In our previous studies, medium- and long-chain triacylglycerols (MLCT), randomly interesterified triacylglycerols containing medium-chain and long-chain fatty acids in the same glycerol molecule, significantly reduced body fat accumulation in humans and rats. To clarify mechanism(s) for this effect of MLCT, we measured energy expenditure and hepatic fatty acid metabolism in rats by comparison with long-chain triacylglycerols (LCT) or medium-chain triacylglycerols (MCT). MLCT, compared with LCT, showed significantly lower body fat accumulation, higher 24-h energy expenditure and acyl-CoA dehydrogenase activity measured using octanoyl-CoA as a substrate, and similar lipogenic activity. MCT, compared with LCT, showed significantly higher energy expenditure, but fat accumulation was comparable. Additionally, MCT exhibited significantly higher lipogenic activity than the other oils. These data suggest that enhancement of energy expenditure and medium-chain fatty acids (MCFA) oxidation without activating de novo lipogenesis are responsible at least for the lower body fat accumulation in rats fed MLCT. The activation of hepatic lipogenesis by excessive intake of MCFA might counteract their preventive effects on body fat accumulation.