Oxidation of tolualdehydes to toluic acids catalyzed by cytochrome P450-dependent aldehyde oxygenase in the mouse liver.

K Watanabe, T Matsunaga, I Yamamoto, H Yashimura

Index: Drug Metab. Dispos. 23(2) , 261-5, (1995)

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Abstract

Mouse hepatic microsomal enzymes catalyzed the oxidation of o-, m-, and p-tolualdehydes, intermediate metabolites of xylene, to the corresponding toluic acids. Cofactor requirement for the catalytic activity indicates that the microsomes contain NAD- and NADPH-dependent enzymes for this reaction. GC/MS analyses of the carboxylic acids formed by incubation under oxygen-18 gas indicate that the mechanism for this oxidation is an oxygenation and a dehydrogenation for the NADPH- and NAD-dependent reaction. Vmax/Km (nmol/min/mg protein) ratios indicate that the NADPH-dependent activity is more pronounced than the NAD-dependent activity. These results suggest that the NADPH-dependent reaction is mainly responsible for the microsomal oxidation of tolualdehydes. The NADPH-dependent activity was significantly inhibited by SKF 525-A, disulfiram and menadione, inhibitors of cytochrome P450 (P450), suggesting the involvement of P450 in the reaction. In a reconstituted system, P450 MUT-2 (CYP2C29) purified from mouse hepatic microsomes catalyzed the oxidation of o-, m-, and p-tolualdehydes to the carboxylic acids, and the specific activities (nmol/min/nmol P450) were 1.44, 2.81, and 2.32, respectively. Rabbit antibody raised against P450 MUT-2 significantly inhibited the NADPH-dependent oxidation of tolualdehydes to toluic acids by 88% (o-), 63% (m-), and 62% (p-) using mouse hepatic microsomes. The present study demonstrated that a mouse hepatic microsomal aldehyde oxygenase, P450 MUT-2, catalyzed the most of oxidative activity of tolualdehydes to toluic acids in the microsomes.