N-nitrosodialkylamines do not function as substrates for liver monoamine oxidase.

Abstract

Since reports in the literature [Rowland, I. R., Lake, B. G. & Gangolli, S. D. (1980) Mutat. Res. 72,[63][64][65][66][67][68][69][70][71][72] have implicated hepatic monoamine oxidase (EC 1.4.3.4) in the activation of N-nitrosodialkylamines to mutagens, it is of basic pharmacological and biochemical interest to determine if these compounds could serve as substrates for this enzyme. The dialkylnitrosamines N-nitrosodiethylamine, Nnitrosodimethylamine, and N-nitrosodibenzylamine were tested and found not to be substrates, competitive inhibitors, or irreversible inhibitors of liver monoamine oxidase. Thus, any role for monoamine oxidase participation in the mutagenic activation of these compounds must be subsequent to an initial conversion of these compounds to their respective secondary amines.The carcinogenic effect of N-nitrosodialkylamines is known to be expressed by metabolic conversion of these amines to reactive intermediates. These reactive intermediates would then interact with DNA to form O-alkyl bases (1), and this alkylation leads to a series of events culminating in carcinogenesis. It is thus of basic interest to identify the enzyme system involved that converts N-nitroso compounds to their activated, carcinogenic forms. Once identified, suitable inhibitors of the activating enzyme system could be used to prevent expression of the carcinogenic activity of the N-nitrosamines.An extensive literature exists to show that N-nitrosodimethylamine (dimethylnitrosamine) can be demethylated by the cytochrome P-450 system to form formaldehyde (2); however, recent studies have shown that the action of mammalian cytochrome P-450 on dimethylnitrosamine does not result in the production of mutagenic species as monitored in the Ames test (3). These results suggested that the mutagenic activation of N-nitrosodialkylamines is effected by an enzyme system distinct from the cytochrome P-450 system.Recent studies in vivo and in vitro (4)(5)(6) Taken together, these studies suggest a role for MAO in the hepatic activation of N-nitrosodialkylamines to a mutagenic form. If MAO were the primary enzyme involved, then N-nitrosodialkylamines might be expected to be reasonable substrates for the purified enzyme. This study was undertaken to investigate this possibility. As shown in this paper, several N-nitrosodialkylamines are shown not to be substrates for MAO, thus demonstrating that the primary step in the metabolic activation of N-nitrosodialkylamines must be catalyzed by another enzyme. METHODS AND MATERIALSMAO was isolated from bovine liver mitochondria according to the method published by Salach (7). The specific activity of the preparation was in good agreement with that published. Rates of benzylamine oxidation were measured spectrophotometrically by following the increase in absorbance at 250 nm due to benzaldehyde formation as published by Tabor et al. (8). The rates of oxygen reduction were determined by using a Yellow Springs oxygen electrode and monitor in conjunction with a laboratory-built voltage suppr...