Separation and purification of anthocyanins by high-speed countercurrent chromatography and screening for antioxidant activity

The all-liquid chromatographic technique of high-speed countercurrent chrom atography (HSCCC) has been applied for separations of anthocyanins. The bip hasic mixture of tert-butyl methyl ether/n-butanol/acetonitrile/water (2:2: 1:5) acidified with trifluoroacetic acid was found to be a suitable solvent system for anthocyanin separation. In some cases, enrichment of the pigmen ts on Amberlite XAD-7 resin prior to HSCCC has been carried out. The anthoc yanin mixtures from red cabbage, black currant, black chokeberry, and rosel le were successfully fractionated using HSCCC. Peak purity control was done by nuclear magnetic resonance spectroscopy as well as electrospray ionizat ion ion trap multiple mass spectrometry. Finally, antioxidant activity of t he purified pigments was determined using the Trolox equivalent antioxidant capacity test.