On the Molecular Mechanism of Metmyoglobin-Catalyzed Reduction of Hydrogen Peroxide by Ascorbate

Hydrogen peroxide induces rapid oxidation of metmyoglobin with an apparent second order rate constant, k1 = 3.4 × 104 M−1 min−1. The product of this interaction is ferrylmyoglobin with an unstable free radical on the globin moiety. This activated form of myoglobin is able: (a) to initiate the peroxidation of erythrocyte membranes and (b) to form intra- and intermolecular covalent crosslinkings. The presence of ascorbic acid in amounts stoichiometric to H2O2 efficiently prevents all the above processes. Moreover, in the presence of ascorbic acid a cyclic process is taking place leading to H2O2 reduction, ascorbic acid oxidation, and unmodified metmyoglobin formation (reaction 1).image The whole process follows pseudofirst-order kinetics with the initial rate of ascorbic acid oxidation dependent only on the concentration of H2O2 and independent of ascorbic acid concentrations as low as the limits of detection. In order to explain the observed kinetics, it is proposed that ascorbic acid is not directly involved in the reduction of ferryl- to metmyoglobin, but it interacts, first with an intermediate in metmyoglobin/H2O2 oxidation process, forming ferrylmyoglobin without the associated globin-centered free radical, and second, with a minor form of myoglobin, with which ferrylmyoglobin is in equilibrium, forming, again, metmyoglobin. The rate-limiting step for the whole process seems to be the conversion of ferrylmyoglobin to this proposed minor form. These findings lead to the view that metmyoglobin, in the presence of ascorbate, may act like a pseudoperoxidase in conditions of oxidative stress in muscle, substituting for the low levels of catalase in this tissue. Copyright © 1997 Elsevier Science Inc.