The aim of this study was to investigate the effects of genistein supplementation in a vitamin E-deficient diet on the genistein concentrations and the lipid oxidation of serum, liver and low-density lipoprotein (LDL) of hamsters. Thirty-six male hamsters were randomly divided into three groups and fed a vitamin E-deficient semisynthetic diet (AIN-76) containing different levels of genistein, i.e., G0 (control group, genistein-free diet), G50 (50 mg genistein/kg diet) and G200 (200 mg genistein/kg diet) for 5 weeks. The concentrations of genistein in serum and liver significantly increased with the increase of genistein supplementation. The vitamin E contents in LDL were higher in hamsters fed G50 or G200 diets than in hamsters fed genistein-free diet. Genistein supplementation to hamsters significantly reduced the propagation rate during conjugated diene formation of LDL oxidation, and the lag time of LDL oxidation in hamsters fed G200 diets was significantly lower than that of G0 diets. In addition, genistein supplementation significantly raised serum total antioxidant capacity and decreased the thiobarbituric acid-reactive substances (TBARS) of LDL and liver in hamsters. However, no significant differences in TBARS were found in serum, irrespective of genistein addition. On the other hand, the relative contents of polyunsaturated fatty acids in LDL were decreased after genistein supplementation. There was a negative correlation between lag time and P/S ratio, and a positive correlation between lag time and vitamin E contents. These data demonstrate that genistein supplementation markedly increased its concentrations in body tissues and reduced oxidative stress of lipid oxidation of serum, liver and LDL.