Dark-adapted, single photoreceptors isolated from the frog retina produce reactive oxygen species (ROS) after about 1 min of illumination with saturating light that we verified by their oxidation of preloaded dihydrorhodamine 123 (DHR) into the fluorescent rhodamine 123 (RHO). In this preparation we tested the antioxidant effects of vitamin E and of melatonin. Melatonin at picomolar and low nanomolar concentrations was determined to be 100 times more potent in inhibiting the light-induced oxidative processes than was vitamin E. On the contrary, both compounds exerted potent prooxidant effects at micromolar concentrations that is above the physiological levels of melatonin. This provides evidence that physiological concentrations of melatonin in a living cell may exert protective actions against a natural oxidant stimulus (light). This helps to define the functional role of endogenous melatonin in photoreceptors, which by their physiological characteristics, are among the marked producers of ROS in the organism.