In this study, chip-based enantioselective open-tubular CEC (OT-CEC) was developed employing BSA-gold nanoparticle (GNP) conjugates as a chiral stationary phase. An immobilization procedure was realized by prederivatization of the glass microchannel with (3-mercaptopropyl)-trimethoxysilane to provide thiol groups, which linked the BSA-GNP conjugates on the inner surface of the microchannels. Incorporation of GNPs into immobilization of BSA selectors greatly increased the BSA phase ratio and favored the BSA stationary phase generated sufficient EOF. Good resolutions of FITC-labeled ephedrine and norephedrine isomers were achieved with 36 mm. effective separation channel length within 250 s. The constructed OT-CEC microdevice exhibited good repeatabilities for run-to-run enantioseparations and kept an enantioselective lifetime of more than 1 month. The effects of pH values and concentrations of a running buffer on the selectivity and resolution of enantioseparations were investigated.