Study On The Hypoglycemic Of Total Iridoid Glycosides Of Cymbaria Dahurica L. On Type 2 Diabetes

Objective: To optimize the extraction and purification process of total iridoid glycosides(XBZG)from Cymbaria dahurica L.,and to study the mechanism of action of total iridoid glycosides of Cymbaria dahurica L.in the treatment of type 2 diabetes(T2DM).Methods: Using the extraction rate of XBZG as an indicator,the extraction process of XBZG was optimized using a single factor combined with response surface methodology.The single factor experiment investigated factors such as material liquid ratio,ethanol concentration,and extraction time.When each factor is a variable in turn,the other factors are invariant for single factor experimental research.On the basis of single factor experiments,the XBZG extraction process was optimized through response surface Box Behnken analysis.Based on the optimization of XBZG extraction process,the refining process was optimized through macroporous resin column chromatography combined with single factor experiments.In the single factor experiment,the main influencing factors were investigated,including the concentration of the treatment solution,adsorption time,adsorbent concentration,adsorbent dosage,treatment solution concentration,and water washing dosage.When each factor is a variable in turn,the other factors are invariant for single factor experimental research.On the basis of optimizing the extraction and refining process of XBZG,a HPLC method was established for the determination of catalpol,aucubin,Leonurin,and cymdahoside A in XBZG.Perform a large amount of extraction and separation according to the above extraction,separation,and purification conditions to provide drugs for the following animal experiments.SD rats were injected intraperitoneally with 30mg· m L-1 STZ for modeling.72 hours later,a fasting blood glucose value of ≥16.7 mmo L indicates successful modeling.SD rats were divided into six groups: normal group,model group,XBZG low,medium,and high dose group,and metformin hydrochloride group.Starting from the second day,the XBZG low-dose group was given a dose of 108mg·kg-1,the medium dose group was given a dose of 216mg·kg-1,the high-dose group was given a dose of 432mg·kg-1,and the metformin hydrochloride group was given a dose of 36 mg · kg-1 of metformin hydrochloride according to the instructions.The normal group and model group were given an equal amount of physiological saline by gavage.Rats were continuously gavaged for 28 days,and on the 29 th day,blood was collected from the abdominal aorta,liver,kidney,pancreas,heart,and spleen.Fasting serum insulin(FINS)and insulin resistance index(HOMA-IR)in serum were detected by microplate reader,and total cholesterol(TC),triglyceride(TG),high-density lipoprotein(HDL-C)Low density lipoprotein cholesterol(LDL-C),glutamic oxaloacetic transaminase(AST)and glutamic transaminase(ALT)indicators.Oil red O staining was performed on the liver of rats.Western Blot method was used to detect peroxisome proliferator activated receptor alpha(PPAR-a)and peroxisome proliferator activated receptor alpha(PPAR-a)in liver γ Peroxisome proliferator activated receiver gamma,PPAR-γ)Protein expression.Results: The optimal extraction conditions for XBZG are as follows(see Figure2-10): ethanol concentration of 59.13%,extraction time of 59.24 minutes,solid-liquid ratio of 1:20.07,and yield of 27.70%.In order to investigate the reliability of the results obtained by the response surface method,the optimized extraction conditions were used to modify the parameter process.The ethanol concentration was 60%,the extraction time was 60 minutes,the solid-liquid ratio was 1:20,and the experiment was repeated three times,with an average value of 26.98%.The research results on the purification process of iridoid glycosides show that the optimal conditions for the concentration of treatment solution,adsorption time,adsorbent concentration,adsorbent dosage,treatment solution concentration,and water washing dosage are 50mg/m L,2 hours,20%,11 BV,4.0,and 7 BV,respectively.The validation experiment results of the optimized process showed that the adsorption rate reached 92.6%,the desorption rate was 68.6%,and the total content of iridoid glycosides was 59.7%.From factors such as the body mass and water intake of rats,XBZG can improve the body weight loss,excessive drinking and urination in T2 DM rats.The fasting blood glucose of the model group was significantly higher than that of the normal group,and the low,medium,and high dose XBZG groups had a reducing effect on fasting blood glucose.From the perspective of organ sensitivity index,the renal sensitivity index of the model group significantly increased compared to the normal group(P<0.01),while the renal sensitivity index of the low,medium,and high dose groups of total iridoid glycosides of Xinba significantly decreased compared to the model group(P<0.05).By detecting TC,TG,HDL-C,and LDL-C in serum,XBZG can reduce TC(P<0.05),TG(P<0.05),and LDL-C levels,while increasing HDL-C levels.The results of AST and ALT indicators were not statistically significant.The results of enzyme-linked immunosorbent assay(ELISA)showed that the XBZG administration group could reduce FINS and HOMA-IR levels(P<0.05).The oil red O staining results showed that a large area of lipid deposition was observed in the liver of the model group rats,and the liver tissue lipids of the low,medium,and high dose XBZG groups and positive control groups decreased.Western blot analysis showed that compared with the normal group,PPAR-a and PPAR in the liver tissue of the model group rats-γThe protein expression level significantly decreased(P<0.01).Compared with the model group,the expression levels of PPAR-a protein were significantly increased in the low XBZG(P<0.05),medium XBZG,and high-dose XBZG groups(P<0.01).The positive group showed no significant increase in PPAR-a protein expression level compared to the model group(P>0.05).Compared with the model group,the PPAR of the XBZG low-dose group(P<0.05)-γ The protein expression level significantly increased.PPAR of XBZG medium dose group,XBZG high dose group,and positive group-γ The increase in protein expression level was not significant(P>0.05).Conclusion: The content of XBZG was determined using UV spectrophotometry and high-performance liquid chromatography.Optimize the extraction process of XBZG using single factor experiments combined with response surface methodology.Separate and purify XBZG using macroporous adsorption resin and optimize the exquisite process.A HPLC method was established for the determination of catalpol,aucubin,Leonurin,and cymdahoside A in the total iridoid glycosides of Xinba.The pharmacological experiments explored the hypoglycemic mechanism of XBZG.The results showed that the total iridoid glycosides of Cymbaria dahurica L.can improve weight loss and polydipsia in T2 DM rats,reduce blood sugar,sensitivity index of kidneys and liver in T2 DM rats,and metabolize lipid components to lower blood sugar.The experimental results can provide experimental data for the in-depth research of Mongolian medicine Cymbaria dahurica L,and provide scientific basis for further development and utilization.