Abstract: AimTo establish chiral separation method for donepezil hydrochloride (E2020) enantiomers by capillary electrophoresis (CE) and determine the two enantiomers in plasma. Methods Alkalized plasma was extracted by isopropanol-n-hexane (3∶97) and L-butefeina was used as the internal standard. Enantioresolution was achieved using 2.5% sulfated-beta-cyclodextrin as chiral selector in 25 mmol·L^-1 triethylammonium phosphate solution (pH 2.5) on the uncoated fused-silica capillary column (70 cm×50 μm ID). The feasibility of the method to be used as quantitation of E2020 enantiomers in rabbit plasma was also investigated. ResultsE2020 enantiomers were separated at a baseline level under the above condition. The linearity of the response was evaluated in the concentration range from 0.1 to 5 mg·L^-1. The linear regression analysis obtained by plotting the peak area ratio (A_s/A_i) of the analyte to the internal standard versus the concentration (C) showed excellent correlation coefficient (r=0.999 2 for R(-)-E2020, r=0.999 7 for S(+)-E2020) and the equations were A_s/A_i=0.024 2+0.289 2C and A_s/A_i=0.010 8+0.273 7C, respectively. The low limit of detection was 0.05 mg·L^-1. The inter- and intra-day precision (RSD) were all less than 20%. ConclusionCompared with CSP by HPLC, the CE method is simple, reliable, inexpensive and suitable for studying the stereoseletive pharmacokinetics in rabbits.