Biphasic recognition recycling high-speed counter-current chromatography (HSCCC) was applied for the separation of amlodipine besilate (ADB) enantiomers. The biphasic recognition system was established by adding hydrophilic sulfobutyl ether-p-cyclodextrin (SBE-beta-CD) in the aqueous mobile phase and lipophilic D-iso-butyl tartrate (D-IBTA) in the organic stationary phase, which is capable of preferentially recognizing (+)-enantiomer and (-)-enantiomer respectively. The two-phase solvent system, which was composed of ethyl acetate-aqueous solution (1:1, v/v) plus the above chiral selectors, was selected as per the distribution ratios and separation factor of ADB enantiomers. Impacts of the type and concentration of chiral selectors, pH value of the aqueous solution as well as temperature on the separation efficiency were investigated. By means of preparative HSCCC, pure enantiomers were obtained through the separation of 30 mg racemates with a purity greater than 97.5% and a recovery rate between 80% and 84%. Experimental results revealed that biphasic recognition recycling high-speed counter-current chromatography offers a promising means to separate racemates with a low separation factor in a highly efficient manner. (C) 2015 Elsevier B.V. All rights reserved.