Kinetic studies of copper-induced oxidation of urate, ascorbate and their mixtures

Urate and ascorbate are the major water-soluble low molecular weight antioxidants in serum. Much attention has been devoted to the effect of these antioxidants on lipoprotein peroxidation in vivo and on their effect on copper-induced peroxidation ex vivo. These studies revealed that urate inhibits ascorbate oxidation in vitro, whereas the effect of ascorbate on urate oxidation has not been systematically studied thus far. The present study addresses mechanistic aspects of the kinetics of copper-induced oxidation of both these antioxidants and their mutual effects in aqueous solutions. We found that: (i) ascorbate becomes oxidized much faster than urate. (ii) Urate inhibits the oxidation of ascorbate but, even in the presence of excess urate, ascorbate becomes oxidized much faster than urate. (iii) Ascorbate, as well as the products of its oxidation (and/or hydrolysis) inhibit the copper-induced oxidation of urate. All these results are consistent with the hypothesis that the rate of ascorbate oxidation is determined by the rate of reoxidation of reduced copper (Cu(I)) to Cu(II) by molecular oxygen, whereas the rate of urate oxidation is governed by the rate of oxidation of urate within a 2:1 urate/copper complex. We think that the mutual effects of urate and ascorbate on each other's oxidation are likely to enhance their inhibitory effect on lipid peroxidation in biologically relevant systems including membranes and lipoproteins.