The current study examined the effects of Artemisia capillaris ethyl acetate (ACE) fraction on oxidative stress and antioxidant enzyme in high-fat diet induced obese mice. C57BL/6J mice were randomly assigned to normal diet group and high-fat diet groups. Normal diet groups were fed with an ACE fraction-free diet or 0.1 g ACE fraction/kg body weight. The high-fat diet groups were fed with an ACE fraction-free diet, 0.05 g ACE fraction, 0.1 g ACE fraction or 0.03 g rosiglitazone/kg body weight. ACE fraction strengthened the antioxidative defense system with an increased activity of superoxide dismutase (SOD) and decreased activities of glutathione-peroxidase (GSHpx) and catalase (CAT) and content of reduced glutathione (GSH). Accordingly, ACE fraction was found to reduce the accumulation of reactive oxygen species (ROS), such as superoxide radical and hydrogen peroxide which decrease the generation of oxidative stress damage substrates, such as the thiobarbituric acid reactive substances (TBARS) and carbonyl value. ACE fraction was found to be excellent for strengthening the antioxidant defense system, reducing the generation of ROS and damaging oxidative substances in the liver of high-fat diet induced obese mice.