A new method for separation of two saponins from Celosiae Semen

Objective:To develop an efficient method for the purity(≥98%) of celosin C and celosin D from Celosiae Semen by high-speed counter-current chromatography(HSCCC) coupled with evaporative light scattering detection(ELSD).Methods :A two-phase solvent system comprised n-butanol-ethyl acetate-methanol-water(3.5∶3.5∶0.6∶10,v/v/v/v) with addition of 0.5%glacial acetic acid was developed by using a computer-aided counter-current chromatography solvent-selection software(CCC-SSS) for separate preparation of the components of the solvent system.Each peak fraction obtained from separation of main saponins was collected according to the ELSD elution profile and determined by HPLC.Results:Using the described method,two saponins,all with purities of over 98%,could be isolated from 150 mg of the main saponins.Their structures were characterized by ESI-MS and 13C-NMR.Conclusions:HSCCC coupled with ELSD and CCC-SSS successfully solved the problem of two saponins' purity for the first time.It is also proved to be a feasible method for separating components without UV absorption from natural products.