Display omitted •Anti-cancer effects of Butein in Colorectal Cancer (CRC): This study systematically explored the anti-cancer effects of Butein in CRC, a natural chalcone found in plants with promising anti-tumor effects across various cancers but largely unclear in CRC.•Comprehensive approach to elucidate Butein’s mechanism of action: The study employed a range of methods including pharmacological, molecular, and in vivo models to comprehensively assess Butein’s impact on CRC cell proliferation, apoptosis, migration, and invasion. Molecular docking and network pharmacology analyses predicted potential target proteins and pathways regulated by Butein.•mplications: The study highlights the multifaceted anti-cancer potential of Butein in CRC and underscores its promising clinical application. Additionally, it reveals the novel role of the p38 pathway. Further research is needed to elucidate detailed molecular interactions and clinical translatability of Butein as a prospective therapy for CRC. Butein, a natural chalcone found in plants, exhibits promising anti-tumor effects across cancers, yet its role in Colorectal cancer (CRC) remain largely unclear. We utilized pharmacological, molecular, and in vivo models to assess its impact on CRC cell proliferation, apoptosis, migration, and invasion. Molecular docking and network pharmacology analyses predicted potential target proteins and pathways regulated by Butein. Experimental validations included western blotting, immunohistochemistry, and functional assays. Butein significantly inhibited CRC cell proliferation, induced apoptosis, and disrupted the epithelial-mesenchymal transition (EMT). Its repression of cell migration and invasion capabilities, suggesting potential anti-metastatic properties. The activation of the p38 signaling pathway emerged as a key mechanism mediating Butein’s anti-tumor effects, supported by network pharmacology predictions and experimental evidence. This study highlights the multifaceted anti-cancer potential of Butein in CRC, emphasizing its promising application and the novel role of the p38 pathway.