Mechanisms of ascorbic acid recycling in human erythrocytes

Vitamin C, or ascorbic acid, is efficiently recycled from its oxidized form s by human erythrocytes. In this work the dependence of this recycling on r educed glutathione (GSH) was evaluated with regard to activation of the pen tose cycle and to changes in pyridine nucleotide concentrations. The two-el ectron-oxidized form of ascorbic acid, dehydroascorbic acid (DHA) was rapid ly taken up by erythrocytes and reduced to ascorbate, which reached intrace llular concentrations as high as 2 mM. In the absence Of D-glucose, DHA cau sed dose-dependent decreases in erythrocyte GSH,NADPH, and NADH concentrati ons. In the presence of 5 MM D-glucose, GSH and NADH concentrations were ma intained, but those of NADPH decreased. Reduction of extracellular ferricya nide by erythrocytes, which reflects intracellular ascorbate recycling, was also enhanced by D-glucose, and ferricyanide activated the pentose cycle. Diethylmaleate at concentrations up to 1 mM was found to specifically deple te erythrocyte GSH by 75-90% without causing oxidant stress in the cells. S uch GSH-depleted erythrocytes showed parallel decreases in their ability to take up and reduce DHA to ascorbate, and to reduce extracellular ferricyan ide. These results show that DHA reduction involves GSH-dependent activatio n of D-glucose metabolism in the pentose cycle, but that in the absence Of D-glucose DHA reduction can also utilize NADH. (C) 2001 Elsevier Science B. V. All rights reserved.