In order to explore the influences of iron overload on lipid peroxidation of rats and the inhibiting effects of antioxidant vitamins such as vitamin E and beta-carotene on the influences induced by iron overload, eighty male Wistar rats were randomly divided into 8 groups according to the weight. The diets of the former 4 groups differed only in iron concentration (50, 200, 350 or 500 mg/kg diet). The diet of the latter 4 groups were consistent with the former 4 groups respectively, but complemented with vitamin E(100 mg/kg diet) and beta-carotene (25 mg/kg.BW) at the same time. After Eight weeks, the levels of serum iron, blood lipid including total cholesterol TC, triglyceride TG, high density lipoprotein HDL, low density lipoprotein LDL, vitamin E, vitamin A, serum malondialdehyde (MDA) and oxidized low density lipoprotein (OX-LDL), the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were determined. Results showed that the iron overload increased the levels of MDA, OX-LDL and LDL in serum, especially in the group of rat with 500 mg/kg diet. The activity of GSH-Px increased irritably and the contents of Vitamin E and Vitamin A decreased significantly. Complementing antioxidant vitamins decreased the levels of MDA, OX-LDL and LDL and increased HDL and the activity of GSH-Px decreased compared with the corresponding iron overload groups. It was concluded that iron overload could promote lipid peroxidation reaction especially in 10 folds of physiologic dose and the activities of antioxidant enzymes increased at the same time to keep the body homeostasis. Antioxidant vitamins may play a role through non-enzymatic reactive system to inhibit the above changes.