The selection of signature peptides is critical for reliable peptide quantification. The purpose of this study was to comprehensively assess the digestion efficiency and stability of tryptic peptides and to identify optimal signature peptides for human hepatic transporters and membrane marker proteins. The plasma membrane fraction of the human liver was digested at different time points (0.5, 1, 2, 4, 8, 16, 24, and 48 h) and the peptides were comprehensively quantified using quantitative proteomics.