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ANALYTICAL CHEMISTRY, VOL. 50, NO. 6, MAY 1978 • 779 Trapping and Determination of Labile Compounds in the Gas Phase of Cigarette Smoke Steven G. Zeldes and Arthur D. Florton' Analytical Ghemistry t7ivision, Oak Ridge National LaGoratory, Oak Ridge, Tennessee 37830 The gas phase of cigarette smoke was trapped and stored on Tenax-GC for subsequent off-site analyses. Specifically, the highly labile compounds isoprene, acetaldehyde, and acrolein were determined quantitatively in the samples which were thermally desorbed In the injector port of a gas chromatograph onto a cooled gas chromatographic column. Optimum con- ditions were determined for adsorption and desorption of the gas phase, and the effects of aging on the trapped gases were studied. It is necessary to chemically characterize the cigarette smoke offered experitnent..'tl animaLs in inhalation bioassays to define the extent and quality of the exposure. It is also of interest to determince the chemical nature of smoke-polluted envi- ronments to assess the possible impact of smoking on nonsmokers. Tenax adsorption followed by thermal de- sorption and gas chromatography has been evaluated as a method for characterizing the volatile organic gas phase constituents of smoke. One of the routine analyses of the gas phase of cigarette smoke is the determination of isoprene, aeetaldehyde, and acrolein (t}t the first because of its close correlation to bio- logical activity of smoke, and the others because of their ciliatoxicity. If these highly labile compounds can be trapped and retained, then the less labile components of interest should most likely he retained also. Breakthrough volumes have been determined (.3, fi, t:3) for a number of compounds, some of which appear in cigarette smoke. Double trapping experiments have shown that our results agree with th<?se authors for compounds of common interest. "Penax-f.C° has been used with some success to trap labile compounds in automobile exhausts (2, 3), ambient air (3, 4), and stack gases (5). The traps used for t.hese samples differ only in size, each consisting of a Pyrex tube packed with Tenax lielc6irr place by glass wool plar;s. The methodology used at this laboratory was adapted from that (rf Zlatkis. L.ichtenstein, and Tishhee N) who used a P' vrex tube I I cm long. 10-mm o.d. and $-rmn i.d. packed with 4 mL of 35 to 60 me:h Tenax. Samples were adsorbed throu h a condenser and desorbed in a mcrdifieci injector port tknttY a cold l)recolutnn. then desorbed a;iecond time c>nto an open tubular coltarnra. r!t thi,, laboratory, the traps were desrrrbed in the rnoditied in,iector port directly onto a packed column carotet.l to ---7t? Qd'. EXPERIMENTAL Adsorbent. Tenax-GrC; (Applied Science Laboratories, Inc., State College, Pa.), a porous polymer, poly-p-2,6-diphenyi- phenylene oxide was selected over other common adsorbents (Porapak, Carbosieve, or activated charcoal) for its several ad- vantages. Its high temperature limit of 450 °C (6) and low re- tention volumes (7) allow high-boiling sample components to be desorbed more rapidly than frorn other adsorbents. In addition, the effect of water vapor on the efficiency of Tenax (8) is in- significant. Preparation of Traps. Traps consisted of Pyrex glass tubing (9-mm o.d., 5-mm i.d.) cut into 5r/.r-inch lengths and fire-polished at each end. One end is ground to a taper to form a seal in the injector port. i15Jf g-inch glass wool plug is placed in the tube at one end, the tube filled, while vilxatin ;, with 60/80 mesh Tenax then topped with a', 8-inch glass wool plug. Traps are conditioned by heating at 250 °C for 30 anin while purging with nitrogen. Conditioned traps are stored in a des- iccator. Sampling Procedure. Samples were collected from weight selected (1C194 f"() mg) Kentucky Reference (II2Ip cigarettes conditioned at 75 °F and 60% relative humidity using an ORNL Single Port Smoking Machine. See Figure 1. Cigarettes are smoked at a rate of I puff per minute (II puffs x35 mLjput`f) using a small vacuum pump to draw each puff through a 0..3~-mL sampling loop. An additional length of tubing is placed before the inlet to the gas sampling valve in order to collect the sample from the middle of the puff. Nitrogen carrier gas flows through 1;'h-inch'Peflon tubing to a solenoid valve which, when activated, directs the flow through the sample loop arxi, when deactivated, allows the flow to h,ypass the loop while the puff is drawn. The carrier then travels through connecting tubing to a stainless steel three-way valve where it is either directed to the modified injector port (4) of the Perkin-Elmer 3920 gas chromatograph or to a ~amphng port to which a trap is attached. See Figure I.. During a standardizing run (wit.hiautt trapping) (Figure 2), the carrier containing the sample is directed t:~ the injector port of the gas chromatograph. A simulated trap, filled completely with glass wool, is placed in the injector port to reduce its volume. :a number of such runs serves to establish the expected level of organic components in an average cigarette. For a run in which the sample will first be trapped on'Penax, then analyzed (Fit;ure :3), a reduced carrier flow (10 mLjmin) is used to purge the sample from the loop into the tapered end of the trap. Carrier gas flow was 12 rnL,tmin (30 psig). Injector port temperature was ry:it) °C; and FID temperature 1,50 °C. Colunrn. G(' C:ralumzn. The column used for thet determinatirm 4 isoprene, acetaldehyde, and acrolein was a modification of one used rnutinelv (1) for this purpo. se. The ~atatianarv ph&;e :;,:"'-Itrintethy lenedis'xyDdil:roropic~nitrile was synthesized by equilibrating a 1:" mixture of acryloinitrile and 1..3-prnpanrdrc7l , Pubh9ht:7 ?978 by the Amerscan Chemical Society

780 • ANALYTICAL CHEMISTRY, VC3L. 50. NO. 6, MAY 1978 ~ :MSrer~i;~k NNER Sy.lAY 1 i i " _.L.Er:oo vr.L•ae. ~ I r~uIA9GE ~ 1 ~ 4!?4.lvME. l : . ................. I I I as+~,~ LC+w ~;CMYTittiL i Af.VE CFNA-VAC PUMP ' ..... LRt.(• : 4 . MRF.1:', - N4Y Figure 1. ORNL single port smoking machine and attached Tenax-Gc trap (l 150PHENE 2' ACC7ALUEHY6E ~® MF4Y.YI. Fcri1t.mTE GfioPIONALOEHtDE .3~ 2-ME.iHYLFUR.AN•E1HY4.f4'HMATE .5) Aff PMYLACETATE ~ ACRSX.EM ~ ,8ACELQNC t ~ I TIME ~ .......:.........._...._.......1_.....-_...._.__.(_.__.,......__2,._..~....1. _:..._...~.~.L_.....__..,._ L_-,i ao sa 'a0 a° 30 zo 1q 0 TIME(mul Figure 2. Standard chrornatogram of the gas phase of a Kentucky reference (1R1) cigarette (not trapped). Vol. of gas phase, 6.7 mL; injector port, 250 °G, FID, 150 °C; flow rate, ^• 12 mL N2/min I y1D 1:40PRF,'NE (23 eC ETALDEHtDE i3) MCTHYL FJRMGTE rROP(aN4LOEHVDE (g 1 1-. ME TNY L F UR e. N• E THYL F0RM4T E (El ME'+HYC4CET4TE Cl`~ ~7( 4CR0LeIN 4CETONE ~ 7o 60 3C) 40 30 TIME (mm) 10 0 Figure 3. Chromatogram of the gas phase of a Kentucky Reference (1R1) cigarette (desorbed from Tenax-GC). Vol. of gas phase, 6.7 mL; trapping flow rate 10 mL/min; desorption time. 10 min ii; 250 °C (trap, 5'/{-in. X 5-mm i.d. packed with 60/80 mesh Tenax-GC. Other conditions same as standard run at room temperature for 72 h, then vacuum distilling and collecting the fraction boiling at 170 °C at a pressure of 1 man (9). A l5-ft X 2-mm i.d. Pyrex column was packed with 15 vrk 'To of the liquid ' on 100 to 120 mesh C:hromosorb W(HP), and _ conditioned overnight at 90 °C and 20 n1L Njrnin. The FID was used for all analyses. C'hrorourtepgraphLc Prrrcedurc. Straig;htt runs used for calibration were performed by sampling as described. The saanple is collected cryothermally at thee head of the column by cooling the column to -70 °C with liquid nitrogen. When sampling is complete, temperature programming begins at 8°C/'min to 10 °(.'., then is held constant for 4 nxin before programming at I°C f rnin to 70 °C. Trapped runs are performed by trapping the sample as de- scribed, then reruoa>ing the simulated trap from the injector port Table I. Trapping Efficiency Relative to Sample Size"~ Isoprene, Acetaldehyde, Acrolein, hp34°C bp21°C bp52.5°C Vol. of gas 4.2 6.7 4.2 6.7 4.2 6.7 phase, mL Av tcg std run 6.4 10.2 10.7 17.2 1.4 2.2 Avµg 6.3 8.0 8.6 1.0..1 1.4 1.9 trapped run Recovered, ~`0 98 78 80 59 100 86 a Trapping flow rate 10 mL/min, desorption time, 10 min. and replacing it with the Tenax trap with its tapered end at tlie column inlet. Note: The sample is always back-flushed from the trap. With the spring arid septum cap in place, the carrier gas is set at 30 psig and the trap is desorbed for 10 min at 250 °C onto the cold column. When desorption is complete, ternperatr.tre programming begins and follows that of the straight run. DISCLISSION •'I`he technique described in this paper requires 12 min for sampling, 10 rnin for desorption, and lZ h for analysis; a taatal of less than 2 h. Straightt runs require 1.0 min less since they do not have to be desorbed. Trapping the sample provides a means for sampling a source which could not be analyzed otherwise. EFFICIENCY Efficiency of the trap is critically related to the sample size. method of storage, and the conditions under which it is desorbed for analysis. These criteria will be discussed in the order of their importance. The capacity of a trap cannot be easily defined. Each constituent has a unique distribution coefficient which de- scribes the ratio of the amount of that constituent adsorbed on the Tenax-GC to the amount contained in the gas passing through the void volume of the trap. The amount of any compound which can be adsorbed on the Tenax (trap capactiy) is a function of the concentration of the total sample. (Concentration in thi..5 system is determined by sample volume and the rate of carrier flow through the sample trap.`r Therefore, trap capacities (breakthrough volumes) of sin ;le components will differ depending upon whether they are trapped as individuals or are contained in a nlulticornponent mixture. Optimutn sample size can be determined by plotting the percentage of the desired component trapped vs. total sample size. The optimum sample size obtained by these criteria was 4.2 mL. Each curve should contain a flat maximum (provided detection limits are not exceeded) along which the sample size is within the capacity of the trap, thus allowing one to choose the optimum sample size for any component. One can assume the trap capacity has not been exceeded if 95% of the desired component is recovered. If a lower percentage is recovered, the trap may be overloaded. See Table 1. Flow rates between 1.7 mL f min and 250 mL/min were tried for the carrier gas flow through the trap during sampling. Multiple determinations were made at 4, 1.3, 15, 20, and 40 mL f rnin. Figure 4 shows the amcbunt. of each constituent trapped vs. the rate of flow through the trap. Arnotuats collected in traps filled at less than capacity are also dependent on the rate of flow but to a lesser degree. The curve tends to have a broader apex at which the efficiency is more de- pendent on the total volume of flow rather than the rate of flow. The apex shifts toward lower flow rates so that the optimum flow is not the same for all sample sizes. Other factors which affect efficient Tenax use are con- trollable by the analystt. Tenax is available in two mesh sizes: :35 f ti0 and G0;'$tl. The finer mesh size proved to be a better

ANALYTICAL CHEMISTRY, VOL. 50, NO. 6, MAY 1978 • 781 600 500 ACETALDEHYDE ISOPRENE 75 65 .__....-.. _. ..... _.......... . 0 5 10 15 20 Nt FLOW RATE DURING TRAPPING (rcofffmin) Figure 4. Trapping efficiency vs. flow rate adsorbent, probably because of increased total pore area. While the trap size is limited to 5t f,; inches by 9-mm o.d. by the dimensions of the injector port, variations are possible for the bore diameter and the volume of packing. The first trap was 7-mm i.d. with about 2 mL of Tenax. Trapping efficiency was poor and gas chromatographic elution peaks were broad and tailing. One milliliter of packing was then tried with even more disappointing results. Double plugs r!f la/2 mL each in a single trap resulted in a sig aificant improvement, but proved to be no better than 3 mL in a single plug. This led to the use of a smaller bore (5 mm) glass tube 51 f.a inches long which was fully packed with Tenax (2.2 mL), and which resulted in a large improvement in resolution without a significant loss of capacity. A 3-mm i.d. trap was tried (0.8 mL'henax) for which the resolution was even better. 1:3ecause of the loss of capacity for the 3-mm i.d. trap, however, further work was done with the 5-mm i.d. fully packed trap. These results lead to the following conclusions: (1) A decrease in the bore diameter of a trap increases the resolution. (2) Greater component retention per unit volume of packing may be achieved with a decrease in bore diameter (7). Desorption. Of all the factors related to the desorption of the trap, the most critical is backflushing of the sample. No recognizable chromatograms were produced without using this technique. The time required for a trap to he desorbed ofitc> the cryothermal column refers to desorption time prior to ini- tiating the temperature program on the chreamatagraph. The trap remains in the hot injector port during the.entire run. The proper time is important for the analysis of labile compounds (see Figure 5). Likewise the temperature of the injector port must be hot enough to release all the components of interest without inducing decomposition of the more re- active ones. An injector port tempe.r::ature between 250 °C and :3t7C1 °C proved to he best for isc,prena::. acetaldehs•de, and acrolein. It became apparc:ntt after surne time that part of the sample was not recovered, even though it was completely adsorbed ,:yn the trap. Four possible causes existed: (I) The time _A¢;ROLEIN 300 4.2m1 OF GAS PHASE I 200 _ f f I 0 5 10 15 20 DESORPTION TIME (min) Figure 5. Trapping efficiency vs. desorption time elapsed between placing the trap in the injector port and replacing the septum cap on the partt allowed some sample to escape. (2) The sample was retained by chemisorption on the Tenax packing. (3) The sample reacted with oxygen in the injector port. (4) The species of interest reacted with other constituents in the sample. An injector device for glass traps machined to attach to the injector port in place of the septum cap allowed purging of air from the injector port before injecting the trap into the hot zone and ensuring that the sample did not desorb pre- maturely nor react with atmospheric oxygen. Experiments performed here and by others (6) indicated that the sample clid not react with the Tenax. Thus, it was concluded that the components of interest were probably reacting with other labile constituents in the smoke. The loss was not large and was rectified by using smaller samples and lowerinl; the in- jector pcartt temperature. Traps were assumed to be reconditioned after desorption. Repeated use of up to several dozen times resulted in no loss of efficiency, as is reported also by Pellizzari, Bunch, Berkley, and McRae (3). Aging. Experiments on the aging of trapped samples up to three days l:arcaduced conflicting data. There was no loss of components in samples aged for 22 h and for 70 h, re- spectivelv, while samples aged only 5 h suffered high losses. Statistical analysis of aging data by the methrrd of least squares using a linear model showed no trend in the plots of peak areas c'tf acrolein and acetaldehyde vs. aging time a.ap to three days. Isvprene showed only a slight downward trend. Traps were capped with number three polyethylene end caps and placed on a shelf in the laboratory for aging. Traps were not protected from rrx,m light. Itt is likely that ;;umc: of the caps produced a better seal than crthers.` Future work on aging should provide a meanfi of capping the ends (if the trap tightly with Teflon seals, orpossihly sealing the t.rap in gl:ass capsules, and storing them in the dark. :kgint (tf samples for over one week has been reported with the trapping of many organic compounds withcsut a signitic°ant. loss of sample (.3). i

782 • ANALYTICAL CHEMISTRY. VOL, 50, NO. 6, MAY 1978 C'f7NC'L[ISIf3NS By use rtf the technique de:,x-rihed herein, itt is E7ussible to trap and recover the mure highly labile c<,rnponents in the gas phase of cigarette smcakes All of the isufaeene and acrolein and 8(} % of the acetaklehycle was recovered. l'attern rece.,l;nitian of the chromatograms indicated that other major components in the gas phase were recovered also. 'I'hee adscrrpttr7n characteristics of many of these on Tenax have already been reported (2-8). Improvements need to he made in the storage of aged samples. Given these, this method should be ap- plicable to remote sampling of cigarette smc7ke and other gases for subsequent analyses at another site. ACKNOWLEDGMENT The assistance of W. H. Baldwin of C)RNI. Chemistry Division and C.-H. Ho of the ORNL Bio f Organic Analysis Section in synthesizinl; the 3,a'-(trimethylenedioxy)di- prcapionitrile is greatly appreciated. C. K. Bayne, Computer Sciences Division, perfcsrmed statistical analyses of the data t . v 011 ahing of tlte gas phase in Tenax. LI'I'ERATURE7 CITEI) (1) A, U. Horton and M. R. Guerin, Tobacco, 176, 45 (19741 (Tab. Sci. No. 19). (2) W. Bertsch, R. C. Chang, and A. Ziatkls, J, Chromatogr. Sc1., 12, 175 (1974), (3) E. D. ReElizzari, J. E. Bunch, R. E. Berkiey, and J. McRae. Anal. Lett., 9 (1), 45 (1976). (4) A, ZIatkEs, H. A. Lichtenstein, and A. Tishbee, Chromatoyraphra, 6(2), 67 (1973). (5) J. S. Parsons and S. Mitzner, Eviron, Sci. Tecdrnol., 9(12), 1053 (1975). (6) R. van Wijk. J. Ghrama.atorr. Sci., 8, 418 (1970). (7) L. CY, Butler and M. F. Burke, J. C;hronkstogr. Sc6., 14, 117 (1976), (8) J. Janak, J. Ruzickova, and J. Novak, J. Chronaatogr., 99, 689 (1974). (9) "The Chemistry of Acxylonitrile", 2nd ed., American Cyanamid Co., 1959. RECEIVED for review November 4, 1977. Accepted February 7, 1978. Research sponsored by the National Cancer Institute, The Council for Tobacco Research--USA, and the Depart- ment of Energy under contract with Union Carbide Corpcr- ratiun. S.G.Z. from Centre College, Danville, Ky. 4U-4''2, was an ORAU summer research participant.