Composite membranes prepared by the interfacial polymerization of L-arginine and piperazine with trimesoyl chloride in situ on microporous polysulfone membrane are reported. The chiral selective layer of composite membranes was characterized by variable angle attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and scanning electron microscopy. The enantioselective performance of membranes was examined and correlated with the composition of selective layer through optical resolution of racemic arginine in reverse osmosis mode. The effects of reverse osmosis process parameters such as trans-membrane pressure, permeation time and concentration of feed on the performance of the membranes were also determined. The observation indicated that membranes having chiral environment performed enantiomeric separation by permeating D-arginine preferentially over 89% enantiomeric excess. The membranes without chiral environment (M1 and M4) though exhibited separation of arginine did not perform enantiomeric separation. The trans-membrane pressure and concentration of feed solution have noticeable effects on enantioseparation. Higher enantiomeric excess resulted from dilute feed and at low trans-membrane pressure. The membranes prepared from L-arginine (2%) and trimesoyl chloride (1%) solutions exhibited highest enantiomeric excess (89%). The enantioselectivity of membranes was independent of time. The separation factor (alpha) for D-arginine achieved as high as similar to 17. (C) 2011 Elsevier B.V. All rights reserved.