1. The ability of ascorbic acid to protect from prooxidant-induced toxic injury was investigated in isolated, intact rat hepatocytes, whose ascorbic acid content had been restored by means of exogenous supplementation. 2. Ascorbate-supplemented and ascorbate-non-supplemented cells in suspension were treated with a series of different prooxidants (allyl alcohol, diethyl maleate, carbon tetrachloride, menadione), and the development of lipid peroxidation and cell injury was evaluated. 3. With allyl alcohol and diethyl maleate, ascorbic acid was able to protect cells from both lipid peroxidation and cell injury. The same protection was offered by ascorbate also in hepatocytes obtained from vitamin E-deficient animals. 4. With carbon tetrachloride, ascorbate supplementation did not affect the initial steps of lipid peroxidation, but nevertheless provided a marked protection against lipid peroxidation and cell injury at later times of incubation. The protection was unaffected by the vitamin E content of cells. 5. With menadione, a toxin which does not induce lipid peroxidation, ascorbic acid did not protect cells against injury. 6. It is concluded that ascorbic acid can act as an efficient antioxidant in isolated rat liver cells, with protection against cell injury. The antioxidant effect appears primarily to involve membrane lipids, and can be independent from the cellular content of vitamin E, thus suggesting that ascorbic acid can play a direct and independent role in the intact cell, in addition to its synergistic interaction with vitamin E described in other models.