Shuganning injection (SGNI), a TCM (traditional Chinese medicine) injection with good hepatoprotective effects, exerted therapeutic effects on hepatocellular carcinoma (HCC). However, the active compounds and effects of SGNI on HCC remain unclear. The objective of this study was to investigate the active compounds and potential targets of SGNI in the treatment of HCC, and explore the molecular mechanisms of main compounds. Network pharmacology was applied to predict the active compounds and targets of SGNI on cancer. The interactions between active compounds and target proteins were validated by drug affinity responsive target stability (DARTS), cellular thermal shift assay (CETSA), and pull-down assay. The in vitro test of the effects and mechanism of vanillin and baicalein was elucidated by MTT, western blot, immunofluorescence, and apoptosis analysis. According to compound characteristics, targets, etc., two typical active ingredients (vanillin and baicalein) were selected as representatives to explore the effects on HCC. Vanillin (an important food additive) bound to NF-κB1 and baicalein (a bioactive flavonoid) bound to FLT3 (FMS-like tyrosine kinase 3) were confirmed in this study. Vanillin and baicalein both inhibited cell viability and promoted apoptosis of Hep3B and Huh7 cells. In addition, both vanillin and baicalein could enhance the activation of the p38/MAPK (mitogen-activated protein kinase) signaling pathway, which may partially explain the anti-apoptosis effects of the two compounds. In conclusion, two active compounds of SGNI, vanillin and baicalein, promoted apoptosis of HCC cells via binding with NF-κB1 or FLT3, and regulating the p38/MAPK pathway. Baicalein and vanillin may be good candidates for HCC treatment on drug development.

舒肝宁注射液（SGNI）是一种具有良好保肝作用的中药注射剂，对肝细胞癌（HCC）有一定的治疗作用。然而，SGNI 的活性化合物和对 HCC 的作用仍不清楚。本研究的目的是研究SGNI治疗HCC的活性化合物和潜在靶点，并探讨主要化合物的分子机制。应用网络药理学来预测 SGNI 对癌症的活性化合物和靶点。通过药物亲和力响应靶稳定性 (DARTS)、细胞热位移测定 (CETSA) 和下拉测定来验证活性化合物和靶蛋白之间的相互作用。通过MTT、蛋白质印迹、免疫荧光和细胞凋亡分析阐明香草醛和黄芩素的作用和机制的体外试验。根据化合物特性、作用靶点等，选取两种典型活性成分（香兰素和黄芩素）作为代表，探讨其对HCC的作用。这项研究证实了香兰素（一种重要的食品添加剂）与 NF-κB1 结合，黄芩素（一种生物活性黄酮类化合物）与 FLT3（FMS 样酪氨酸激酶 3）结合。香兰素和黄芩素均抑制细胞活力并促进 Hep3B 和 Huh7 细胞凋亡。此外，香草醛和黄芩素均可增强p38/MAPK（丝裂原激活蛋白激酶）信号通路的激活，这可能部分解释了两种化合物的抗细胞凋亡作用。总之，SGNI 的两种活性化合物香草醛和黄芩素通过与 NF-κB1 或 FLT3 结合，调节 p38/MAPK 通路，促进 HCC 细胞凋亡。黄芩素和香草醛可能是药物开发中 HCC 治疗的良好候选者。