VITAMIN-E RECYCLING IN HUMAN ERYTHROCYTE-MEMBRANES

Vitamin E, the major lipid chain-breaking antioxidant in erythrocyte m embranes, is present in low concentration, suggesting that mechanisms should exist to protect against its loss. Enzymatic pathways for the r ecycling of vitamin E from its tocopheroxyl radical have been observed previously in inner membranes of mitochondria and microsomes. These p athways use electron transport enzymes and their substrates to regener ate vitamin E. Erythrocyte membranes also contain significant NADH-cyt ochrome c reductase activity, as well as cytochrome b5, the function o f which is not yet known. Using an enzymatic oxidation system composed of lipoxygenase and arachidonic acid, free radicals were produced in human erythrocyte membranes, and their reaction with chromanols was fo llowed by ESR and high performance liquid chromatography (HPLC). Since the endogenous vitamin E content of the membranes is very low, we use d a vitamin E homologue lacking the hydrocarbon chain (2,2,5,7,8-penta methyl-6-hydroxychromane) as a probe molecule for ESR measurements. Ho wever, parallel HPLC determinations of lipid hydroperoxides and of end ogenous vitamin E confirmed the results obtained by ESR. It was found that protection against the loss of vitamin E can be provided either b y NADH-cytochrome b5-dependent enzymatic recycling or by a nonenzymati c pathway involving ascorbate and dihydrolipoic acid.