Effect of glutathione levels, sulfate levels, and metabolic inhibitors on covalent binding of 2-amino-3-methylimidazo[4,5-f]quinoline and 2-acetylaminofluorene to cell macromolecules in primary monolayer cultures of adult rat hepatocytes

Abstract

The covalent binding of radiolabeled 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-acetylaminofluorene (AAF) to cell macromolecules was studied using primary monolayer cultures of adult rat hepatocytes. A time course for covalent binding was determined, and revealed similar levels of binding for both chemicals. Inhibition of glutathione synthesis by L-buthionine sulfoximine (1 mM) enhanced binding of both AAF and IQ with a greater increase observed for IQ. Addition of excess glutathione (10 mM) to the medium resulted in a slight decrease in IQ but not AAF binding. Addition of the P-450 inhibitor, 2-[2,4-dichloro-6-phenyl)-phenoxy]ethylamine (DPEA, 0.1 mM), resulted in almost total (94%) inhibition of IQ binding, with a lesser effect (42%) on AAF. Methimazole (1 mM), a competitive substrate of the flavin-containing monooxygenase, had no effect on the binding of either compound. Pentachlorophenol, an inhibitor of sulfate conjugation, decreased AAF binding substantially but produced a much smaller decrease in IQ binding. Addition of excess sulfate did not change the binding levels of either IQ or AAF. Cell density had little effect on IQ or AAF binding levels.