Stabilization of extracellular ascorbate mediated by coenzyme Q transmembrane electron transport

Ascorbate is a major water-soluble antioxidant found in body fluids of mammals, where it acts as a scavenger of noxious soluble-free radicals or lipid peroxidation-initiating radicals in the aqueous phase, as well as a regenerator of other antioxidants (e.g., vitamin E) in the lipid phase of biological membranes and lipoproteins. Ascorbate oxidation is a two-step process involving the one-electron intermediate ascorbate free radical (AFR) or semidehydroascorbate, and the two-electron, oxidized form dehydroascorbate (DHA). Intracellular reduction of ascorbate has been documented to occur both from AFR and DHA through different enzymatic reactions, with the participation of reduced nicotinamide adenine dinucleotide (NADH) and reduced glutathione (GSH) as electron donors, as well as from ascorbate acting at different cell locations. Among these mechanisms are the AFR reductase found in the chromaffin-vesicle membrane and in the outer mitochondrial membrane, the thioredoxin reductase, and glutaredoxin. The two spectrophotometric assays described in the chapter (i.e., ascorbate measurement at 265 nm and AFR signal at 360 nm), together with the measurement of NADH consumption at 340 nm, are valuable and valid tools to address this issue.Supported by Grants No. BMC2002-01602 and BMC2002-01078 of the Spanish Ministerio de Ciencia y Tecnología.Peer reviewe