Panax ginseng is a precious and ancient medicinal plant. The completion of its genome sequencing has laid the foundation for the study of proteome and peptidome. However, the high abundance of secondary metabolites in ginseng reduces the identification efficiency of proteins and peptides in mass spectrometry. In this report, strong cation exchange pretreatment was carried out to eliminate the interference of impurities. Based on the charge separation of proteolytic peptides and metabolites, the sensitivity of mass spectrometry detection was greatly improved. After pretreatment, 2322 and 2685 proteins were identified from the root and stem leaf extract. Further, the ginseng peptidome was analyzed based on this optimized strategy, where 970 and 653 endogenous peptides were identified from root and stem leaf extract, respectively. Functional analysis of proteins and endogenous peptides provided valuable information on the biological activities, metabolic processes, and ginsenoside biosynthesis pathways of ginseng.