Identifying natural variation of health-promoting compounds in staple crops and characterizing its genetic basis can help improve human nutrition through crop biofortification. Some varieties of sorghum, a staple cereal crop grown worldwide, have high concentrations of proanthocyanidins and 3-deoxyanthocyanidins, polyphenols with antioxidant and anti-inflammatory properties. We quantified total phenols, proanthocyanidins, and 3-deoxyanthocyanidins in a global sorghum diversity panel (n = 381) using near-infrared spectroscopy (NIRS), and characterized the patterns of variation with respect to geographic origin and botanical race. A genome-wide association study (GWAS) with 404,628 SNP markers identified novel quantitative trait loci for sorghum polyphenols, some of which colocalized with homologues of flavonoid pathway genes from other plants, including an orthologue of maize (Zea mays) Pr1 and a homologue of Arabidopsis (Arabidopsis thaliana) TT16. This survey of grain polyphenol variation in sorghum germplasm and catalog of flavonoid pathway loci may be useful to guide future enhancement of cereal polyphenols.