芪葛汤有效成分降脂的体外初筛实验

Preliminary Screening of Effective Components of Qige Decoction with Lipid-lowering Effect

【目的】体外筛选芪葛汤降脂的有效成分。【方法】采用300μmol/L油酸+150μmol/L棕榈酸+100μmol/L亚油酸体外构建大鼠BRL肝细胞脂质堆积模型,体外筛选黄芪甲苷、葛根素、川陈皮素、3’-羟基葛根素、毛蕊异黄酮、柚皮素、槲皮素等7个芪葛汤的有效成分。以四甲基偶氮唑盐(MTT)实验确定芪葛汤有效成分对肝细胞的毒性剂量。预给药干预48 h,分别使用氯仿与甲醇、异丙醇抽提细胞脂质,检测甘油三酯(TG)含量,采用油红O染色法观察细胞脂质堆积情况。【结果】经MTT实验确定100μmol/L黄芪甲苷、1 mmol/L葛根素、40μmol/L川陈皮素、500μmol/L 3’-羟基葛根素、100μmol/L毛蕊异黄酮、100μmol/L柚皮素、20μmol/L槲皮素为干预肝细胞的无毒性剂量。氯仿与甲醇抽提脂质发现100μmol/L黄芪甲苷、1 mmol/L葛根素、40μmol/L川陈皮素、100μmol/L柚皮素能降低肝细胞TG含量。而异丙醇抽提脂质结果发现100μmol/L黄芪甲苷、40μmol/L川陈皮素、100μmol/L柚皮素、20μmol/L槲皮素能降低肝细胞TG含量。油红O染色结果提示模型组细胞大量脂质堆积,而黄芪甲苷,川陈皮素、柚皮素的脂质堆积稍有减少,其他组则改善不明显。【结论】芪葛汤有效成分黄芪甲苷、川陈皮素、柚皮素可降低大鼠BRL肝细胞脂质堆积。

Objective To screen the effective lipid-lowering components of Qige Decoction in vitro. Methods Firstly,lipid accumulation model of Buffalo rat liver(BRL)cells was established by using 300 μmol/L oleic acid,150 μmol/L palmitic acid and 100 μmol/L linoleic acid in vitro. In vitro,7 active ingredients of Qige Decoction including astragaloside Ⅳ,puerarin,nobiletin,3’-hydroxy puerarin,calycosin,naringenin,quercetin were screened. methyl-thiazolyl-tetrazolium(MTT)assay was used to determine the toxic dose of active ingredients of Qige Decoction for liver cells. After 48 hours of pre-administration intervention,chloroform and methanol,and isopropanol were used to extract cell lipids,respcetively,and oil red O staining method was used to observe the accumulation of cell lipids. Results MTT assay results showed that 100 μmol/L of astragalosideⅣ,1 mmol/L of puerarin,40 μmol/L of nobiletin,500 μmol/L of 3’-hydroxy puerarin,100 μmol/L of calycosin,100 μmol/L of naringenin,20 μmol/L of quercetin were non-toxic doses for intervening liver cells. The results of chloroform and methanol extraction showed that 100 μmol/L of astragaloside Ⅳ,1 mmol/L of puerarin,40 μmol/L of Nobiletin and 100 μmol/L of naringin reduced the content of TG in liver cells. Isopropanol extraction results showed that 100μmol/L of astragaloside Ⅳ,40 μmol/L of nobiletin,100 μmol/L of naringin and 20 μmol/L of quercetin reduced the TG content of liver cells. Oil red O staining results showed that a large amount of lipid accumulation in the model group,and the lipid accumulation in astragaloside Ⅳ,citrinin and naringin groups was slightly reduced,however the improvement was not obvious in the other groups. Conclusion The effective components such as qigedecoction,astragaloside Ⅳ,tangerinin and naringin is effective for reducing lipid accumulation in BRL cells.