Preparative isolation and purification of isobenzofuranone derivatives and saponins from seeds of Nigella glandulifera Freyn by high-speed counter-current chromatography combined with gel filtration

Although the medicinal plant and food Nigella glandulifera Freyn has been researched for decades, isobenzofuranones have never been isolated before. Two isobenzofuranone derivatives and two saponins were Successfully separated and Purified from seeds of N. glandulifera Freyn by high-speed counter-current chromatography (HSCCC) with the optimized two-phase solvent system, n-hexane-ethyl acetate-methanol-water (7:3:5:5, v/v). Salfredin B-11 (22.1 mg, HPLC Purity 95.3%), 5, 7-dihydroxy-6-(3-methybut-2-enyl) isobenzofuran-1(3H)-one (18.9 mg, HPLC purity 97.3%) and crude sample 2 (555 mg) were separated from 600 mg of ethyl acetate extract of N. glandulifera Freyn. Following a cleaning-up step by chromatography on Sephadex LH-20, hederagenin (12 mg) and 3-O-[beta-D-xylopyranosyl-(1 -> 3)-alpha-L-rhamnopyranosyl-(1 -> 2)-alpha-L-arabinopyranosyl]-hederagen in (45 mg) were separated from sample 2. All of the fractions before peak 11 were collected and Subjected to a Sephadex LH-20 column and eluted by methanol, two of triterpene saponins (12 mg of hederagenin and 45 mg of 3-O-[beta-D-xylopyranosyl-(1 -> 3)-alpha-L-rhamnopyranosyl-(1 -> 2)-alpha-L-arabinopyranosyl]-hederagenin) were isolated. The structures of peak fractions were identified by IR, electron ionization MS. H-1 NMR and C-13 NMR. 5, 7-Dihydroxy-6-(3-methybut-2-enyl) isobenzofuran-1(3H)-one was isolated for the first time from higher plant and salfredin B11 was isolated for the first time in this plant. (C) 2009 Elsevier B.V. All rights reserved.