On a reversed-phased Hypersil C₁₈ column, 10 bile acid derivatives were separated using 2-(12-oxobenzo[b]acridin-5(12H)-yl)ethyl 4-toluenesulfonate (BAETS) as pre-column derivatization reagent by high-performance liquid chromatography with fluorescence detection and mass spectrum identification in conjunction with a gradient elution. Optimum derivatization, giving the corresponding sensitively fluorescent derivatives, was obtained by the reacting of bile acids with BAETS at 95 ℃ for 30 min in the presence of basic catalyst potassium carbonate in dimethylsulfoxide solvent. The optimum excitation and emission wavelengths were at λ_ex/λ_em＝272 nm/505 nm. The percent ionization δ values in aqueous acetonitrile and aqueous methanol were in the range of 0%～88.83% and 0%～89.15% respectively. The identification of bile acid derivatives was obtained on the basis of an atmosphere-pressure chemical ionization source at positive model detection. Correlation coefficients are ＞0.9995, and detection limits at signal-to-noise of 3 are 0.76～1.62 ng/mL for the labeled bile acids.

Key words: high performance liquid chromatography/ion-trap mass spectrometry, percent ionization, 2-(12-oxobenzo[b]acridin-5(12H)-yl)ethyl 4-toluenesulfonate (BAETS), bile acid