Isolation of Anthocyanin from Hibiscus sabdariffa by High Speed Counter-current Chromatography
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Abstract:
The calyx of Hibiscus sabdariffa was extracted with 40% ethanol at room temperature, and poured through D101 resin. A solvent system of water-n-butanol-methyl tert- acetonitrile-trifluoroacetic acid (6:3:1:1:0.001, V/V) was used for isolation and purification by high speed counter-current chromatography (HSCCC) using upper phase as stationary phase and lower phase as mobile phase ,with the flow rate of 2 mL/min and injection volume of 120 mg. 21.8 mg anthocyanin 1 power was obtained with the purity and the recovery rate being of 97.8% and 95.3%, respectively. Anthocyanin 2 was isolated twice with purity of 96.2% (5.3 mg) and recovery rate of 92.3%. The chemical structures of the two anthocyanins were identified as delphindin-3-O-sambubioside for anthocyanin 1 and cyanidin-3-O-sambubioside for anthocyanin 2 by ESI-MS and NMR. The method is simple, reproducible, which is suitable for preparation of anthocyanins from Hibiscus sabdariffa, and also provides basis for further pharmacological studies and quality control of anthocyanins
