Light-emitting diode (LED) systems have emerged as a promising and sustainable technology, with significant biotechnological applications in horticulture. Numerous studies have highlighted their use in greenhouse farming and food safety. This study aimed to investigate the influence of red LED lights on the postharvest accumulation of bioactive compounds in tomato (Solanum lycopersicum) fruits and the impact of maturity stage on this process. Tomato fruits harvested at the Breaker and Pink stages were exposed to daily cycles (12 hours) of red light for 14 days (20 °C), followed by 7 days of dark storage (20 °C). The results revealed changes in color parameters (a* and a*/b*) in Breaker fruits, with lycopene levels increasing by 52% and 53% in both Breaker and Pink stage fruits, respectively, by the end of the storage period. No significant differences were observed in firmness, soluble solids, and titratable acidity parameters. Additionally, the expression levels of genes involved in the biosynthetic pathways of lycopene, vitamins C and E, and folate were analyzed using qPCR assays. The SlGGP1 and SlHPPD1 genes, which regulate vitamin C and E biosynthesis, respectively, showed changes in relative expression in Breaker stage tomatoes stored in the dark. The SlPSY1 gene, involved in lycopene biosynthesis, exhibited altered expression only in Breaker stage tomatoes after 14 days of treatment. The SlGCHI gene, a regulator of folate biosynthesis, increased its relative expression in both Breaker and Pink stages during the treatment. These findings suggest that the Breaker stage is particularly suitable for red light treatments, as it results in significant increases in lycopene levels while maintaining the organoleptic quality traits of tomato fruits.