title: Comparative studies on performance of CCC and preparative RP-HPLC in separation and purification of steroid saponins fromDioscorea zingiberensisC.H.Wright.
abstract: Steroid saponins fromDioscorea zingiberensisC.H.Wright were separated for the first time using two chromatographic methods for comparison: counter-current chromatography (CCC) coupled with evaporative light scattering detector (ELSD) and preparative reversed phase high-performance liquid chromatography (RP-HPLC) with an ultraviolet detector. Ethyl acetate-n-butanol-methanol-water (4:1:2:4, v/v) was chosen as the two-phase solvent system for CCC, while the acetonitrile-water (25:75 for the first step and15:85 for the second step, v/v) was used as the mobile phase in the preparative RP-HPLC. The following five steroid saponins were purified by theses two chromatographic methods, in one-step operation by CCC and by two-step operation in preparative RP-HPLC: 1) 26-O-β-D- glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside (compound A), 2) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 4) 26-triol-3-O-[β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)]-β-D-glucopyranoside (compound B), 3) 26-O-β-D-glucopyranosyl-(25R)-furost-5-en-3β, 22ζ, 26-triol-3-O-[α-L-rhamnopyranosyl-(1→4)]-β-D-glucopyranoside (compound C), 4) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-{α-L-rhamnopyranosyl-(1→4)-[β-D-glucopyranosyl-(1→3)-β-D-glucopyranosyl-(1→2)]}-β-D-glucopyranoside (compound D) and 5) 26-O-β-D-glucopyranosyl-(25R)-furost-5, 20(22)-diene-3β, 26-diol-3-O-[β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosy-(1→2)]-β-D-glucopyranoside (compound E). The purities of these five steroid saponins separated by both methods were over 95%, and structural identification of these compounds was performed by ESI-MS, and13C NMR. Comparison of these two established approaches revealed that CCC required a longer separation time but with less solvent consumption, whereas preparative RP-HPLC gave a shorter separation time but with higher solvent consumption. These results demonstrated that either of these two methods of different separation mechanism is feasible, economical and efficient for rapid preparative isolation and purification of steroid saponins fromDioscorea zingiberensisC.H.Wright.
authors.affiliation: Biomedicine Key Laboratory of Shaanxi Province, Northwest University, 229 Taibai North Road, Xi'an, Shaanxi 710069, China.
authors.forename: Xinxin
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Biomedicine Key Laboratory of Shaanxi Province, Northwest University, 229 Taibai North Road, Xi'an, Shaanxi 710069, China.
authors.forename: Jinru
authors.initials: J
authors.lastname: Liang
authors.affiliation: Institut Parisien de Chimie Moléculaire, Université Pierre et Marie Curie-Paris 6, 4 place Jussieu, 75005 Paris, France.
authors.forename: Yongmin
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Biomedicine Key Laboratory of Shaanxi Province, Northwest University, 229 Taibai North Road, Xi'an, Shaanxi 710069, China.
authors.forename: Jianli
authors.initials: J
authors.lastname: Liu
authors.affiliation: Biomedicine Key Laboratory of Shaanxi Province, Northwest University, 229 Taibai North Road, Xi'an, Shaanxi 710069, China.
authors.forename: Wenji
authors.initials: W
authors.lastname: Sun
authors.affiliation: Laboratory of Bioseparation Technology, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Ito
keywords: CCC; Dioscorea zingiberensis C.H.Wright; preparative RP-HPLC; steroid saponins