Cytochrome c oxidase reduces oxygen, the natural substrate, to water upon receiving four electrons from cytochrome c. Hydrogen peroxide is a two-electron reduced form of oxygen so that it can also be reduced to water by a two-electron reduction process. The rate of reduction of oxygen by cytochrome c oxidase strongly depended on pH whereas that of hydrogen peroxide did not. In the presence of excess ascorbate and TMPD which constantly supply electrons to cytochrome c, the reduction level of cytochrome a was much higher for the reduction of oxygen than for hydrogen peroxide. In addition a ferryl oxo species was observed in the reduction of hydrogen peroxide. This suggests that electron transfer from the low potential site to the binuclear site prior to the binding of oxygen is rate limiting in the reduction of oxygen and the thus formed peroxy species is protonated. On the other hand, binding of hydrogen peroxide to the oxidized binuclear site is rate limiting in the reduction of hydrogen peroxide and the conversion of the peroxy species to the ferryl oxo species is extremely fast. Plausible mechanisms for the reaction of cytochrome c oxidase with oxygen and with hydrogen peroxide are proposed to account for these observations.