Pharmaceutical College, Nanjing University of Chinese Medicine, Nanjing 210023, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2013 Sep 15;935:70-4. doi: 10.1016/j.jchromb.2013.07.012. Epub 2013 Jul 26.
A method of using high-speed counter-current chromatography (HSCCC) was established for preparative isolation and purification of puerarin glycosides from the crude sample after enzymatic glycosylation of puerarin. Four fructosyl puerarins were successfully purified for the first time by HSCCC with a two-phase-solvent system composed of n-butanol-acetic acid-water (4:1.5:6, v/v/v). A total of 5mg of puerarin (1), 20mg of β-d-fructofuranosyl-(2→6)-puerarin (2), 41mg of β-d-difructofuranosyl-(2→6)-puerarin (3), 18mg of β-d-trifructofuranosyl-(2→6)-puerarin (4) and 15mg β-d-tetrafructofuranosyl-(2→6)-puerarin (5) were obtained in one-step separation from 100mg of the crude sample with purities of 98.5%, 98.3%, 98.9%, 97.8%, 97.5% and 97.2%, respectively. Among them, compounds 2-5 are novel compounds, and their chemical structures were identified by HRMS, (1)H NMR, (13)C NMR and 2D NMR. Pharmacokinetic analysis showed that β-d-fructofuranosyl-(2→6)-puerarin (2) was able to maintain higher plasma concentrations and have a longer mean residence time in the blood than puerarin.
建立了高速逆流色谱（HSCCC）方法，用于从葛根素酶糖化后的粗样品中制备分离和纯化葛根素苷。首次使用由正丁醇-乙酸-水（4：1.5：6，v/v/v）组成的两相溶剂系统，通过 HSCCC 成功地纯化了四种果糖基葛根素。从 100mg 的粗样品中，一步分离得到 5mg 的葛根素（1）、20mg 的β-d-果糖基-（2→6）-葛根素（2）、41mg 的β-d-二果糖基-（2→6）-葛根素（3）、18mg 的β-d-三果糖基-（2→6）-葛根素（4）和 15mg 的β-d-四果糖基-（2→6）-葛根素（5），纯度分别为 98.5%、98.3%、98.9%、97.8%、97.5%和 97.2%。其中，化合物 2-5 为新化合物，其化学结构通过高分辨质谱（HRMS）、（1）H NMR、（13）C NMR 和 2D NMR 进行了鉴定。药代动力学分析表明，β-d-果糖基-（2→6）-葛根素（2）在血液中的血浆浓度和平均滞留时间均高于葛根素。
