Scutebarbatine A (SBT‐A), a diterpenoid alkaloid found in the root of Scutellaria barbata D. Don, has been reported to induce the apoptosis of A549 cells. In this study, we investigated the antitumor activity of SBT‐A in human hepatocellular carcinoma (HCC) cells and the potential underlying mechanisms. Our results showed that SBT‐A inhibited the growth of HCC cells in a dose‐dependent manner. SBT‐A treatment caused cell cycle arrest and decreased the expression of cyclin B1, cyclin D1, p‐Cdc2, and p‐Cdc25C. SBT‐A triggered cell apoptosis via a caspase‐dependent pathway, and cell viability was partially restored by pretreatment with the pan‐caspase inhibitor Z‐VAD‐FMK. In HCC cells, treatment with SBT‐A increased the phosphorylation of extracellular signal‐regulated kinase 1 and 2 (ERK1/2), c‐Jun N‐terminal kinase 1 and 2 (JNK1/2), and p38 mitogen‐activated protein kinase (p38 MAPK). Moreover, SBT‐A activated endoplasmic reticulum (ER) stress through the upregulation of protein kinase RNA‐like ER kinase (PERK), activating transcription factor 4 (ATF‐4), and CCAAT‐enhancer‐binding protein (C/EBP) homologous protein (CHOP). Our data indicate that SBT‐A inhibits the proliferation of HCC cells and triggers their apoptosis via the activation of MAPK and ER stress. SBT‐A is a potential agent for the treatment of HCC.