We herein investigated the in vitro effect of hypoxanthine on the activities of antioxidant enzymes such as catalase (CAT), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD) in erythrocytes, as well as on thiobarbituric acid-reactive substances (TBA-RS), in the plasma of rats. Results showed that hypoxanthine, when added to the incubation medium, enhanced CAT (10.0 μM), GSH-Px and SOD (8.5 μM and 10.0 μM) activities in erythrocytes of 15-day-old rats, reduced CAT activity (10.0 μM) and enhanced GSH-Px activity (10.0 μM) in erythrocytes of 30-day-old rats, reduced CAT activity (10.0 μM) and enhanced GSH-Px activity (8.5 μM and 10.0 μM) in erythrocytes of 60-day-old rats, as compared to controls. In addition, hypoxanthine (10.0 μM) enhanced TBA-RS levels in the plasma of 30- and 60-day old rats. Furthermore, we also tested the influence of allopurinol, trolox, and ascorbic acid on the effects elicited by hypoxanthine on the antioxidant enzymes and TBA-RS. Allopurinol and/or administration of antioxidants prevented most alterations caused by hypoxanthine in the oxidative stress parameters evaluated. Findings suggest that hypoxanthine alters antioxidant defenses and induces lipid peroxidation in the blood of rats; however, in the presence of allopurinol and antioxidants, some of these alterations in oxidative stress caused are prevented. Data indicate that, in humans, antioxidant administration might serve as a potential adjuvant therapy for ameliorating the damage caused by hypoxanthine.