Flesh extract of Mangifera indica possesses anti-proliferation effect on different types of cancer cells. However, the effect of its seed kernel extract (MSP) on the growth of human colorectal carcinoma cells (CRC) has not yet been evaluated. Phenolic species of MSP were extracted and measured by colorimetry. Two CRC cell lines (Colo 320DM and SW480) were treated with MSP and assessed for viability by trypan blue exclusion, for cell cycle distribution by flow cytometry, for apoptosis by annexin V labeling, for mitochondria potential by rhodamine 123 staining and for changes in the levels of proteins involved in cell cycle control or apoptosis by immunoblotting. MSP inhibited the proliferation (12.5 μg/mL -50 μg/mL) of Colo 320DM and SW480. MSP inhibited proliferation by blocking cell cycle progression at G1 (SW480) or S (Colo 320DM) phase and inducing apoptotic death. Western blotting indicated that MSP-blocking cell cycle was associated with cyclin levels. MSP-treated Colo 320DM and SW480 also showed activation of caspase 8, 9 and 3. MSP induces cell cycle arrest and apoptotic death in two CRC cell lines. The results indicate that MSP is a potential novel chemoprevention and treatment agent for colorectal cancer.