Traditional medicinal herbs are an untapped source of potential pharmaceutical compounds. This study aims to determine whether the proliferation of breast cancer cell lines could be inhibited by germacrone, a natural product isolated from Rhizoma curcuma. Germacrone treatment significantly inhibited cell proliferation, increased lactate dehydrogenase (LDH) release, and induced mitochondrial membrane potential (Delta psi m) depolarization in both MCF-7 and MDA-MB-231 cells in a dose-dependent manner. Germacrone induced MDA-MB-231 and MCF-7 cell cycle arrest at the GO/G1 and G2/M phases respectively and induced MDA-MB-231 cell apoptosis. Furthermore, germacrone treatment significantly increased Bok expression and cytochrome c release from mitochondria without affecting Bcl-2, BcI-xL, Bax, and Bim protein expressions. In addition, germacrone treatment induced caspase-3, 7, 9, PARP cleavage. We concluded that germacrone inhibited the proliferation of breast cancer cell lines by inducing cell cycle arrest and apoptosis through mitochondria-mediated caspase path