Rats were fed semipurified diets containing 10% fat with a constant polyunsaturated/monounsaturated/saturated fatty acid ratio of 1:1:1. This was performed by mixing vegetable oils. Linoleic acid was the sole polyunsaturated fatty acid in the linoleic acid group. The a-linolenic and arachidonic acids were contained at the 1 % level in the diet at the expense of linoleic acid in the α-linolenic acid and arachidonic acid groups, respectively. The concentration of serum triglyceride, phospholipid and liver triglyceride were significantly lower in the α-linolenic acid and arachidonic acid groups than in the linoleic acid group. Hepatic phospholipids were significantly higher in the α-linolenic acid and arachidonic acid groups as compared to the linoleic acid group. Compared to the linoleic acid group, the percentages of arachidonic acid in the phosphatidylcholine (PC) and phosphatidylethanolamine (PE) in the liver and heart were significantly lower in the α-linolenic acid group; in contrast, they were significantly higher in the arachidonic acid group. Dietary arachidonic acid, but not α-linolenic acid, resulted in a marked reduction of linoleic acid and increased arachidonic and n-6 docosapenta-enoic acids in heart cardiolipin. Platelet thromboxane (TX) A₂ production tended to be lower in the α-linolenic acid group as compared to the linoleic acid and arachidonic acid groups. There was no difference in TXA2 production between the linoleic acid and arachidonic acid groups. The aortic production of prostacyclin (PGI₂) was the same among the three groups. The ratios of TXA₂ and PGI₂ in the linoleic acid and arachidonic acid groups were comparable, whereas they were significantly higher than that in the α-linolenic acid group. The results demonstrate that dietary arachidonic acid enriches the arachidonic acid content in tissue phospholipids more effectively than linoleic acid, whereas the effect on the production of TXA₂ and PGI₂ was comparable under these experimental conditions.