Abstract: OBJECTIVE To establish method for enantioseparation of 3-n-butylphthalide(NBP) and 6-Br-NBP on Chiralpak IC column by HPLC. METHODS HPLC had been developed for the chiral separation of NBP and 6-Br-NBP on a Chiralpak IC column under normal phase mode. The influences of organic solvent, column temperature and flow rate were investigated and the enatiomeric separation was optimized systematically. RESULTS NBP and 6-Br-NBP could be separated completely. In the circumstance of hexane-ethanol(98∶2), flow rate 0.8 mL·min^-1, temperature 35 ℃, to NBP, the separation factor α was 1.07 and the resolution R_s reached 2.20. As hexane-isopropanol(70∶30), flow rate 0.8 mL·min^-1, injection volume 10 μL, temperature 20 ℃, to 6-Br-NBP, the separation factor α was 1.10 and the resolution Rs reached 2.67. CONCLUSION This method is suitable for the chiral separation of NBP and 6-Br-NBP.