alpha-Tocopherolquinone (TQ), a product of alpha-tocopherol oxidation, can function as an antioxidant after reduction to alpha-tocopherolhydroquinone (TQH2). We examined the ability of human NAD(P)H:quinone oxidoreductase (NQO1) to catalyze the reduction of TQ to TQH2 in cell-free and cellular systems. In reactions with purified human NQO1, TQ was reduced to TQH2. Kinetic parameters for the reduction of TQ by NQO1 (Km = 370 microM; k(cat) = 5.6 x 10(3) min(-1); k(cat)/Km = 15 min(-1) x microM(-1)) indicate that NQO1 can efficiently reduce TQ to TQH2. A comparison of the rate of reduction of TQ and coenzyme Q10 by NQO1 showed that TQ is reduced more efficiently than coenzyme Q10. Experiments with either Chinese hamster ovary (CHO) cells stably transfected with human NQO1 or CHO cell sonicates demonstrated a correlation between NQO1 activity and TQ reduction to TQH2. CHO cells with elevated NQO1 generated and maintained higher levels of TQH2 after treatment with TQ relative to NQO1-deficient CHO cells. TQH2 generated from NQO1-mediated reduction of TQ prevented cumene hydroperoxide-induced lipid peroxidation in rat liver microsomes. In addition, cumene hydroperoxide-induced lipid peroxidation was inhibited more efficiently by TQ in CHO cell lines with elevated NQO1 activity. These data demonstrate that NQO1 can reduce TQ to TQH2 and that TQH2 can function as an efficient antioxidant. This work suggests that one of the physiological functions of NQO1 may be to regenerate antioxidant forms of alpha-tocopherol.