Our previous transcriptomic study found that methionyl-methionine (Met-Met) exerts an anti-inflammatory effect in the bovine mammary epithelial cell (MAC-T) at a molecular level. However, evidence of whether the metabolic production of Met-Met confers protection was scarce. To investigate the inflammatory response and metabolite changes of Met-Met in lipopolysaccharide (LPS)-induced inflammation of MAC-T, mass spectrometry-based metabolomics and qPCR were conducted. The increased levels of IL-8, TNF-α, AP-1, and MCP-1 were reduced by pretreating with 2 mM Met-Met after LPS exposure. Metabolomics profiling analysis demonstrated that LPS induced significant alteration of metabolites, including decreased tryptophan, phenylalanine, and histidine levels and increased palmitic acid and stearic acid levels as well as purine metabolism disorder, whereas Met-Met reversed these changes significantly. Pathways analysis revealed that overlapping metabolites were mainly enriched in the cysteine and methionine metabolism, fatty acids biosynthesis, and purines degradation. Correlation networks showed that the metabolic profile was significantly altered under the conditions of inflammation and Met-Met treatment. Collectively, Met-Met might relieve MAC-T cell inflammation via hydrolysate methionine, which further changes the processes of amino acid, purine, and fatty acid metabolism.