Melatonin, a natural plant hormone, has recently gained notable attention for its multifunctional roles in animals and plants, particularly in extending the shelf life of perishable food products. As an endogenous antioxidant and regulator of physiological processes, melatonin is potentially responsible for delaying senescence, mitigating oxidative stress, and enhancing the overall quality of harvested products. This chapter explores the mechanistic understanding of melatonin’s crucial role in extending the shelf life of perishable food products, focusing on its biochemical properties and interaction with reactive oxygen species (ROS), which are primary factors of food spoilage. This chapter explores the role of melatonin’s influence on post-harvest physiology, especially in modulating cellular processes such as gene expression, membrane integrity, and ethylene production, which are crucial for delaying ripening and spoilage. The discussion will also review recent research demonstrating melatonin’s effectiveness in various food systems, including fruits and vegetables. This chapter also highlights the probable integration of melatonin with other preservation technologies and the potential for genetically engineered crops with enhanced endogenous melatonin levels. This comprehensive overview of the molecular/biochemical mechanism and practical melatonin application in different food systems may aid researchers and industries in developing food products with minimal post-harvest losses and ensuring food security.