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Objective To establish a simultaneous quantitative analytical method of 10 anthraquinone derivatives in Rheum officinale based on ultra-high performance liquid chromatography (UPLC) for the quality evaluation and control. Methods The UPLC method was performed on a BEH C18 column (100 mm × 2.1 mm, 1.7 μm) with a binary gradient mobile phase consisting of a mixture of methanol and 0.1% aqueous phosphoric acid with column temperature at 34 ℃. Using a gradient elution method; The flow rate was set at 0.20 mL/min; The injection volume was set at 2.0 μL; The detection wavelength was set at 410 nm. This study focused on the influence of analytical parameters on the separation efficiency of five anthraquinone glucosides in R. officinale. To obtain robust chromatographic resolution of the five anthraquinone glucosides, relatively important chromatographic parameters including the retention time, resolution, theoretical plate number, and symmetry factor were investigated by altering the column temperature and flow rate. Results The optimal column temperature was found to be 34 ℃, and the optimal flow rate was 0.20 mL/min. A new UPLC analytical method was developed for simultaneous quantification of 10 anthraquinone derivatives in R. officinale based on these optimal parameters. This UPLC-based analytical method was successfully applied to quantitative determination of 10 anthraquinone derivatives in Rheum tanguticum and Rheum palmatum. The analysis showed that there were significant content differences between aloe-emodin-8-O-β-D-glucoside (more than 60-fold) and rhein-8-O-β-D-glucoside (more than 77-fold) in the two species of rhubarbs tested. Conclusion The UPLC method established for the simultaneous determination of 10 anthraquinone derivatives in R. officinale based on the optimal parameters is accurate and sensitive, which will provide a more comprehensive evaluation for the chemical characterization of medicinal materials of R. officinale from different sources.
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Objective To evaluate the ability of the antiplatelet aggregation of ten anthraquinone derivatives in Rhei Radix et Rhizoma by using bioassay method, and to screen for the indicators that can be used to control the quality of rhubarb wine processing product. Methods Platelet aggregation instrument was used to determine the platelet aggregation rate induced by ADP in vitro and calculate the antiplatelet aggregation rates of 10 anthraquinone derivatives (aloe-emodin, rhein, emodin, chrysophanol, physcion, aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, emodin-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, and physcion-8-O-β-D-glucoside) at different concentrations. The biopotency was calculated by bioavailability software. In order to verify the accuracy of the bioassay results, the estimated inhibition constant of rhein and chrysophanol-8-O-β-D-glucoside in P2Y12 protein receptor were determined by molecular docking software. Results The bioassay results showed that the antiplatelet potencies of rhein and emodin were significantly higher than those of aloe-emodin, chrysophanol and physcion. Compared with the antiplatelet biopotency of aspirin, the antiplatelet potencies of rhein and emodin were 5.02 and 5.15 times higher than that of aspirin, which indicated that rhein and emodin have strong inhibitory effect on ADP-induced platelet aggregation. However, the antiplatelet potencies of aloe-emodin, chrysophanol and physcion were equivalent of that of aspirin. The antiplatelet potencies of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, emodin-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, and physcion-8-O-β-D-glucoside were higher 4.13, 4.46, 9.31, 5.46, and 7.80 times than that of aspirin, respectively, which indicated that the five anthraquinone glucosides had a strong ability to antagonize ADP-induced platelet aggregation. The results of molecular docking showed that P2Y12 protein had different selectivity to 10 anthraquinone derivatives, especially for rhein, chrysophanol-8-O-β-D-glucoside, and the estimated Ki value were 5.73 and 2.51 μmol/L, respectively, indicating that rhein, chrysophanol-8-O-β-D-glucoside produced a strong inhibitory effect on P2Y12 protein at a lower concentration level. Moreover, the activity of chrysophanol-8-O-β-D-glucoside was stronger than rhein, which was consistent with their measured intensity of antiplatelet aggregation. Conclusion All results showed that there were some differences among antiplatelet potencies of 10 anthraquinone derivatives, and finally the screening of rhein, emodin can be used as evaluation indicators to control the quality of rhubarb wine processing product.
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Objective To analyze the main chemical constituents and their contents in aqueous extract of Polygoni Multiflori Radix (PMR, root of Polygonum multiflorum), and to elucidate the effects of aqueous extract of PMR and its main constituents on the expression of the mRNA of CYP1A2, CYP2C9, and CYP2E1 in human liver L02 cells. Methods The main chemical constituents and their content in aqueous extract of PMR were determined by HPLC. The cytotoxicity of aqueous extract of PMR and its main constituents on L02 cells was determined by MTT assay. The mRNA expression of CYP1A2, CYP2C9, and CYP2E1 in L02 cells were detected by quantitative real-time PCR. Results There were four main well-separated chromatographic peaks standing for tetrahydroxy stilbene glucoside, emodin-8-O-β-D-glucoside, emodin and physcion in aqueous extract of PMR. The content of thesecomponents in aqueous extract of PMR was (1.14 ± 0.03)%, (0.106 9 ± 0.001 6)%, (0.010 8 ± 0.000 9)%, (0.003 55 ± 0.000 19)%, respectively. The cytotoxicity of aqueous extract of PMR and emodin on L02 cells at 24 h was dose-dependent, and the concentration of 50% inhibition was 7.290 mg/mL and 0.082 mmol/L respectively. Tetrahydroxy stilbene glucoside, emodin-8-O-β-D-glucoside and physcion did not show significant cytotoxicity on L02 cells in the experimental concentrations. Aqueous extract of PMR and emodin significantly inhibited the expression of mRNA of CYP1A2, CYP2C9, and CYP2E1 in L02 cells. Emodin-8-O-β-D-glucoside inhibited the expression of mRNA of CYP1A2 and CYP2C9. Tetrahydroxy stilbene glucoside inhibited the expression of mRNA of CYP1A2 but activated the expression of mRNA of CYP2C9. Physcion inhibited the expression of mRNA of CYP1A2 and CYP2C9 in a dose-dependent manner, but inhibited the expression of mRNA of CYP2E1 in low concentration and activitated the expression of mRNA of CYP2E1 in high concentration. Conclusion The inhibition of aqueous extract of PMR on the expression of mRNA of CYP1A2, CYP2C9, and CYP2E1 in L02 cells is the combined effect of all components in it. The main four components all inhibit the expression of mRNA of CYP1A2. The anthraquinone is the main component inhibiting the expression of mRNA of CYP2C9. The free anthraquinone is the main component inhibiting the expression of mRNA of CYP2E1.
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0.05). The generation of ROS increased obviously after the cells were incubated with them for 12 h, and the increase of ROS reached the peak treated for 24 h. The levels of ?? m were time-dependently decreased after treating with four compounds for 12, 24, and 48 h. Conclusion The grouth of both MDR KBv200 cells and parental drug-sensitive KB cells were inhibited to the treatment of four anthraquinone derivative in vitro. The mechanism of their effects is associated with the increase of the cellular ROS level and the decrease of ?? m.