A 56-days feeding trial was conducted with tiger puffer Takifugu rubripes (average initial body of 14 g), in order to investigate the effects of dietary bile acid supplementation on the fatty acid composition and anti-oxidative capacity. The feeding trial was conducted in indoor flow-through seawater system. Two diets with suitable (8.5%, diet C) or high lipid level (12.5%, diet HL) were prepared. BA as porcine bile extract mainly composed of hyodeoxycholic acid was supplemented into the two diets at the level of 0.02% to formulate diet BA and diet HLBA, respectively. A fifth diet was prepared by supplementing an excess level of porcine bile extract (0.10%) into diet C (diet HBA). Each diet was randomly fed to triplicate tanks. The results showed that the MUFA contents in liver and muscle were reduced by dietary bile acid supplementation, significantly lower in group HBA than in the control group (P<0.05). The n-6 PUFA contents in groups HLBA and HL were significantly lower compared to other groups. In the groups with moderate lipid level, the contents of C20:0, C16:1n-7, C18:1n-9, C18:2n-6, C20:4n-6, C18:3n-3, C18:4n-3, C20:5n-3 and C22:6n-3 in the liver tended to decrease with increasing bile acid levels. The serum activities of glutathione peroxidase and superoxide dismutase tended to increase with increasing bile acid levels, and were significantly higher in high-lipid groups than in the control group (P<0.05). However, the glutathione reductase activity showed an opposite pattern. The relative mRNA expression in liver of the control group was significantly higher than that in other groups, while the gene expression of peroxiredoxin 1 in group BA was significantly higher than that in the control. The bile acid supplementation in high-lipid diets significantly increased the hepatic gene expression of catalase. In conclusion, the dietary bile acid supplementation reduced the contents of a series of long-chain fatty acids in both diet and fish tissues,and the effects of dietary bile acid on serum activity of anti-oxidative proteins were different from those on the hepatic expression of anti-oxidative genes.