葡萄糖氧化酶法筛选西洋参干预胰岛素抵抗脂肪细胞模型的活性成分

Screen the Active Constituents that American Ginseng Impact the Model of Adipocyte Insulin Resistance by Glucose Oxidase Method

目的:通过葡萄糖消耗试验测定西洋参单体成分对IR脂肪细胞模型葡萄糖消耗量的影响,筛选西洋参干预胰岛素抵抗的活性成分。方法:将西洋参的根部用70%EtOH回流,将提取物进行AB-8型大孔树脂柱色谱分析,按不同浓度洗脱,得到西洋参95%醇提组、西洋参60%醇提组、西洋参30%醇提组和水提组四个部分。本课题组的前期药效学试验已证明,西洋参60%醇提组为活性部位。将西洋参60%醇提部位用硅胶柱色谱及HPLC等现代分离技术与方法分离和纯化,通过理化常数及光谱数据确定其结构,共得12个化合物。将3T3-L1前脂肪细胞诱导分化为成熟的脂肪细胞,给予高糖高胰岛素(25mmol·L-1葡萄糖,1×10-6mol·L-1胰岛素)刺激,建立胰岛素脂肪细胞模型,随机分为空白组、模型组、罗格列酮组、西洋参单体成分组。药物干预培养24h后,通过葡萄糖消耗试验测定西洋参单体成分对IR脂肪细胞模型葡萄糖消耗量的影响,筛选西洋参干预胰岛素抵抗的有效成分。结果:与模型组细胞葡萄糖消耗量比较,罗格列酮组、人参皂苷R3组、20(R)人参皂苷Rb1组、20(R)人参皂苷Rb2组、拟人参皂苷F11组细胞葡萄糖消耗量有不同程度的升高,其中人参皂苷R3组、20(R)人参皂苷Rb1组、20(R)人参皂苷Rb2组、罗格列酮组葡萄糖消耗量明显升高,差异极显著(P<0.01),拟人参皂苷F11组差异显著,(P<0.05)。其余化合物组与模型组比较,葡萄糖消耗量无统计学差异。结论:20(R)人参皂苷Rb1,20(R)人参皂苷Rb2,人参皂苷R3,拟人参皂苷F11可能是西洋参干预IR脂肪细胞模型的活性成分。

Objective：To screen the the active constituents that American Ginseng impact the model of adipo-cyte insulin resistance by Glucose consuming experiment showed the compound constituents significantly effect the glucose mechanism in adipocyte cells. Methods：The roots of American Ginseng is extract with 70% EtOH to get extractive. Then extractive is eluted according to different concentrations by AB -8 type macroporous resin column chromatography to get Ginseng 95% alcoholic extraction group,Ginseng 60% alcoholic extraction group,Ginseng 30% alcoholic extraction group and aqueous extract group. It is proved that Ginseng 60% alcoholic extraction group and is effective component by pharmacodynamic study. Then the 60% alcoholic ex-traction group isolated by silica gel column chromatography and HPLC preparation chromatography. Their structures were determined by physical and chemical properties ,as well as spectrographical identification. There are12 compounds. Later,effect the preadipocyte cells by high glucose and high insulin in order to modelling adipocyte insulin resistance. Then divided randomly into control group,model group ,rogridone group ,compound constituents of Ginseng group. Screening the the active constituents that American Ginseng impact the model of adipocyte insulin resistance by Glucose consuming experiment showed the compound constituents significantly effect the glucose mechanism in adipocyte cells. Compared with model group,other group rats GLUT -4 mRNA expression is escalated. There was an extreme difference between roglitazone group,ginsenoside R3 group、20（R）ginsenoside Rb1 group、20（R）ginsenoside Rb2 group and model group（P 0. 01）. There was a significant difference between pseudoginsenoside F11 group and model group（P 0. 05）. Conclusion：Ginsen-oside R3、20（R）ginsenoside Rb1 group、20（R）ginsenoside Rb2、pseudoginsenoside F11 may be the active constituents.