Tobacco Documents Online

I RECENT STUDIES IN NICOTINE CHEMISTRY - III Jeffrey I. Seeman Philip Morris Research Center P. O. Box 26583 Richmond, VA 23261 . . RECEIVED MAR 23 1qq3 ~~,.. ~.G.''•~_::ri~ 1.~ a

r ABSTRACT The synthesis in our laboratories of a wide variety of nicotine analogues has served as the basis for a range of chemical and pharmacological investiga_ tions. We have examined the effect of substituents on nicotine's structure, conformation, and chemical reactivity. A chemical model for nicotinic activity has been developed by a thorough evaluation of the methylation of these analogues, thereby simultaneously quantifying the relative and absolute nitrogen nucleophilicities of these nicotinoids. Implications and extensions of this work are discussed. . V

1. INTRODUCTION Nicotine (1) is undoubtedly the most well known tobacco constituent to the lay person, if not .the only constituent known to both the tobacco consumer and nonconsumer alike. This alkaloid may well have received more scientific attention than any other compound isolated from tobacco.1-11 N Nicotine has a long and splendid history (see Table 1). It was named after Jean Nicot, a French ambassador to Portugal, who in the mid-sixteenth century is said to have introduced tobacco seed and leaf to the royal courts r of France.2'S' Nicotine was first isolated in 1828, before the• isolation of such other important -alkaloids as codeine, atropine, papavarine and physo- stigmine.5 The correct structure of 1 was proposed by Pinner in 1893 and it was ~first synthesized by Pictet in 1895.5 The chemical literature of the nineteenth century is replete with reference to nicotine's reactivity. For example, the famous August Kekule reported the ethylation of 1 in 1853,12 some twelve years before he proposed the structure of benzene. Nicotine clearly has had a life of its own, independent of its inherent relationship. with tobacco and tobacco products. There have been many interestiRg chemical results which originated during studies on nicotine and related compounds. In perhaps a larger sense, nicotine has played' a central role in the investigation of mammalian neurophysiology and neurochemistry.

2 A major classification of the autonomic nervous system is the designation of the cholinergic synapses and neuroeffector junctions as either nicotinic or muscarinic, based on which of these two alkaloids (nicotine or muscarine) is activating.13 Nicotine continues to be an important "scientific tool in phys- iological and pharmacological studies of the nervous system".2 Importantly, the very concept of receptors was postulated in 1905 by Langley as part of his research on nicotine pharmacology.2 While nicotine was once used in the treatment of various human diseases, it no longer has any therapeutic applica- tions in man, though there remains some veterinary uses for the material. , J Probably its major utility is as an insecticide and a fumigant, where there remain specific applications for this natural and biodegradable compound.2 We shall first give a modest overview of the current state-of-the-art in nicotine. pharmacology. Second, to place our work in perspective, we will present a particular postulate in nicotine structure-activity relationships (SAR) which we will examine and evaluate in the course of the presentation of our results: Third, we will discuss the conformational properties of the nicotine analogues of pertinence to this study. We will then discuss chemical reaction modelling as it relates to nicotine pharmacological activity. Finally, we willi put together our physical and chemical evidence and suggest an important feacture in nicotine analogue SAR. II. NICOTINE PHARMACOLOGY: AN OVERVIEW ,13,14 Nicotine has numerous direct and indirect pharmacological activities which ace described in literally thousands of research reports and numerous review aptic!•es and books. Nicotine acts on both the peripheral and central nervous systems (PNS and CNS, respectively) as well as on the cardio- vascular system, the gastrointestinal tract, and the endocrine glands. Its

3 activity is particularly complex because it can have both stimulant and depres- sant phases of action. It acts at cholinergic synapses of autonomic ganglia and at neuromuscular junctions, first stimulating and then depressing (or blocking) activity. Nicotine can cause the release of catecholamines in a variety of organs, and it is this property combined with its stimulation of sympathetic ganglia which results in vasoconstriction and increases in blood pressure. Nicotine also stimulates the central nervous system, though its mechanisms of CNS action are less well understood than its PNS actions.2,13,14 . Many of the classical pharmacological tests for nicotinic activity relate to its PNS properties, e.g., contraction of the guinea pig ileum and frog rectus muscle, which when combined with the appropriate blocking or antagonism studies results in information regarding nicotinic activity. Other tests, e.g., contraction of rabbit aortic strips, are more complex in that nicotine may induce the release of adrenergic mediators which in turn are responsible - for the observed pfjarmacological events. Still other tests, e.g., blood pressure, may simultaneously incorporate the actions of a variety of mech- anisms involving both the PNS and CNS. Lastly, in the derivation of SAR, one is interestcd in determining relative toxicity (LD50) of the compounds under investigation,. and these certainly represent the effects of numerous systems.2,13,14 One cannot quantify or even describe the CNS properties of compounds in the same fashion that PNS activity is characterized. Behavioral pharmacol_ ogy has become a major tool in the evaluation of CNS properties of nicotine10 and many. other compounds of medicinal interest. In this field, the emphasis is om evaluating the effects of the compound on a variety of behavioral tasks, the end result being a behavioral profile from which CNS actions can be predicted'. ,

4 The actions of nicotine are mediated through its binding and subsequent interactions at nicotinic receptors located throughout an organism. It is evident that all nicotinic receptors need not be identical. For example, hexamethonium acts as an antagonist at nicotinic cholinergic ganglionic sites while decamethonium is inactive at the ganglia; the converse antagonistic activity for hexamethonium and decamethonium is observed at the neuro- muscular junction.14 Much work, has been done in recent years in the isolation, identification, and characterization of nicotinic receptors, and these studies have been particularly successful with regard to the peripheral J nicotinic cholinergic receptor from the electric organ of the Torpedo fish and electric eel.15 A wide range of receptor binding studies has been performed, both with purified receptors and with inhomogeneous mixtures, e.g., rat brain homogenate. 8 An important and crucial issue in 'the development of SAR is the relationship between receptor binding studies and in vivo prop- erti es . Until recently, ,one of the serious deficiencies in the development of SAR for nicotine: has been the lack of suitable analogues, particularly those incorporating the nicotine (or other tobacco alkaloid) ring system. For exampl~, very interesting and thorough classical pharmacological studies were reported some fifteen years ago by Barlow16 and Haglid17 and their collab- orators in which a large number of aminoalkylpyridines 2 were examined. A modest series of nicotine analogues was prepared by Yamamoto as part of his work on nicotinoids as insecticides.6 Of course, the tobacco alkaloids them- selves are structurally related to nicotine and have been the subject of many pharmaco4ogical tests. N iR1 ~n \R 2

5 To achieve our goal of developing a SAR for the tobacco alkaloids, we have defined the following subobjectives: 1. To synthesize a variety of nicotine analogues which incorporate different electronic, steric, and stereoelectronic features. 2. To determine a wide range of physical and chemical data for these analogues which would (hopefully) correlate with pharmacological results. 3. To choose for our syntheses specific analogues which can aid in other aspects of our work, e.g., in potential nicotine receptor isola- tion studies. ~ III. THE EXAMINATION OF A NICOTINE SAR HYPOTHESIS: THE HAGLID PROPOSAL In 1967, Frank Haglid reported that 4-methyinicotine (3) was pharmacol- ogically inert in a number of tests.18 He suggested that the inactivity of 3 . was due to its inab$lity to adopt specific molecular conformations necessary for receptor binding.18 Of course, this is only one of a number of possible explanations fdr low activity, e.g., an alternative possibility is steric bulk around,, the pyridine ring. We decided to examine Haglid's hypothesis in light of our own interest in this area, and we prepared a series of pyridine substituted nicotinoids in addition to 4-methylnicotine: 2-methyl, 5-methyl, and 6-methylnicotine (4-6),19-21 ~ CH3 `N.--~ CH ~ CH 3 3 3 5 VV 6 VV N

6 Figure I illustrates the activities of these compounds and nicotine with re ard to three tests: LD 22-23 g so. guinea pig ileum, and rat blood pressure. In all cases, S-methylnicotine and 6-methylnicotine were equiactive, within an order of magnitude, to nicotine while both 2-methylnicotine and 4-methylni- cotine were significantly less active than nicotine. These results confirm Haglid's pharmacological data and are consistent with his hypothesis,1$ since one would anticipate that a methyl substituent at nicotine's C2 would cause . the same across ring interactions with the pyrrolidine moiety as a methyl group at C4. But consistency is insufficient proof when we are dealing with J so few compounds. We will further illustrate our work in the area of nicotine SAR by examining the Haglid postulate in terms of the conformational and chemical properties of these and other analogues, with a goal of further defining the limits and value of this particular SAR. IV. THE CONFORMATION OF NICOTINE . 11 Specification of. nicotine's conformation requires definition of three stereochemical features: (a) the orientation of the pyrrolidine N'-methyl group relative to the pyridine ring (cis as in 7, trans as in 8); the conforma- tion of ithe five membered pyrrolidine ring; and the orientation of the pyridine ring relative to thee pyrrolidine ring, conveniently described by the dihedral angle t(H2'C2'C3C2) (c.f., 7). By conformation we mean "any one of the infinite number of momentary arrangements of the atoms in space that result from rotation about single bonds. "24 If we strictly keep to this definition, it may be argued, and in fact has been argued' by this author, that the orientatianal. properties of the N'-methyl group in nicotine is a configurational question, not a conformational one.25-27 This is a subtle distinction about which: there remains some debate among practicing stereochemists, and we will not concern ourselves about it in this context.

7 CH3 A CH3 7 8 VV ~ A. The Orientation of the N'-Methyl Group . In the x-ray analyses of both nicotine dihydroiodide2$ and nicotine- salicylic acid' complex (1:1),29 the N'-methyl group was found to be trans to the pyridine ring, as in 8. However, this result does not necessarily reflect solution. or gas phase conformational propensities, since if is well known that crystal lattice forces are not always suitable models for alternate environ- ments. Furthermore, the crystalline samples were undoubtedly prepared under conditions in .~which equilibria between 7 and 8 were operative (c.f. Scheme I), and,/it is theoretically possible that the acid salt of a minor component could have crystallized preferentially. A,number~of theoretical studies have also suggested that the N'-methyl group is trans, at • least for an isolated molecule in the gas phase.22,30-34 That 8 would be more stable than 7 is intuitively reasonable on the basis of steric hindrance. However, Chynoweth, Ternai, Simeral and Maciel reported in the first solution phase experimental data on this subject "that the N- methyl group is preferentially on the same side of the pyrrolidine ring as the pyridine ring."35 Their study involved' the observation of an intra- molecular nuclear Overhauser effect (NOE) on the protons attached to C2 and C4 (H2 and H4 respectively) when the resonance of the N'-methy) group

8 was irradiated. Chynoweth, et al. based their conclusion on three factors: (a) the observation of the NOE; (b) an NOE can be observed only when the proton of the irradiated resonance and the proton of the observed resonance are spacially closed to each other; and (c) the N'-methyl group is close to H2 and H4 only in conformation 7,35 One factor which Chynoweth, et al. did not consider was the effect of the nitrogen inversion process on the NOE experiments.35 Scheme I il- lustrates the equilibria involved. Let us assume that 8 is the more stable isomer. If nitrogen inversion (7 F 8) is fast relative to 1/T1(8), where T1 , ~ ~ is the proton relaxation rate, then during the NOE experiment, there would be sufficient time for the methyl group of 8 to invert to 7prior to relaxation and return to spin equilibrium. The net effect could be the observation of an NOE. even though the resonance irradiated was that' for the N'-methyl group in 8 and not 71 Alternatively, the major isomer could be 7 as concluded by Chynoweth, et al. though based on inconclusive informa- tion.25,35 To differentiate between these two alternatives, we determined the NOE for nicotine under conditions in which the rates of interconversion 7 F 8 were mtlch slower than 1/T1(8).25 This condition was obtained when nicotine was dissolved in a very strong acid (e.g., trifluoroacetic ac4d), resulting in the diprotonated nicotine salts 11-12 '(see Scheme I and Fig. 2). Under these conditions, we were able to establish that deprotonation at the pyrrol- idine nitrogen was very slow on the NMR timescale, and' certainly consider- ably slower-than 1/T1(8). Table II indicates that no NOE was observed at