The effects of jolkinolide B on HepG2 cells as revealed by ¹H-NMR-based metabolic profiling

Jolkinolide B (JB), which is isolated from the dried root of Euphorbia fischeriana Steud., has been reported to possess various therapeutic effects, such as treatment of edema and abdominal distention and protection against acute lung injury, and it has also been reported to have anti-inflammatory even antitumor properties. Thus, JB has always been considered a promising anticancer drug candidate. In the current work, a cellular metabolomics evaluation based on the nuclear magnetic resonance (NMR) approach was applied to investigate the mechanism of JB in HepG2 cells. In addition, biological assays such as the MTT assay, DAPI staining and the Annexin V-FITC/PI assay were implemented to evaluate cell viability and apoptosis in JB-treated cells. Subsequently, we used multivariate statistical analyses, such as principal component analysis (PCA) and orthogonal projection to latent structure with discriminant analysis (OPLS-DA) to identify metabolic biomarkers. In total, 36 metabolites in the cell extract samples and 30 metabolites in the cell culture media samples were clearly identified to be altered after the treatment. Variations in the specific metabolites suggested that HepG2 cells that were exposed to JB displayed the disordered effects in multiple metabolic pathways, such as the tricarboxylic acid cycle, amino acid metabolism, GSH synthesis and pyruvate metabolism. NMR-based cell metabolomics provided a holistic method for the identification of JB's antitumor mechanisms and the exploration of its potential applications in preclinical and clinical studies.