title: Mechanistic aspects of the inhibition of microsomal drug oxidation by primaquine.
abstract: The kinetics of inhibition of microsomal drug oxidation (as aminopyrine N-demethylase activity) by the antimalarial agent primaquine were found to be concentration-dependent. Lower concentrations of primaquine (0-40 microM) elicited slope-hyperbolic, intercept-hyperbolic noncompetitive (mixed) inhibition with an inhibitor equilibrium-dissociation constant (Ki) of 21 microM. On the other hand, primaquine concentrations greater than 40 microM elicited essentially simple competitive inhibition as judged from Lineweaver-Burk and Dixon analysis with appropriate replots (Ki = 23 microM). The coincident Ki values suggest that the same enzyme-inhibitor complex is involved in inhibition over all concentrations of primaquine tested. The apparent change in kinetics was accounted for in terms of a four-step interaction scheme involving a ternary enzyme-substrate-inhibitor complex that catalyses substrate oxidation at a slower rate than the binary enzyme-substrate complex. Competitive inhibition reflects the likelihood that the ternary complex does not form at all, presumably due to reduced accessibility of the active site to substrate. A good correlation was found between the Ki values for the inhibition of aminopyrine N-demethylase activity (21 or 23 microM) and the modulation of aminopyrine binding (26 microM) by primaquine. These findings suggest that the inhibition of aminopyrine N-demethylase activity by primaquine is mediated via an interaction with the oxidised form of cytochrome P-450. In addition, the apparent change in inhibition kinetics suggests a concentration-dependent change in the capacity of primaquine to modulate substrate binding to cytochrome P-450 as well as the formation of a P-450-aminopyrine-primaquine ternary complex.
mesh_terms: D000633:Aminopyrine N-Demethylase / Q000037:antagonists & inhibitors*; D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D007700:Kinetics; D008297:Male; D008433:Mathematics; D008862:Microsomes, Liver / Q000201:enzymology*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011319:Primaquine / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
chemical_list: D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D000633:Aminopyrine N-Demethylase; D011319:Primaquine