ERYTHROCYTE ASCORBATE RECYCLING - ANTIOXIDANT EFFECTS IN BLOOD

Ascorbic acid is an important antioxidant in human plasma, but require s efficient recycling from its oxidized forms to avoid irreversible lo ss, Human erythrocytes prevented oxidation of ascorbate in autologous plasma, an effect that required recycling of ascorbate within the cell s. Erythrocytes had a high capacity to take up dehydroascorbate, the t wo-electron oxidized product of ascorbate, and to reduce it to ascorba te. Uptake and conversion of dehydroascorbate to ascorbate was saturab le, was half-maximal at 400 mu M dehydroascorbate, and achieved a maxi mal intracellular ascorbate concentration of 1.5 mM. In the presence o f 100 mu M dehydroascorbate, erythrocytes had the capacity to regenera te a 35 mu M ascorbate concentration in blood every 3 min. Ascorbate r ecycling from DHA required intracellular GSH. Depletion of erythrocyte GSH by more than 50% with diamide did not acutely affect the cellular ascorbate content, but did impair the subsequent ability of GSH-deple ted cells to recycle dehydroascorbate to ascorbate. Whereas erythrocyt e ascorbate recycling was coupled to GSH, an overwhelming extracellula r oxidant stress depleted both ascorbate and or-tocopherol before the GSH content of cells fell appreciably. Recycled ascorbate was released from cells into plasma, but at a rate less than one tenth that of deh ydroascorbate uptake and conversion to ascorbate. Nonetheless, ascorba te released from cells protected endogenous oc-tocopherol in human LDL from oxidation by a water soluble free radical initiator. These resul ts suggests that recycling of ascorbate in erythrocytes helps to maint ain the antioxidant reserve of whole blood. (C) 1998 Elsevier Science Inc.