Mutagenic activation of 2,4-diaminoanisole and 2-aminofluorene by isolated rat liver nuclei and microsomes

Abstract

The carcinogenic hair-dye component 2,4-diaminoanisole (2,4-DAA) is converted to a mutagen when incubated with isolated liver nuclei from β-naphthoflavone(BNF)-induced rats in the presence of NADPH. No significant mutagenic activation of 2,4-DAA could be seen with nuclei from untreated or phenobarbital(PB)-pretreated animals, indicating the involvement of cytochrome P-448 associated mono-oxygenases in the activation process. On a protein basis, rat liver microsomes were markedly more active than rat liver nuclei in producing mutagenic 2,4-DAA intermediates. BNF-pretreatment increased 2,4-DAA mutagenicity with microsomes far more than PB-pretreatment. Metabolic activation of the model hepatocarcinogen 2-aminofluorene (AF) showed differences from 2,4-DAA in response to inducer treatment. When incubated with isolated nuclei, AF was much more mutagenic than 2,4-DAA, especially with control nuclei, while PB- and BNF-pretreatment only resulted in a slight increase in mutagenicity. With microsomes, however, this pattern was changed; control preparations showed highest mutagenic activity with AF, whereas BNF-treatment led to reduced mutagenicity compared to the controls. This difference in mutagenic activity between control and BNF-preparations increased with increasing microsomal concentrations, suggesting either variations in the involvement of several inducible metabolic pathways for AF leading to the formation of both mutagenic and non-mutagenic intermediates, or a relative increase in the concentration of nucleophiles serving as traps for mutagenic metabolites. Cobaltous chloride, an inhibitor of cytochrome P-450 enzymes, reduced mutagenic activation of both arylamines. α-Naphthoflavone (ANF), a selective inhibitor of cytochrome P-448 mediated reactions, had a much more pronounced effect on 2,4-DAA mutagenicity than on AF mutagenicity with nuclei from BNF-pretreated rats. It could also be shown that 2,4-DAA was activated to irreversibly protein-bound products by nuclear and microsomal fractions. The present findings indicate the involvement of separate mono-oxygenases in the mutagenic activation of 2,4-DAA and AF.