Separation of binary amino acids from aqueous solutions by solvent extraction (SX) and liquid surfactant membrane (LSM) was compared, in which the acidic extractant di(2-ethylhexyl)phosphoric acid (D2EHPA) was selected as an active carrier. The distribution ratios were measured by batch SX experiments. Important variables affecting LSM permeation experiments, including the pH in the aqueous external phase, surfactant concentration, the volume ratios of W/O emulsions to aqueous external phase and that of organic to aqueous internal phases, and HCl concentration in the internal phase, were systematically studied. It was shown that the separation was promising only when the amino acids had sufficiently different isoelectric points. Compared to batch SX results, the optimal pH for effective separation shifted to a lower level and the selectivity was enhanced in LSM process, which was a result of the counter transport of hydrogen ions.