Isolation and purification of three flavonoid glycosides from the leaves of Nelumbo nucifera (Lotus) by high-speed counter-current chromatography

Semi-preparative high-speed counter-current chromatography (HSCCC) was successfully used for isolation and purification of flavonoid glycosides from the leaves of Nelumbo nucifera (Lotus) by using a two-phase-solvent system composed of n-hexane–ethyl acetate–methanol–water (1:5:1:5, v/v/v/v). The targeted compounds isolated, collected and purified by HSCCC were analyzed by high performance liquid chromatography (HPLC). A total of 4.6 mg of isoquercitrin, 9.1 mg of hyperoside and 3.0 mg of astragalin with the purity of 95.8%, 97.5% and 98.3%, respectively, were obtained in one-step separation and less than 6 h from 80 mg of crude extract from the leaves of N. nucifera. The chemical structures of all the three compounds were identified by MS, 1H NMR, 13C NMR. Astragalin was obtained from N. nucifera for the first time.