Objective  The study aims in investigating anti-tumor activity of ginsenoside, resveratrol, ganoderma polysaccharides and cordyceps sinensis in human cancer cell lines, and whether the combination of all four phytochemicals (GRGC) exerts synergistic anti-tumor effect. Methods  Human cancer cell lines (A549, MCF-7 and SW480) were treated with single formulation of Ginsenoside, Resveratrol, Ganoderma polysaccharides or Cordyceps sinensis, or the combined GRGC. All treated cells were analyzed for proliferation by CCK8 assay. Analysis of cell cycle arrest and apoptosis were conducted in cells treated with combined GRGC by flowcytometry. Furthermore, the ultrastraccture of A549 cells treated with GRGC was investigated by transmission electron microscope. Results  Ginsenoside, resveratrol, ganoderma polysaccharides and cordyceps sinensis exerted dose-dependent inhibition of proliferation in A549, MCF-7 and SW480 cells, while the mixed GRGC demonstrated significant synergistic effect in antitumor activity. After treatment with the GRGC, A549, MCF-7 and SW480 cells were arrested in the G0/G1 phase. Flowcytometry analysis revealed that the mixture of GRGC could induce apoptosis in A549, MCF-7 and SW480 cells in a dose-dependent manner. Electronphotomicrograph of GRGC treated A549 tumor cells showed that the cell membrane and nuclear envelope were broken, the nucleus concentrated and aggregated along the border, indicating massive or crescent like apoptosis. Conclusions  The mixture of the four phytochemicals GRGC exhibits significant synergy in inhibitory activities against tumor cells (A549、MCF-7、SW480), induces cell cycle arrest in the G0/G1 phase to inhibit tumor cell proliferation and to induce apoptosis of tumor cells (A549、MCF7、SW480). The combination of all four phytochemicals (GRGC) have obvious synergistic anticancer effect, which provides a solid foundation for the comprehensive treatment of cancer.