 7(, 14. Lbtrke, J. W.:: Slouplmn, W. D. J. Plrye. (.riern. 1962. 66. 506. Merqs. Y. In "lonb UpJei". lnmeny D.; LarwYq, D. G. FA.; PMOn Nomenclature Pnu:[pw York. 1981;p 97. Mereu, Y. "Intr°6ucaon tuL1qW SINe Chrrb4y-I Wter: Wsr Yak. 19 7T, pp 197-254. a = activity Mebwr, H. P.; Kuk, C. L AlrilE J. 1972, ie, 661. A, B = van. Laar coefficients AI, Bi= coefficients M.wner. H. P.: TeeMr, J. W. L.l. Eng. O'wn. Plnceus Du. Der. 1972, 11.128. ^ Ar = concentration of water in alcohol phase in equilibrium with pure water E = percent error in correlation /, (" J" = unit correction factors F~= bound water concentration Fr= free water concentration . VoL 6: Msl°r, J. W. "ConpMrntM Trealhe of Incr9aMe Ca.nYSey Lonpnrn: LaMdi,.1956; pp 550-670. Meyer, R. J.; Plelsrhe, E. "OmeYn IYndbla'a der AnmOenWS C1rIW": Vslp-Qix,Ye, weAUYn, 1936; Vcl 113. pp e16-6Je.. PICer. K..S. J..Pbyt. Chem. 1973, 77, 268. Prr.r, K.S.; Palper. J. C. J. Mp. Q'nm. 1960, 61,.1N. Prex, K.8.; S9xsUr, L F. J. S°b. Chem. 1976, 5. 269. Ptmr. K. S.; Ray, R. N.; S6wetr. L F., J. Am. Chem. Soc. 1977. 99. H = linear coefficient. 4930. K= distribution coefficient, chemical equilibrium coefficient N = number of experimental points Rencn, H. In "Fartla6en °f Carnp,aerAMed CnenYcal Prepw t)eYpl-. MM,. R S. H.: SNOar. w. D., Ecl.; 6qrrMq Founoetl6n: New. Ycrk, 1961; p 53. X= mole fraction or molality in aqueous phase Y = mole fraction or molality in alcohol phase Creek Letters T = activity coefficient e = standard deviation Steplen, 1t: Steplrn. T. "Sphb6fy ol. InayrYc Corryouqs"Perpem°n Prea: Cat°r4 1964; Vol. 2, Pan 1, Tab. 2320-2374. SxpruWN E. MSe. TM.4, Tetlwa~IsnelInaeMe of Toclnobgy. 1376. Received jor review December 22, 1983 Accepted September 10, 1984 Influence of Water on Gualacol Pyrolysis J. R. Lawaont and M. T. I(leln' Peparlmaa a Clwnka/ 6pMMtyl LiNaney o7 rJSawaro.: Newrk- Oa/eirere 19716 The Nflumwe of water on pualacol pyrolysb was examYnedtfrou0h a series of pyrdyses spannkg reduced water densl0es from 0.00 to 1.6 at 383 °C. Neat gualacol pyroysls yielded catechol arld dlar as major products and mGlor products.Including phena.and ocresol; methanol was not observed. Water provided a paraqel reaction palfwiay that was fomlay eqtlvalant to fHalaod hydrdysis to oatedld andmefflerlol. The overal: reaction 6eleclfuMy to frycroysb was a oontkKlous function of water Aeasny at the ccrld6ons eramYled. The observed product spectra arld yuslltatke MneOcs were sumnadked In temis of free-radk.at steps for neat pyrolysis, to w11k21 guamcol salvatbn and hydrolysis steps were added to account for the Influence of water. Introduction The appeal of using supercritical fluid (SCF) solvente in separation processes (Paulaitis et al., 1983) has en- couraged their use in the extraction of volatile products from macromolecules, such as biormsaa (Koll and Metzger, tE. I. du Pont de Nemours & Co., Inc., Wilmington, DE. 87119364 1978; Olcay et aL,.1983) and coal (Wbirtehead and Willianu, 1975;Roes and Nguyen, 1983; Squires et.aL, 1983;Bleasing and Ross, 1978;dezko et aL,1982; Barton, 1983; Vasilakos et aL, 1983;. Fong et al, 1983; Maddocks et aL, 1979; Smith et aL, 1983; Amestica and Wo1f,.1984). Product yields are generally dependent on both the resources and the SCF solvent extrectant, and Squires et al.. (1983) have recently demonstrated the importance of thermal decomposition 0196-4313/85/1021-020350t80/0 ® 1985 American Chemical Society (~ C'/c t ` a
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ANSWER 1 AN CA82'..(3).:14.151..e TI Essential oil of tobacco. Phenpl fraction AU ~Devreus:, Mic:hel; Viart, Pierref, Esnault, Daniel: LO Go SC DT CO PY LA AB A Tab., Sect. 1, 11, 12 11--(Plant Biochemistry) J ATF:tCAA 197a Fr Essential NaC1 soln. i s er h extd. from,tobacco.powder by steam distn. from a the powder was suspended. The distd. oil was fractionated t thinr-1'ayer chro content of the commonly in v after fermn. appeared as Phenols in eval+_rzti on Appro;:. 18 ANSWER 2 ve a phenoli fractioniwhich was analyz.edd by gas and tog. Theyieldl of oils was .apprx.1'l% and the phenol. eta oce 5 was .apprx.1%. Among the pbenolsfound someoc.cur le materials, both in freshly harvested or later es. Xylenols wer-e.e alsofound'o but these usually A reeult he toba+ a~y b phenols w AN CAS5'~( 9 )'.. : 59f]49s TI Studies on compositions of smoke components of lamina and midrib cigarettes. Part I. Comparison~of phenols in smokes of lamina an midrib of flue-cured tobacco ALI I.shiguro,, Shi.geog Sato, Shizuko; Sugawara., Shiro; k:abLU-aki, Yoichi CS Cent.. Res. Inst., Japan Tvb. and'~Sal!t Public Corp. LO Yok:ohama,. Japan 60 Agric. Biol. Chem., 40(5), 977-82 SC 11-7 PPlant Biochemistry) DT JCO Ar:CHA6. PY 1976 LLA Eng AE To reveal the difference of the main strea.m.smoke.compnm between l.amina~~and midribof flue--cured tobacco, the yield of neutral, basic.,, acidic, and phenolic fracti~onsofthe smoke condensates of 1'amina and midrib cigarets were compared.. The compn. of the phenolic fractions were also compared by glass capillary gas chromatog. Neutrall and basic fractions were dominant in laminaA smoke condensate,, whereas an Et20-insol. fraction was dominant and formed ..apprx.5R"/% of the midribs,nol!:e c~~ndensate. For thesemiquant. ana.l. of phenols with gas chromatog. mona- and di.hydroxybenzene_s were extd. fromthesmoke condensates and loss of these compds, by oxidn. was prevented by addh. oi DL-ascorbic acid. After trimethylsilylatiom they were simultaneor.rsly e::amd. with gas chromatog. Apparently, 4-Me, 4-Lt,, and'4--vinylcatechol in lamina smoke were much more rich than those in midribsmok:e, whereas coniferyl alc. was more concd. in midrib smoke. ] of special curing processes, e.g. fire curi.ng.. :o ar-ebel i eved of i mportance f or qural i ty e related to.the phenols appearing in the.smof:~.ee ere obsd.
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~ TA6181(Canpeued) Cqmppuy,d pyro{Jbl 3-Melhoryutedd 8-MethylnUChul 1-4eproYyMatechd yY,ane., fhlnPMnet Tetrahydroturan 2,6-DinmlhylNnn 2,BBenzotuiau oibekatu+ut ThiopMne N•Mfrmcyc4s Pyrrolidin. ]-Plndiue Piperidine Pyrmle eCaibmole 9'Ethyltarbuok Een.imidstle Indde 41Jethy6ndole 3-Methylindole Pyridine 2-Methylpyndine 3Ethylpyridine 9,6-Dimethylpyridine B,6-DimethylpyrldNe 2,7,8-Trimethylpyddirm 2,4,6-TeimethylPyridine 2-PYridinole qhinolhn Pynune 9-Methylpyraeine 2,6-DimrthylPyrWna NieuUna p-Nirotyrine M.bmine Hannen 226 87119420 TADt.E I (CunNnued) Cemment Cnmpound Ccmmnd Amiua 'feded quanUtetirely wilh TA 98 and TA I0G 1-AmMoLuwe TeeYd quanUlatively with TA 30U tode at 30 pmol/plete (me Rewlt.) I-Amino-7-methylFropme Teesed quanWaUvdy with TA 100 and TA TeRed quantitatively with TA 100 1637; tazk at 3UrmoUplate 2-Aminupentane 8-Amhmpropen. 8enaytamine T.aed quanUtall.ely with TA 100 ad TA 1637 T.Ned quan4tativNy with TA 200 up to Aniline Tmted quanutathely with TA 1537 3 ymNJplat. 3-MethylanfliM Tlrted quautitethrtly with TA 98 and TA 100 8-6blhylm9iue Tuted qumtilatiMy with TA 98; toale at e-Methylm9iw Tmted quantltaUVely with TA 100; tave at 60 pmollplate 30rmeuWne 2.3-Dimethylsn8iqe TMed quandtaUvely with TA 100; toxic at TWed quuUtaYVely with TA 100 tll mstad concentntimu 2,6-DlmethylanuNe 9,&Dimslhyl.nl9ne Teated quanUtaUrdy with TA 1636 and TmNd quanUYUVeIy with TA 100 np to TA 1637 3 ymd/plae y.d,8-Tdmetbylsniiine Tutedquentitatiwly with TA 10U; ta.ie at 3opnmhpiete 2Ethylanilina TeaYd quutUYdvdy with TA 1535 T.Wed quentitaUvely Mtlt TA 98 FEthylan0;na Tmted quentittetively with TA 100, TA 1636 N-EU.ylaniline end TA 2537; prceipitrtm at 30 pioolfPlete DlDhenylemine Preelpitatee Prmipitetm e-Nayhthy/amine B,41-Dla.ainotolwne TeM1ed qutntlhtlvtly with TA 98; toxie at 30.md/plrte Of6ermmyuunde Ta4d quantitatiwly with TA 90; Mdcat DimeNyLdfosldt 30Mmd/plete Tesmd yuantitetireb with TA 99', tode at 30Mm01/plete Tulad quantit.tively with TA 100 Tuted qu.ntitatively with TA 100 Tat.d quentiteUVely wiW TA 98 and TA 100 tme Reeult.) Tmted 9uenutaUvety wlth TA 100 Teded qumntetively with TA 96 Teetud quantitatbely with TA 98 and TA 100; lazie tl. 3 ymnl/plale; proeipit.tea at ]0 µmul/pleW. 15 pmolJplate but gave no effect without prelnnuhation. When pteincub9ted witb 69A qtlinoline we1 found to be very toxic at eoncentntlons above I pmol/plate, and no mutagenic aotivity could be demonstrated at lower concentrations. It is pruible, howerer, that a mutsgenic activity can he demonstrated by testing different 8-6 preparations at varying eoncentrm tions [4,16]. (c) Harman, an indo) derivative which did not exhibit mute- genic activity, has bean reported to have a co-mutagenle effect (16). The hydrocarbons which gave negative results are detailed in Table 11. Since .il hydrocaubons initially gave a negative response in spot tes6 using S-9A, and since it is known [171 that 3-toethylcholanthrene is a better inducer of hydrocarbon metabolizing P-q50 enzymea than is Aroclor, each hydroearbon was also tested wing S-OM- In view of the fxts that we experienced some difCtculties rclated to the limited solubility of the hydr0- 227
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' Materials and methods Tester strains and media ' 5 histidine-dependent strains of Salmonella typhimurium, TA1535, TA100, TA1537, TAl_ 538 and TA98, were obtained from Dr. B.N. Ames (Biochemistry Department, University of California, Berkeley, CA). The characteristics of these strains, the media, and the experimental procedures are is described in the literature (Ames et al., 1975).. Chemicals I L-Histidine • HCI, biotin and nicotinamide adenine dinucleotide phosphate (NADP), were purchased from Sigma Chemical Co.,l St. Louis, N-Methyl-N'- nitro-N-nitrosoguanidine (MNNG), 2-aminoanthrat•ene (ANTH), 2-nitro- fluorene (NF), 9-aminoacridine (9-AA), and 2-aminofluorene' (2-AF), were supplied by Aldrich Chemical Co., San Leandro, CA. Dimethylsulfoxide (DMSO), spectrophotometric grade, came from Eastman Chemical Co., Rochester, NY. The supernatant of Aroclor 1254-induced rat-liver homogenate (S9) was obtained from Litton Bionetics Laboratories, Inc., Kensington, MD. Sources of the test chemicals are listed in Table 1. Testprocedures The test compounds were dissolved in DMSO except for the cresols, gluco- isosaccharinic acid and 2,5tlichloro-3,6ilisulfohydroquinone, dissolved in water. Each substance was tested over a wide dose range as described previously (Ames et al., 1973, 1975; Nestmann et al., 1979a, b). According to the muta- genic and lethal responses observed, in a second experiment, doses of each com- pound were extended to a lethal range or to the limit of solubility. The Aroclor 1254-induced rat4iver preparation (S9) dosage was standardized with 3 promu- tagens (ANTH, benzo[a]pyrene, and 2-AF). The optimal dosage for activation of mutagens was found to be 50 µl/plate of the S9 liver fraction and was used subsequently in screening. ln each experiment, controls were run to determine the number of spontaneous revertants and to determine the response of the strains to known strain-specific mutagens in the presence and absence of S9. Results 9Z66IT48 Among 48 compounds tested (Table 1), only tetrachloropropene and penta- chloropropene were mutagenic in the standard Salmonella assay (Table,2). Tetrachloropropene showed dose-related increases of base-substitution muta-s tions in strains TA1535 and TA100 and of frameshift reversions in strain TA98. Strain TA1538, containing the same hfs- allele as TA98 but lacking the R plasmid, did not show elevated mutation. A weak response was found in strain TA1537, which reverts bya different frameshift mutation. Fig. 1 shows the dose-related mutagenic effect of tetrachloropropene in strain TA100. Pentachloropropene induced reversions in strains TA1535 and TA100 only. Dose-response curves of pentachloropropene in strain TA100 are plotted in Fig. 2. The presence of S9 greatly reduced the mutagenicity of both com- pounds. pounds. TABLE 2 MUTAGENICITY OF TETRACHLOROPROPENE AND PENTACHLOROPROPENE IN Snlmnnefta Irphlmurlum STRAINS -Compaund Doaa (M/piatc) Hii revertants(Dlate TA1585 TA100 TA1637 TA1538 TA98 -39 +S9 59 +59 -69 +59 -S9 +S9 -59 +59 1.1.2,8- 0 45 41 144 177 6 12 8 16 21 25 Tetnehleeo-2- 6 78 38 249 114 9 13 8 18 33 26 Dropene 60 296 61 1314 260 7 9 9 13 78 29 260 168 59 2610 773 14 10 10 15 241 31 ' 600 120 84 2868 1342 2B 66 9 13 207 66 1000 7 20 413 1726 4 a 0 28 0 48 1 1,1.2.3.2- 0 17 25 114 124 a 7 6 23 25 37 Pentechloro- 10 132 74 146 134 18 13 12 28 34 42 ptoVene 60 189 122 268 134 7 8 13 28 31 37 150 243 151 788 202 11 6 18 28 39 37 400 19 26 166 127 7 7 6 23 27 34 SINNO 6 1814 - 8188 - 1 9-AA 125 - - - - 1896 - - 2-NF 50 - - - - - - 2489 - 1166 - ANTH 6 54 620 181 1426 - - 10 2221 24 1683 2.AF 60 - - - - 8 1438 Volatile compounds ethylene dichloride (1,2-dichloroethane)-1,1,1-trichloro- ethane, and methylene chloride (dichloromethane) were non-mutagenic following the standard plate-incorporation method of Ames et al. (1975). (1) 1,2-Dtchloroethane, When plates containing ethylene dichloride (3,6 and 9 mg/plate) were incubated in desiccators, dose-related increases in the numbers of revertants were found in strains TA1535 and TA100, with and without S9. The maximum yield in strain TA100 was similar to that reported previously by McCann et al. (1975b), i.e. only 20 revertants above background. The maximal response in strain TA1535 was a doubling of mutant colonies. w a ` 3000 ~ lizooo W ~ ¢ 1000 (+S9) ij'e/" , b1-59) ~• 0 - 100 300 500 '/ Io00 TETRACHLOROPROPENE (pg/plo/e) FIe. 1. Muteeenic response of 1.1,2,8-tetraehloro-2qropene in strain TAIOO, using the SWnoneOahnam- maWn-mtcroaome test. -
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Al.ltrat'1' LIiV.aS b]11 H)R IIIV 'I'Jlt) 20... S-F,lGy(p•yrocnfcrl!n!: -- n:. I- 1:3.. h: ltuols.snft: 1-fi i t IEnurs). A-7(IL dae I. c:. i\u c-ItunOin'.. d: 'l.uuc1: 1--:1 with tlnslained W w(alklp slaiucd cltrotnusulncs. in :1 typicnl pt'ncLt'omosnnlr alainin;;. r: C-tnitusis in'3-7, ~rratt< Ccudi in :L.and 7. cnntplctrc-milnsis in 5. :3--ic uftett utollled chromnzomr+. j: PraRUlruls vcrv rarc: in the I'ullowing cuneentralions 11)11--Y01) cclle.lcerc ~crntinizedtcillt tltefullu.v- 111}JrCSllll: enuren!rtqInm\L. : s u In il 12 1:1 4 hnlll's . 11. 11 ll 0 . 11 I O I dny ...... - 1 ll 11 LJ /l miln.is,in'!-5 lv.illi bu.e ntvlapltcmc:wd tuultlcd c'.hrumusumrs.in a nnd ?I. OrcinrrL--u: 1_.-1:3.. 6:16~n:Is,nCt.' I pl huurl...2--:Ollltom-sl. 1(1' dnv). c: 4--7. d: -'Lnnc I: 1 2. with slnc:lrg oRgltltiirdinn. c•:' (:- 4. In I Ihcrc occurrcd ullcr I tl:ny n t-rrc characlcriilic Vpcrol• c-nlilusis witlt +Iron,>;W cunlrnclcd chromn.,unlc._ lhe annc ud tehich o•crrv:dmnst ,plli•ri¢ul. 'I'Lcsocllr,mum,wrs Nvcrv oi :ln e:lu:J !nuRicdnusv Nci111 Ihc. t•xcrptiat ul. IGc deeply .lainr:ld c'.uh'omcrir region. (`. 5:.1 I'ragmenls tcorc timnd.. but une c:lw. ul µ.rud"chi:t,uta in a)t '!2. 'Pliynrahytlruqluun/tc. - 'c'. : I:1. ?oull :t wt•rctiis."l•.cd in heal Iluf• un cu~rlin~:. rncslalA pnuciPiictlyd. I krpt clr:v. h: It~~rnl. .n(t: n--73 !:} huurs1L •l--5 U1 d,ni. IY'.S •l,n'•b, c: ti-14. 4: 2 and 3 have I)ycuntic, deeply lained clvutnn.n!ne•. n: ~:i1liWisis slark im::. -l Im.s cscIusiccIy Irtll tut•lap6acc.: ailica :1~ aclull iulO uninuc!c:n~ Iclaltlfa.rv,. ,,nl nr /chich chruuln.u uue :ilms.cslrr~:~• nfl :!nJtlnd. 5 hac :d.. :~nm- plctc c-milusis.Lut uluuturr oi:Jfunr.Ivl:r. l'.iu'omn.~~fnry rIl:n .omc- lvliui' un:lllcd in ir. In I}. . 7. tniwti r-nlil'm r• :utd normal ntilnsrs. j: (1••urrulrannn \U: . . !P. 1" 1! 1_ LI 4 Ituurs .... I) u. I u 1 0!1 1'rIP 'Inc....... . ... Il 0 I !) II R. METHOXYPHENOLS. •ia. (:,:.,i.rrnl in-mcthus3 'phrn•:I~ .eu~-I_-.1:1. b.:Ilnols s!'tl: I.-.r2 finslnnlly)l a(1 davl. 4 1•12 r!` Lom•i: 1--42 laillt wrakly >.loined, disSUllvith. cltrouu~sumc,: ~+Dli•n ~-villi distiuCL internal .,trncturc_ r: C-tnilnsiaunl!t' in ;1.--1, in Ihe. (nrmcr cnnccntrali(wn :1 cll:!rnclcri.vlir I',cpc.vith .c:lllercd-oul.. %amonqiy rnnir;tcted c-p:lira. smine- tiulcs uudlihala.r unaplln.us. j: oMJ -lo i ~.II b,b.l7O)D~b> I Fr 133 d: PY. CI11 31q t+n. Fr hm mo c: 1 CVs CVW Fr
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CHROMOSOME FRAI:MENTATIOK467 uilosis In (i, c have see p. s:une• -A (2 urring Ylrallg made litnvis. Lhsul 2 la 0 t J.. Quihhydrone. - u: 1--1a. h:ltools sufL. 1-2 (2 hours), 71-7 (4.hours):, 8(1 day): c: 10--I'1. d: 1- a hnnrstlnul;toxio-effects with agglutinalion off the chrolnolin: uuclcoli nnd helerochromatin often deeplystnine•d. 5: i-ID Il:rte parliol c-luitosis„ often chemqed by stickiness. /: Fragmcnlsin IhcI'ullowing: f.nurenllatl(nn Sr. .u rl. II 1_ 13 Fr .°o'' .......... ... 1 4 .`I l 'J 10, a-:Ynpfltllol. -a: 3-Iti: h: Itoots sufl: 3-li (1 hour), 7 (3 hnurs)', 8(I day). c: 10-12, d:: 3-5. rhromosomes in different stages of dissolution. Differential slainin.r ol'tlur cr•nfromeriehelerochromatin in a. c: C-mitosis in 6-9. 8 haz complete c-efl'ect, 4 has a strikiut;lyhigh frequency of multipnlar spindir.. 1: Fragments are extremely rare. In 9 and 10, however, somc clear causs of 17•agmcntation wl•re observed. In 11 several cases of Ituudncldnsmnla were found. It. /1-Nnphlho(,.- a: 4-lfi-, li: Rouls solt: 1-5 (30 min.), li I/ hour), 7 (1 ctay), 8 (2 dRVs). c: Smoll r-hunoursN in U-10. c1:. 4-5. e: C-initoses in tL--B, in 8 extremely ntanynndtipolar anaphuses: /: No fragments and no pseudochiasnmtu alsertrd. 12. Nhld+/hure:vorcinoL -,r. f--1:1. fr lirluls sufl: 1-2 (inslmilly.!. 3(~5 min.), 4-5 (30 min.), 0-8 iI elal.i. c'I'enclencies to c-lumnnrs.in 9. d: 1--1. [n 1-2 nuclci awd chrumu,nmes ngltlulinatcd, in :f---a thin,, slender chromosomes wilh clear internal slrtrclure.. r: /:-miloses begin ta:appc:lrin a nndnu dowu In Y/: l'ncloubtrd ir:l;mculsucc.ur in a couple (il' treatments, (Mi--I3 rdtcr 4 huur. :utd9-10, 13 :dler I dac werr :cnah•sed).: a% iil ri.. I ', in itl :111cr L hours. In addilion zome cusrs uf nltached fr:lgment.c.nnd Irsencfnchi.umaL•r. 13.. :Ili:rvin.. - a: 'PIIe cnncrniralion zonr Icsh'd st:u•ts with 0.m2 g/I /lI/ ee, whleh cornsponds Ier u.,.o-- mul'I. "fhic ~~ns dissulced. whett heatecl... On.cooling, precipitalecr.as fuenled holh in tLis solutiou and in Ghe ucxG lowcr nne. lioughlp entnded. ihr eowcentralicru r.omr vmploycd cnrrc.pouds Icl ,nlr luwn<bcrs 8-1 ii. Illc puinl nD s:duraliun lyin;; bctwr•cn !1 :uld 10. !r: Routz sofl: &--!1 tt davlt r: No r-lulnuun. cl---c: \rilher immcdi:rtely killing zone nor sirinsnP c-ulilnsis tverr sern. These zoncs tvould r•t-idrnl1V he situated nhln"c Ihc sutur:dion point uf nlieariit. j: Fra;;menialiou is very cntnmon in 8-ill; i. c. in x:dur:uecl and.ne:lrlv suluratc•.d soluliou. In lf no rc:d fra;;Im~nt. nccur. but oltou pycudo- chiasneda. '1'iLr fl+lpnleldntiun reaetion is rlf :I special iype in Ihis .nlulhner. uuin-" to .tir•kiuvnehein-1 , 1 rly' o.rulmon in ionr :i. Llydinnn• ~x~nIt1 -.
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Sl.m:nT I.r:I:t\ ANo JuM nIN 'rJln mclnpllascs nnd nuaplmscx hacc :drrnd'}. rcnchurl lclUphasc- r: I:-ntilosis hacs not In!cn idrnlifird ill :I-a utt'ing Gt Iack oP mclnphasc.. in 6, tnce.rc:dl mitnlie.sL•Il;cs nccttr•.tnuM Jic,isions urc uunn:d, 6td'sume Ilnrc c-lcmkncivs. j: Pra;uu•.nls uccur ill Illr 1'ullnwin;; 17otluencics iscrl p. 41ia..bclnw/. In 1Y:tLt•t•e o•vrc rccnrdrrl ?2 u1t:IchrJ fragtm•uls, thc s:rnlc tcns the cuac in CS.. lal t{• 15 and 16 uu tra-I'mt•utswrre sccn... 6. 1lydroxyhydroquinunr•. - n:', I--15. fr: Runts sul't: I---0 12 hnursl.,5 ISLuura! Il.-.1(1 f"1'.6our,t.. r: 1:-Imnours r:lr(-, UW uccurrin/; all thron>;h lhe scrics jtt, 7• 8. 151. . d: "/.unc I: l- _t::r{glulilr:rlinv.strong iu 1-5. c: C-ntilnsia ill 2--a... llwuqit nnl~° weak Ivndt•ncies, mnde tutclc:tr by Illv slrong ,.+lickillc,s. .\L.o, ill 'i Ncc:ik irndcncipn'trn c-nlitnu4t. f: ill the lullmvin., c•:jncs, incucL ccll !tctrr Innrc lh:nl y2 1 r:r"mcuts: t'.nu.rnlroliun\n• ~j -1 s ; ill 11 1. IS It If. Fr ?, ........ 7 /S I ll 14 ?I] 0 I - 1) sl . -. Plrloroyfuci:rnl. n: I t:t3 b: linnh aulC l-a 14 uuru's.l, 't--:r Itolq•] !t dari. Ilrrserics wllurluu:dr•Iy' nnt Crslcd lonp;rrlh:nr onc d:q:. r: 1': ,~ e: C.j t~ n- tumom's nerur in 6--la d;.3 ;;jvr, a IYlticnt picture: mctaplL:ue.nud ~ fragnt•. ;i nuaph:ne clvolnusomc, ire rittr•n ,I unlc.s 1.111 shuta tSettrdilul stainings 1• Id Illr Itt•li•InrllrumaIin .11,n Ji..ulutumn ul Iht c6nmin,ome •irmc i, -'ttT 3(3m ;: fuuncl, resullm„ ill Ilcu rcnIrumttu. r: \ltutd}' ill I lrnd~ncics.ln ~ y d c-milosis :Irc Councl. 2 and .S :,nc. Irpicnt Lou roc Li- trud .Inapliasr a, in tllin, .~ :f.ill utitusc. trculr.hpr 13t-brm :r nn Itun c unlusc, uccur 6cd t.lnai~4 begin• `i C-1CinlCllc'itrS~O ttit•IItPI' Ilrtt~"n. /.'. 1 nP I~.rnntCnt~ ~r -, , Ntti/• nuCOrUPU: 111 1.r 1 1 dqp. ! .oulc i ~~1 1 11 Il I U II t ' ,. I'1 hours ~. 2 - _ . . . l. .~. 1 I~t: •. 1 1 11 . IIP dU 30 l 21 y .... ..~ - _. ~ l/nlz gj ! I'srnduclu I,in.Jn tc r rrvrrn m rl ;ruJt? tl lcr d houi s.. Ili qctl I g, p-lirn nqttinnne - rr: 1- I b' linnts suft~• I 1/ Itutn'+L ?- 7 ,~tlte m ~ I Immsl. 8 11 doe.l... cAn e.tum ur. t.;•te tLnrrccd. d 1-+n huce nn eorre5l ` imnlr.dlalc killing ac•lion,.:md all +1Lo++' .Irun-1 a;;`Ittliitnliun. C:Ilrrnnu- 4.:md solncs rnfleu ruml)Iclrlcv drslninvd c:r tnutllcd. r: C.ntilusis uccurs ill immed i 3. S•, nutchr.rc cmnplcla. hlll inlrrtviuL1lyd tvilh uurnlnl ulilo.cs. (: I'rak- crnldrl . nlctllao'cresceq. ill Il,e' 4id4utcin", Irr:dnlruh: ;~ f: PraF ,; Sltllr8t i 1s ._ I:t clliRltll i! 12 .. 11 m ~ all6slae i! ~t N l:n W
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F 1000 a H CI-L-C_O~ct '-ol-S9) ~~ •(+59) o 100 200 300 400 PENTACNLOROPROPENE (yq/plote) Fie- 2. Muptenklty of 1,1.2,3,39.ntachloraD+opene In stnin TA100, usin` the Seimoneas/memmNlen- mkrewme tnf. (2) Dichloromethane, With incubation in desiccators of plates containing methylene chloride, non-mutagenic results still were observed. However, when 0.5 ml methylene chloride was added to an open glass dish within a desiccator also containing Salmonella plates (rather than added directly to the plates), mutagenic effects were observed. The numbers of revertants were doubled in strain TA1535 and increased 6-fold in strain TA100. A similar response in strain TA100 with this modified technique was reported by Simmon (1977). (3) 1,1,1-7)richloroethane. Dose-related increases in revertants were found in strains TA1535 (up to 6-fold) and TA100 (up to 2.5-fold) but not TA98 when 0.1, 0.5 and 1.0 ml volumes were added to open glass dishes inside desic- ca€ors which also contained plates seeded with bacteria. The mutagen was present for 16 h, and the plates were incubated for a total of 72 h. Simmon et a). (1977) have reported a similar response for strain TA100. The remaining 43 compounds were not mutagenic in the standard Salmo- nella plate-incorporation assay (data available from authors upon request), in that no increases of revertants were observed at concentrations of these agents which resulted in lethality or, in the ease of 7 compounds, at the limits of solubility (Table 1). i,1,2,2-Tetrachloroethane, reported to be mutagenic in strain TA1530 using a modified Salmonella assay for volatile mutagens (Brem et al., 1974), was not mutagenic in the 5 conventional strains using the same or 2 other modified procedures, For 9 of the remaining compounds found to be non-mutagenic in this study, previous studies using Salmonella have been reported. Non-mutagenicity was reported after testing with only one dose for 2,6-dichlorophenol, 2,4,5-tri- chlorophenol, 3,4,5-triehlorocatechol (Rasanen et al., 1977) and guaiacol (Ferretti et al., 1977). The entire dose range was lethal in tests with acenaph- thene (Gibson et al., 1978). More complete studies have shown the non-muta- genicity of toluene (Anderson and Styles, 1977), eugenol (Green and Savage, 1978; Swanson et al., 1979) and syringol,(MeMahon et aL, 1979). Using liver microsomes and cytosol (S13) for metabolic activation, trans-anethol was found to be mutagenic (Swanson et al., 1979). &M1sTT4s ! t i Discussion Tetrachloropropene and pentachloropropene now are added to the list of bacterial mutagens known to be found in complex pulp- and paper-mill effluent mixures. Others include neoabietic acid (Nestmann et al., 1979b), benzyl chloride (McCann et al., 1976a), bromo-p-cymene and dichlorop. cymene (BjQrseth et al., 1979), 1,1,2,2-tetrachloroethane and 1,2-dichloro- ethane (Brem et al., 1974), dichloromethane (methylene chloride; Simmon, 1977), and 1,1,1-trichloroethane (Simmon et al., 1977). The last 3 of these compounds were detected as mutagens in this study by using variations of the standard Salmonella test that enable experimentation with volatile chemicals. Also present in pulp-mill effluents are 3 carcinogens (safrole, chloroform and carbon tetrachloride), that previously were found non-mutagenic in Salmonella (McCann et al., 1976a). In yeast, however, safrole is recombinagenic, and chloroform and carbon tetrachloride induce gene convertants, recombinants and gene revertants (Callen et al., 1980). It is of interest to note that both tetra- and penta-chloropropene, as well as all other mutagens identified to date in pulp- and paper-mill effluent, are direct- acting in Salmonella, not requiring metabolic activation by liver homogenates to show mutagenicity. Thus they differ from other mutagenic halogenated olefins, such as chlorinated ethylenes, which require biotransformation presum- ably via the formation of epoxide- derivatives for mutagenic activity (Greim et aL, 1975). Although certain alkyl halides, such as vinyl chloride, can act directly in Salmonella, their mutagenicity can be enhanced greatly by liver homogenates (Bartsch et al., 1976). In contrast, the results presented here show that the activity of tetra-'and penta-chloropropene is reduced by microsomal preparations, suggesting that these compounds are metabolized to inactive metabolites and/or bind to microsomes in such cell-free preparations. A similar situation exists in the soil environment, where single alkyl halides are known to be degraded chemically or metabolized by biological systems via dehalogena- tion as a first step) (Castro, 1977). This may be an explanation for the reduc- tion in mutagenic activity after experimental bio-treatment of pulp and paper mill effluent (CPAR, 1979). The mutagenic natural wood product neoabietic acid is ieadily biodegradable even in receiving waters (Hemingway and Greaves, 1973; Leach et al., 1977), but many chlorinated organics formed during the process of pulp bleaching may not be. Accordingly, without bio-treatment or some other effective process, such compounds may persist in the receivtng water and may even be bioaccumulated in the food chain (Landnei et al., 1977) to present a greater threat to environmental quality. Many pulp-mill effluent constituents remain to be tested, and possible additive and synergistic effects also must be considered. Screening programs, such as the present study, are useful in the identification of mutagens and potential carcinogens. However, the threat of these compounds to human health must be evaluated by in vivo mammalian assays that are more appro- priate for human risk assessment<
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STEt'ENa AND O'rHSas.: EsPECroaAh'TS. [Cen. xx1 a. r. Feb.14R. voi. 48 ON TIIE EXPECTORA.'TACTION OF CREOSOTE AND THE GUAIACOLS• Bl' M.1sY E'. STEVENS, 9LICE K. RONAN, T. S. .SoIIRKES.AND ELDON .til. BOrn, M.D. Department of Pharmacology, Queen's University, Kingston, Ont. CREOSOTES.and guaiacols have been used as expectorants for approximately one hundred years. They have been included in the.leading pharmaeopceias and are presentin the current editions of the British Pharmacopeeia and United, States Pharmacopmias. They were ae- cepted: into New and Non-Offlcial Remedies for 311 years. In recent years, however, there has been a rising tideof scepticism as to their actual value as expectorants, scepticism evident in several textbooks of pharmacology and materiamedica, in clinical usage, and especially in the rejection of such drugs from. New and Non- Official Remedies by the Council on Pharmacy and Chemistry of the American Medical Asso- ciation in 1938.1 The reasons for this declining popularity of the guaiaeols are partly the lack of experimental proof of any action upon the respiratory tract' and partly a lapse of interest. The latter is understandable to students of the history of drugs; many drugs come and go for no apparent reason except perhaps that the practising physi- ciann can keep in mind a knowledge of a limited number of drugs only and hee naturally employs those drugs which are.of contemporary interest and popularity. Experimental studies on the guaiacols have yielded conflicting data. Fellotve° concluded after an extensive series of studies that calcium creosotate is not excreted in the bronchial fluids nor in expired air, and that it does not affect the volume;, character, or ease of expectoration of sputum in patients with pulmonarytuber culosis. Using a radiological method, Gordonoff and Janett2 deduced from their observations that creosote and guaiacols decrease the volume of bronchial secretion in man, and Gordonoff,' in an extensive review of expectorants, classified these drugs as"secreto-motor" expectorants or drugs which stimulate expulsion but. not the production of bronchial secretions. Conncll, Johnston and Boyd' found that guniacols in- crr..ise the water content of the respiratory tract • This paper xas pro.aonted in prcliminarvformhe- fnrothc Canadinn Medienl Aremeiation at ~its Annnal .ltecting in Winnipng, Jnnc, 1941. of albino rats,, and especially of the trachea, and suggested that their evidence indicated an in- creased output of respiratory tract fluid. Perry and Boyd° demonstrated in the rabbit that guaiacol glycerol ether (resyl) increases the out- put of thee respiratory tract fluid.. In the more recent literature from Europeann authors,. Peytral,r for example, reports a decreasedout- put of sputum in tuberculous patients with the same drug,, while the data of Hofmann' would suggest an increased output of respiratory tract fluid with this drug given to patients with bronchitis and pneumonia. There are other relovant data of recent origin, but they add little to the eonfused, picture which indicates, at its face value, that under certain conditions guaia- cols may decrease the output of respicatoryy tract fluid,, under other circumstances mav have noeffoct, and under a third set of conditions may increase the output... Whether or not thetherapeutie value.of these drugs.is such as to warrant their extended study is a moot question. However; for some years investigations in this laboratory have been directed toward a study of expeetorants and itwas deeidedto examine further the effect of guaiacols upon the output ofrespiratory tract fluid using thee method of Perry andBoyd?This method, briefly,., consists in measuring the output of respirator,vtract fluid from a tracheal cannula in urethanized animals when thee in- -- haled air is warmed to body temperature.and saturated with water vapour. The results ob- tained indicate that the guaiacols studied do in- crease the output of respiratory tract fluid in anasthetized catsand rabbits if the drugs are given in sufficiently large doses. The mechanism of this reaction has been further found to be partly due to a reflex tYromthestomaclu and partly to an effect after absorption intoo the blood stream, possibly upon the brain stem or directly upon the sccretory glands of the respira- tory tract itself. EtPERTTIruNT.1L Cnniocol.-Guaiacol nms washed into t~he stomach of rabhits thrnngh a rubbrr calheter, ...., ... ,w ce (-mt.ctea Ey t,¢, rigli: rW.H (iiie 12
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CIiROAIO9plli Fll\nJIKRTA'1'IUv he frag- rcnctiun •s nf' dle 'tretches crv loav hul Ihcir dc.lh is of gmenl in itc; samc u• nt 1hc nilnsis is. r ill snme 1 o-nilrn- recorded :ul attack e•v.idetltiyy lary con- ed in this ~ cffect, is' en iu the : ragments .urnts arc c•.cll I :kn lu is Ihr : fact, Illc .1 >imil••Ir i Pi;{. ? rl l. cadl, Ihere. :1•rheaped I leuds. I f •iut-effrcts :It :111:11'k, (n sum2 c iu Ihcse acrordinq. ,umbcr nl .nLa:wnr. Alnong thee substaneeslchich eshibil Ilr:lgnmenls in more Ihan hulf of the lesled cnneetltralions the lullowinq hnrr Ihr higllesl fr °„: IllyIle.t rr "rn l. Py'rogallnl .. . ........ ... .......... .. ..... li5 3. Hydroquinouc .... .............. 47 a• p-Phenelenediaminr 31i 4. Pyrncatechol . ......... .... . . ..... .... 24 5. p-rlutinophenol Sy (I. Itydruxt'hvdrnqninone•. t I 7. I3enznqninnne .................._....... 12 This list i:ss r:dhrr significanl in unr rcvpcct: Cnmmon lo Nas. 1--ti is thefact' that they are readily oxidized. No,. 7 may in lhc living cell funn a system hydroquinone ~ par•Iquinone. 'I-he lislincludes all Ihose phcnnls lvhich are most easily oxidizalrle, liz. those rehich have their fNl in ortho- or par:l-position. Pheuol, rrsorcinal and phloro;lncinolarrl decidedh•y lesss active,, even Ihough they:dl give some fTagments: All lhese facts go Io shoWIlnlf the11cl1VItC. is in snule way connected with lhe oxidation of the substances. Either lhey may interfere at some significant point with the nnrnud oxid:dion-reduclion st•stem of•f the licing, ecll.. orr their action is brou);lil ahoul ilysome product arising at their osiiln.linn, cnnsrqueullc a.ub.tancl• beinrnore artivethe.ensii•r this product iss fnrmed.771e pt'uetlt IYagnnentuliuun rccu•lion is ~Irikiul;ly 1'euliniscevrt ol' X-rayel'feels and mustard );as.clfccls: Our prcliminarv resulls indic:ltr, hmvever. that considerable dill'crrncc, are also preseni. .11ios•c all. it scems th;N the induction ul'tranalucaiiuns (rcuuiun nl' hrolcenendsl is much rarer in our reaction than inX-r:lrIrealm--nls• 11us may he ctuc loo the chemically induccd hrraia, ot the chromosonle thread LminG spread ntrr a long period u•hile lhr \-rny. hrratcs appear s.imultaneouslv. 13ruken ends have a greater cllanrc nf iindinl; eacir other in the.l:lller ¢ase. Treatments with Inltct;lyd g:Iti (\'pl$NF.t1GF.)t atld LrV•1V, lnlpllb- Iichedt within Ihe .ame malcria Jm«, that Iranslocations f'nrnx•d a large fraction ot the induced c•hnn-les. •Phe ehunfics induced in Altinnl rout lijwfi' y mu.hmd gas did nol :II all reach lllrs:lnlotrequenev nn. Illn fra;;nleutatiunss in our mosl active snLslatrces. kt is evident thatibt•>e high freqprnrivs uGprol'uwld drslrut•Ainu nf li!Ir c•hrrnnnsume m:Jcrinl,, without killing nf the crll ;md tissoe Irc•atv(L nlahe omr snbslanrev. hi,,hlv ..inl•.•cr.lin_ in relediun lo ihc ranccr prnbll-ma; w,-IL P.tkallsTlr:R a ,a
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212 Wende, C.C. (1976) The toaefty of pury and paper m(tl efauente md eorraep"ndtne meoureme"t pe.e. dunt, Water Ree., 10, 8&9=864: Wilaon, D., and B. Hwtflord (1976) The fate of tutpentlne In aented 1. ~ T195-Iei. eOOM' Pulp p°D°r Ce".• 14. It.ta((on Research, 79 (1980) 213-221 C ElwrierlNorth-Holland Biomedical Press ,1C~YATION OF THE MUTAGENS OF BEEF EXTRACT IN VITR L'i t'WO /tI.HO DOL\ARA 1'2, ROBERTO BARALE 3, SANDRA MAZZOLI t anfUINO l1tF.'fTl ~ 213 tdwmtu di Farm ologia e ToaaicofoPia~ Uniuersitt7 di Airenze (ital ,° Center for the b• :.•n ol Na(ural stems, Washington Uniueraiy, St. Louu, M(U.S.A.), 3 Laboratorio !. Krnrtira, Uniuerait 1Pida (Italy) and 4 Laboratorio GentR(pA., Pisa (Italy) M.re:red 26 February "K0) ~ . u.,.. L4Cn1 ~ auepted 9 June 1980) >ummary the activation in vitro of the my ens isolated from beef extract, when .r,rd with Salmonella typhimuriu/n s'n TA1538, requires the presence of ~ fractions from fivera of PCB- or 3-meth cholanthrene•induced rats. S9 frac- ., at from uninduced rats were'unable to ac 'vate the mutagens, but no induc- t.,n was necessary with Swiss albino mice. G- mice had intermediate activa• .•:n capabilities, which increased after the additio f 0.75% BHA to their diet. F4.man S9 were poor activators. No activation was o erved with whole homog- n~..atrs. The active metebolites formed were unstab at 37°C and firmly 5. -md to protein, / tthen isolated liver's were perfused for activation, no ' herently active n.a¢ens were rele;rsed into the pcrfusate. Similarly, no a've mutagens .ae found in the,t'trme of mice after administration to them of b f extract u.a,lens p.o. or~ S p. Intrasartguine host-mediated assay did not s w any nCailcant muta mc effect. The possibility that genotoxic effects o hese .,:n:/~suods mi t be confined to metabolically eompetent cells of ind ed e._mals is disc sed. (zPsttr~e t-Pe°rted by a arent from C.N.R. (C7910282766), by e Vaat from U.S. National Inftitute of eenen INIEIIS) (PR01-ES•0i964), by a contract with the U.S. National Cancer Inlt(tute (NOl-CP• t5a521. bv LeP Tumoif, Sezlone al Firenze md by contract No.267177/1 ENV (Commission of
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:t1.HENT I.F:V.tN %YII J(lli IIIN TJI() AIour recurcling of tlte IParmmtil, it w:ts necessarvtu distingnislt between free h'agmenls and ,nch ns nre stilPnftnched to thecLrmua- sulne. There at'c. all kinds uf Iran,itinns Lrl.reeu changes which re- presenl unlp'sm:dl cllllSlrlcllntlc ntl the cltrolnusomes and such where nmst nl' the chrmnosutne<Irm Iru Jicapprarrd between [lie fragment :utel the chromusonm but where unc I`.tinth• slnined thread is slill cuu- nccling Ihc fraFtnenl with lltr thrunuos:nue: Oflen the same ehrotuu- sonme artu is eroded :dunrils whuM lem;lh: Iltee result 6eing a ruu;;h uulline :utd au alternation ul ,I:linrd and stuinlcss purlions within llte chrumusomc Fig. 2n1; .1t ntelaph:ue il often happens thal nnechrutnalid iseruded r.ight thrnu;;lt ahilr Ihr ofhel' h:ts unlp developed a conslricliou nt thc samc Ievrl ~Pig. I'r,. 1^rcc I'rat;ntcnls :irc alrcadv prc,enl at tuclaphosc. 11'. hutvrtrr, uulc :our c•.lir(:roatid is severed, thr I'ra;mrnl will remain pnirrd willt its siatrr se;;ment until annphnse slart.. Therefore. frrr.lnrlaph:lnr fr:l,tncnts h•ing.nulside Ihe equtdnrial plate urc cntnpuruticely r:ln•: Ihep :Ire d:.ubla:md comisl ul' 6ollt sister etirntn:dids lFig. 1. hl: \lvlkqlha,:• i, ruu,/•qtnnll}' nu favouroblc sla{ie fur cuuntingg fruglnenls. 111 v:mr litles, however: vt•ry manv chromu- <uutes, tvcre seen In lie frn^mrnled alir:tdv :d' Inrlnplutse;; in sotnc cases tvr I'uund Ih:tt a fraGmenL•ttion uflrn nccurred at about Ihe +urne level, :tl :t little disldmce 17om the rhrmmosnule cnd IFi;;. I dt', R'c d:tre nul rlaiut., hutccvcr. 16n1 Iltis iss duc lu :t ecrtain cltruwnsnmc rr;;iun livinr; .:vpociadlc susceptihle In fen,mrnl:Himt• bul we havt• uur utlentiunu directed to thiss pruLlcm tnc tulurc:+naly,is• .\t annpfiase ilte distinctinnn b0tveou 11•ce aud :ulaclicd fraSmenl,. :Ind lurrely rruded cln•unrdids iv m:l(Ic npParrnl. Frce frap;menh rrtn::iit. :It [lleeqtl:diu'- m-hrn.r:l, the nli:te}hoJ Ir:t;mrnl, arr- pullyd liv tlloil.r chramusuntes Inw,nrds Ihe pulv,. II is rtuclintcs tlitiicult lo ere tLe, . alt:lclrtnettt tGrend..Lul Ilwpo.illiun ul" a.v.rrlein Irugmrnt nfL!u tiivon an1 inJicntiun as to Wltrlher it i. In— • ur nnl. 'Lltc Ilrcr fr:lgmcr,ts usilnlh- nricnlalev lht7nselves lransvrrs:tlIv nn Ihr ~piudh•. tvhile lhe ulthcln•d'fi'onFImruls...nfcuursc. are silu:tlel lcn;;lhaise. In nur rrcordv mtlv . un- duut.ledly frcr fragmcul.. nt'c tnunlcd.. utn- cnuuts Iharrl'nt•r ;;ivin;; miuiinuur ft'eqttencic,.od fra;;utcut:Jiou: Since Ihe.Incu fr::l"rneul-Ivpes ncrvtuqrmg vaclr olhrrIhrmlRit :Jl Imalutrnls Ilu•.A' arr cunsidt.rrd lon•prrseut different asperts nl' IL,• .:nne re:tclinu. i'rrllnps n Irsa savere :Ittar.l: on.tLe•f clu•nnwsumr Ihre:ul rrsuliS in u crnulrie•.linu, while a suntr_ wlutl'. hordrr tdluclc ::ices :t. Inr.iraglnrnl. 'I'hr I7rv t'r:lgmenls utnvitavt•:,nv sizc linnt verY .m:11Pall. - utiutdr, In vldit':: rLranfonoulo .tnuls. AecnrdiuGlv, Ihe renlt•uuu•rir ~ prr Sm pro ,: ,-: are rl Ifi.., lha attr: ont- Fig pra Fy.., bec, . vvhi' ° sus c6rt (ST pha title tlrcr recr: han chr. pnrl Itvn arm a m Itrrt sum rvm: nac autb arc: sTEI Ircal Irenl Areo cnu" /ltr I 1 r ~'3
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9 rr C111/0711)SOSIG FIIaG\IES'fXl'IOti 4 1iSl tU. rn-Cresof.-rr.. I--I:3. 1r: Itunls sutt:1-2 (iustantlv ), 3 /l 11ourF, d(-{ 1)oorsl, 5 l2 rL•i vsj. c: icuIlcroJ'c•-IUtnottt:c iu li-1:?. rl.ancl r:.l- -rcc closely s•illt preccding ulbst:wrr. Chrt• 17uc ccnU•omcric sininingss in 2 nnQ~:3. c-milosis ill 4-5, weak Irndr•ncirs to c-mila.eis in :3 .md 6. d fs clwractcrized by star uud b:rll mclalph:lscx nncl Lr thc Iypicnl mottlinp;nf the cltromasumes. f: Fiagnocul:dinululs mtty- been air.erved ill 5.. where I fr.tgutcnt occurred in 1011 crlis studicd. 17. p-Crrxol. - a: l--Ci. b: l3unls soll: Il- 2(inslantIt•), 3 f I lwurp, 4 (4 hoursI. 5(2 clm's)L r: 6--1 LL d und e: Sss in prrcr'<liu;- Iwo sub- slunees. Stained centromerc hrlerochrurmdin in 2, c-mitosis3-!). 4 has mottled chromosomes: f: Coumninrlinn \n. e ,. s ir IU II 12 Il Fr Q . ........... 4 5 Il /1 11 q 0 In 1l there occurred ou(- rasl• nl' interlorlccd' ring chrumalicls i-frag- menL in une :utaplmse. In 12 IGcrr were seen pscuducllihsmatu. 18. Y,.5-A,r/lcnol, - n:. 1--I:1. U: Rurds nnfi: L---t liusluuliv). :r (I d;ty): c: No c-ttm)ourss seen. rl: Zone I: 1--4. Oflcu.n distinct chruum- vrmee structures, lLeteroc•Lromnh• ai litrprnchromusumrs cerr pro- nrnwccd... r: C.-mili)sis in -1--i. -1 has unR~. ))•rak Il-ndmtcirc 1o c-miln.is. the met.•Iphusic clvomusnntns :trr b,nl liltlr rnntrnctnd :md arc nflru slaiuudonly at the rrntromerrs. lu 5Ihoomilosis is nmrc typii•u! with slruol-I clu-oumsumocunlruc'Iirmand ;rrunntntnrd innlllinri.. /: f.~rncrnhminn Yn. „ 1' r .',; . .......... I i r 14 11 13 11 211.? 0 :I ij.~ n .lfter I duy' nu fragmrnls ccerr inclt wilhin 1t)0 annlvsedd crll> ill r::~II uf the conrentralions 8-13. 19. 7YrymrL -- a: -t-17. M1: IOun,/. .nl'1: 1-- ir il nrin.ll, li I:i uuu.'. .--8 (1 duyb `/ (2 rl:rest. c: Tt'ndracics to c-truuow:s in 13. d: '/.noc.. 1:. d--li. f;l)rrnnosmltes taintly stainv.d. uttvn 'r.ith rii•iincl inlornu! slrur'- turc. slur)rinG dvidcd cln-ulunlid.. r:. (:-miloii< iu 1i--9. in 7- full; r- milosiswitli strnn.,ly y cuniracterl.chromnaoutrs, in 7--Nlntoltir•tl clu-onw- ;utmv~_ f: Prc:,Ipn:nls rn:curnod Lrtl e\Irc:irelv rurrly: l-r ,~~ 11 huurs~ ..., U It tl ,. ~" I I J:rY U In tt ICiS. O.r 0 0 q O II n
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0 and carcinogenicity [3), an array of short•tenn mutagenicity test systems have been developed employing both psokaryotic and,ukaryotic oSganisms. The moet widely used of these is the Ames' system 141 which measures the reversion frequency to prototrophy of a series of histidine-auxutrophs of Salmoneffa ryph/murtum carrying well defined mutations. Tobacco smoke condensates are mutagenic in the Ames' system and the mutagenicity is generally dependent on the presence of an activating system from (rat) liver, the ao called SS9 fraction [5]. However, the mutagenic potential of tobacco smoke condensates is much higher than can be accounted for by its content of known mutagens [6] such as benz[a]pyrene. An obvious reaton for this is that the mutagenic activity of the vast majority of the more than 3000 compounds identified in tobacco smoke [7,g] is unknown. The present work was therefore undertaken where we have used the Ames',eystem to study the mutagenicity of 239 compounds selected primarily on the basis of their abundance in tobacco and tobacco emoke. MATRRIAIS AND MCTHO0.4 Compounds Most of the compounds tested sre major constituents of the gsseous or temivolatile phases of tobacco smoke. They were subdivided according to their functional groups into 12 different classes to facilitate comparison of the activity of chemically related compounds. Although most of the com- pounds are commercially available, some wem synthesized or isolated from tobacco. AB compounds have been checked for purity using thin-layer chromatography, gas chromatogmphy, and NMR; those containing more than 3% impurity have been purified by means of liquid chromatography, recrystallization or dist0letion, The structures of some of the compounds studied were-confrrmed by ['H]- and (t^C]NMR, e.g. the substitution pattern of multieubstituted ammetic substances and the hrera;hing pattern of inethylsubstituted long chain alkyl derivatives. Spectroscopic grade ethanol and dimethyl sulfox(de were used to prepare the solutions of the compounde used in the biological teting. Mutagenicity, testing The 4 bacterial strains used (TA 98, TA 100, TA 1535 and TA 1537) are histidinerrequiring mutants of S. typhimuAUm LT-2. They were obtained from Dr Bruce N. Ames (University of California, Herkeley, USA). The strains have been described previously in detail [9,101. Initially, cultures were grown In Difco nutrient broth. Since this medium is suspected to have a weak mutagenic activity [Ames, B.N., persm eomm.], it was substituted for Oinid nutisent broth No. 2 in later experiments. Revertants were scored on glucosenminimal sslts medium [11] supplemented with 0.05 pmol h6tidine and 0.05 pmol biotin. Plates used for viable counts contained 10 pmol histidine (and 0.05 pmol blot(n). The experiments were carried out essentially as described by Ames [4). 220 8'7119417 The following controls were made for each experiment: --the viable count was determined; -the number of spontaneous revertants was measured; -the presence of the rfa-mutation was checked by crystal violet inhibi= tion; -the pseesence of the plasmid pKM 101 In strains TA 98 and TA 100 was checked by resistance to ampicillin; , -theresponse to the positive controls N-methyl-N'-nitro-N-nitrosogueni- din (not requiring meta6olic activation) and 2-aminoanthmcene (requiring activation) was checked. 8-9 The S-9 fractions for metabolic activation were_ prepared as described previously [4]. Aroclor 1254 (S-9A) or 3-methylcholanthreoe (S•9M) (both suspended in com oil) were used as inducing agents. Aroclor induction was performed on male Sprague--Dawley nta (-200 g), by giving each rat a single i.p. injection of 500 mglkg 5 days before decapitation, wh9e 3-methyl- cholanthrene induction was accomplished by giving a daily injection i.p: of 20 mg(kg during 3 days before decapitstion. The S-9 fraction was prepared by centdfugation of a liver homogenate at 9000 g for 10 min: Aliquots of the supernatant S-9 were etored at -70°C: We have used 4 different prepara- Kons of S-9 from Aroclor-ioduced rats, each derived from a pool of 3 animals, while the same preparation of S-9 from 3-methylcholanthrene- induced rats was used throughout the experiments. The activity of the S-9 preparations was tested using 2-aminoanthrscene. The amount of S9 used in all experiments was the dose found optimal for metabolic aetivation of this aubstance. The S-9 mix contained per ml: 100 pmol sodium phosphate buffer (pH 7.4), 8 pmol MgClr, 33 pmol KCI; 6 pmol glucose 6-phosphate, 4 pmol NADP, and 0.03--0.1 ml S-9. Ilg9UL79 Most of the compounds tested were not found mutagenle. All substances except the hydrocarbons, which gave a negative response are listed in Table l. The results presented here require some comments: (a) Pyroga0ol is probisbly a weak niutagen. In some expedments a 2--2.Sfold (ncrease of revertants wae found in quantitative tests using strain TA 98 both with and without addition of 8-9 (conc. range 0.3-3 ymolfplate). (b) Qulnoline is known to be cercmogenic to rats [12) and it has recently been reported to show a mutaggenic effect with strains TA 98 och TA 100 using S-9 from Aroclor induced rat. The duse range employed was 0.2-8 pmol/plate and mutagenicity hasbeen observed both with [13] and without [14] preincuba- tion. However, in our experiments quinolinc gave a negative responm In spot teab. It was therefore tested quantitatively at concentrations of 0.01- 221
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REFERENCES I E.L.WynderandS.D.bS.Ilman,CancerReay87(1B7t)4608. 2 E.L. WyaLer and D. Herfman, &lence, 162 (1968) 889. 3 B.N. Amea, W.E. Duntoe, E. Yemuakl and 7.D. twe, Prte. Natl. And &1., 70 t197S)2281. 4 R.N. Ame/, J. MaCann md E. Yamwkl, Muta. Raa, 31 (1976) 847. 6 L.D. Kier, E. Yamwkl and B.N. Ames, Proe. Nau. Acad. gel., 71 (1974) 4169. 6 T. Sueimura, M. Na9ao, T. K.wechl, M. Hunde, T. Yahql, Y. 8siea, S. 8am and N, Mateukura- Muta¢eniarclnoyem in P6od, with eyecW nfennm to highly muta- eenic pymytic producte in broiled foode, In H:H. Hlaeq J.D. Watmn and J.A Wimtm (Edz)..Urieire of Numan Cenar, Cold Spdne Harbor Confm.nia nn all prolifmation 1977. p. 1581. 7 CR. Green, Neutral (ezyeanated) eompnund. in elgar(tM1 anoke and Ihair pomble prenunurc, Reeent Adr. Tobaeeo &1., 3 Q9771 91. 8 R.A. Heckman and F.W. Beat, Abatract, S2nd Tobacco Chamiet'a Resevtlt Con- ference,htontreY,Canada,197e. 9 R:N: Ames, E.G. Oueney, J.A. Mlller md H. Barted,, Proc. Nail. Aud. &1., 60 (1992) 3128. 10 J. MRlann, N.E. Bpineam, J. Robod and E.N. Ame. Proc. Natl. Aeed. &i:, 72 (1976) 979. li J.Spizuen,Pres-Nau.A<ad.ScV.,44(1958)1072. 12 K. Nhao, Y. ShinoTUa, H. Teude, S. FukueMma, M. Takahashi and N. Ito, Cancer Ra., 36 (1976) 329. IS M. Nasan, T. Yahqi, Y, eeb,q T. Bushrnra and N. Ito, Mutat. Ru., 49 (1977) 666. 14 R. Taleatt, M. HoW teie and E. We6 Blnehem. Phmm., 26 (1976)1825: 15 J. A.hby and J.A. Stylel Natun, 271 (19781452. 38 K. Umnawa, A. Shirai, T. Mat.uahima and T. Bu9lmora, Proe. Natl. Aead. &1., 76 (19]8)928. 17 A.P. Alearez, D.R. Bicken and A. Kappm, Proe. Nal. Aead. Set., 70 (1878) 1321. 18 E. El.enetadt, Bacaed.l mutannlelty teatlee: 8ome pnctied nonzidentlone, In W.K: MeElheay and S. Abr.hameon (Ed.). Arexine clumic.l mut.yene: The riek to humans, Banbury Repmt, Vol. 1,1979, p. tl. 19 J. 61cCann and B.N. Amae-Proe. NaN. And, &I., 78 (1976) 960. 20 E. K.n.r, Dtseh. AGOth. Zte.,109 (1989) 1290. 2] A. laeaaeayne, N.P. Buu;Hei, F: Z.jdela and D. Erv74Lamy, C.R. Aead. Sci., P.rie, 25; (1961) 826. 22 B.L. Van Duunn, A. Bieaq L. Lenpxth, B.M. Geldaebmldt and A. Sagd, N.Y. tYncm Inq, MaMr., 29 (19") 173. 23 G: Grimm.r, Dtseh. ApotB. Zt`.,108 (1968) 529. 24 G. Gdmmeq 2. Kmbdorech., 69 (1967) 2233. 25 J. McCenn, 6. Chol, E. Yanvaakl and B.N. Ames, Proe. Nail. Aead Sel., 72 (1976) 6t66. 26 1&Amdtz and D. Hoftmann, Chem. Rev., 77 (1977) 295. 27 A. Pilotti. K_ Ancker, E. Arrhanlua and C. Enzell, Taxicalop 5(1976) 49. 232 87119423 Toaiccloey, l6(1980)238-242 0 Elsevie./North-Holland &ientific Publirhm Ltd. BIOLOGICALEFFECTSOFSURFACfANTS,IV.EFFECPS OF NON-IONICS AND AMPHOTERICS ON HeLe CELLS ROBERT ERNBT and JOe6PH ARDtTTI Pepertmenl o f Devdopmenl and Cell9iulup, Unitanily of CaBfomM, trw,e, Cdi! In 92717 (UB.a.) (Received Decembm 10th,1979) (ACceptad Mareh 17th, 1990) ' SUMMARY Surfectanta, representing 3 non-ionic and 1 enphoteric series of homo- )ogs exhibited marked differences in their lethal and non-to:ic effects on Helr ceiL. Toxicity of non-ionics generally decreased inreraely with increasing hydruph9ic chain length and irtcreaeed with increasing sice of the lipaphile. Lethal levels of the surLctants coincided with surface tension reduction of the media to 45 dynes em" or below. Surface tensions of non-toxic coneentmtions were substantially higher than tJtaee for toxic )eveh. Nun.toxie doees were, therefore, below the critical micelle concen- tr.tion of the surfectantr evaluated. The data suggeet that physical properties are a principal cause of the toxic effects of these detetgents on HeLa cells. INTRODUCTION Limited information te avsilatile un the re)atfonehip between chemical ettucture, physical properties and toxicological elfects of surface aetiye egents (surfactanta) despite their wide industriai, household and agricultural use. Therefore, several homologous series of surfactante with .arying charges, molecular weights,'and hydrophiiic/Hpophidc balance (HIB) were prepared and pudfied. In our prior work (1) with there agents, employing the fresh water coelentemie Hydna attenuote as b)oueay organism, lethal concentia- tiona were always coincident with surface tensions of 49 * 4 dynes or the culture edutione: It was possible to establish toxicity trends with respect to the Rpophilic isdicds of the Ionic species ecreened. However, the AbbrMatlons: CMG criC al mic.ll. concentntlon.; HLB. hydropbiBePipeyhilic bdaoee, tlT,eurraeetenabm. 233
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.1 B.plgna411i, nal6 reasons for these variations, the pH oEthe stomach contents of the rats was determined and was found to range from 1.87 to 5.75 (mean. t S:D.:3.34 t 1.44; n = 9). It wasfurther noted that 4 out of 9 stomachs that contained solid dietary constituents tended to show an elevated pH (4.8-5.7). It is therefore assumed that the variations of NPRO excretion in rats dosed with precursors dissolved in water may be due to differences in stoma& pH, which is possibly affected by in- take, rate of emptying of the stomach or other physiological factors involving gastritacid secretion. Comparison of the effects ofphenolic compounds onn nitrosa- tlon of prollne invivo and in vitro The influence of phenolic compounds on formationn of NPRO in vivo and in.vitro iss compared in Table I: Inn vivo N- nitrosation was catalysed by resorcinol,, catechint phenol and p-nitrosophenol and inhibited by chlorogenic acid in a man- ner similar to that observed in in vitro experiments, although the magnitude of the effects in vivo was smaller in general, 27-72%of those shown in vitro. In the absence of phenolic compounds and when the concentration of nitrite was in- creased from 10 to 20 µmol, (experiments no. I and 4), the yield of NPRO formed in vitro increased 4-fold (7.2 to 28.9 nmol in 15 min). As with the kinetics of the nitrosation of proline (30), formation of NPRO was dependent on the square of the nitrite concentration. In the corresponding in vivo experiments (Table l, experiments no. lI and 4), the amount of NPRO excreted in 24-h urine increasedd only 2.5-fold, corresponding to concentrations formed within 4-7 min in vitro at pH 4 and 37°C. Lower yields of NPRO in vivo were noted earlier (5) and may be attributable to the rapid disappearance of sodium nitrite from the rat stomach by absorption, decomposition or reaction with gastric com- ponents (31,32). The catalytic effect exerted on in vivo nitrosation by resor- cinol and catechin increased.when the ratio of [nitrite):[cata- lystg changed from, 10 to 4(resorcinol). (Table 1, experiments 2, 5) or from 10 to 5(eatechin) (experiments 3, 6), inagree- ment with results obtained in vitro (FigureA). Discussion Endogenous nitrosation wasstimatedin rats by applying a new method developed in this laboratory (5) which monitors NPRO excreted in. the urine after sequential administration of proline and nitrite. The results demonstrate, to our know- ledge for the first time, that resorcinol,.catechin, phenol and p-nitrosophenol can catalyse and chlorogenic acid can inhibit the nitrosation of proline in rats in vivo. The extent of catalysis and inhibition are dependent on the pH and on the relative concentrations of nitrite and of the phenolic com- pound. These dataa confirm those of previous studies (14-17), indicating that phenolic compounds, such as resor- cinol,.which can readily form C-nitrosoderivatives by reac- tion with nitrite, can also catalyse the formation of N-nitroso compounds, in our case from proline. By inference, this may explain the observed catalytic activity of catechin, a molecule which carries a resorcinol moiety. In contrast, those phenols that are easily oxidized by nitrous acid to quinones act as scavengers; they may therefore inhibit nitrosation by reducing the nitrosating agentt to nitric oxide. Such a mechanism may explain the inhibitory action of chlorogenic acid. A possible mechanism has been. proposed for the catalysis of nitrosationby some of our phenolic compounds (16,17): 1048 during an initial step, C-nitroso derivatives may be fortned which further react with the nitrosating species to generate a more powerful nitrosating agent (thought to be a nitroso quinone oxime derivative). Its.reaction with the nitrosatable - substrate produces the nitroso compound'6 i.e., NPRO in our studies. This postulated mechanism implies that assay systems containing increasing amounts of a catalytically active .. phenolic compound lead to increasing concentrations of the ~ C-nitroso intermediate, and consequently to a reduced con- i centration of the nitrosating species. This mechanism would I- explain the optimum ratio for nitrite to phenolic compound ~ that we observed. Accordingly, a large exoess of phenolic compound'would inhibit nitrosation, because the nitrosating agent would be completely used up in forming the catalysr ~ and nonawould be left to conthtue the catalytic reaction. The potential importance of phenolic compounds as. modifiers of endogenous nitrosation stems from their wide- spreadoccurrence in human foodstuffs and beverages (24,25, 33,34). Phenol itself has been reported to be present in beer and in heavily smoked bacon at levels of up to 300 p.p.m. (24,25,27); nitrite is also added as a preservative to bacon. Catechin and chlorogenic acid are both important consti- tuents of the human diet (33,34),, and may be present simultaneously,.for example inn tea and fruits. Concentrations of catechin in edible foodstuffs: vary considerably, and values in the range of 1-75(mg/100 g fresh material) in fruits, up to 800 in some berries, have been reponed (33). Extrapolation of these results, obtained in rats and in vitro, to man in vivo is.difficult, because the reaction is dependent on the relative concentrations of nitrate,.nitrite, nitrosatable substances and of agents.that can either enhance or inhibit nitrosation. In addition, in vivo concentrations of these com- pounds vary with the amounts ingested and conditions prevailing in thee saliva (e.g., capacity to reduce nitrate or nitrite) and in the stomach (e.g., pH and rateof gastric emptying). The daily intake of total phenolic compounds pre- sumably exceeds the mean concentration of nitrite in the stomach. However, the modifying effect of these compounds on the endogenous nitrosation may depend on numerous fac- tors such ass the ratio between potential catalysts and in- hibitors in the diet and their absolute concentrations.available in vivo, the relative rates of their reactions with nitrite, the se- quence and timing of intake as well as the bio-availability in the stomach. It would be impossible to predict the outcome of reactions occurring within such mixtures solely on the basis of estimated intake of catalysts and of inhibitors. Despite this complexity, it has been shown, by applying the NPRO methodd to a human volunteer (7,35), thatt endogenous nitrosation is efficiently inhibited bysimultaneous intake of vitamin C and also, to a lesser extent, of vitamin E. The in- hibitory effects of these compounds were found to be iden- tical in rats (5), in which a catalytic effect of thiocyanate on the nitrosation of NPRO has also been demonstrated (36)..adl the studies carried out inn this laboratory indicate a good qualitative agreement between data obtained on the nitrosa- tion of proline in vitro, in rats and those obtained ina human subject. Results from the present study on phenolic com- pounds confirm this notion, as the modifying effects on nitrosation in vitro were qualitatively similar to those found in vivo, although when compared quantitatively they were 1/3 to 2/3 smaller in rats. Taken together, it can be concluded that (a) use of in vitro assays to screen complex mixtures and pure compounds present in the human diet for their catalytic N; W
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IOee i a p r l s vTwsfela,. Fg: a Do.e rnponae to 2,edl.minotoluene of TA 9S abr metabolie acdvation. chrysene towards TA 100 and perylene and cnrunene towards TA 98,aB after 5-9N activation. Coronene is not previously known as a mutagen and a dose response curve is given in Fig. 1. Of 5 indoles tested, 2, 2,3dimethyl- and 2,3,5-trimethylindole, showed mutagenic activity (towards TA 98), wh0e the other 3, indole, 2-methyl- and 3-methylindole, were inactive. The above 2 mutagenic indoles were the only representatives of the 239 com- pounds studied which were mutagenic without activation (Fig. 2). Of the 2 aminotoluenes examined, 3,4diaminotoluene was without effect, whereas 2,6-diaminotoluene was mutagenic to TA 98 and TA 100 after activation with S-9A. A typical dose-response curve is shown in Fig. 3. The other mutagenic amine, 0-naphthylamine, was without effect in spot tests and in quantitative tests using &9A, but was mutagenic to TA 100 after activation with 5-9bt. Others have reported activation of p-naphthylamine with S-9A [91. DISCUSSION Out of the 239 compounds examined in this study, only 9 were found to be mutagenic. The large number of substances made it necesaary to screen for potential mutagens by spot tests, which .re certainly less sensitive than quantitative experiments. Spot tests also suffer from the disadvantage that strain TA 100 is difficult to ute because of its high background (150-- 200 colonies/plate) [4] and possibly also from a more marked influence of poor solubllity. instabflity and/or toxicity of various substances. An additional factor of uncertaintyy in a11 the experimenta perfonned is the snmetimea large variations found in the S-9 prepamtions with respect to activstion of various chemical classes of substances [151. Nevertheless, we believe that screening by spot testing can be used to identify the most potent mutagens among the compounds tested. 230 8'7119422 Not unexpecledly, the highest proportions of mutagens was found among the hydrocarbons. Four of them, benx[o/anthracene, chrysene, bene[o]- pyrene and perylene ate known carcinogens and mutagens [18,19]. The fifth hydrocarbon, coronene, is a prevriously not documented mulagen 1201; it has been shown to be inactive after subcutaneous injection in mice [21] and to produce only skin pap0lomas after topical application [221. It is rather surprising that itc mutagenic activity has not been revealed earlier, since its mutagenic potency is companble to that of bena]a]anthracene, and since coronene is a common air po8utant in industrisl areas [231 and has been found in soot [23], amoked fish and meat (24] and vegetables [231. Of the mutagenic amines encountered in this study p-naphthylamine is a previously known mutagen-carcinogen [25], while the mutagenic activity of 2,6-disminotoluene was not known. The latter result was not unexpected in view of the mutagenic activity previoualy described for 2,4-diamino- toluene [25], although 3,4-diaminotoluene was inactive. It is known that the mutagCrlic (and carcinogenic) activity of close chemical relatives often differs substantially [191. To our knowledge, methylated indoles have not previously been shown to be mutagenir tn contrast to the other mutagens found here, the indolea do not require metabolic activation for mutageniclty. We have now investi- gated a series of methylindoles, and it seems that 2,3-dimethyl substitution is essential for the mutagenic activity [Florin, I. et al., unpublished). While these indoles are present in tobacco smoke 126], where they may arise from proteins or other N-contsining compounds by pyrolysis, studies of tobacco amoke fractions containing these compounds only show mutagenic activity sfter metabolic activation by S-9 prepamtions [6]. It is presently not possible to account for all mutagenic activity of tobacco smoke condensates in terms of well defined chemical entities and this may be ascribed botlr to inadequate knowledge about the compounds present and probable synergia- tic effects. Although identification of biologically hannful substances in tobacco smoke may point to ways of removing some of these, it is evident that before such measures can be contemplated, more data from several in vitro systems are required. We are presently examining the feasibility of this approach by studying the biological effects of tobacco and tobacco smoke constituents in several systems. So far, results in addition to those presented here are ava0able for the inhibition of cell growth on Ascites samoma BPS cells [271, integrity and permeability of ceB membranes of lung fibroblasts [Thelestam, M. et al., unpublished), and noradrenaline stimulated respiretion in hamster brown fat tissue [Pettemsuo, B. et al., unpublished). ACKNDWLEDCEMBNT9 The authors are indebted to Ms. K. Bernholm and Ms. E. Kasemi-Vala for skBfW technical assistance. 231
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@ 126 ''rEVEKS AND OTHN.RS: ESPEC't'ORASIti ether (resyl), was given by intramuscular injeo- tion to a total of 64 cats in, doses of from 0 to. 0.5 grams per kilo body weight. There were noo significant changes in the output of respiratory tract fluid in the controls receiving water only. All of the groups receiving the guaiacolateex- hibited an increased output of respiratory tract fluid, an increase which in the case of the higher doses reached as much as 100%. These results suggestt that the drug would normally act after absorption into the blood stream as well as through a gastric reflex. There is, of course,.the possibility that the intramuscularly injected drug was excreted into the stomach and acted reflexly. From what is known of the metabolism of guaiacols, they are excreted mostly in urine as glyeuronatess and, sulphates9 and the same applies to guaiacol glycerol ether.'° Guaiacol glycerol ether has been occasionally injected intramuscularly in clinical therapy (see,, for example, Hofmann"). Dlsoossxov Guaiaeol derived its name from guaiac resin from which it was first isolated in 1826 by IInverdorben." It is present in rather small amounts in guaiac resin and probably con- tributed little to the reputed therapeutieaetions of that drug which was broughtto Europe in the 16th century from the guaiaetrees of San Domingo and Jamaica and was used up.to the latter part of the 19th century as a stimulant, diaphoretic, alterative and emmenagogue." The subsequent history of guaiacol is bound more closely to that of creosote, off which it is the chief constituent. Creosote wass isolated in 1830by Reichenbach who obtained it as one of his products of the destructive distillatiom of ic"u.d. Creosote quickly found its way into the ph;n•nmeopceias of London and Edinburgh which, withthe Dublin pharmaeopmia, were.thee precursors of the British PharmacoptEia. Thus by18d2, Christison" wrote that "creosote has become an important article of the materia medica". The claims put forward at that time forthis(t.hen)r newdrttgremind one of the variegated claims putfmtivard for many ne"v dru_s today-apparently this cnstom has not changed! Thus creosote was stated too be an irritant, a styptic, aa narcotic, a sedntive,, an anndcne, :ln antiseptic. a diuretic: and tirhc uw.in.l for tilcel:v: burns, con.lum'tivitis,., drnnm•- rhmn. Irprosy, psoriasis. toothaelle, rhnlcra. ha•mopt;asis. phthisis, mrnorrha-,ia and hron- [c°e..t.a.J. Fen.19{3;voL44 chitis!" Christison also noted that, "An old, German nostrum for chronic diseases of the chest, called Aqua Binelli, seems tocontain creosote for its only active ingredient." Creo- sote was taken internally 100 years ago for many of the above conditions in doses.of 1 or 22 dropsdispensed as Mistura creazoti, Ed. Ph.,, which was asyrapy mixture. Ottt.of this initial multitude of indications for the use of creosote,., the field: for internal thera- peutics gradually became limited to its use as an expectorant in chronic bronchitis, bronchicc- tasis. and especially in pulmonary tuberculosis. At this point a complicating factor was intro- duced in that some authors postulated, uponn theoretical reasoning, that creosote had an anti- septic action towardd the tubercle bacilli in the lung. Thiss idea has been promulgatedm throughmany decades and is occasionally referred to. even today. The idea lacks proof, and this. has been repeatedly pointed out for half acentury.. Thus in 1909 Stevens" wrote: "So long ago as Addison's time,, creosote was used as a remedy for phthisis. This use was revived by Bouchard and Gembert in 1877, and again by Sommer- brodt in 1887. While it has been shown con- clusively that the drug has noo specificc influencee on the tubercle bacilli in the lung, neverthelesss the. testimony of many observers is convincing that it has a positive value in. allaying cough,, lessening expectoration and lowering tempera- ture." About the beginning of the 20th century, a large number of derivatives of creosote ap- peared on the market, mostly from tlree various pharmaceutical companies which were thenn becoming large and acquiring increasing in- fluence. These newer derivativesincluded ereo- snte carbonatc,, guniaeol, t,niaiacnl carbonnt.e. guaiacol potassium sulphonate (e.y. thiocol), guaiacnl benzoate (e.g. benzosol), gnaiacol phos- phite, guaiacol cinnamate (e.g. styt:2col)- guaia- col glycerol ether (e.g. gtmiamar, resyl),. etc. The chief advantage claimed for thcse dru!ss over rttaiacol and ercosote iras that they were lesrs irritnting totheo gastrointestinal tract. Tin Illany Illstallees, however, the ndvertisincr litrrn- llvr est.ravacantly extended the rlaims for thrrapentie tls.dnlnrss, and as thrse rlaims weree trnduallY fmwd too bc u•ithout fonndatimi effart:, a sreptirism arnse in theminds of phssi- rinns as to Ihr value of these drues under :ury rilrunivtanvrs. This has been the evolution of rreosote mtd I
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L L Gt.asnuem ively). and the purenl compound, anisole. Of the monosubstiluled compounds, stimulation was marked with allylbenzene and ally( phenyl ether,, intermediate with rrpropylbcnzene and absent with /Lmethylstyrcne. It must 6e emphasized that these findings pertain to the dbsages screened (rable I): more massive levels werc not investigated. It was shown earlier (Gcrshbain, 1975) that the action of n-propylbenzene on the Rgenerating liver was border- line and that styrene an&phenetole were without in-Ilucntx alihe very high doses studied It may be men- tioned that the methylenedioxyfienzene, piperine, which occurs in black pepper, was present in the pep- per, oil screened at too low a level to be effective (Table 2). Toxicological studies on anisoles, allylbenzene and', Ilxthylencdioxybcnzenes have.dealt withh the deter- minationn of the oral LDyo (JcnneL Hagan, Taylor, Cook & Fiizhugh, 1964) and with the evaluation of microscopic changes,, the most striking of which was hepa5c celtenlargement (Hagan er al., 1965; Taylor et al., 19641- The increase in size of the hepatic cells varied from slight withsafrolc and isosafrole to moderate with dihydrosafrole. Other consistent alter- ations were minimal degrees of fatty. Inetamorphosis, bile-duct proliferation and architectural irregularity. The degree of liver enlargement was slight with also extended to increases in wet and dryiiver.ce;y expressed in relation too bodyy weight in intact r::. , Thc results with intact andd partially hepat.w:mnw.. i. m:des were also checked im the femalc t+ul no > t ditTcrena in the liver paramcterscould he dix.~-_: with the agents tested. The latter work was unc, i taken btxausn of a report by one group.(Homhar.. i er uf., 1962) that fibrosis and ceroid deposits in ,. i :.~ livers of female rats fed a 1°o safrole diet were i., marked than those observed in the malc fuiii I). ioi he pn. ru Gers dr Ir. Gers rel he Gen eugenol. Because of the short period of investigation used in the present study, morphologicalfmdings in the liver were not remarkable even though the doses administered were massive. The cffccl on liver regeneration exerted by. com- pounds with an azulene structure was ascertained in this study. Azulene as such has been reported to stimulate the restomGon tif liver weight when given pharrrc Jr.,18. 601. by the oral and se routes (Gershbeih: 1975) and the Cekan,. Z_ Herout, V. & Sorm. F. (1954C On r.-. current se fiddings. (Table 1), indicate that the fully LXII. lsolalion of the prochamazulene from Ilar.-- aromatized structure is reypired for the activity. Azu- chamomilla. L.. a further compound of the _oir- . Ienes occur in. ,4Jatric'uria, cfruuuanf/la L in cubeb, group. Colfn C:ech. chrm. Cormn m r. EJL EJU ly.' , +cormwood, guaiac wood and vetivcn oils and: in yar- Cukun,. Z.. Prochazka, V.. Herout, V. & Sorm. F. ..- row flowers, amongg others. and except for the blue On ltqxnes.CL Isolation and consatuuon of nur~ ' chamomile oils and the tea, the findings were negalive another gua;anolide from camomile t:llarriruru ,- 4\'lltl I11e52 materials as well aSN'Ith the pale oils with nulld L.I CoCn Crrch. c. Cwwmm. 6yf. EdnJJ.'.t'-! low azulenc content from. Andnvnfs nohifis L. Ciamiciun, G. u. Silber. P. fltr90f I%ber die Comn:.. a des Apiols und semun Demac Bec dt dimr G.~ Whereas the liydro-derivative: guuirne, obtained from -1Zg3. . the sesquiterpene alCohol. guaiol, was ineffective, the Craig. J. O. (1953), Poisoning by the volatile oils in aromatized molecule, guaiazulcnc, was very active by hood Arc'hs DieChildh. 28. 475. i the se route in an overall dosage of 1065 mg/kg (series Dandiya, P, C. & Shurma: J. D. 114624 Studies on t. 19E; Table 1) as were the Hungarian chamomile oils, cnlmnu.x Part V: Phurmacnlogical actionc nf avror. __ H41 and H-5. The. latter should contain about 5"` (J-asarone on central nervous system. 6iJfan J, „,_ ;.. • of the chamazulene,. although some oils have been 50•'16- repor[ed with levels up to IS°; It is thought that Elcombe, C. R.• Bridges, J. W:, Gray, T. J. B.. Sc: • Smith, R. FI. & Nener. K. 1..(1975J,.Inh+:uuum,u ne chamazulenedocs not occur in the plant as such but with rat hepade microsome> Biochrnr. Plruns-, :. results from enzymatic action on the component ses- 1427_ yuitcrpenes (Ruhemann & Lewy, 1927) or from Epstein. S. S, Fujiii K_ Andrea. J.. & Mantel. \. ," another derivative or precttrsor (Cckan, Herout & Carcinogenicity'testingmf selected food':Wdai+e. ~. Sorm, 1954: Cekan, Proeh:izka, Herouf & Sormt enteral administration to infant micc. To.d:. up,•. r. ~ 1959- K h 1942, Givrn, by the oral routc . Haitian nua: 16. 321. oc . atc. Ji:,n: Je Fricdmun, M. A AmoIJ. F Buhop. Y''. A Lp t v o. vcnvert nd, the alcohol mixture vetiveroli and Q9 guaione, like the Matricaria tea andoils, stimulaa+d f19714 Addiuvcand s)aerbrsuc mhiMnun ol m'ar . ~ m+crosumal en>yntc functions hy pijwnnJ bL- t liver rc6dlcr.ruon,.al[hou>••h the Bourbon vefiven oil ,~ safrole und other n+cthylumdroxyplinnl d.m,.- proved' negative ula level of D35"„',. These findings Esfmrrmrin 27.IB5?. f~ indicate that significant arumatization of the hydro- Fritmh, P., de Saint Bhmyuat..G..:, Ikrachc. R. i.~ :¢ulenes.might occur in the gastro-intestinal tract. Absorption gasoo-inte>linale chee le rat. dc I'amr•. _+4 With few exceplions. the stimulalory response rrans-anutholc, du butylhydroxyenisolc et do sdrr.: engendered byseveral agents on. the regenerating liver Co.wvr. Tosicof. 13. 359. S~ t 4 Aeknoudedyrnrrats-Theauthor wishes to thank the<. ge, porations that supplied him with various oils and it_ ' Gr• products. He is especullyindebted to Mr. R. 1. Ei.r: . by of Frirzmhe Dodge & OlcotL. lne-. Mr P. H. Manh;.:.. ! pa of J. Manheimer, Inc. and Mr. C. H. Ziumiceki af S.: (ia•ri Penick.& Co. for their untiring aid and co-operati.c. sul I REFERENCES Borchcrt.. P., Miller, J. A., Miller. E C. &Shir.x i. e. : (i1973a) I'-Hydroxysafrole, a proximate cardn.;c'. - metabolite of safrole in the rat and mouse. Cmrrcr 3. Grac lu; atc G ucr' D- Hag4 33.59tL ' To Borcherh. P.. Wislocki, P. G.. Miller-. J. A.. & 61111c. . C(1973b} The metabolism of the naturallq .xr.". • hepatocurcioogen safrole to 1'-hydro.ys:drolear.c :. clectrophilic reactivity of I'-acetoxysafrole. C,rnev 33.575. Bouchardat. G. ct Tardy- t 1g964 Sur I'esuncc d'u:- - Russin C.r..L.hJ. Semic .1rad. Sci.. Pmii 122. o-• Caujolle: M. F. cl Mq'nier, D. (In(A4 Acti+itr h•.r. crmiunte dana Ics series de fcugenol eU du vfrole. ;
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~• 372 Ci'cqz"~ J. Ap-k. foa!dChem. 1882. 30..372-374 q:./'t.(.12'.Q.w ., '1'~'rolytic Studies on the Contribution of Tobacco Leaf Constituents to the Formation of Smoke Catechols William S. Schlotzhauer,• Robert M. Martin, Maurice E. Snook,;and Ralph E. Williamson Pyrolysee of different tobacco leaf varietiee,underconditions designed to simulate cigarette smoke formation, demonstrated a correlation between leaf polyphenol levels and pyrolyzate catechol. Bright tobacco varieties contained significantly higher levela of polyphenols as compared to identically cured Burley tobacco varieties and produced correspondingly higher pyrolyzate catechol yields. Leaf con- stituents, including polyphenols, lignin, cellulose, and sugars, were pyrolyzed to determine potential conversion to catechol and alkylratechola. Chlorogenic acid, the polyphenol generally present in highest amounts in tobacco leaf, produced the highest pyrolytic conversions to catechol and 4-ethylcatechoL Flavanoids (ruf.itt and quercetin) produced lesser amounts of catechol and 4-alkylcatechols. The polymeric phenolic leaf constituent, lignin, produced significant yields of catechol. Catechol (1,2-dihydro:ybenzene), considered an active cocarcinogen (Van Duuren et al.,. 1973)) and the most abundant phenol in cigarette smoke, has been identified (Schlotzbauer et al., 1978; Hecht et ai,1981) as thee prin- cipal component of the weakly acidic fraction of cigarette smoke condensate which has been shown to posseas tu- mor-promoting activity (Wynder and Hoffmann, 1964; Bock et eL, 1969, 1971). A number of compounds, known to be present in tobacco leaf, have been shown to produce catechol under various pyrolytic conditions. Zane and Wender (1963) produced catechol and alkylcatechola by thermal degradation of the leaf tannin, chlorogenic acid, and of the flavanoids, rutin and quercetin, at reported melt temperatures of 600 °C. Caffeic acid, a structural com- ponent of chiorogenic acid, was reported (Jones and Schmelt.z, 1968) to produce catechol (31.60 mol % yield) by hot tube(700°C)pyrolysis under nitrogen. Sinceneither of these studies attempted to closely simulate actual conditions occurring in a burning cigarette, no re- alistic quantitative relationships between these leaf con- stituents and' smoke catechol levels were determined.. Recently,.Catmella et al. (1980) extracted tobacco leaf with hexane, eliloroform, benzene,.and methanol and concluded, through pyrolysis studies, that the catechol precursors resided mainly in the methanol extract, which contained chlorogenic acid and sugars. Using a method designed to simulate the formation of cigarette smoke (Schlotzhauer et a1,1979). Schlotzhauer and Chortyk (1981),.bypyrolysis of extracts obtained with solvents of increasing polarity, estimated that half of the catechol content of tobacco amoke is attributable to the ethanol extractable poly- phenols of tobacco leaf, withthe remainder of smoke catechol precursors residing in the lignin-cellulose-con- taining leaf fibers. Since type„cultivation, and curing of the tobacco plant have been shown to result in considereble variation in levels of chlorogenic acid and total polyphenols (Tso, 1972), we felt it of importance to detenninethe relationship between leaf polyphenol content and smoke catechol levelefrom different tobacco varieties:In this study, a series of Bright and Burley varieties, with known polyphenol contents [aa determined by colorimeti•ic Tobacco Safety Research Unit,.Agricultural Research Service, US. Department of Agriculture,.Athens,.Georgia 30613 (W.S.S., R.M.M.,.and M.E.S.),.and Tobacco Re- search Laboratory, Agricultural Research Service, U.S. Department of Agriculture, Oxford, North Carolina 27565 (R.E.W.). (Williamson, 1975) and high-pressure liquid chromato• `" graphic(Snook and Chortyk, 1981) analysesJ were pyro- lyzed and catechol yields were determined. Repreeentative" leaf polyphenols and additional compounds reported to _ produce phenolic degradation products, including lignin`(Clark, 1968) and carbohydrates (Bell et a1, 1966), were pyrolyzed under identical conditions to determine their contributionato the yield of smoke catechol. EXPERIMENTAL SECTION . . Tobacco Leaf Samples. Fourteen tobacco leaf varieties (nine Bright and five Burley) grown at the Tobacco Re- search Laboratory, Oxford, NC, in three replicate plots, were selected for these studies. For preservation of uni- fortnity, all plants were cultured, harvested, and cured as for conventional flue-cured tobacco. After being cured, all harvests from a plot were bulked, the midribs were re- moved, and a portion of the resulting leaf.materialwae converted to standard cigarette shred. Prior to pyrolysis, the shredded samples were equilibrated to 10% moisture content by placing them in a chamber containing a tray of saturated aqueous sodium bromide solution for 48h. A second portion of leaf materialwas ground to pass a 40-mesh (1-mm) screen to produce samples for high- pressure liquid chromatographic determination of leaf polyphenol content. Standard Leaf Compounds. Chlorogenic acid,.mp 207-209 °C, and caffeic acid;.99%+, predominantly the trans compounds, were obtained from Aldrich Chemical, Inc. Rutin (crystalline trihydrate), cellulose (a-cellulose fiber), S-D-(-)-fructose (crystalline), and sucrose (Grade 1, crystalline) were obtained from Sigma Chemical Co• Quercetin (dihydrate) was obtained from Fluka A.G. As the best representative plant lignin, a Kraft softwood Ggaro was obtained from the Southeastern Forest Experiment Station, U:S. Department of AgricultureForeetServicei Macon, GA. Pyrolyses. Samples (25 g) were pyrolyzed by using apparatus (Smith et a1.,.1975, Higman et al., 1977) and experimental parameters (Schlotzhauer et al., 1979) pre' viously described. Pyrolysis products were collected in traps containing chloroform-methanol (9:1 v/v, distilled- in-glass grade).. The pyrolyzate solutions were concen- trated under vacuum on rotary evaporators (30 °C) to an accurate volume, genelally25 ml., sufficiently concentrated for subsequent gas chromatographisanalyses. Glass Capillary Gas Chromatography (GG2). AL' quots.of the pyrolyzate solution were combined with a knownn quantity of the internal standard,.p-(sec-butYl)- Whit _ . Spei Non Can. Nor, Virg Spei~, ' 401 Eff Cok Ken° Gr- War~' Ker H4 ave avel ° All values 0 (1976). ° Det, method more 5 Table II.. lda)o con chlor °, ®' querc ~ rittin 1 caffe lignir cellu sucr. fruc, j °Indetermii phenol, in a r bis(trimethy) Co.) was add heating bloci were directh 5720A gas ch detectors an( column whic on a1.,.1980). T Q jection portFi programmed ' b+ gas (helium)) t0 was 100mL/t - w Uhder thess µ catechol elu it b o tained w tern, were b: thentic cate High-Prt Quantitatior by the metl Thie article not sublect to U.S. CopyriyFd. PubBShed 1982 by the Ame1can Chemlcal Society
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TABLE H EFFECTS OF TOBACCO AND TOBACCO SMOKE CONSTITUENTS ON CILIABY ACf191TY Alcohols 1 Methanol 2 Ethanol 8 1-Hexanole 4 1-Heptanold 5 1-Octanol 6 1-Nonanole 7 1-Decanole 8 1-Pentadennole 9 2-Propanol 10 2-Hexen-1o1 11 Tetnhydro`enniol 12 Gennlol 13 Fameeol 14 Menthol 16 u-Terpineol 16 0•lonole 17 Propylene glycol 18 Furfurylalcohol 19 Benzylalcohol 20 2-Phenylethanol Ethen 21 Aniwle 22 8-Methylaniwle 23 4-Methylaniaole 24 2,8-Dimethylaniwle 25 8,6-Dimethylani.oie 26 2,8,6-Telmethylaniwl: Time (min) to dlioetaila at 5 mM concentration No. compound Time (min) to dliostasie at 6 mM copcentntion 27 2,4,6-Trimethylanleole 14 > 60 28 Ethylphenylether 28 >60 29 Diphenylether 20 >60 80 1,2,8-TMmethoxybenzane > 60 28 81 2-Methoxynaphthalene'd >60 10 32 2-Sthozynaphthalene 19 17 >60 >60 > 60 Acids 8 88 Methanolc add 20 22 34 Bthanolcadd >60 6 35 Hexanolcadd >60 10 88 Heptanolcaddd >60 11 37 Octanoic acid 4 17 88 Nonanolc acid 11 16 39 Decanoic add° 2 > 60 40 Dodecanolc add° • 80 > 60 41 Pentadennoicaddcd >60 >60 42 8-Phenylpcopenoicacid >60 >60 43 Benzoicadd >60 44 2,6-Dihydrozybenaoicadd >60 45 8,6-Dimethoxy-4-hydroxy- >60 18 henzoic add 5 48 8-Pyridine carhoxylic add > 60 4 47 Indole-8-ecetic edd > 80 14 48 4-HydroryQuinaldicacid >60 28 (0.8 mM) 32 49. p-Phthalic add > 60
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w, 204 Ind. Eng. Gbem..FUndsm., Vol. 24. Nb. 2. 1985 Table 1. Summary of Condltioaa for Gualacol Pyrolysia in Water T, °C Cr„ox 10~, moi/®~ Cw.o X 104, mol/cm° Pr, • Cer X 104; mol/cmt reaction time, min 3831 1.5 3.62 0 0 1.44-2.24 5-9g 363t L.5 3.67 7.52 0.04 0.97-1.85 5-90 383 * 1.5 3.62 70.9 0.40 1.02-1.90 5-90 383 t 1.5 3.62 137.0 0.77 1.80-2.25 5-9g 3&9 t 1.5 3.62 282.0 1.59 0.57-2.80 5-180 363 3 1.5 1.80 282.0 1.59 1./g-2.22 5-90 383 t 1.5 3.62 36110 2.08 1.22-1.71 15-L'lu ' Based on TSw a' 374.1 °C;P~w - 3206 pd4 Vcw = 56S cmt/mol; reactor volmm - 0.65 cm+k reactions during extractions at temperatures in excess of 350 °C. It thus seems likely that the high yields of coal volatiles (Scerrab,.1983) and low yields of eelluloeic char (Modell, 1977) observed in supercritical water (T° = 373 °C) extractions ere a result of both pyrolytic fragmentation and mass transfer steps. This motivated our use of the model compoundgusiacol as a probe into the pyrolysis pathways and kinetics contributing to the extraction of volatiles from lignin with supercritical water. The details of a macromolecule's pyrolysiss pathways;, kinetics, and mechanisme are usually obscured by the complexity of both its structure and conversion product epactra.. However, the relative simplicity of a model com- pound's structure and reaction product spectra permits quantitative analysis of reaction pathways, which can be used to model the related reactions of the macrantolecule'e pyrolysis. Lignin is a polymer of single-ring phenolic monomers (Freudenberg and Neish, 1968; Harkin, 1967; Glasser and Glasser, 1974), moet of which contain at least one methoxy substituent in a ring position ortho to the hydroxyL. Guaiacol (o-methoxyphenol) thus models key aspects of lignin etructure. Previouss guaiacol pyrolysee (Ceylan and. Bredenberg,1982; Bredenberg and Ceylan,1983; Vuori and Bredenberg, 1984; Shaposhinikov and Kosyukova, 1965; Kravchenko et aL,1970; Connors et aL, 1980; Klein and Virk, 1981) indicate that its maJor low molecular weight products ere catechol, phenol, cresols, meth