| Abstract: | 本研究以原產於中國大陸內蒙古地區之甘草(licorice, Glycyrrhiza uralensis Fisch. rhizomes)為材料，經不同溶劑萃取，選擇對醣解酵素α-amylase及α-glucosidase活性最佳抑制能力的50%乙醇萃取物及其純化活性成分，利用小鼠FL83B肝細胞株進行細胞試驗，探討不同濃度甘草萃取物及純化物質對細胞之毒性以及對細胞葡萄糖擬似物攝入量及肝醣合成量之促進活性。首先將甘草分別以乙醇、50%乙醇以及熱水萃取，測定各萃取物對醣解酵素活性的抑制能力。結果顯示50%乙醇萃取物對兩種酵素具較佳抑制效果，對α-amylase及α-glucosidase抑制率分別為15.64%及51.05%。以此萃取物依序經Amberlite XAD-7、Cosmosil 75 C18-OPN及HPLC等液相層析分離純化後，以紫外-可見光吸光、質譜及1H、13C核磁共振等光譜分析，鑑定出主要組成分為 4'-hydroxy-flavanone-7-O-glucoside (1)、4-O-β-D- glucopyranosyl- 2', 4'-dihydroxy-chalcone (2) 、 4',7-dihydroxy flavanone (3)、 ononin (4)、7-hydroxy-4'-methoxy-isoflavone (5)等5種類黃酮。其中以純化物質2、 3、5的含量較多，而醣解酵素抑制活性測試結果顯示，純化物質2的抑制率最佳，因此被選擇進行後續之細胞試驗。在XTT細胞毒性測試中，添加200 ppm甘草萃取物以及純化物質2均未對小鼠肝臟FL83B細胞產生毒性。在添加50、100及200 ppm甘草50%乙醇萃取物及其純化物質2時，葡萄糖擬似物(2-NBDG)細胞的帶入量及細胞內肝醣合成量均提升。本研究結果初步顯示甘草50%乙醇萃取物可能藉由降低腸胃道中糖解酵素作用及增加肝中葡萄糖帶入與儲存，協助細胞調節葡萄糖之恆定性。
Licorice (Glycyrrhiza uralensis Fisch) is a legume that is mainly grown in southern Asia and Europe. Licorice, also known as sweet root, is one of the most widely used medicinal herbs in traditional oriental medicine, where it has been used to treat bronchitis, coughs, arthritis, adrenal insuf?ciency and allergies. In this study, the active components were isolated and identified from the 50% ethanol licorice extract, and were tested for their hypoglycemic activities in vitro. 50% ethanol licorice extracts showed strong inhibitory properties against both α-amylase and α-glucosidase. The extract was further separated successively by Amberlite XAD-7, Cosmosil 75 C18-OPN, and reversed HPLC chromatographies to obtain five components. The isolated components were identified as 4'-hydroxy-flavanone-7-O-glucoside (1), 4-O-β-D-glucopyranosyl-2',4'- hydroxy-chalcone (2), 4',7-dihydroxyflavanone (3), ononin (4) and 7-hydroxy-4'-methoxy-isoflavone (5) by MS, UV, and 1H,13C-NMR analyses. Among them, the isolated component 2 showed the most contents and the strongest inhibitory activitis toward α-amylase and α-glucosidase, with inhibition rates of 52.55% and 31.00% at the concentration of 1mg/ml. Hypoglycemic activities of the 50% ethanol licorice extract and isolated component 2 were further verified in a murine FL83B hepatocyte line. Both the 50% ethanol licorice extract and isolated component 2 had no inhibitory effects on the growth of FL83B cells at the concentration of 200 ppm by the XTT assays. In the presence of the extract or isolated component, uptake of glucose analogue 2-NBDG was significantly enhanced when FL83B cells were rendered resistant to insulin by a prior incubation with high concentration of glucose. Glycogen contents both in normal and in insulin resistant cells were also increased by adding the 50% ethanol licorice extrat or isolated component 2 in a dose-dependent manner. Our results demonstrated the 4-O-β-D-glucopyranosyl- 2',4'- hydroxy-chalcone from licorice may inhibit α-amylase and α-glucosidase, and promotes glucose uptake and storage in hepatocytes, implying a hypoglycemic activity in vivo. |
