Preparation, Evaluations And In-vivo Hypoglycemic Activity Of Dihydromyricetin Phospholipid Complex

Dihydromyricetin?DMY?is the main active ingredient in vine tea with the content over 30%,which has been found to have a variety of pharmacological activities,such as anti-inflammation,hypoglycemia,improving insulin resistance and lipid metabolism.However,DMY has poor solubility,poor absorption in vivo,as well as low oral bioavailability.Therefore,preparation technology is needed to improve its oral bioavailability.Up to now,the novel formulations of DMY reported include inclusion complexes,liposomes,microemulsions,nanomicelles,etc.Although the solubility of DMY was improved in some extent,pharmacokinetics and pharmacodynamics experiments were lack to proving the improving bioavailability of DMY.Therefore,this project aims to prepare DMY phospholipid complex,characterize in vitro and in vivo,which might lay a foundation for the development and clinical application of DMY as new drug.The main research contents of this program are divided into three chapters:Chapter 1 Preparation and characterization of DMY phospholipid complexesObjective:To prepare and characterize the DMY phospholipid complex in vitro.Methods:Firstly,a HPLC method for quantitative determination of DMY in phospholipid complex was established.Secondly,DMY phospholipid complex was prepared by solvent volatilization method,and the preparation process was optimized with DMY combination rate as the index.Thirdly,DMY phospholipid complex was identified by X-ray powder diffraction?XRPD?,differential scanning calorimetry?DSC?,scanning electron microscope?SEM?,fourier transform infrared spectroscopy?FT-IR?.Finally,drug loading,solubility and characteristics in aqueous solution of DMY phospholipid complex were also investigated.Results:?1?Under the method,there is no interferential peak signals for the quantitative determination of DMY.Good liner relationship of DMY was observed within the concentration range of 5.61⁵6.1?g/mL.The limit of detection?LOD?and limit of quantitation?LOQ?of DMY were 0.0561?g/mL and 0.1683?g/mL,respectively.The RSD values of the precision and repeatability experiments were 0.62%and 1.09%,respectively?n=6?.The RSD values of sample recovery ratewere 1.02%,1.23%and0.48%,respectively;?2?In the phospholipid complex,DMY was fully dispersed in the phospholipid with an amorphous form.At the same time,there was a certain interaction force between the hydroxyl group of DMY and the polar group of the phospholipid.?3?The average drug loading of DMY in the phospholipid complex was 25.92%.When compared to the free DMY,water solubility of DMY phospholipid complex was increased by 1.68 times.In aqueous solution,particle size of DMY phospholipid complex was?231.8±7.65?nm,the polydispersity index?PDI?was?0.206±0.024?,Zeta potential was?-60.78±5.86?mv.Conclusion:?1?The estabilished HPLC method has a good specificity,good linear relationship,high precision and accuracy.?2?The successfully prepared DMY phospholipid complex can effectively improve the solubility of DMY.Chapter 2 Pharmacokinetic study of DMY phospholipid complex in healthy and T2DM ratsObjective:To investigate the pharmacokinetic characteristics of DMY phospholipid complex in healthy SD rats and T2DM rats,and to investigate the changes of pharmacokinetic parameters of DMY and DMY phospholipid complex in T2DM rats.Methods:High-fat diet feeding combined with intraperitoneal injection of streptozotocin?STZ?was used to establish the T2DM rats model,and the blood concentration of DMY in healthy SD rats and T2DM rats after intragastric administration was determined by the estabilished LC-MS/MS method.Results:?1?In healthy SD rats,when compared with the free DMY group,the maximum blood concentration(C_max)and the area under the curve?AUC?of DMY phospholipid complex group was increased significantly,and the in vivo clearance rate?CL?and apparent volume of distribution?Vd?were decreased significantly after oral administration at a sigle dose of 100 mg/kg.The same changes were also observed in T2DM rats.?2?Compared with healthy SD rats,C_maxax and AUC of free DMY was increased significantly in T2DM rats,and CL and Vd were decreased significantly after oral administration at a sigle dose of 100 mg/kg.DMY phospholipid complex showed the same change rules.Conclusion:Phospholipid complex technology can improve the oral bioavailability of DMY,which is expected to be developed as a novel drug delivery system for DMY.In addition,in the state of T2DM,the pharmacokinetic parameters of DMY and DMY phospholipid complex significantly changed,the inside mechanism needs to be further studied.Chapter 3 In-vivo hypoglycemic activity of DMY phospholipid complexObjective:To investigate the in-vivo hypoglycemic activity of DMY phospholipid complex in T2DM mice.Methods:T2DM mice model was established by high-fat diet feeding combined with intraperitoneal injection of streptozotocin?STZ?.After the successful modeling,T2DM mice were given DMY,DMY-phospholipid physical mixture,DMY phospholipid complex,phospholipid and metformin at low or high doses once a day through gavage.The changes of body weight for T2DM mice were monitored once a week,and the changes in fasting blood glucose?FBG?of T2DM mice were measured every two weeks.On the basis,glucose tolerance test was carried out.Serum insulin and lipid levels of the mice were detected finally.Results:The body weight of mice in the healthy mice group was increased,and there was no significant change in fasting blood glucose?FBG?in healthy mice.While body weight loss in T2DM mice was observed.After long-term drug intervention,metformin,DMY and DMY phospholipid complex could significantly alleviate body weight loss and reduce fasting blood glucose in T2DM mice.Metformin,DMY and DMY phospholipid complex could significantly improve T2DM-induced glucose tolerance abnormalities,as well as insulin resistance and blood lipid metabolism in T2DM mice.Conclusion:DMY and DMY phospholipid complex can improve T2DM to some extent,and it is expected to develop as a new drug for the treatment of T2DM...