1625 were chosen because these groups are essential components of proteins which are known to act as donors in molecular complex formation in biological reactions with vitamin K I , which, e.g., are responsible for the maintenance of the normal blood coagulation .9 Since it is believed that the charge transfer in these complexes is accomplished by the unpaired electron of the nitrogen to the quinone group of the vitamin K I (or a-tocopherolquinone), this effect should be enhanced by the presence of additional n-butyl groups. The fact that the opposite was observed, namely that K , decreases with the number of n-butyl groups present, is therefore not the result of an inductive effect, but could be explained in terms of a steric hindrance which makes the formation of the molecular complexes with the higher n-butyl substituted donors more difficult. Vitamin D) has been known as a good electron donor in biological molecular complex formation." The results of the present study where the formation constants K , for vitamin K I-vitamin D3 complexes have been assessed in cyclohexane solution (see Figure 2 and Table I) confirm this observation and suggest very similar donor capabilities for vitamin D? and mono-n-butylamine. The strong reactivity displayed by a-tocopherolquinone, which is believed to be one of the major products in the oxidation process of vitamin E, toward Ps may open up an interesting possibility of studying the oxidation process of vitamin E. While a-tocopherolquinone reacts very rapidly with Ps, the rate constant is 2.15 X 1O'O M-I s-I in benzene, and is even relatively reactive in its complex with indole (Kob,d = 0.85 X 1 0 ' O M-' s-I), the nonoxidized vitamin E shows (in benzene solution) hardly any reactivity toward Ps. Thus this drastic difference in the behavior of the oxidation product of vitamin E and vitamin E itself could be, utilized for further studies of
the biologically interesting autoxidation process of vitamin E, which supposedly inhibits the autoxidation of the unsaturated fatty acids.2 Summarizing, it can be said that these initial investigations of molecular associations in biological systems by positron annihilation techniques seem to support the feasibility of this new technique in the study of biological reactions. References and Notes Work partially supported by the U S . Energy Research and Development Administration. For examples, see A. Szent-Gyorgyi,"Introductionto SubmolecularBiology", Academic Press, New York, N.Y., 1960;J. Rose, "Molecular Complexes", Pergamon Press. Elmsford, N.Y.. 1967;A. Pullman and B. Pullman, "QuantumTheory of Atoms and Molecules in the Solid State", P. 0. Lowdin, Ed., Academic Press, New York, N.Y., 1966, p 345; S. Shifrin, Proc. Natl. Acad. Sci. U.S.A., 58, 148 (1967):S. Yomssa, Proc. Theor. Phys., Suppl., 40, 249 (1967). Y. C. Jean and H. J. Ache, J. Phys. Chem., 80, 1693 (1976). For general references, see (a)J. Green and J. Lee, "Positronium Chemistry", Academic Press, New York, N.Y., 1964: (b) V. I. Goldanskii, At. Energy Rev., 6, 3 (1968);(c)J. D. McGervey in "Positron Annihilation", A. T. Stewart and L. 0. Roellig. Ed., Academic Press, New York, N.Y., 1967, p 143; (d)J. A. Merrigan,S. J. Tao, and J. H. Green, "Physical Methods of Chemistry", Vol. I, Part 111 D, A. Weissberger and B. W. Rossiter,Ed., Wiley, New York, N.Y., 1972; (e)H. J. Ache, Angew. Chem., Int. Ed. Engl., 11, 179 (1972);(f) J. H. Green, MTP lnt. Rev. Sci., 8, 251 (1972): (g) V. I. Goldanskii and V. G. Virsov, Annu. Rev. Phys. Chem., 22, 209 (1971). W. J. Madia, A . L. Nichols, and H. J. Ache, J. Am. Chem. SOC.,97, 5041 (1975). V. I. Goldanskii and V. P. Shantarovich, Appl. Phys., 3, 335 (1974). P. Jansen, M. Eldrup, B. Skytte Jensen, and 0. Mogensen, Chem. Phys., I O , 303 (1975). B. Levay and P. Hautojarvi, J. Phys. Chem., 76, 1951 (1972). For quinone and Its biological review, see R. A. Morton, "Biochemistry of Quinones", Academic Press, New York, N.Y., 1965. For examples, R. S. Snart, Biochim. Biophys. Acta, 88, 502 (1964);D. E. Laskowski, Cancer Res.. 27,903 (1967);G. Saito and Y. Matsumoga, Bull. Chem. SOC.Jpn., 44, 1788 (1971);L. M. Amzei, R. J. Paljak, F. Saul, J. M. Varga, and F. F. Richards, Roc. Natl. Acad. Sci. U.S.A., 71, 1427 (1974). T. L. Williams and H. J. Ache, J. Chem. Phys., 50, 4493 (1969). A. Szent-Gyorgyi in ref 2.
Regiospecific and Enantioselective Horse Liver Alcohol Dehydrogenase Catalyzed Oxidations of Some Hydroxycyclopentanes' a,b Anthony J. Irwinlc and J. Bryan Jones* Contributionfrom the Department of Chemistry. University of Toronto, Toronto, Canada M5S I A I . Received July 13, 1976
Abstract: Horse liver alcohol dehydrogenase (HLADH) has been shown to have the ability to retain its enantioselectivity while effecting regiospecific oxidation of only one of two unhindered hydroxyl groups within the same molecule. This provides a synthetically useful combination of properties which cannot be duplicated in a single step by traditional oxidation methods. AI1 reactions were performed on a preparative (up to 1 g) scale and proceeded in good yields. HLADH-catalyzed oxidation of (f)-cis-2-(2'-hydroxyethyl)-3-cyclopenten- 1-01 (1) was regiospecific for the primary alcohol group. The reaction was also enantioselective, and I R,2S-cis-2-carboxymethyl-3-cyclopenten1-01 lactone (4, 49% optical purity) and unchanged IS,2R-1 (23% optical purity) were isolated. The same regiospecificity, but no enantioselectivity, was observed with (f)-cis-2-(2'-hydroxyethyl)cyclopentanol (2) as the substrate. Both enantiomers of the prostaglandin synthon 4 were subsequently obtained (37 and 47% optical purities) via HLADH-catalyzed oxidation of the racemic hemiacetal precursor of 4. For the 1,3 isomer of 2, (f)-cis-3-(2'-hydroxyethyl)cyclopentanol (3), secondary alcohol regiospecificity is manifest with 3-(2'-hydroxyethyl) cyclopentanone (9) being the major product. The enantioselectivity of this reaction is very high, giving 3s-9 and recovered 1R,3R-3 of 97 and 70% optical purities, respectively. The regiospecificities observed were as predicted by the diamond lattice section of the active site. The model is more equivocal, but still useful, in analyzing the enantioselectivities of the above reactions.
In recent years, the requirements of synthetic chemists for reagents capable of effecting selective or asymmetric transformations have increased dramatically. Enzymes present Irwin, Jones
unique opportunities in this regard, and the exploration of their properties as chiral catalysts is now receiving considerable attentiom2 One of the great synthetic attractions of an enzyme
/ HLADH-Catalyzed Oxidations of Hydroxycyclopentanes
1626
is that the various facets of its specificity can endow it with the potential for effecting highly controlied and selective transformations in a single step which would otherwise involve several operations if more traditional reagents were used. In this paper, alcohol dehydrogenase catalyzed oxidations are explored in which regiospecificity and enantiospecificity operate concurrently. This combination is one whose asymmetric synthesis value is immediately apparent. Several alcohol dehydrogenases, which catalyze C H ( 0 H ) C=O oxido reductions, are synthetically usefuL2 Of these, the NAD+ 3-dependent enzyme from horse liver, HLADH,3 is the best d o c ~ m e n t e d ,but ~ , ~the question of whether it has the capacity to effect regiospecific oxidoreduction of only one of two or more hydroxyl or carbonyl groups within the same substrate while retaining enantioselectivity has been virtually ignored. The recently r e f i r ~ e ddiamond ~.~ lattice5 section of the active site was used to select suitable substrates for evaluating this aspect of HLADH specificity, and, in keeping with our overall to extend the synthetic utility of the enzyme, the dihydroxycyclopentane compounds 1-3 of more organic
H L A D Y 20 C , pH 9
NAD' recycling 45 "/- oxidation (6 h)
(46% opt pur)
a:>o (-)-lS, 2R-4 50% (37%opt pur)
Regiospecific enzyme-catalyzed oxidation of only the hydroxyethyl group was also observed for the saturated diol ( f ) - 2 . Again only a trace of the hemiacetal tautomer of the initial aldehyde product was detectable in the reaction mixture, and the products isolated (Scheme 111) were unchanged diol Scheme Ill
OH
1
Scheme II
(f)-2
3
2
than biochemical interest were selected for this initial study. '
Results The racemic diol 1 was subjected to HLADH-catalyzed oxidation at pH 9 using FMN recyclingh of catalytic amounts of the NAD+ coenzyme. The reaction was worked up when GLC analysis showed it to be half complete.' Oxidation was found to be totally regiospecific for the hydroxyethyl group, and the overall process was also enantioselective to a considerable degree. The results are summarized in Scheme I.8 The Scheme I HLADH.
(*)-l
m
"c. PH B
NAD+ recycling
*
OH
40% oxidation ( 4 h) 7Cnl
(+)-1R, 2s-4 4Wo
( 2 3 k o p t pur)
(4Woopt pur)
(-)-lS,2R-1 I c)yo
initially formed aldehyde did not accumulate owing to the facility with which its hemiacetal 5 underwent subsequent HLADH-mediated oxidation to the lactone 4. A small amount (4%) of 5 was detected (by GLC) in the reaction mixture immediately following termination. However, this material was removed by the subsequent isolation and purification procedures. The optical purities and absolute configurations of the Scheme I products were assigned using literature data.' The lactone 4 and its epoxide are important prostaglandin ~ynthons.~-'* Disappointingly, the 1R,2S configuration of the sample of 4 obtained from the Scheme I reaction is the opposite of those of the naturally occurring prostaglandins. However, this disadvantage was overcome using the hemiacetal 5 as a substrate. HLADH-catalyzed oxidation of ( f ) - 5 proceeded enantioselectively as shown in Scheme 11. Enantiomerically enriched 1 R,2S-4 was isolated directly from the enzymemediated reaction while the lactone possessing the natural 1S,2R configuration was readily obtained by silver oxide oxidation of the recovered hemiacetal IS,2R-5. No oxidation of (f)-5 occurred under the Scheme I1 conditions in the absence of enzyme. Journal of the American Chemical Society
/
99:5
/
HLADK NAD+ recycling 20 "c, pH 9
30% oxidation ( 4 h )
*=+a>, OH
(+)-lS,2s-6
(-)lR, 2R-2
90% (
