There is no consensus yet on the role of oxidative stress in the nutritional outcome of chronic ethanol feeding and the status of cellular antioxidative defense systems against ethanol toxicity. In this study, chronic alcohol consumption in humans was reproduced in Sprague-Dawley rats to investigate the effect of ethanol ingestion on the regulation of oxidative stress in liver with a special focus on glutathione. Adult male rats were given 36% of total energy as alcohol in the Lieber-DeCarli liquid diet for 6 wk. The control group was pair-fed the diet containing isocaloric dextrin-maltose instead of ethanol. Chronic ethanol ingestion enhanced expression of cytochrome P450 II E1 in the liver, but did not significantly alter either the level of hepatic thiobarbituric acid reactive substances or the carbonyl group content of proteins. The hepatic concentrations of total and reduced glutathione and the activities of catalase, glutathione reductase and glutathione S-transferase were significantly higher in the ethanol group than in the control group. The activities of glutathione peroxidase and glucose-6-phosphate dehydrogenase were significantly lower in the ethanol group than in controls. Chronic ethanol consumption by well-nourished rats for 6 wk increased enzyme activities related to the recycling and utilization of glutathione in the liver. Such an enhancement in the activities of the hepatic antioxidative defense system may be one of the protective mechanisms of the body against oxidative tissue damage caused by ethanol-induced free radicals.