Oxygen free radicals have the potential to mediate cell injury. Defenses against such radicals include the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-PX). The purposes of this study were (1) to develop an in vitro model using human cells in which to investigate a potential pharmacologic agent as an inducer of these antioxidant enzymes; (2) to investigate the phenylurea derivative N-[2-(2-oxo-1-imidazolindinyl)ethyl]- N-phenylurea (EDU) in this model with paraquat (PQ) serving as the positive control; and (3) to determine if induction of the antioxidant enzymes by EDU occurs in vivo. Human gingival fibroblasts (Gin-1) were used as the target cell in vitro; PQ and EDU, an inducer of SOD and CAT activities in plants, were evaluated as antioxidant enzyme inducers. Total SOD activity in Gin-1 cells increased 2-fold ( p < 0.05) in the presence of 1.0 m m PQ for 18–48 hr compared with untreated controls. Gin-1 cells incubated with 0.25–2.0 m m PQ for 24 hr had significantly increased total SOD (1.5 to 2.0-fold; p < 0.05). CAT activity increased with 1.0 and 2.0 m m PQ ( p < 0.05). In the presence of PQ, GSH-PX activity decreased ( p < 0.05) in a concentration-dependent manner, indicating inactivation of this enzyme. No toxicity, indicated by lactate dehydrogenase released into the incubation medium, was noted at PQ concentrations below 5.0 m m. In the presence of 0.125–2.0 m m EDU, total SOD activity in Gin-1 cells significantly increased (1.5 to 2.0-fold; p < 0.05). CAT activity significantly increased in a dose-dependent manner ( p < 0.05), while GSH-PX activity remained constant following exposure to 0.125–2.0 m m EDU. Intraperitoneal administration of EDU to rats twice a day for 2 days at 100 mg/kg induced SOD activity in heart, liver, and lung compared to controls ( p < 0.05). CAT activity increased in the liver 56% and in the lung 36% ( p < 0.05). GSH-PX activity remained constant. Our findings indicate that Gin-1 cells are a useful model in which to study inducers of antioxidant enzymes in vitro and that the phenylurea compound EDU induces SOD and CAT activities both in vitro and in vivo.