Enzyme specificity in the metabolic activation of N-nitrosodimethylamine to a mutagen for Chinese hamster V79 cells

Abstract

In order to elucidate the mechanism of the metabolic activation of N-nitrosodimethylamine (NDMA), the relationship between NDMA demethylase (NDMAd) and NDMA mutagenicity in Chinese hamster V79 cells was investigated. The microsome-mediated activation system produced NDMA mutagenicity similar to the S9-mediated activation system, suggesting that microsomes are solely responsible for the activation process. Pretreatment of rats with ethanol- or acetone-induced microsomal NDMAd activity, and such treatment also enhanced microsome-mediated NDMA mutagenicity 6-7-fold. The patterns of NDMA activation by ethanol- and acetone-induced microsomes differed distinctly from that by phenobarbital-induced microsomes for both NDMAd and the mutagenicity. The former type of microsomes had a low Km for NDMA, but the latter appeared to have very high Km values, and NDMAd was highly positively related to NDMA mutagenicity. Purified cytochrome P-450 isozymes from acetone- and phenobarbital-induced microsomes, P-450ac and P-450b, respectively, were effective for the activation of NDMA to a mutagen in a reconstituted monooxygenase system. In parallel fashion to NDMAd activity, P-450ac was effective at low substrate concentrations, whereas P-450b exhibited appreciable activity only at high NDMA concentrations. The results demonstrate clearly that NDMAd, which is effectively catalyzed by a specific P-450 isozyme inducible by compounds such as ethanol, acetone, and isopropanol, is primarily responsible for the activation of NDMA to a mutagen.