Rapid Separation of Three Proanthocyanidin Dimers from Iris lactea Pall. var. Chinensis (Fisch.) Koidz by High‐Speed Counter‐Current Chromatography With Continuous Sample Load and Double‐Pump Balancing Mode

Introduction - The dried seeds of Iris lactea have been used in traditional Chinese medicine. Previous studies have been focused on irisquinones while other chemical components are rarely reported. Objective - To establish an efficient high-speed counter-current chromatography (HSCCC) separation method with continuous sample load (CSL) and double-pump balancing (DPB) mode to isolate proanthocyanidins from I. lactea. Methods - Firstly, an ethyl acetate extract of I. lacteawas pre-fractionated by silica column chromatography for the enrichment of proanthocyanidins. Secondly, the enriched proanthocyanidins sample ( EPS) was further fractionated by HSCCC with a two-phase solvent system ethyl acetate: n- butanol: water ( 9: 1: 10, v/ v/ v) using DPB mode. The flow rate of the two phases was 2.2mL/ min, the revolution speed was 900 rpm, the separation temperature was 30 degrees C and the detection wavelength was 280nm. Finally, the structures of the three isolated proanthocyanidins were elucidated by spectroscopic methods and compared with published data. Results - Under the optimized conditions, 600mg of the EPS with six continuous injections ( 100mg/ time) was fractionated, yielding 57mg of prodelphinidin B3, 198mg of procyanidin B3, and 162mg of procyanidin B1, at purities of 97.2%, 98.1% and 97.3%, respectively. Conclusions - The HSCCC separation method with CSL and DPB proved to be rapid, convenient and economical, constituting an efficient strategy for the isolation of proanthocyanidins. Copyright (c) 2015 John Wiley & Sons, Ltd.