In vitro interactions of sulfur-containing compounds with the hepatic mixed-function oxidase system in mice: Effects on paracetamol activation and covalent binding

Abstract

The effects of five thiols ( N-acetylcysteine, cysteamine, cysteine, glutathione, 2-mercaptopropionylglycine) and methionine on various metabolic stages leading to the covalent binding of paracetamol to hepatic microsomes were examined using mouse liver preparations. All five thiols decreased paracetamol covalent binding at concentrations 10- to 100-fold smaller than those needed to affect significantly the apparent dissociation equilibrium of substrates with oxidized cytochrome P-450, the flavoprotein-mediated reduction of the cytochrome or the oxygenation of cytochrome P-450 substrates. Thus, N-acetylcysteine, cysteamine, cysteine, glutathione, and 2-mercaptopropionylglycine decreased the covalent binding of [ 3H]paracetamol-derived radioactivity by 50% at concentrations of 0.06, 0.01, 0.03, 0.13, and 0.11 m m, respectively. Only cysteamine and cysteine (5 m m) affected the spectral interactions of paracetamol (>1 m m) with cytochrome P-450. Cysteamine also markedly decreased the affinity of cytochrome P-450 for ethylmorphine and aniline. N-Acetylcysteine, cysteamine, glutathione, and 2-mercaptopropionylglycine (1 m m) decreased NADPH-cytochrome c reductase activity by <20%. This effect was augmented in the presence of paracetamol. N-Acetylcysteine, cysteamine, cysteine, glutathione, and 2-mercaptopropionylglycine inhibited the oxygenation of at least one of the mixed-function oxidase substrates acetanilide, benzo[α]pyrene, biphenyl, and ethylmorphine up to 80%. Methionine was ineffective in all the assays examined and the mechanism of its known effects in vivo must involve metabolites such as homocysteine, cysteine, and glutathione which are not produced by an in vitro microsomal system. These data suggest that the effects of sulfur-containing compounds on the formation of a reactive paracetamol metabolite are of lesser importance than effects on its subsequent covalent binding, although they may influence the overall efficacy achieved.