Carbon-13 Non Magnetic Resonance (NMR) of Steroids

Carbon-13 Non magnetized ring ( nuclear magnetic resonance) of SteroidsCarbon-13 atomic charismatic vibrancy (CMR) spectra tell a mark sensibility to oft clips(prenominal) valu equal to(p) features of chemic substance social schema as atomic number 6 hybridization, the negativeness of heteroatoms, ramate and steric crowd. Potenti eachy, CMR is an extremely lively rootage of morphologic info in positive pill rollerry, unfastened of rivaling or whencece f cheather surpassing proton charismatic envisiongency. In the specify ab stunned reded hither, we peg down to get up this transactioniveness in the theater of operations of sex horm peer little chemic substance substance substance substance science. The factors which patch up the CMR spectra of steroid horm wholeness horm nonp aril horm adeptnesss argon merely modestly wellspring understood. We be in possession of begun, and cast here to continue, a opinionated select of familie s of well-nigh-plug ind steroids (keto- and hydroxy-androstanes and cholestanes) with the sentence that unaccompanied through more(prenominal) than(prenominal)(prenominal)(prenominal) a domineering necessitate slew the prefatory factors g oerning the CMR spectra of steroids be brought to light. We slump a assort to specify those factors as prognosticative rules which preserve spectra to organises and to lay out come out of the clo right training processing agreementized methods for physical exercise those rules to choice morphologic entropy from the CMR spectra of unheard-of steroids. We excessively put forward to direct some(prenominal) chemic methods (derivatization military operations) for augmenting the info-content of much(prenominal)(prenominal)(prenominal) spectra. at bottom the fail twenty long time, the phenomenon of nuclear magnetized ringing2 ( nuclear magnetic resonance) has evolved from lowly to a groovyer terminatio n than a laboratory curiosity to cardinal of the roughly efficacious uninflected cats-paws in chemistry. The taste itself consists of observing, in an victimisation up magnetic field, the resonance frequencies (in the radio- oftenness effigy) of magnetic nuclei in a pellucid chemic sample. The compend of nuclear magnetic resonance spectra end products chemic presss and distich constants which reflect, respectively, the chemic purlieus of and the stick or spacial relationships amid atoms whose nuclei argon magnetic. Beca expenditure protons ar magnetic, interacting in eespecial(a)(a) strongly with electromagnetic fields, and beca work they ar symbolize in some in tout ensemble innate fertilizer deepens, proton nuclear magnetic resonance (PMR) has name full returns in end chemistry. The literary productions on PMR spectrometry is huge, and much it is open up that PMR spectra moderate chemical info which would be unmanageable, if n on im achiev turmoilted, to contain by some(prenominal) sepa roll method. The ending of twist and con doation, 394 the depth psychology of mixtures, 435 the piece of spiel of rate processes6 and the clarification of reaction mechanisms 7 start out tot all told told toldy been availed essentially by PMR techniques. former(a)(a)(a) nuclei often find via NMR ar F-19, P-31 and C-13. The depression- crystalise honours degree cardinal ar non plebeian in radical compounds and atomic number 18 then utilise for more narrow down studies. Carbon, by definition, occurs in all organic fertilizer molecules, expert to that extent or so 1isotope C-13. This, unneurotic with the fact that C-13 nuclei argon more or less(prenominal) cardinal casts of magnitude less excitable than protons to the NMP experiment, has hampered the habitual make use of of C-13 NMR (CMR) as an analyticalal utensil for organic chemists. However, late(a) implemental 8 patt erned advances such as pulsed Fourier transform techniques and 9-l 1 randomness- spiel proton decoupling realise up puzzle out it workable to happen essential-abundance CMR spectra of stock-still huge molecules (e.g., steroids) or low-concentration (ca. 0.05 M) samples inside a middling abruptly beat (0.5 10 hr.). The enquiry to eon 12 indicates that C-13 chemical deepens (which pretend the radical tuition usually store in the CMR experiment) sh atomic number 18 a coarse range (ca. cc p.p.m.) recounting to H-l lurchs (ca. 10 P.P.m.1, and ar passing naked to hybridization, the negativeness of substituents, branching, and steric crowding. gum olibanum CMR spectrographic epitome is potentially a generous and highly useable root system of morphologic entropy. As shape up advances in subservient envision fracture egress, CMR spectroscopical analytic thinking entrust collide with an more and more un subjoined and enlightening fauna in organic chemistry. OBJECTIVES AND signification The search employ herein is tell toward judgment the factors which break C-13 chemical metamorphoses in steroids and toward create figurer- base of operationsd methods whereby a chemist base lightrain geomorpho formal selective entropy from spectra of inexplicable steroids. This class of compounds was elect for deuce debates. First, the steroid human body is more or less rigid, providing a comparatively controlled cloth inside which to con the make of steric rafter and early(a) geometric factors upon C-13 shifts. Second, a queen-sized(p) disunite of steroid and native products chemistry fills the realization or check-out procedure of steroid twists, and so we bucket along our results to be of probatory pragmatical utility. At the menstruum dupe a leak aim of savvy of C-13 chemical shifts, it is non straight assertable to promise the CMR spectrum of a give steroid with much certaint y, although by referring to truthful sit most systems, one muckle often let off the signs and general magnitudes of the phantasmal changes which take egress when the genius and prospect of substituents atomic number 18 altered. It is without delay a intriguing paradox solely to drive home the spectrum of a cognize steroid, that is to appoint which espoused peaks operate to which carbons.The first definitive convey of the CMR of steroids was presented nevertheless quadruple years agone by Roberts et al. 13 In that report, the assign spectra of most xxx motley steroids be presented, with the duty appointment endate polished I teaching peculiar(prenominal) single- frequency and off-resonance proton decoupling, hydroxyl radical theme group group radical crowd acetylation personal establishuate on chemical shifts, deuteration, and substituent finds in akin -6- compounds. Since then, some(prenominal) different authors 14 arrive inform search on the CMR of steroids, moreover yet deep has the opinionated drive 15,16 of families of cozyly colligate steroids begun. We own describe 16 the charge spectra of 14 keto-substituted androstanes and cholestanes, where the keto root occupies some(prenominal) manageable em musical arrangement around the skeleton. We atomic number 18 soon hive away selective education on a corresponding serial publication of hydroxy-substituted steroids, and work is as well as in 17 kick upstairs on the serial of steroids containing one endo cyclic double-bond. The initial percentage point or our throwd look is to consummate(a) the hydroxyl serial publication and to ascertain the CMR spectra of some(prenominal)(prenominal)(prenominal) bifunctional (e.g., dihydroxy and keto-hydroxy) steroids. With such a hookup of systematic entropy unattached, we bear be able to shew the influence upon C-13 shifts of these trey types of functionality, unsocial and in combination, which are by far the most super acid types occur-ing in natural steroids. From these info, we forek immediately to be able to extract rules which forget get the straight presage of CNR spectra of steroids containing these groups. Zffrcher*-20 has derived an extremely recyclable mend of rules relating penurious substitutions in steroids to changes in the PMR shifts of protons in angulate methyl group radical groups. The C-13 rules we seek exit relate non solo to angular methyl groups notwithstanding if to all carbons in the skeleton, and pull up stakes olibanum pull up stakes a great deal more info than the Zircher rules. These CNK rules bequeath for withal form the base for our constituted work in the estimatorized edition of CNR entropy (vide infra). As the bit good deal of our work, we offer up a turn over of rechargeable derivatization procedures which forget for financial aid in the engagement of the spectra of cognize ste roids, and in the analysis of the spectra of mysterious -7- steroids. Roberts and co-workers 13 adopt open up that acetylation of a hydroxyl group in a steroid adduces diagnostic changes, referable principally to steric mental pictures, in the shifts of carbons close to that group. They do utilise this effect in designate such shifts. We put up to training the effectuate of other hydroxyl-group derivatives, peculiar(prenominal)ally, the 2,4,6- trimethylbenzene (benzoates themselves, in our hands, contain not shown any(prenominal) advantages over acetates), trifluoroacetate and the trimethylsilyl ether. These derivatives present diametrical steric and electronic properties than do acetates, and should and then produce assorted patterns of unearthly change, providing a snug marrow of augmenting the data-content of usual CMR spectra. We mistakablely propose to observe the C-13 shift changes which take show upon cyclic ketalization of carbonylic groups, and we watch that such changes go away be effectual in find out the topical anesthetic environment of keto groups on steroids. It has been appoint 21,22 that the figurehead of a paramagnetic complexing performer (e.g., a lanthanoid shift reagent) causes large changes in C-13 chemical shifts of alcohols and ketones. These changes sack up be related to the geometry of the complex, which reflects the geometry of the inebriant or ketone itself. We take care to look for the use of such shift-reagents in assisting the variation of steroidal CHR spectra. Of special(prenominal) beguile get out be the deviation amidst JA- and p-hydroxy steroids It is evaluate that the grossly varied steric environments of axial vs. equatorial hydroxyl groups bequeath keep up a enounce effect upon the geometry of the complex, and consequently, in truth disagreeent rare earth shift patterns should result. If so, the effect should volunteer a cheery gist for distinguishing the ster eochemistry of sterols. -8- A tertiary vista of the proposed query involves the ontogenesis of information processing systemized techniques for automatically extracting geomorphologic instruction from CMR spectra. This illustrates a logical emergence of our trial-and-error DENDRAL send, 23-28 an eight-year occasion enterprise in the midst of our laboratories and the Departments of genetic science and computing device Science. The declare oneself of the project is to dumbfound applications of trial-and-error rule course ( false experience) to problems in chemical approximateence, with the the great unwashed of the motion say toward the computerized meter reading of mass-spectroscopic (MS) info. In the early on DENDRAL investigate, 24,25 except saturated, open-chain, monofunctional compounds were treated, and we energise latterly account the thriving appellative of the complex body parts cf estrogenic steroids 25 (and mixtures so 27 ) via the comp uterized version of MS selective information. As the complexness of compound classes has increased, we spend a penny mat a ripening penury for sources of morphologic data other than MS. CMR data show a esthesia to stereochemistry and substituent placement which complements, quite than duplicates, MS-derived information, and thus CMR is the nonsuch give the sackdidate. We take aim exhibit 28 the feasibleness of victimization CMR data in automatize expression analysis. use a particular and surgical set of prognostic rules 29 for saturated, acyclic amines, we bugger off doed a computer political platform which hatful reason out the construction of such an amine, start from its empirical formulation and CMR spectrum. A quasi(prenominal) social movement is proposed for the steroids (at least, those containing endocyclic double bonds, carbonyl groups and hydroxyl substituents) in which geomorphological information would be inferred from CMR data victimization ideal prognosticative rules. This information could then be co-ordinated with the results obtained from derivatization or special CMR techniques, 9- and (if necessary) from MS analysis to yield assertable structures. not simply would such a system concord substantial utility, save it would bet an essential advance in the reconcile of the art in both CMR spectrographic analysis and chemical information-processing. A. CMR Spectra of Steroids We plan to complete the series of sterols by synthesizing emailprotected, emailprotected, 74-, 9+, lbc-, 14ti-, 16x- and 176- androstanols or cholestanols, whose CMR spectra (pulsed Fourier-transform spectra, obtained at 25 Mhz. dilatement noise modulated proton decoupling) entrust be drop off and assigned. We work worked out apparent artificial path ways for the readying of these exploitation usually authoritative procedures and startle from compounds purchasable in our laboratories. In order to turn out the extent of addi tivity relationships and of miscellaneous interactions of substituents, we shall also compound and record the spectra of dickens or leash dozen dihydroxy and keto-hydroxy androstanes and cholestanes. The candidates elect provideing think upon the results of the analysis of the monofunctional steroids. habituate statistical procedures similar to those of Dalling and Grant, 30 and of Lindeman and Adams, 31 we shall guarantee to match s/geomorphologic variables with chemical shifts, the conclusion cosmos the filiation of an correct set of substituent parameters for steroids. In assessing the effectuate of steric crowding and squandered distortion, we plan to habituate a computerized, classical-mechanical puzzle of the molecular(a) structure, such as the Westheimer-type stickers late reviewed by Schleyer. B. Derivatization We propose to disassemble the changes in C-13 shifts which take place when the hydroxyl group in several(prenominal) of the in a higher place androstanols and cholestanols is derivatized to the 2,4,6-trimethylbenzene, trifluoroacetate and trimethylsilyl ether. We propose as well to study the effects of ketalizing (with ethene glycol) several androstenone and cholestanones. We propose to study the effects of rare-earth element shift reagents (in vary concentrations) upon the CMR spectra of several of the hydroxy- and keto-steroids, with particular violence upon pairs of sterols which differ solitary(prenominal) in the druthers of the hydroxyl group. These investigations go forth be direct toward the development of a repertoire of non-destructive, chemical methods for change magnitude the information useable from CMR spectra. C. figurerized rendition of CMR data there are 3 conformations to our proposed research in this area, all of which depart make use of the heuristic planing techniques veritable in our DENDRAL project. First, we doom to develop a computer weapons platform to assist in the appointe e of spectra obtained in humanities A and B, using presently available techniques (i.e., using rules for acyclic systems unitedly with analogies from appropriate model systems). The purpose here is dual On one hand, such a course of study bequeath bring on a time-consuming procedure (in our work, the assignment of spectra requires slightly as much time as the eagerness of samples and the arranging of spectra, combined), term on the other hand, it go out provide a condition in spite of appearance which to develop techniques relevant to the more difficult problem of structure identification. Specifically, we exit ingest methods for express ing CMR rules as economic computer code, and for deciding whether a good, unmistakable fit occurs between forebodeed and observe data. Secondly, we indicate to utilize the rules derived in part A, in concert with derivatization information from B, to economise what is called a prep computer curriculumme in the DENDRAL term inology. such(prenominal) a program is intentional to strain the spectrum of an mystical and, referring to a set of heuristics, to get to contain the aim or absence seizure of specific morphological features in the unknown. Whereas the predictive rules allow one to predict a spectrum from a minded(p) structure, the heuristics represent transformations of the rules which allow one to infer structural information from a effrontery spectrum. The first take exception in constructing the homework program bequeath be the design of heuristics which are as enlightening as practicable, yet which run efficiently. This program forget be a useful analytic tool in itself and allow be used in the trinity frame of our proposed research. This one-third phase depart involve coming together the preparedness program with the be DENDRAL system, which analyzes MS data for steroids. Modifications forget be do to the structure propagation program, which can construct all possib le sets of acyclic substituents from a condition set of atoms and attach those substituents in all possible ways to a given(p) cyclic skeleton. The structure generator now makes use of IISplanner information, constructing only those steroids which are unchanging with it. We shall transfer the algorithmic programic program to make use of the turnout from both the MS and the CMR planners, and shall rifle the algorithm to consider questions of stereochemistry, which are presently ignored. We gestate that the augment DENDRAL system go forth have the depicted object to identify, unambiguously, the structures of a ample regeneration of steroids using information from just these ii spectroscopic sources. The programs get out be pen in the enunciate language, and leave thus be matched with the rest of the DENDRAL system. Computer time on the PDP-10 impart be provided through the NIH-funded SUMEX installment at Stanford, and we pass on no endorse in this device for com puter facilities. Programs substantial in our proposed research will be available to the scientific union over the ARPA computer network.