PROTECTION BY VITAMIN-C OF OXIDANT-INDUCED LOSS OF VITAMIN-E IN RAT HEPATOCYTES

Studies with liposome model membranes demonstrate that vitamin C (asco rbic acid) can regenerate vitamin E (alpha-tocopherol) but in vivo exp eriments have yielded equivocal results concerning this recycling path way. In vitro evaluation of vitamin C-vitamin E recycling in cell syst ems is lacking. We investigated the capacity of vitamin C to spare or regenerate vitamin E from loss during oxidative stress in primary rat hepatocytes. Livers from 3-month-old male Sprague-Dawley rats were per fused with collagenase and the hepatocytes incubated for 180 min in pH 7.4 Hank's balanced salt solution with 0.1% bovine serum albumin and the free radical generator 2,2'-azo-bis(2-amidinopropane) dihydrochlor ide (AAPH; 10-100 mM), a water soluble compound, or 2,2'-azo-bis-2,4-d imethylvaleronitrile (AMVN; 250-900 mu moles/L cell suspension), a lip id soluble compound, or a vehicle control. Hepatocyte alpha-tocopherol decreased by 75% in a dose-dependent fashion following AAPH treatment and was completely consumed following AMVN exposure. Loss of vitamin E was more rapid after treatment with AMVN than the AAPH (50% loss by 30 and 180 min with 900 mu M AMVN and 10 mM AAPH, respectively) while declines were 30% or less under control conditions after 180 min. The addition of 2 to 6 mM vitamin C to the media after 15, 30, or 60 min p revented futher loss of cellular vitamin E after AAPH or AMVN treatmen t. Vitamin C decreased the accumulation of alpha-tocopherolquinone by 35% following AMVN treatment, suggesting a recycling rather than an ex clusive sparing action of vitamin C on vitamin E. Regardless of its sp ecific mechanism of action, vitamin C can protect against the oxidant- induced loss of vitamin E in vitro. (J. Nutr. Biochem. 9:355-359, 1998 ) (C) Elsevier Science Inc. 1998.