Triterpenoids from<i>Ziziphus jujuba</i>induce apoptotic cell death in human cancer cells through mitochondrial reactive oxygen species production

Ziziphus jujuba var. inermis Rehder is an edible fruit-producing species of the Rhamnaceae family. In the present study, we isolated eight triterpenoids (1-8) from the fruits of Z. jujuba var. inermis and evaluated their apoptotic cell-death-inducing activities in human cancer cell lines (A549, PC-3, and MDA-MB-231). The structures of compounds 1-8 were determined by spectroscopic methods. Among these, four isomers of coumaroyl alphitolic acid showed potent cytotoxic activities on these cancer cells: 3-O-cis-p-coumaroyl alphitolic acid (3), 3-O-trans-p-coumaroyl alphitolic acid (4), 2-O-trans-p-coumaroyl alphitolic acid (5), and 2-O-cis-p-coumaroyl alphitolic acid (6). Moreover, compounds 3-6 induced apoptotic cell death in a concentration-dependent manner. We further investigated the apoptosis-inducing effects of compound 4 in PC-3 cells which triggered the cleavage of procaspase-3, procaspase-7, procaspase-8, bid, and PARP. Compound 4 increased both the mitochondrial reactive oxygen species (ROS) production and the phosphorylation of p38 MAPK (mitogen-activated protein kinase), but decreased the mitochondrial membrane potential. Pretreatment with Mito-TEMPO (a specific mitochondrial-targeted antioxidant) or a specific p38 inhibitor (SB203580) attenuated apoptotic cell death triggered by compound 4 which suggests that compound 4 may induce apoptotic cell death in these cancer cells by increasing the mitochondrial ROS production as well as the subsequent p38 MAPK activation. The study findings provide a rational base to use Ziziphus extracts for cancer treatments in traditional oriental medicine.