Constant darkness stimulated the secretion of the pineal hormone melatonin. The intensity of melatonin synthesis, which has a strong antioxidant effect, can change significantly under changing light conditions. A free-running rhythm of melatonin secretion revealed in constant darkness affects the antioxidant system. In this study were investigated the effects of constant darkness, melatonin and its receptor antagonist luzindole on retinol and tocopherol contents in the tissues and organs of adult male rats. The rats (7 months old) were divided into two groups and kept in constant darkness (DD) or standard lighting (control, 12:12 LD) for 14 days. Animals of each group were divided into three subgroups: control, receiving melatonin 22 mg/kg of body weight or luzindole at a dose of 10 mg/l. The levels of retinol and tocopherol were determined by HPLC. The study showed that staying in the dark did not affect the tocopherol and retinol content in rat tissues and organs. The greatest changes in the content of vitamins A and E under the influence of all the studied factors were observed in the liver and skeletal muscle. The relationship of retinol and tocopherol with melatonin in most cases had a mutually compensatory nature, i. e. an increase in the content of one antioxidant induced a decrease in another so that the dynamic balance of the antioxidant system was maintained.