INTRODUCTION: Prostate cancer (PCa) is a malignancy characterized by abnormal cell proliferation in the prostate gland, a critical component of the male reproductive system. Atractylodes lancea DC. (ALD), a medicinal herb commonly used in traditional Asian medicine, is highly regarded for its antioxidant, antidiabetic, and anticancer properties. Virtual docking stud-ies have identified Atractylenolide II and III as active components of ALD, demonstrating strong binding potential to inhibit androgen receptor (AR) activity, with docking scores of -8.9 and -9.3, respectively. These findings suggest that ALD may exert a synergistic effect comparable to or greater than that of enzalutamide (ENZ) in inhibiting AR. How-ever, its specific anticancer and anti-metastatic mechanisms in prostate cancer remain unclear. METHODS: The cytotoxic effects of ALD were evaluated on PC3 and DU145 prostate cancer cells, as well as on the normal prostate cell line BPH-1. Cell viability was assessed using the EZ-Cytotoxic kit, while colony formation assays and TUNEL staining were used to meas-ure proliferation and apoptosis, respectively. Apoptosis was further analyzed through an-nexin V-FITC/PI staining and quantified by flow cytometry (FACS). Western blotting was performed to elucidate the underlying molecular mechanisms. Additionally, mito-chondrial membrane potential (ΔΨm) and intracellular calcium levels were measured to evaluate mitochondrial function, while reactive oxygen species (ROS) generation was assessed with and without pretreatment with N-acetylcysteine (NAC) . RESULTS: ALD selectively reduced the viability of PC3 and DU145 prostate cancer cells while spar-ing BPH-1 normal prostate cells, demonstrating cancer-selective cytotoxicity. ALD dis-rupted mitochondrial function by reducing ΔΨm and increasing intracellular calcium lev-els. A concentration-dependent increase in ROS generation was observed in PC3 and DU145 cells, which was completely inhibited by NAC pretreatment, confirming a ROS-mediated mechanism. Colony formation assays revealed a significant reduction in prolif-eration, while TUNEL and annexin V-FITC/PI staining indicated enhanced apoptosis. Western blot analysis showed that ALD modulates critical survival pathways, leading to apoptotic cell death. DISCUSSION: These findings demonstrate that ALD exerts potent anticancer effects against metastatic prostate cancer cells through ROS-mediated apoptosis and mitochondrial dysfunction, while exhibiting minimal cytotoxicity toward normal prostate cells. The presence of ac-tive compounds such as Atractylenolide II and III suggests a synergistic interaction that enhances AR inhibition and promotes apoptosis. ALD's ability to engage multiple path-ways highlights its therapeutic potential as a selective and multifaceted treatment for ag-gressive prostate cancer.