Formation of N-(glutathion-S-methylene)-4-aminoazobenzene following metabolic oxidation of the N-methyl group of the carcinogen, N-methyl-4-aminoazobenzene.

A major biliary metabolite of the hepatocarcinogen, N,N-dimethyl-4-aminoazobenzene (DAB), in the rat was identified as N-(glutathion-S-methylene)-4-aminoazobenzene (GS-CH2-AB). This conjugate was prepared synthetically by a Mannich condensation of 4-aminoazobenzene (AB), formaldehyde (CH2O) and glutathione (GSH) and has been characterized by chemical analysis and by ultraviolet, visible and 13C-NMR spectroscopy. The same conjugate was also formed in vitro by incubating N-methyl-4-aminoazobenzene (MAB), NADPH, NADH and GSH with rat hepatic microsomes. Evidence is presented that GSH reacted with an intermediate resulting from a cytochrome P-450-dependent oxidation of the N-methyl substituent. This reactive intermediate is presumed to be either an N-methylol or a methimine derivative of AB. The significance of this detoxification mechanism is discussed. The presence of an additional major aminoazo-dye GSH conjugate is also noted.