Spectrophotometric determination of Melatonin content involves measuring the absorbance of the compound at a specific wavelength using a spectrophotometer. Melatonin is a natural compound produced by various organisms, including bacteria and eukaryotes which plays a crucial role in regulating the sleep-wake cycle (circadian rhythm) and is involved in various physiological processes, including immune function, antioxidant activity, and regulation of reproductive hormones.