Rates of glucuronidation and sulfation of 7-hydroxycoumarin were studied in perfused livers from normal chow-fed rats, or in livers from rats that had been fed liquid control or ethanol-containing diets. During infusion of 100 microM 7-hydroxycoumarin, rates of glucuronidation were similar in livers from chow-fed or control diet rats, but were 34% less in livers from ethanol-fed rats. These rates of glucuronidation in perfused livers could not be explained by changes of UDP-glucuronyltransferase activity, which was highest in hepatic microsomes from ethanol-treated rats and lowest in microsomes from chow-fed rats. The low rates of glucuronidation in livers from ethanol-treated rats were correlated with low hepatic concentrations of UDP-glucuronic acid, which were less than 70% of the levels measured in the other treatment groups. However, the diminished UDP-glucuronic acid levels could not be explained by alterations in adenine nucleotides, NAD+/NADH ratios, glycogen, UDP-glucose, or activity of UDP-glucose dehydrogenase. Rates of sulfation declined during prolonged 7-hydroxycoumarin infusion in livers from ethanol-treated rats, but not in livers from rats that had received the control diet. Similarly, hepatic concentrations of adenosine-3'-phosphate 5'-sulfatophosphate (PAPS) also decreased with time only in livers from ethanol-treated rats. Thus, chronic ethanol feeding impairs glucuronidation and sulfation in perfused livers as a result of diminished availability of the required cofactors for these conjugation pathways.