Fluorescein isothiocyanate amino acid separation

Wu S, Dovichi NJ (1989) High-sensitivity fluorescence detector for fluorescein isothiocyanate derivatives of amino acids separated by capillary zone electrophoresis. J Chromatogr 480 141-155... 

Miniaturized columns have provided a decisive advantage in speed. Uracil, phenol, and benzyl alcohol were separated in 20 seconds by CEC in an 18 mm column with a propyl reversed phase.29 A19 cm electrophoretic channel was etched into a glass wafer, filled with a y-cyclodextrin buffer, and used to resolve chiral amino acids from a meteorite in 4 minutes.30 A 6 cm channel equipped with a syringe pump to automate sample derivatization was used to separate amino acids modified with fluorescein isothiocyanate.31 Nanovials have been used to perform tryptic digests on the 15 nL scale for subsequent separation on capillary Electrophoresis.32 A microcolumn has also been used to generate fractions representing time-points of digestion from a 40 pL sample.33 A disposable nanoelectrospray emitter has been... 

Amino acids derivatized with 9-fluorenylmethyl chloroformate (90) were separated by CE and determined by LIF with a pulsed laser LOD 0.5 nM (SNR 2)322. A sensitive technique for amino acids is capillary zone electrophoresis (CZE) combined with LIF of their fluorescein isothiocyanate (133) derivatives. Not all amino acids give good resolution. LOD for proline and arginine were 0.3 and 0.5 nM, respectively323. 

The thiohydantoin derivatives of amino acids obtained from 4-(4-dimethyaminophenyl-azo)phenyl isothiocyanate (141) and fluorescein isothiocyanate (133) can be separated by CZE. Lowering the absolute detection limits of thiohydantoin derivatives of the amino acids is a basic requirement for the development of highly sensitive protein sequencer based on Edman-like processes. Thus, the absolute LOD of thiohydantoin derivatives are at present of the order of 1CT16 mol for 141 and 10-21 mol for 133331. 

The chromatographic separation of a mixture of 0.5 p.m fluorescein-5-isothiocyanate (FITC)-derivatized amino acids on a 2.5 cm x 100 xm x 5 fim LC microchip column is shown in Fig. 6.3, indicating a sharp resolution within 40 seconds. 

Ceriotti and Verpoorte [20] integrated a fritless column for NCEC with conventional stationary phases, which was used for the separation of fluorescein isothiocyanate (FITC)-labeled amino acids. The chips were fabricated in poly(dimethylsiloxane) using deep-reactive-ion-etched silicon masters. The... 

Harrison and co-workers were the first to show quantitative analysis of six fluorescein isothiocyanate (FITC)-labeled amino acids on a microdevice, with separations possible in less than 5 s [21], The concentrations they examined... 

The separation power and sensitivity of CE is illustrated in Figure 21.23. A mixture of 18 amino acids is separated in 30 min, at quantities ranging from 2 to 7 attomoles. The amino acids were derivatized with fluorescein isothiocyanate (Fl i C) to form fluorescent derivatives, and a fluorometric detector system was employed. The detection limits ranged down to 10""2o mol in 1 nL, corresponding to 10 " M and 6000 molecules ... 

Capillary zone electrophoresis with microchip-based SCCE has been used to separate fluorescent dye from degradation products and to separate fluorescein isothiocyanate (FITC) labeled amino acids. Capillary zone electrophoresis (CZE) in fused-silica capillaries has been used to separate the racemic mixtures of (a-hydroxybenzyl)methyltrimethylammonium and (2-hydroxy-l-phenyl)ethyltrimethylammonium with f)-cyclodextrin as the chiral pseudo-stationary phase. L-Phenylalanine and L-phenylalanine-ring-D5 (the hydrogens in the aromatic ring were substituted with deuterium) and another separation of the closely related amino acids phenylalanine and tyrosine have been resolved using CZE with capillary-based SCCE. ... 

Enhancement in Separation Resolution Having established the flow characteristics of our p-FCCE device, its resolving power for fluorescein isothiocyanate (FITC)-labeled amino acid samples was determined. In Fig. 8a, an... 

---