The hormone synthesized by the pineal gland, melatonin, has been shown to be a direct free radical scavenger both in vivo and in vitro. Thus, it potently protects cells from the damage induced by oxidative agents. In this study, we demonstrate that melatonin increases glutathione peroxidase activity in several tissues from chicks. This stimulation is time dependent and maximal increases are seen 90 min after melatonin injection (500 micrograms/kg intraperitoneally), although enzymatic activity is still elevated 135 min after its administration. No significant increases were detected 45 min after the injection. Glutathione peroxidase is generally considered to be an important antioxidative enzyme because it metabolizes hydrogen peroxide and other hydroperoxides. Thus, melatonin not only is a direct scavenger of toxic radicals but in an avian species, as in mammals, it stimulates the antioxidative enzyme glutathione peroxidase. The ability of melatonin to increase glutathione peroxidase activity is consistent with its general role as an antioxidant.