DEVELOPMENT OF A HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHOD FOR THE ANALYSIS OF ENATIOMER ENANTIOMER INTERACTION IN OXIDATIVE-METABOLISM OF BUNITROLOL IN RAT-LIVER MICROSOMES/

A high-performance liquid chromatographic method for the assay of enan tiomeric 4-hydroxybunitrolol (4-OH-BTL) formed from racemic bunitrolol (BTL) in rat liver microsomes was developed. Racemic bunitrolol was i ncubated with rat fiver microsomes fortified with an NADPH-generating system. Metabolites extracted with ethyl acetate were converted to ace tyl derivatives with acetic anhydride in pyridine. The derivatives of 4-OH-BTL were well separated by the liquid chromatography equipped wit h a chiral column. Using this method, the metabolic interaction of BTL enantiomers was examined, The 4-OH-BTL-forming activities from enanti omeric BTL were higher than those from racemic BTL in rat liver micros omes, while the formation of ratios of 4-OH-BTL enantiomer to its anti pode were the same under the two conditions. The K-i values obtained f rom kinetic studies using each BTL enantiomer as an inhibitor of its a ntipode were almost the same (ca 0.9 mu M), which were close to their Michaelis constants (K-m values). Oxidative activities of enantiomeric and racemic BTL were almost equally inhibited by debrisoquine and qui nidine, a typical substrate and a selective inhibitor of the CYP2D sub family, respectively. These results indicate that a BTL enantiomer is a mutual metabolic inhibitor of its antipode and BTL enantiomers compe te for the same CYP2D isozyme in rat liver microsomes. (C) 1994 Academ ic Press, Inc.