This study was designed to investigate the antioxidative activity of a substance isolated and purified from culture broth of Bacillus sp. The antioxidative substance was identified as 2,6-bis(1,1-dimethylethyl)phenol (Phen-251). The antioxidative activity of Phen-251 was higher than that of dl-tocopherol and BHA against low-density lipoprotein (LDL) oxidation by thiobarbituric acid reactive substance (TBARS). Phen-251 inhibited the copper-mediated oxidation of human LDL in a dose-dependent manner with almost complete inhibition at 40 µg/ml. Phen-251 at a concentration of 60 µg/ml also inhibited oxidation of LDL induced by J774. LDL oxidized by copper-mediated or cell-induced oxidation was degraded at a much greater rate than native LDL. These results suggested the importance of further research on Phen-251 in the investigation of atherosclerosis and free radical-induced injury.