An efficient method for the preparative isolation and purification of alkaloids from <italic>Gelsemium</italic> by using high speed counter-current chromatography and preparative HPLC.

AbstractWe established an efficient method using high-speed countercurrent chromatography (HSCCC) combined with preparative high-performance liquid chromatography (prep-HPLC) for isolating and purifying Gels<italic>emium elegans</italic> (<italic>G. elegans</italic>) alkaloids. First, the two-phase solvent system composed of 1% triethylamine aqueous solution/<italic>n</italic>-hexane/ethyl acetate/ethanol (volume ratio 4:2:3:2) was employed to separate the crude extract (350 mg) using HSCCC. Subsequently, the mixture that resulted from HSCCC was further separated by Prep-HPLC, resulting in seven pure compounds including: 14-hydroxygelsenicine (<bold>1</bold>, 12.1 mg), sempervirine (<bold>2</bold>, 20.8 mg), 19-(R)-hydroxydihydrogelelsevirine (<bold>3</bold>, 10.1 mg), koumine (<bold>4</bold>, 50.5 mg), gelsemine (<bold>5</bold>, 32.2 mg), gelselvirine (<bold>6</bold>, 50.5 mg), and 11-hydroxyhumanmantenine (<bold>7</bold>, 12.5 mg). The purity of these seven compounds were 97.4, 98.9, 98.5, 99, 99.5, 96.8, and 85.5%, as determined by HPLC. The chemical structures of the seven compounds were analyzed and confirmed by electrospray ionization mass spectrometry (ESI-MS), 1H-nuclear magnetic resonance (1H NMR), and 13 C-nuclear magnetic resonance (13 C NMR) spectra. The results indicate that the HSCCC-prep-HPLC method can effectively separate the major alkaloids from the purified <italic>G. elegans</italic>, holding promising prospects for potential applications in the separation and identification of other traditional Chinese medicines. [ABSTRACT FROM AUTHOR]