Effects of low corn oil, high corn oil, and high fish oil diets on altered hepatic foci development in female Sprague-Dawley rats were investigated. Rats assigned to Groups 1-4 were initiated with saline as the control and those assigned to Groups 5-7 were initiated with diethylnitrosamine (DEN 15 mg/kg) at 24 hours of age. After weaning, all rats, except those in Group 1, received 500 ppm phenobarbital (PB) in their diet as tumor promoter for three months. Altered hepatic foci development was significantly lower in DEN-initiated rats fed the high fish oil + PB diet than in DEN-initiated rats fed the high corn oil + PB diets. Liver weight and relative liver weight were significantly greater in rats fed the high fish oil + PB diet than in rats fed the other diets, and hepatic biotransformation/detoxification enzyme activities were greater in rats fed the fish oil + PB diets than in rats fed the other diets. These results suggest that the effect of a high fish oil diet on altered hepatic foci may occur through regulation of hepatic biotransformation/detoxification enzyme activities, leading to alteration in the tumor-promoting action of PB. Dietary lipid significantly affected the hepatic phospholipid fatty acid composition of rats. n-3 polyunsaturated fatty acids were incorporated into membrane phospholipid at the expense of n-6 polyunsaturated fatty acids. A high fish oil diet caused greater oxidative stress in rats, as measured by plasma vitamin E level, red blood cell glutathione status, liver lipid peroxidation, and hepatic glutathione reductase activity. Pearson's correlation analysis indicated that the foci number was negatively correlated to the liver thiobarbituric acid-reactive substance and 7-pentoxyresorufin O-dealkylase activity, and the foci area was negatively correlated to the liver thiobarbituric acid-reactive substance activity (p < 0.05) in rats of groups that developed foci. These results suggest that the type of dietary lipid is the more important determinant for gamma-glutamyl transpeptidase-positive foci development than the amount of dietary lipid when rats consumed approximately the same amount of calories in all the dietary groups, and the underlying mechanisms may be partially ascribed to the antioxidant/oxidation status and biotransformation/detoxification system of rats.