Ginsenosides are the major active components in ginseng. In this study, we prepared 4 kinds of protopanaxadiol-type ginsenosides with different sugar residues, and observed their anti-proliferative activities on human colorectal cancer cell lines HCT-116 and HT-29. The results suggested that the native protopanaxadiol-type saponinRc had a dose-dependent inhibitory effects both on HCT-116 and HT-29 cells. With the number of sugar residues decreased, the anti-proliferative effect of intermediate product Mb increased significantly. However, the biotransformation products Mc and Rg3 with two sugar residues had slight inhibitory effects on both two human colon cancer cells. HT-29 cells were more sensitive to all these ginsenosides than HCT-116 cells. When the glucosidic linkages of ginsenosides were specifically cleaved by glycosidases, their anti-proliferative effects changed notablely. Our results suggested that the number of sugar residues might mainly influence the anti-proliferative functions of ginsenosides. Our results provide some useful information about the correlation between the anti-proliferative activities and the structures of ginsenosides. The protopanaxadiol-type ginsenosidesRc, Mb and Mc might be exploited as potential drug candidates for application in medicine or pharmaceutical industry.