Analytical Sciences

Separation and Determination of Fluorescein Isothiocyanate-Labeled Amino Acids by Capillary Electrophoresis with Laser-Induced Fluorescence Detection

Keita TAKIZAWA and Hiroshi NAKAMURA*

Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Science University of Tokyo, Ichigaya-funagawara-machi, Shinjuku, Tokyo 162-0826, Japan

A method for the separation and determination of amino acids by capillary electrophoresis with laser-induced fluorescence detection has been developed. At first, the condition for the derivatization of amino acids with fluorescein isothio-cyanate (FITC) was studied; it was proved to be optimal at 50ûÃ for 5 h. Next, the separation of 20 FITC-amino acids was attempted by using an unmodified fused silica capillary. Actually, 18 kinds of FITC-amino acids were separated with 96 mM sodium borate buffer (pH 9.5) containing either 25% acetone (separation system 1) or 10% methanol (separation system 2). The resolution of 19 FITC-amino acids was attained with 80 mM sodium borate buffer (pH 9.2) containing 45 mM a-cyclodextrin (separation system 3). Finally, we succeeded in separating 20 FITC-amino acids by combining two migration systems (2 and 3). The FITC-amino acids were detected by argon-ion laser-induced fluorescence (Ex 488 nm, Em 520 nm). The detection limits of the amino acids obtained with separation systems 1 and 2 were 0.052 - 1.01 fmol. (Keywords: Fluorescein isothiocyanate, laser induced fluorescence detector, capillary electrophoresis, amino acid, a-cyclodextrin)

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