Background: Melatonin, a neurohormone produced by the pineal gland, has recently been reported in foods, mainly of plant origin. Melatonin provides a number of benefits for human health. It has many important health benefits because it is a potent antioxidant and anti-inflammatory hormone. Objectives: The aim of this work is to investigate the relationship between malnutrition and food deprivation on melatonin levels in normal individuals in comparison with people suffering from gastrointestinal diseases. This study will focus on the nutritional factors apart from the intake of tryptophan that affects melatonin levels in humans. Materials and Methods: This study reported interests regarding the optimization, validation, and application of analytical liquid extraction and high-performance liquid chromatography (SYKAM) coupled to a fluorescence detector for the determination of melatonin in rice grains and blood samples. Results: In the present study, the results showed that there were significant differences between the three study groups, where the lowest values for melatonin hormone concentration were found in Group C as 2.033 ± 0.69 pg/mL which includes people suffering from famine, whereas the results of Group B showed clear differences of concentration than that in Group (C) as 3.520 ± 0.62 pg/mL which includes patients who were in hospital suffering from diseases and disorders of the digestive system, but at the same time their values were less than that of the healthy control Group A as 6.457 ± 0.59 pg/mL. Conclusion: Based on the present study, it can be concluded that melatonin hormone levels are highly deficient in people who are suffering from poverty, deprivation, and starvation compared to people with problems and disorders of the digestive system.