Elsevier

Biochemical Pharmacology

Volume 25, Issue 19, 1 October 1976, Pages 2153-2162
Biochemical Pharmacology

Warfarin—stereochemical aspects of its metabolism by rat liver microsomes

https://doi.org/10.1016/0006-2952(76)90127-1Get rights and content

Abstract

The R- and S-enantiomers of warfarin were differentially metabolized by hepatic microsomes prepared from male Wistar and Sprague-Dawley rats. These studies were not only carried out with two strains of rats but were conducted independently in two laboratories employing different techniques. Although minor differences were observed, the same stereoselectivity was found for the microsomal transformations produced by both strains. The formation of 7- and 8-hydroxywarfarin was stereoselective for the R-enantiomer and in addition this enantiomer was metabolized more rapidly than the S-enantiomer. The converse stereoselectivity was found for the process of 4′-hydroxylation in Sprague-Dawley rats but it could not be conclusively shown for Wistar rats. A Michaelis-Menten analysis of the metabolic products of R- and S-warfarin formed by liver microsomes from male Sprague-Dawley rats is reported. The Km for the processes of 6-, 7-, and 4′-hydroxylation for both isomers and the Km for 8-hydroxylation of the R-isomer were all of the order of 0.03 to 0.11 mM and were not statistically different. The Km for 8-hydroxylation of the S-isomer, 0.20 mM, was significantly greater. The Km for benzylic hydroxylation of both isomers appeared to be still greater but was less precisely determined. The Vmax for each of the enantiomeric pairs of products was statistically different. The kinetic data are interpreted as being inconsistent with the supposition that an arene oxide (6–7 and/or 7–8) may serve as the intermediate in the formation of 7-hydroxywarfarin from either isomer. Further, if product formation is assumed to be rate limiting, the data provide evidence for at least three distinct enzymatic processes which may or may not be distinct hemoproteins. Reduction of the side-chain ketonic function of warfarin to the corresponding diastereomeric warfarin alcohols by the 105,000 g supernatant fraction displayed both a high degree of stereoselectivity (R-isomer) and stereospecificity (S-reduction). This reduction was best catalyzed by NADPH rather than by NADH. Determination and quantification of the metabolic products obtained after incubation of R- and S-warfarin with the 10,000 g supernatant were consistent with the summation of those independently produced by the microsomal pellet and the 105,000 g supernatant.

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    Present address: Laboratory of Chemical Pharmacology, National Heart and Lung Institute, National Institutes of Health, Bethesda, MD 20014.

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