This study employed a quantitative sarkosylation group analysis based on 4D rapid DIA to investigate the changes in protein sarkosylation in mouse primary hepatocytes. For the accuracy of the results, we first conducted protein quantification, followed by the detection of sarkosylation. The experiment included four treatment groups, with three biological replicates in each group, totaling 12 samples. After protein extraction, enzymatic digestion, and sarkosylation peptide enrichment, the samples were analyzed using liquid chromatography-tandem mass spectrometry. The generated data set included raw mass spectrometry files and processed quantitative data, which could be used to identify differentially sarkosylated proteins and explore the potential biological functions of lysine sarkosylation