Large-scale separation of alkaloids from Corydalis bungeana Turcz. by pH-zone-refining counter-current chromatography

pH-Zone-refining counter-current chromatography (pH-zone-refining CCC) was successfully applied for the large-scale separation of alkaloids from Corydalis bungeana. The crude extract was separated by a two-phase solvent system composed of petroleum ether-ethyl acetate-methanol-water (5:5:2:8, v/v) where triethylamine (10 mM) was added to the upper organic stationary phase as a retainer and hydrochloric acid (5 mM) to the aqueous mobile phase as a displacer. As a result, 285 mg of protopine, 86 mg of corynoloxine, 430 mg of coryno1ine, and 115 mg of acetylcorynoline were obtained from 3.0 g of crude extract in a one-step separation. The purities of these compounds were 99.1%, 98.3%, 99.0% and 98.5%, respectively, as determined by HPLC. The chemical structures of these isolated compounds were confirmed by ESI-MS, ¹H-NMR and ¹³C-NMR.