The nature of nitrosamine denitrosation by rat liver microsomes

The nature of the denitrosation of nitrosamines by rat liver microsomes was investigated. The rates of NADPH-dependent nitrosamine demethylation and denitrosation were compared in the same incubation mixture using several types of microsomes and inhibitors. Pretreatment with isopropanol, pyrazole, phenobarbital, and 3-methylcholanthrene had parallel effects on the microsomal demethylation and denitrosation reactions. Nitrite was produced with N-nitrosodimethylamine, N-nitroso-N-methylethylamine, N-nitroso-N-methylbutylamine, N-nitroso-methylbenzylamine, or N-nitroso-N-methylaniline as a substrate. With control microsomes, the rate of the denitrosation reaction was 9-39% that of demethylation depending on the type and concentration of nitrosamines used. Using nitrosodimethylamine as the substrate, the Km of denitrosation was about twice that of the demethylation reaction. Several polar organic solvents such as ethanol and isopropanol inhibited the denitrosation and demethylation reactions and each solvent inhibited both reactions to about the same extent. In the presence of cumene hydroperoxide, microsomes can catalyze the denitrosation of nitrosodimethylaime which is also accompanied by demethylation. Studies with a reconstituted system and with inhibitors indicate that the denitrosation reaction requires the presence of both cytochrome P-450 and NADPH-cytochrome-P-450 reductase. The results suggest that the denitrosation is closely linked to the demethylation reaction.