Thiophenol and 4-aminothiophenol were used to study levels of toxicity in human red blood cells. Thiophenols caused conversion of oxyhemoglobin to methemoglobin. Reduction of corresponding disulfides by intracelluar glutathione caused cyclic reduction/oxidation reactions, resulting in increased oxidative flux. Three levels of oxidative stress were observed in these experiments: the lowest level resulted from incubation with 0.25 mM thiophenol; the intermediate level with 0.50 mM thiophenol or 0.25 mM 4-aminothiophenol; the highest levels with 0.50 mM 4-aminothiophenol. Methemoglobin formation increased with increasing level of oxidative stress. Glycolysis and the hexose monophosphate shunt were inhibited at the intermediate and highest levels of stress, respectively. Above the highest level of stress non-intact hemoglobin was formed and cell lysis occurred. These metabolic responses were reflected in cellular levels of NADH, NADPH and reduced glutathione. At the lowest level of oxidative stress, both glycolysis and hexose monophosphate shunt were increased such that near-normal levels of NADH, NADPH and reduced glutathione were maintained and methemoglobin formation was kept to a minimum. The response of red cells to 0.25 mM thiophenol appears to represent a level of oxidative stress to which the cell is capable of adaptive metabolic response. Glycolysis contributes approximately one-quarter of the total reducing equivalents from glucose metabolism in response to the oxidative challenge by thiophenol. The results suggest that the metabolic response to autoxidation of endogenous thiols is thiol exchange with glutathione and reduction of resulting glutathione disulfide by the hexose monophosphate shunt.