This study investigates the antioxidant potential of over 20 sorghum genotypes, spanning popular lines, inter-specific hybrids, and inter-generic crosses. Parameters such as free radical scavenging, flavonoid and phenolic content, and nitrous oxide (NO) neutralization were meticulously assessed. Leading the pack, ISC2020-C and SPV2612 demonstrated exceptional DPPH and ABTS radical scavenging, highlighting their prowess against oxidative stress. ISC812-C stood out for its highest total phenolic content, linking phenolic abundance to overall antioxidant strength. Positive correlations between NO scavenging, DPPH, ABTS, and total phenolic content emphasized the synergistic role of bioactive components in colored sorghum grains. Principal Component Analysis (PCA) revealed distinct patterns, with ISC202-C, ISC812-W, 27B, and SPV2612 displaying potent antioxidant profiles. UPGMA analysis of solvent extracts unveiled clusters, hinting at untapped diversity in inter-specific and inter-generic crosses. ISC304-C shared similarities with SM2288-G, while ISC812-C formed a unique cluster, suggesting unexplored bioactive profiles. In conclusion, this exploration showcases colored sorghum grains as rich sources of bioactive antioxidants. Utilizing inter-specific and inter-generic hybridization strategies can enhance sorghum’s nutritional value, fostering the development of safe and functional food products. The research sets the stage for optimizing sorghum breeding and processing techniques, maximizing antioxidant potential for practical applications in human health and food security.