Analysis on Changes of Chemical Composition Contents and Efficacy of Improving Insulin Resistance in Type 2 Diabetic Mice of Anemarrhenae Rhizoma Before and After Processing with Salt-water / 中国实验方剂学杂志

ABSTRACT

Objective:By comparing the changing of chemical composition contents and the effects of improving insulin resistance in type 2 diabetic KKAy mice， to explore the processing principle of Anemarrhenae Rhizoma processed with salt-water. Method:Ultra-performance liquid chromatography-tandem mass spectrometry （UPLC-MS） was established for determining the contents of seven saponins and mangiferin in raw and salt-processed products of Anemarrhenae Rhizoma. The mobile phase was 0.1% formic acid aqueous solution （A） and acetonitrile （B） for gradient elution （0-1 min， 90%-80%A； 1-2 min， 80%-78%A； 2-5.5 min， 78%-70%A； 5.5-10.5 min， 70%-40%A； 10.5-12 min， 40%-20%A； 12-12.1 min， 20%-90%A； 12.1-13 min， 90%A）. The flow rate was set at 0.3 mL·min-1. The mass spectrographic analysis employed electrospray ionization （ESI） and negative ion acquisition mode. The acquisition range was m/z 100-1 200. The experimental type 2 diabetic KKAy mice were divided into model group， Anemarrhenae Rhizoma group （7.2 g·kg-1·d-1） and Anemarrhenae Rhizoma processed with salt-water group （7.2 g·kg-1·d-1）. C57BL/6J mice were considered as normal group and were given the same volume of saline. There are nine mice in each group， once a day for 21 consecutive days. The fasting blood glucose （FBG） was measured once a week. Three hours after the last administration， the blood samples of mice were collected by drawing eyeballs and were centrifuged to separate serum for further experiment. The fasting insulin （FINS）， leptin （LEP）， glycated hemoglobin （HbA1c）， glycated albumin （GA）， glucagon-like peptide-1 （GLP-1） levels in serum were detected by enzyme-linked immunosorbent assay （ELISA）. The insulin sensitivity index （ISI） and the homeostasis model assessment insulin-resistance （HOMA-IR） were calculated. The expressions of phosphatidylinositol 3-kinase （PI3K）， phosphoenolpyruvate carboxylkinase （PEPCK） and peroxisome proliferator activated receptor gamma co-activator1 （PGC1） mRNA in hepatic and adipose tissue of mice from each group were detected by real-time fluorescence quantitative polymerase chain reaction method （Real-time PCR）. Result:After being processed with salt-water， the contents of 8 chemical components in Anemarrhenae Rhizoma were increased， among which the contents of timosaponin AⅢ， timosaponin BⅡ， timosaponin BⅢ， anemarrhenasaponin Ⅰ， anemarrhenasaponin Ⅰa， mangiferin were significantly increased， and increased by 43.78%， 38.77%， 25.84%， 28.21%， 22.51%， 24.04%， respectively. Compared with the model group， raw and salt-processed products of Anemarrhenae Rhizoma could significantly decrease the levels of FBG， FINS， HOMA-IR， HbA1c， LEP， GA （P<0.05， P<0.01）， increase the levels of ISI， GLP-1 （P<0.05， P<0.01） in serum of mice with type 2 diabetes， and significantly increase the expression of PI3K and PGC1 mRNA in hepatic and adipose tissue （P<0.05）. It is worth noting that salt-processed products is better than that of raw products. Conclusion:Raw and salt-processed products of Anemarrhenae Rhizoma have obvious hypoglycemic effect. And the hypoglycemic effect of Anemarrhenae Rhizoma can be promoted after being processed with salt-water by promoting insulin secretion and improving insulin resistance. Incremental components are the probably material basis for enhancement of hypoglycemic effect of Anemarrhenae Rhizoma after being processed with salt-water.