ONE-ELECTRON REDUCTION OF VANADATE BY ASCORBATE AND RELATED FREE-RADICAL GENERATION AT PHYSIOLOGICAL PH

The one-electron reduction of vanadate (vanadium(V)) by ascorbate and related free radical generation at physiological pH was investigated b y ESR and ESR spin trapping. The spin trap used was 5,5-dimethyl-1-pyr roline N-oxide (DMPO). Incubation of vanadium(V) with ascorbate genera ted significant amounts of vanadium(IV) in phosphate buffer (pH 7.4) b ut not in sodium cacodylate buffer (pH 7.4) nor in water. The vanadium (IV) yield increased with increasing ascorbate concentration, reaching a maximum at a vanadium(V): ascorbate ratio of 2:1. Addition of forma te to the incubation mixture containing vanadium(V), ascorbate, and ph osphate generated carboxylate radical (.COO-), indicating the formatio n of reactive species in the vanadium(V) reduction mechanism. In the p resence of H2O2, a mixture of vanadium(V), ascorbate, and phosphate bu ffer generated hydroxyl radical (.OH) via a Fenton-like reaction (vana dium(IV)+ H2O2 --> vanadium(V) + .OH + OH-). The .OH yield was favored at relatively low ascorbate concentrations. Omission of phosphate sha rply reduced the OH yield. The vanadium(IV) generated by ascorbate red uction of vanadium(V) in the presence of phosphate was also capable of generating lipid hydroperoxide-derived free radicals from cumene hydr operoxide, a model lipid hydroperoxide. Because of the ubiquitous pres ence of ascorbate in cellular system at relatively high concentrations , one-electron reduction of vanadium(V) by ascorbate together with pho sphate may represent an important vanadium(V) reduction pathway in viv o. The resulting reactive species generated by vanadium(IV) from H2O2 and lipid hydroperoxide via a Fenton-like reaction may play a signific ant role in the mechanism of vanadium(V)-induced cellular injury.