硅胶柱色谱/RP-HPLC/LC-ESI-MS分离纯化鉴定拳卷地钱中芹菜素-7-O-β-D-葡萄糖醛酸苷

Isolation,Purification and Identification of apigenin-7-O-β-D glucuronide in Marchantia convoluta with Silica Column Chromatography,RP-HPLC and LC-ESI-MS

目的 :建立分离纯化鉴定拳卷地钱中芹菜素 -7-O -β -D -葡萄糖醛酸苷的硅胶柱色谱 /RP -HPLC/LC -ESI -MS方法。方法 :拳卷地钱叶经过 80 %乙醇提取后 ,减压蒸馏 ,得粗提物。拳卷地钱叶粗提物用甲醇溶解后上硅胶柱 ,用甲醇 -氯仿配比为 2 5∶1,2 5∶2 ,2 5∶3…… 2∶2 5 ,1∶2 5混合溶液洗脱 ,各洗脱液进行RP -HPLC分析 ,较纯的洗脱液进行LC -ESI-MS分析 ,为制备分离黄酮类化合物单体提供指导。结果 :氯仿与甲醇配比为 2 5∶15、2 5∶16、2 5∶ 17、2 5∶18的洗脱液经过RP -HPLC分析为单一组分 ,tR 为12 1min ,与对照品芹菜素 -7-O -β -D -葡萄糖醛酸苷共注射进行RP -HPLC实验 ,发现峰高增加 ,tR 为12 1min ,UV[λmax为 3 3 6/ 2 96(sh) / 2 67nm]和IR光谱与芹菜素 -7-O -β -D -葡萄糖醛酸苷基本一致。LC -ESI -MS测定结果表明 ,与对照品芹菜素 -7-O -β -D -葡萄糖醛酸苷的分子量相同为 44 6。结论 :由此可以鉴定该洗脱组分为芹菜素 -7-O -β -D -葡萄糖醛酸苷。

Objective :The methods of silica column chromatography,RP HPLC,UV,IR and LC ESI MS for isolating,purifying and identifying apigenin 7 O β D glucuronide in Marchantia convoluta was established in this article. Methods :The crude extracts of Marchantia convoluta leaves was obtained after Marchantia convoluta leaves were extracted with 80% ethanol and the extracts were concentrated under reduced pressure.The obtained crude extracts dissolved with methanol were subjected to silica gel column and eluted with the successive eluting solvent mixture of methanol chloroform.The eluted fraction was analyzed by RP HPLC.The further degree of purity were analyzed with UV,IR and LC ESI MS. Result :The elution chloroform methanol 25∶15,25∶16,25∶17,25∶18 were indicated to consist of only one kind of constituent by HPLC and UV,which retention time(t R) and λ max (nm)were 12 1 min and 336/296(sh)/267nm respectively.The HPLC experiment of the co-injections of this fraction with the standard of apigenin 7 O β D glucuronide resulted in an overlapping and accumulating effect with the similar retention time.The UV and IR spectra of the test solution were similar to the apigenin 7 O β D glucuronide.On the other hand,the LC ESI MS experiments indicated that the molecular weight is 446. Conclusion :So the fraction can be identified as apigenin 7 O β D glucuronide.