Efficient methods for isolating five phytochemicals from Gentiana macrophylla using high‐performance countercurrent chromatography

Abstract

Efficient high-performance countercurrent chromatography methods were developed to isolate five typical compounds from the extracts of Gentiana macrophylla. n-Butanol-soluble extract of G. macrophylla contained three hydrophilic iridoids, loganic acid (1), swertiamarin (2) and gentiopicroside (3), and a chromene derivative, macrophylloside D (4) which were successfully isolated by flow rate gradient (1.5 mL/min in 0-60 min, 5.0 mL/min in 60-120 min), and consecutive flow rate gradient HPCCC using n-butanol/0.1% aqueous trifluoroacetic acid (1:1, v/v, normal phase mode) system. The yields of 1-4 were 22, 16, 122, and 6 mg, respectively, with purities over 97% in a flow rate gradient high-performance countercurrent chromatography, and consecutive flow rate gradient high-performance countercurrent chromatography gave 1, 2, 3 (54, 41, 348 mg, respectively, purities over 97%) and 4 (13 mg, purity at 95%) from 750 mg of sample. The main compound in methylene chloride soluble extract, 2-methoxyanofinic acid, was successfully separated by n-hexane/ethyl acetate/methanol/water (4:6:4:6, v/v/v/v, flow-rate: 4 mL/min, reversed phase mode) condition. The structures of five isolates were elucidated by H, C NMR and ESI-Q-TOF-MS spectroscopic data which were compared with previously reported values.

Application of off‐line two‐dimensional high‐performance countercurrent chromatography on the chloroform‐soluble extract of Cuscuta auralis seeds

“…CCC boasts several advantages over conventional, solid‐based column chromatography, including the nonadsorption of target molecules into the stationary phase, high‐recovery rate, and short run times. Therefore, CCC methods have been widely applied to isolate natural products from medicinal plants and agricultural crops .…”

An overview of the two-phase solvent systems used in the countercurrent separation of phenylethanoid glycosides and iridoids and their biological relevance

“…The least polar system t-butyl methyl etheracetonitrile-water (30:30:40) was used for the isolation of non-glycosidic iridoids. Some variations in the mode of operation of the CCC and CPC columns were observed during the survey, including step gradient elution (de Juliao et al 2010;Leitao et al 2015), enhancement of flow rate (Liang et al 2013b;Rho et al 2016), two-step CCS (Lei et al 2001b;Lemus et al 2015;Li et al 2008), elution-extrusion CCS (Chen et al 2011;Liang et al 2018) and recycling CCS (Chen et al 2011).…”

“…A mixture of n-butanol-0.1% (aq) trifluoroacetic acid (1:1) was successfully employed for the isolation of loganic acid, gentiopicroside and swertiamarin from Gentiana macrophylla Pall. under consecutive flow rate gradient conditions (from 1.5 to 5 mL/min) (Rho et al 2016).…”

Analytical Methods of Phytochemicals from the Genus Gentiana

“…Subsequently, through purification of semi-preparative HPLC from Fra2, Fra3, and Fra4, which were produced from HPCCC, Compounds 4 , 9 , 8 and 42 were obtained. Besides, Compounds 4 , 8 , 1 and 42 of the extracts of G. macrophylla were also successfully isolated by HSCCC using an n -butanol/0.1%aqueous trifluoroacetic acid (1:1) system [ 60 , 134 ]. A one-step HSCCC method was established to separate 8-hydroxy-10-hydrosweroside, compound 8 and 15 from the crude extract of G. scabra .…”