This study aimed to decipher the biochemical changes of milk fermented by the culture starter, Streptococcus thermophilus. Streptococcus thermophilus S10 possesses good fermentation properties; thus, it was the chosen strain used here. The metabolomic profiles were generated by ultra performance liquid chromatography coupled to a quadrupole-time-of-flight mass spectrometry (UPLC-QTOF MS), and data were analyzed by multivariate analyses methods like principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA). A total of 39 significant differential metabolites were identified between milk and fermented milk samples, including peptides, amino acids, fatty acids and related metabolites, vitamins. Thus, it is likely that some of the currently identified differentially abundant metabolites also contribute to the unique flavor, taste, and aroma of fermented milk. The bitterness of fermented milk are likely to be due to some of these amino acids, whereas the formation of S-methyl thioesters from short chain fatty acids, providing a unique flavor to fermented milk. What's more, small peptides detected in fermented milk maybe have bioactivities and beneficial functions. Some of these changes might contribute to the unique sensory and nutritional qualities of fermented milk. Our data based on LC-MS metabolomics are valuable references for future development and improvement of fermented dairy products.