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Sensitive indirect photometric detection of inorganic and small organic anions by capillary electrophoresis using Orange G as a probe ion


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Johns, CA, Shaw, MJ, Macka, M and Haddad, PR 2003 , 'Sensitive indirect photometric detection of inorganic and small organic anions by capillary electrophoresis using Orange G as a probe ion' , Electrophoresis, vol. 24, no. 3 , pp. 557-566 , doi: 10.1002/elps.200390063.

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This study addresses the two major problems in the use of dyes as highly absorbing probes for indirect photometric detection in capillary electrophoresis (CE). First, effective electroosmotic flow (EOF) modification or suppression to allow separation and detection of a wide mobility range of analytes is not straightforward when electrolytes containing increased dye concentrations are used. The suppression of EOF to less than + 5×10-9 m2V-1s-1 was achieved with a combination of a poly(ethylenimine) (PEI)-coated capillary and the addition of the neutral polymer hydroxypropylmethylcellulose (HPMC) to the background electrolyte. Second, the deterioration of baselines due to adsorption of the dye probe to the capillary wall is generally a problem. In this work, baseline quality at higher probe concentrations was significantly improved by a rather unusual but highly effective combination of a simultaneous application of a slight overpressure (25 mbar) at the injection end during the separation, and the use of a relatively narrow capillary of 50 m inner diameter. Both measures would appear to be counterproductive. Optimisation of the probe concentration with regard to signal-to-noise ratio resulted in an electrolyte of 4 mM Orange G, 0.05% HPMC buffered at pH 7.7 by the addition of 10.0 mM histidine isoelectric buffer. Very high separation efficiencies of 128 000-297 000 plates were made possible by the relatively high probe concentration. Combined with excellent detection sensitivity, even with the introduction of hydrodynamic flow and a reduced optical path length, these measures resulted in limits of detection ranging from 0.216 to 0.912 M with a deuterium lamp light source (248 nm) and from 0.147 to 0.834 M with a 476 nm blue light-emitting diode (LED) light source. Reproducibility over 30 consecutive runs without changing the electrolyte was excellent, with relative standard deviation (RSD) values of 0.14-0.80% for migration time, 1.27-3.36% for peak area and 0.88-5.12% for peak heights. The optimised electrolyte was used for the analysis of inorganic anions in air filter samples, providing good agreement with results obtained by ion chromatography.

Item Type: Article
Authors/Creators:Johns, CA and Shaw, MJ and Macka, M and Haddad, PR
Keywords: Anions • Capillary electrophoresis • Detector linearity • Dyes • Indirect photometric detection • Isoelectric buffer • Light-emitting diode
Journal or Publication Title: Electrophoresis
ISSN: 0173-0835
DOI / ID Number: 10.1002/elps.200390063
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