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Objective: To employ the response surface methodology (RSM) to optimize the preparation conditions for ginsenoside Rh4 (GRh4) and Rk3 (GRk3) using the yields as index. Methods: High temperature hydrolysis combined with RSM was used to prepare GRk3 and GRh4 from ginsenoside Rg1 (GRg1), and the reaction conditions were optimized at the same time. Results: The optimal preparation conditions of GRk3 and GRh4 were confirmed using Design Expert 7.1.6 software as followss: acid concentration of 0.02%, reaction temperature of 116.4℃, and reaction time of 2.22 h, respectively. Under the above conditions, the yields of GRk3 and GRh4 were 123.56 mg with conversion rate of 61.78%. Conclusion: The preparation conditions obtained from RSM optimization are suitable for large scale preparation of GRk3 and GRh4.
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AIM: To investigate the apoptosis and molecular mechanism of human hepatocellular carcinoma HepG2 cells induced by ginsenoside Rh4.METHODS: Human hepatocellular carcinoma HepG2 cells were treated with ginsenoside Rh4 at doses of 10, 20 and 40 μmol/L, and the inhibitory effect of ginsenoside Rh4 on HepG2 cell viability was measured by MTT assay.The apoptotic rate of HepG2 cells was analyzed by flow cytometry.The morphological changes of the HepG2 cells were observed by Hoechst 33258 and TUNEL staining.The expression of apoptosis-related proteins Bax, Bcl-2, caspase-3 and caspase-9 was determined by Western blot.RESULTS: Ginsenoside Rh4 promoted apoptosis of HepG2 cells in a dose-dependent manner.TUNEL and Hoechst 33258 staining showed that the cells appeared obvious shrinking, swelling and rupture after treated with ginsenoside Rh4 for 24 h.The results of Western blot showed that with the increasing concentrations of ginsenoside Rh4, the expression of pro-apoptotic proteins Bax, cleaved caspase-3 and caspase-9 increased, while anti-apoptotic protein Bcl-2 decreased gradually.CONCLUSION: Ginsenoside Rh4 induces apoptosis of human hepatocellular carcinoma HepG2 cells, and the main mechanism may be related to down-regulation of Bcl-2 and up-regulation of Bax, cleaved caspase-3, and caspase-9.
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UPLC-QTOF-MS/MS was used to identify metabolites in rat blood, urine and feces after the administration of n-butanol extract derived from steamed notoginseng. The metabolic process of saponins came from steamed notoginseng was analyzed. The metabolites were processed by PeakView software, and identified according to the structural characteristics of prototype compounds and the accurate qualitative and quantitative changes of common metabolic pathways. Four saponins metabolites were identified based on MS/MS information of metabolites, namely ginsenoside Rh₄, Rk₃, Rk₁, Rg₅,and their 15 metabolites were verified. The metabolic pathways of the four ginsenosides in n-butanol extract included glucuronidation, desugar, sulfation, dehydromethylation, and branch loss. The metabolites of main active saponin components derived from steamed Panax notoginseng were analyzed from the perspective of qualitative analysis. And the material basis for the efficacy of steamed notoginseng was further clarified.
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Objective: A new, environment-friendly and efficient method for the preparation of rare ginsenoside Rg6, F4, Rk3, and Rh4 was established, which provides a theoretical basis for preparing rare ginsenosides. Methods: Rare ginsenoside was prepared by hydrolyzing ginsenoside Re using aspartic acid as the catalyst, through semi preparative HPLC, the target compounds Rg6, F4, Rk3, and Rh4 were rapidly separated from the degradation products, quantitative analysis, and structure identification by HPLC and NMR. Results: Ginsenoside Re was hydrolyzed by aspartic acid according to the ratio 10∶1 at 120℃ for 1 h, the conversion rate of ginsenoside Re was 100%, the yields of rare ginsenoside Rg6, F4, Rk3, and Rh4 were 11.2%, 13.1%, 20.6%, and 24.3%, respectively, and the purity of the four compounds were all above 99%. Conclusion: The method is simple, low-cost, and non-pollution for environment, the research has important application value for the development of green environmental protection of rare ginsenosides drugs and health food.