Bitter orange peel extract induces endoplasmic reticulum-mediated autophagy in human hepatoma cells

[Display omitted] •IC50 of BOPE on Hep3B cells for 24–72-h exposure is 48.95–27.18 μg/mL.•BOPE can promote ROS and trigger ER-mediated autophagy rather than apoptosis.•TAN and NOB are polymethoxyflavones existing in BOPE.•CI values show the ratio of TAN and NOB in BOPE can work synergistically in the antiproliferation. This study was aimed at investigating the anti-cancer effect of bitter orange peel extract (BOPE) on Hep3B, a human hepatoma cell line. Upon treatment of Hep3B cells with 6.25–100 μg/mL of BOPE for 72 h, its antiproliferative effect was shown to increase in a concentration- and time-dependent manner. The observations, such as concentration-dependent elevation of LC3 fluorescent puncta, increased conversion of LC3-I to LC3-II, no characteristic apoptotic nuclear fragmentation, and no significant effect on percentages of annexin V-positive apoptotic cells, confirmed the autophagy triggered by BOPE in Hep3B cells. Moreover, BOPE promoted ROS generation and induced endoplasmic reticulum stress through calcium ion-related AMPK/TSC2/mTOR and DAPK/Beclin-1 pathways, leading to autophagic cell death. Furthermore, our analysis showed that the major components of BOPE, tangeretin and nobiletin, exhibited a synergistic effect on the growth inhibition of Hep3B cells. Thus, these results suggest that BOPE may be used as a functional food against liver cancer.