Application of 2-(11H-benzo[a]carbazol-11-yl) Ethyl Carbonochloridate as a Precolumn Derivatization Reagent of Amino Acid by High Performance Liquid Chromatography with Fluorescence Detection

On a reversed-phase Hypersil BDS C18 (200 mm × 4.6 mm, 5 μm) column, 20 amino acids, which were derivatized using 2-(11H-benzo[a]carbazol-11-yl) ethyl carbonochloridate (BCEC-Cl) as precolumn derivatization reagent, were separated in conjunction with a gradient elution. Optimum derivatization was obtained by reacting amino acids with BCEC-Cl at room temperature for 5 min in the presence of sodium borate catalyst in acetonitrile solvent. The fluorescence excitation and emission wavelengths were 279 and 380 nm, respectively. The identification of amino acid derivatives from hydrolyzed bovine serum albumin and bee pollen was carried out by postcolumn mass spectrometry with electrospray ion (ESI) source in positive ion mode. Linear correlation coefficients of the amino acid derivatives are > 0.9993, and detection limits (at signal to noise of 3:1) are 1.49–19.74 fmol for the labeled amino acids.