甘草有效成分的提取及协同氟康唑抗真菌活性研究

Extraction of active ingredients from roots of Glycyrrhiza uralensis Fisch.and their synergistic antifungal activity with fluconazole

目的:研究甘草中主要活性成分三萜皂苷和黄酮类化合物的抗真菌活性,拓展甘草在抗真菌治疗中的应用。方法:通过硅胶柱层析、凝胶柱层析技术从甘草粗提物中分离得到甘草查尔酮A、异甘草素、甘草素和甘草酸4种成分,甘草酸水解得到甘草次酸。5个单体化合物经LC-MS、1 H-NMR和13 C-NMR鉴定。采用微量稀释法分别测定5个化合物体外协同氟康唑抗耐药白念珠菌0304103、0304385、0304876,热带念珠菌0306849,克鲁斯念珠菌ATCC19297的最低抑菌浓度(MIC_(80))和部分抑菌浓度指数(FICI)。结果:甘草查尔酮A、甘草次酸和异甘草素可协同氟康唑抗耐药白念珠菌0304103、0304385、0304876,FICI均<0.5,其中甘草查尔酮A联合氟康唑抗真菌效果最为显著。但5种单体化合物不具有协同氟康唑抗热带念珠菌或克鲁斯念珠菌的活性。结论:甘草中的甘草查尔酮A、甘草次酸和异甘草素具有协同氟康唑抗白念珠菌活性。

Objective：To investigate the synergistic antifungal activities of triterpenoid saponins and flavonoids fromGlycyrrhiza uralensis Fisch.,so as to expand the application of Glycyrrhiza uralensis Fsich.in antifungal therapy.Methods：Four chemical constituents（licochalcone A,isoliquiritigenin,liquiritigenin and glycyrrhizinic acid）were isolated from the crude extract of G.uralensis by silica gel chromatography and gel column layer chromatography.Glycyrrhetic acid was obtained by hydrolysis of glycyrrhizinic acid.Then,structures of the five compounds were all confirmed by LC-MS,1 H-NMR and 13 C-NMR.Microdilution test was performed to identify the minimal inhibitory concentration（MIC80）and fractional inhibitory concentration index（FICI）of the five active compounds against fluconazole-resistant Candida albicans 0304103,0304385,0304876,Candida tropicalis 0306849 and Candida kruseii ATCC19297 in vitro.Results：Licochalcone A,glycyrrhetic acid and isoliquiritigenin exhibited synergistic antifungal activities against fluconazole-resistant C.albicans 0304103,0304385,0304876 and the FICI values were less than 0.5.Of all the constituents,licochalcone A showed the most potent synergistic antifungal activity,however,the five compounds didn＇t display any antifungal effects against C.tropicalis 0306849 or C.kruseii ATCC19297.Conclusion：Licochalcone A,glycyrrhetic acid and isoliquiritigenin exhibited synergistic antifungal activities combined with fluconazole against C.albicans.