Castration may decrease off-odors and improve meat flavor. Meat flavor is generated through complex chemical reactions that involve hydrophilic and hydrophobic flavor precursors. In this study, we investigated the flavor precursors in psoas major muscles of castrated and intact sheep using lipidomics and targeted metabolomics. Castration decreased testosterone levels and increased intramuscular fat content. Six hundred fourteen lipid molecules confirmed showed a separation between castrated and intact sheep based on principal component analysis. Fourteen lipid species and 224 lipid molecules increased in castrated sheep. Targeted metabolomics analysis showed that 18 hydrophilic metabolites were affected by castration; however, only hypoxanthine significantly increased in the castration group. Among 45 volatiles identified, 1-octen-3-ol and hexanal were significantly higher in castrated sheep. These results revealed that lipids, hydrophilic metabolites, and volatile compounds in lamb were affected by castration, which might be beneficial in lamb quality.