知母盐制前后化学成分含量及改善2型糖尿病小鼠胰岛素抵抗作用的差异分析  

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

作　　者：郑威 闫丽 高慧[1] ZHENG Wei;YAN Li;GAO Hui(Chinese Materia Medica Processing Engineering arid Technology Center of Liaoning Province,Key Laboratory of Processing Principle Analysis,National Administration of Traditional Chinese Medicine(TCM),School of Pharmacy,Liaoning University of TCM,Dalian 116600,China)

机构地区：[1]辽宁中医药大学药学院,国家中医药管理局炮制原理解析重点研究室,辽宁省中药炮制工程技术研究中心,辽宁大连116600

出　　处：《中国实验方剂学杂志》2020年第22期140-147,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81102810);辽宁省自然科学基金项目(2018010410-30)。

摘　　要：目的:通过比较知母盐制前后化学成分的含量变化,以及对2型糖尿病KKAy小鼠胰岛素抵抗的影响,探讨知母盐制后降糖作用增强的炮制原理。方法:采用超高效液相色谱-串联质谱法(UPLC-MS)测定生知母、盐知母中7种知母皂苷及芒果苷的含量,色谱条件设定为流动相0.1%甲酸水溶液(A)-乙腈(B)梯度洗脱(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),流速0.3 mL·min-1;质谱条件为电喷雾离子源(ESI),负离子采集模式,采集范围m/z 100~1200。建立2型糖尿病KKAy小鼠模型,将成模小鼠随机分为模型组、生知母组、盐知母组,另设C57BL/6J小鼠作为空白组,每组9只。各给药组灌胃给予相应药液(7.2 g·kg-1·d-1),空白组、模型组给灌胃同体积生理盐水,每日1次,连续21 d。每周检测各组小鼠空腹血糖(FBG),于最后1次给药3 h后眼球取血,利用酶联免疫吸附法(ELISA)检测各组小鼠血清中的空腹胰岛素(FINS),瘦素(LEP),糖化血红蛋白(HbA1c),糖化白蛋白(GA)和胰高血糖素样肽-1(GLP-1)水平,计算胰岛敏感指数(ISI)和胰岛素抵抗指数(HOMA-IR);采用实时荧光定量聚合酶链反应法(Real-time PCR)检测各组小鼠肝组织和脂肪组织中磷脂酰肌醇3-激酶(PI3K),磷酸烯醇式丙酮酸羧激酶(PEPCK),过氧化物酶体增殖物激活受体γ共激活因子1(PGC1)mRNA的表达。结果:知母经盐制后,所测的8种化学成分含量均有升高,其中知母皂苷AⅢ,知母皂苷BⅡ,知母皂苷BⅢ,知母皂苷Ⅰ,知母皂苷Ⅰa和芒果苷的含量升高显著,其含量依次增加了43.78%,38.77%,25.84%,28.21%,22.51%,24.04%。生知母、盐知母均能明显降低2型糖尿病小鼠FBG,FINS,HOMA-IR,HbA1c,LEP和GA水平(P<0.05,P<0.01),均能明显升高ISI和GLP-1水平(P<0.05,P<0.01),均能明显上调肝组织及脂肪组织中PI3K,PGC1 mRNA的表达(P<0.05),且盐知母作用优于生知母。结论:生知母、盐�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 100-1200.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 coactivator1(PGC1)mRNA in hepatic and adipose tissue of mice from each group were detected by real-time fluorescence quantitative polymerase chain reaction method(Re

关 键 词：知母 炮制 化学成分 2型糖尿病小鼠 胰岛素抵抗 降血糖 物质基础 

分 类 号：R587.1[医药卫生—内分泌] R943.1[医药卫生—内科学]