Rational approach to solvent system selection for liquid-liquid extraction-assisted sample pretreatment in counter-current chromatography

•A rational method to solvent selection for liquid-liquid extraction was established.•The solvent selection based on K values of target compounds.•The impurities can be removed with the lowest loss rates of target compounds.•The separation of two target flavonoids from the extract was presented as an example.•The present method was efficiency to raise yields of HSCCC separation.

A rational liquid-liquid extraction approach was established to pre-treat samples for high-speed counter-current chromatography (HSCCC). n-Hexane-ethyl acetate-methanol-water (4:5:4:5, v/v) and (1:5:1:5, v/v) were selected as solvent systems for liquid-liquid extraction by systematically screening K of target compounds to remove low- and high-polarity impurities in the sample, respectively. After liquid-liquid extraction was performed, 1.4 g of crude sample II was obtained from 18.5 g of crude sample I which was extracted from the flowers of Robinia pseudoacacia L., and then separated with HSCCC by using a solvent system composed of n-hexane-ethyl acetate-methanol-water (1:2:1:2, v/v). As a result, 31 mg of robinin and 37 mg of kaempferol 7-O-α-l-rhamnopyranoside were isolated from 200 mg of crude sample II in a single run of HSCCC. A scale-up separation was also performed, and 160 mg of robinin with 95% purity and 188 mg of kaempferol 7-O-α-l-rhamnopyranoside with 97% purity were produced from 1.2 g of crude sample II.