alpha-Hydroxylation of nitrogen-15 labelled N-nitrosamines by isolated hepatocytes

The principal pathway of nitrosamine metabolism has long been considered to be alpha-hydroxylation. For N-nitrosodialkylamines, this hypothesis requires that a molecule of molecular nitrogen be released for every molecule of nitrosamine that is alpha-hydroxylated. Thus, the quantitative determination of nitrogen formation should provide a measure of the importance of this pathway. This method was applied earlier to the doubly-labelled nitrogen-15 compounds, N-nitrosodimethylamine (NDMA), N-nitrosomethylphenylamine (NMPhA) and N-methyl-N-nitrosourea (MNU), using both a 9 000 X g supernatant fraction of liver and the intact animal as metabolic systems. The in-vitro results were quite different from those obtained in vivo. The majority of the NDMA (67%) and the MNU (88%) were converted to nitrogen in vivo, while NMPhA gave considerably less nitrogen (52%). These results differed by a factor of approximately two from those obtained in vitro (NDMA, 33%; NMPhA, 18.8% and MNU, 96%). Since such differences may be a result of the loss of cellular architecture, we have extended the work to include isolated hepatocytes. It had been shown previously that isolated hepatocytes constitute a practical alternative to in-vivo systems, even though the correlation with in-vivo metabolism appears to depend on the substrate analysed. The values obtained using this system (NDMA, 47%; NMPhA, 23%; and MNU 105%) reconfirm that metabolism may be substrate dependent. As in our previous studies, no mixed nitrogen (15N14N) or labelled nitrogen oxides were found. The data are all consistent with the hypothesis that at least one demethylase for each of the nitrosamine substrates is associated with a cell membrane.(ABSTRACT TRUNCATED AT 250 WORDS)