Enantioselective enzymatic catalysis.:1. A novel method to determine the enantiomeric ratio

Abstract

Abstract The enantioselective properties of a large number of enzymes are conveniently characterized by the enantiomeric ratio, E = ( k cat / K M ) s /( k cat / K M ) R . Chen and coworkers [ Chen et al., J. Am. Chem. Soc. 104 , 7294‐9 (1982)] have shown that the value of E can be obtained by measuring the enantiomeric excess, ee , as a function of the degree of conversion, ξ, in kinetic resolutions of a racemic substrate. We noticed that determination of E from ee = f( E , ξ) is not practical when: ( a ) low amounts of enzyme or substrate are available; ( b ) analytical procedures for the determination of ee values of substrates or products are cumbersome or inaccurate; ( c ) complications arise from maturation of reaction mixtures. Direct evaluation of E from the kinetic parameters, on the other hand, requires both enantiomers to be available in enantiomerically pure form. We show that the initial reaction rates for mixtures of enantiomers at fixed substrate concentration under otherwise identical conditions, are related by: ( v s , v R and v x represent the initial reaction rate of homochiral S ‐enantiomer, homochiral R ‐enantiomer and mixtures with (molar) fraction x of S ‐enantiomer, respectively). Analytical procedures for the determination of E , v s and v R by this approach are presented. The merits are demonstrated for the lipase‐catalysed kinetic resolution of racemic glycidol ester and for the resolution of racemic alcohols by a quinoprotein dehydrogenase and an NAD‐dependent dehydrogenase.