Use of dyes as indirect detection probes for the high-sensitivity determination of anions by capillary electrophoresis
Doble, PA and Macka, M and Haddad, PR, Use of dyes as indirect detection probes for the high-sensitivity determination of anions by capillary electrophoresis, Journal of Chromatography A, 804, (1-2) pp. 327-336. ISSN 0021-9673 (1998) [Refereed Article]
High sensitivity for indirect detection was achieved by utilising highly absorbing species as the displaced co-ion (or probe). Two highly absorbing dyes, bromocresol green and indigo-tetrasulfonate, were investigated as potential probes in the determination of small organic and inorganic anions. The concentration of these probes was kept as low as possible to ensure the signal-to-noise ratios were reasonable and the background absorbance was within the linear range of the detector. Four different protocols for buffering the electrolyte with such low probe concentrations were investigated. Buffering with agents that introduce co-anions [acetate or 2- (cyclohexylamino)ethanesulfonic acid (CHES) buffers] proved unsuitable as detection sensitivities were diminished due to competitive displacement by the analytes and system peaks were also induced. Buffering without introduction of co-anions was achieved using the buffering base, diethanolamine, or the use of ampholytes, lysine and glutamic acid. For separations performed with these two buffering approaches, migration time reproducibilities were less than 1% R.S.D. for most analytes. Minimal detectable amounts were in the low attomol region (1-10-18 mol), corresponding to sub-μM vacuum injected solution concentrations. These were an order of magnitude lower than the general detection limit reported for indirect photometric detection, and were comparable with detection limits achieved with indirect fluorescence detection. Finally, the detection limits were further improved by approximately three times for anions analysed with indigo-tetrasulfonate as the probe when a Z cell was employed as the detection cell.