Vol. 26, Issue 7, 681-688, July 1998

Enantioselective, Mechanism-Based Inactivation of Guinea Pig Hepatic Cytochrome P450 by N-(-Methylbenzyl)-1-Aminobenzotriazole

Christopher J. Sinal, Maurice Hirst, Christopher D. Webb, and John R. Bend

Department of Pharmacology and Toxicology, University of Western Ontario

N-Aralkylated derivatives of 1-aminobenzotriazole are well-established, mechanism-based inhibitors of cytochrome P450 (CYP or P450). In this study, the kinetics of inactivation of CYP2B-dependent 7-pentoxyresorufin O-depentylation (PROD) and CYP1A-dependent 7-ethoxyresorufin O-deethylation (EROD) activities by enantiomers of N-(-methylbenzyl)-1-aminobenzotriazole (MB) were compared. The racemic mixture (±)-MB, as well as the enantiomers ()-MB and (+)-MB, produced a time-, concentration-, and NADPH-dependent loss of PROD and EROD activity in hepatic microsomes from phenobarbital-treated guinea pigs. The rates of PROD inactivation by ()-MB were significantly faster than for (+)-MB. Consistent with this, the derived maximal k_inact was also significantly greater for ()-MB than for (+)-MB (0.49 vs. 0.35 min¹). In contrast, the concentrations required for the half-maximal rate of inactivation (K_i) were equivalent for ()-MB and (+)-MB, whereas the degree of competitive inhibition of PROD activity was greater for (+)-MB. No significant differences were found among ()-MB, (+)-MB, and (±)-MB with respect to mechanism-based inactivation (k_inact = 0.18, 0.16, and 0.17 min¹, respectively) or competitive inhibition of EROD activity. No differences were found for the maximal extent of PROD or EROD inhibition or the loss of spectral P450 after an extended 30-min incubation with the inhibitors. We conclude that mechanism-based inactivation of guinea pig CYP2B, but not CYP1A, isozymes by MB occurs in a stereoselective manner, most likely as a result of a difference in the balance between metabolic activation and deactivation for the MB enantiomers.