| Aim:To investigate a more efficient method of the synthesis and puration of hexamethylmyricetin(HMM), meanwhile, to develop a specific and sensitive HPLC detection method.Methods: HMM was synthesized by methylation of myricetin using Me2SO4 as methylation reagent. The reaction process was also further optimized by the factors of reactant ratio, temperature and reaction time. After reaction started, samples were taken at different time points. The ultraviolet absorption of the mixture was detected by HPLC to judge the progress of reaction. At the end of the reaction, the solvent was separated and a raw solid product formed. Then wash the solid mixture with water and recrystlize with alcohol. The product structure configuration was identified by MS, 1H NMR, 13C NMR and then compared with formerly reported data.Result:①The product was confirmed as HMM by MS, 1H NMR, 13C NMR;②Synthesis of HMM by the optimized process gave a product of 53 % final yield and purity≥98.84 % (HPLC area%).③The optimized reaction conditions: the ideal reactant ratio, temperature and reaction time are 1:7:9(myricetin∶Me2SO4∶K2CO3), 50℃and 9 hours, respectively.Conclusion: The modified procedure of synthesis was convenient, economical and produced less pollution. Optimized reaction conditions could promise a high yield and purity of HMM product.PartⅡEffects of HMM on the hKv1.5 channel in HEK293 cellAim: To investigate the effects of HMM on the hKv1.5 channel in HEK293 cell.Methods: HEK293 cells are transfected and stable expressed human Kv1.5 (IKur current) gene, a whole-cell patch clamp technique is used to record the current from each cell. Result: 10μM verapamil decreased ultra rapid actived delayed rectified potassium current by 87±5%, and this effect could be partially reversed by washout verapamil, which proved the current recorded was IKur, which was inhibited by the application of 0.1,0.3,1,3μM HMM and was partially reversed by washout HMM. Statistically, 3μM HMM can significantly suppress IKur . The IC50 of HMM effect on IKur is 1.79μM.Conclusion: Hexamethylmyricetin significantly inhibited hKv1.5 channel currents. Aim: To investigate the effects of FGF-21 manufacturered by GROST on KKAy and ob/ob diabetic mice.Methods:①KKAy mice were housed individually in cages, which were fed a high-fat diet ad libitum, while, ob/ob mice were housed in cages, each cage contains 5 mice, which were fed a normal chow diet ad libitum. Both KKAy and ob/ob mice were located in a room with controlled temperature (23±2℃), humidity 50%, lighted on from 7:00-19:00 and were allowed water ad libitum.②Blood was collected from the toe of KKAy and ob/ob mice by eye scissors, when the volume of bleeding is about 0.6μl, blood glucose was proceed by Roche glucometer, the value and the time were recorded.③oral glucose tolerance test (OGTT, oral glucose tolerance test). ob/ob mice were fasting for 2h, collected 0 min blood glucose as control standard, then oral administration 2g/kg glucose solution, blood samples were taken after administration 2g/kg glucose solution 30min, 60min and 120min.Results:①FGF-21 show no obvious effect on KKAy mice;②FGF-21 low-dose (0.6 mg/kg/d) can decrease fasting blood glucose on male ob/ob mice, though,the difference was not statistical significant; FGF-21 high dose (2.5 mg/kg/d) have a lowering postprandial blood glucose effect on male ob/ob mice, which on the 7th day after administration, the difference was statistical significant;③About OGTT results, FGF-21 low-dose (0.6 mg/kg/d) have a certain role in the improvement of impaired glucose tolerance on male ob/ob mice. Conclusion: FGF-21 display a certain role in the anti-diabetes action which as below: reduce blood glucose level on male ob/ob mice, correct insulin resistance. Therefore, FGF-21 is likely to be a useful clinical drug to prevent and treat diabetes. There are many further researches need to be proceeded. |
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