Declines in oxidative and thermal stress tolerance are well-documented in aging systems. It is thought that these alterations are due in part to reductions in antioxidant defenses. While intracellular thiols are major redox buffers, their role in maintaining redox homeostasis is not completely understood, particularly during aging, where the reliance on antioxidant enzymes and proteins may be altered. To determine if thiol supplementation improved the antioxidant enzyme profile of aged animals after heat stress, young and old Fischer 344 rats were treated with N-Acetyl-Cysteine (NAC; 4 mmol/kg i.p.) 2 h prior to heat stress. Liver tissue was collected before and 0, 30 and 60 min after heat stress. Aging was associated with a significant decline in tissue cysteine and glutathione (GSH) levels. There was also an age-related decrease in CuZn superoxide disumutase (CuZnSOD) activity. Heat stress did not alter liver GSH, glutathione disulfide (GSSG), or antioxidant enzyme activity. With NAC treatment, old animals took up more cysteine than young animals as reflected in an increase in liver GSH and a corresponding decrease in glutamate cysteine ligase activity. Catalase activity increased after NAC treatment in both age groups. CuZnSOD activity did not change with heat stress or drug treatment, while manganese SOD activity was increased in old animals only. These data indicate that GSH synthesis is substrate-limited in old animals. Further, aged animals were characterized by large fluctuations in antioxidant enzyme balance after NAC treatment, suggesting a lack of fine control over these enzymes that may leave aged animals susceptible to subsequent stress.