Cancer chemotherapy remains a serious challenge, and new approaches to therapy are urgently needed to build novel treatment regimens. The methanol extract of the stem of was used to synthesize biogenic zinc oxide nanoparticles (ZnO-NPs) that display anticancer activities against colorectal cancer. Biogenic ZnO-NPs synthesized from methanol extract of stem (ZnO-NPs TM) were tested against HCT-116 cell lines to assess anticancer activity. UV-Vis, FTIR, XRD, SEM, and TEM analysis characterized the biogenic ZnO-NPs. To see how well biogenic ZnO-NPs fight cancer, cytotoxicity, AO/EtBr staining, Annexin V/PI staining, mitochondrial membrane potential (MMP), generation of reactive oxygen species (ROS) analysis, and caspase cascade activity analysis were performed to assess the anticancer efficacy of biogenic ZnO-NPs. The IC50 values of biogenic ZnO-NPs treated cells (HCT-116 and Caco-2) were 31.419 ± 0.682μg/ml and 36.675 ± 0.916μg/ml, respectively. qRT-PCR analysis showed that cells treated with biogenic ZnO-NPs Bax and P53 mRNA levels increased significantly ( ≤ 0.001). It showed to have impaired MMP and increased ROS generation. In a corollary, our study showed that biogenic ZnO-NPs have an anti-tumour effect. Biogenic ZnO-NPs TM showed both and anticancer effects that could be employed as anticancer drugs.