Preparative Separation of Anthocyanins from Purple Sweet Potatoes by High-Speed Counter-Current Chromatography

Abstract

Abstract A two-step high-speed counter-current chromatography (HSCCC) method was successfully developed to separate and purify anthocyanins from purple sweet potatoes. In the first step, a mixture of two anthocyanin compounds was separated from the extract of purple sweet potatoes by HSCCC. A two-phase solvent system composed of n-butanol/ethyl acetate/0.5% acetic acid (3:1:4, V/V) was used with the upper phase as the stationary phase and the lower phase as the mobile phase at a flow rate of 2.0 mL min−1 and an injection volume of 300 mg. Subsequently, the two compounds were further separated and purified by a refining HSCCC run, with a two-phase solvent system of 0.2% trifluoroacetic acid/n-butanol/methyl tertiary butyl ether/acetonitrile (6:5:2:1, V/V) with the upper phase as the stationary phase and the lower phase as the mobile phase at a flow rate of 1.5 mL min−1 and an injection volume of 100 mg. By this two-step HSCCC method, 63 mg of compound 1 was obtained from 450 g of purple sweet potatoes with a purity of 98.5%, and 48 mg of compound 2 was obtained with a purity of 96.7%. Furthermore, the chemical structures of these two compounds were analyzed by UV-VIS, ESI-MS/MS, and NMR. Compound 1 was identified as peonidin 3-O-{6-O-(E)-caffeyl-2-O-[6-O-(E)-p- hydroxybenzoy-β-D-glucopyranoside]-β-D-glucopyranoside}-[5-O-(β-D-glucopyranoside)], and compound 2 was identified as peonidin 3-O-{6-O-(E)-caffeyl-2-O-[6-O-(E)-ferulyl-β-D-glucopyranoside]-β-D-glucopyranoside}-[5-O-(β-D-glucopyranoside)]. The two-step HSCCC method is a stable, efficient, and reliable technique for the isolation of anthocyanins from purple sweet potatoes.