Schiff Bases of Amino Acid Esters as New Substrates for the Enantioselective Enzymatic Hydrolysis and Accompanied Asymmetric Transformations in Aqueous Organic Solvents 1,2 Virinder S. Parmar,* ,† Amarjit Singh, † Kirpal S. Bisht, † Naresh Kumar, † Y. N. Belokon, ‡ K. A. Kochetkov, ‡ N. S. Ikonnikov, ‡ S. A. Orlova, ‡ V. I. Tararov, ‡ and T. F. Saveleva ‡ Department of Chemistry, University of Delhi, Delhi-110 007, India, and A.N. Nesmeyanov Institute of Organoelement Compounds, 28 Vavilov, 117813 Moscow, Russia Received October 17, 1994 (Revised Manuscript Received December 1, 1995 X ) The enzyme (lipases and chymotrypsin)-catalyzed hydrolysis of Schiff bases derived from racemic amino acid esters and aromatic aldehydes has been investigated. The reactions were successfully carried out in different aqueous organic solvents at ambient temperature, but the aqueous acetonitrile (5.4% water content by volume) was the solvent of choice. The L-amino acid (ee 98%) precipitated out from the solution as the reaction progressed, and the liberated aldehyde and unhydrolyzed D-ester (ee 40-98%) remained in the solution. The range of substrates included amino acids having different types of side chains. The addition of an organic base (DABCO) into the solution resulted in the racemization of the remaining D-ester and the additional hydrolysis of the substrate, thus leading to the effective asymmetric transformation of the initial ester. Upto 87.5% of the initial racemate was converted into the L-enantiomer. Introduction Enzymes have been widely used for the resolution of racemic organic compounds. 3,4 The use of acylases derived from different sources for the resolution of R-amino acids is well documented and broadly employed both in chemical laboratories and industry. 5 Recently it has been shown from our laboratories and by other groups of workers that some proteases and lipases are good catalysts for the enantioselective hydrolysis of amino acid derivatives. 6 Another traditional proteolytic enzyme, chymotrypsin, was used to resolve hydrophobic N-pro- tected amino acids esters and free amino acids esters, 7 but its application was limited in scope due partly to high instability of the free amino acids esters; after a week, more than 50% of the free amino acid ester was converted to diketopiperazine and other polycondensation prod- ucts. 8 We put ourselves a task of elaborating a new resolution procedure based on the use of chymotrypsin and lipases as catalysts and Schiff bases derived from aromatic aldehydes and racemic amino acids esters as substrates. The envisaged advantages of these substrates are the lability of the N-protective group, their increased solubil- ity in organic and aqueous-organic solvents, and high R-C-H bond acidity of the amino acid moiety. The latter properties might be employed to bring about the racem- ization of the unhydrolyzed enantiomer and thus induce asymmetric transformation 9 of the initial racemic Schiff base during the course of hydrolysis. This paper reports a successful application of this approach to the resolution of Phe, substituted Phe, Ala, nor-Val, and R-Me-Phe. It has been shown that the asymmetric transformation of Phe could be carried out, and L-Phe was obtained in 87.5% yield from the racemic Schiff base derived from p-chlorobenzaldehyde and D,L-Phe ester when the chy- motrypsin-catalyzed hydrolysis was carried out in the presence of DABCO in aqueous organic solvents. Results and Discussion The Schiff bases were prepared according to the literature procedures from the corresponding amino acid ester and the aldehyde. 10 The Schiff bases of 4-F-D,L- Phe, and 2-F-D,L-Phe were prepared by alkylation of the corresponding Schiff base with 4-fluoro- and 2-fluoroben- zyl bromide, respectively. The Schiff bases of D,L-Ala, D,L- nor-Val, D,L-Phe, 4-F-D,L-Phe, 2-F-D,L-Phe, and D,L-R-Me- Phe ethyl esters with benzaldehyde were also prepared by direct alkylation of the Schiff bases derived from the corresponding aldehyde and the glycine ester 11 (Scheme 1). The Schiff bases of D,L-Ph-Gly-OEt, D,L-nor-Val-OEt, D,L-Trp-OEt, D,L-4-F-Phe-OEt, and D,L-2-F-Phe-OEt have been prepared for the first time. The reaction protocol was a very simple one. Chymot- rypsin (10 -7 -10 -6 mol) or a lipase was added as an insoluble powder to the aqueous organic solvent (1:19, † University of Delhi. ‡ A. N. Nesmeyanov Institute of Organoelement Compounds. 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Chem. 1996, 61, 1223-1227 0022-3263/96/1961-1223$12.00/0 © 1996 American Chemical Society