A rational route based on liquid-liquid-refining extraction and high-speed counter-current chromatography for separation of target compound from Toona sinensis

Abstract

Gallotannin, 1,2,3,4,6-penta-O-galloyl-β-d-glucose (PGG), exhibited a wide range of biological activities. In the present study, a method combined liquid-liquid refining extraction and high-speed counter-current chromatography was established for improving the separation yield of PGG from Toona sinensis. Liquid-liquid refining extraction was used to enrich the target compound, the impurities with high polarity and low polarity can be removed with appropriate two-phase solvent systems, and a series of equations were developed to determine the times of extraction and loss rate of target compound. As a result, the two solvent systems composed of n-hexane–ethyl acetate–methanol–water (1:2:1:2, v/v) and ethyl acetate–water (1:1, v/v) were selected to remove the low polarity impurities and high polarity impurities, respectively, and 78.08 % by weight of the crude extract was removed, while the loss rate of PGG was only 12.09 % after extraction. Then, the PGG enriched samples were further separated by high-speed counter-current chromatography with a two-phase solvent system n-hexane–ethyl acetate–methanol–water (0.4:5:0.4:5, v/v). PGG (75 mg) with purity of 97.33 % was isolated from 350 mg of the PGG enriched samples. This study proved that the liquid-liquid refining extraction was a very effective method to enrich the target compound from complex mixture and a good assistant for large-scale preparation of natural products.