The effects of ubiquinol and vitamin E on ascorbate- and ADP-Fe3+-induced lipid peroxidation were investigated by measuring oxygen consumption and malondialdehyde formation in beef heart submitochondrial particles. In the native particles, lipid peroxidation showed an initial lag phase, which was prolonged by increasing concentrations of ascorbate. Lipid peroxidation in these particles was almost completely inhibited by conditions leading to a reduction of endogenous ubiquinone, such as the addition of succinate or NADH in the presence of antimycin. Lyophilization of the particles followed by three or four consecutive extractions with pentane resulted in a complete removal of vitamin E and a virtually complete removal of ubiquinone, as revealed by reversed-phase high pressure liquid chromatography. In these particles, lipid peroxidation showed no significant lag phase and was not inhibited by either increasing concentrations of ascorbate or conditions leading to ubiquinone reduction. Treatment of the particles with a pentane solution of vitamin E (alpha-tocopherol) restored the lag phase and its prolongation by increasing ascorbate concentrations. Treatment of the extracted particles with pentane containing ubiquinone-10 resulted in a restoration of the inhibition of lipid peroxidation by succinate or NADH in the presence of antimycin, but not the initial lag phase or its prolongation by increasing concentrations of ascorbate. Malonate and rotenone, which prevent the reduction of ubiquinone by succinate and NADH, respectively, abolished, as expected, the inhibition of the initiation of lipid peroxidation in both native and ubiquinone-10-supplemented particles. Re-incorporation of both vitamin E and ubiquinone-10 restored both effects. In the vitamin E-supplemented particles, the duration of the lag phase and its prolongation by ascorbate were dependent on the amount of vitamin E incorporated and were decreased upon pre-incubation of the particles at 37-degrees-C in an aqueous buffer medium. In the ubiquinone-10-containing particles, lipid peroxidation was inhibited by succinate or NADH and antimycin even when these were added after it had reached maximal activity following initiation. Ascorbate, in a concentration that prolonged the lag phase in either the native or the vitamin E-supplemented particles, had no inhibitory effect when added after lipid peroxidation had reached maximal activity. It is concluded that ubiquinol is capable of inhibiting lipid peroxidation in beef heart submitochondrial particles directly, without the mediation of vitamin E. It is pointed out that ubiquinone is the only known lipid-soluble antioxidant that can be synthesized de novo in animal cells, and for which there exists an enzymic mechanism-the mitochondrial electron transport system-that can efficiently regenerate the active antioxidant from its oxidized form, the ubisemiquinone, resulting from its inhibitory effect on membrane lipid peroxidation. These features, together with the general occurrence of ubiquinone in biological membranes and in conjugation with low-density lipoprotein, suggest a highly important role of ubiquinone in cellular defense against oxidative damage.
