Chemiluminescent Detection of Amino-Acids using In-Situ Generated Ru(bpy)(3)(3+)
by Jackson, Warren Alan; Bobbitt II, Donald R.
A technique for the detection of underivatized amino acids based on the chemiluminescent (CL) reaction between in situ generated Ru(bpy)(3)(3+) and the amino acid is described. Glassy carbon was found to be an ideal material for the in situ generation of Ru(bpy)(3)(3+) from Ru(bpy)(3)(2+) in alkaline pH in the presence or absence of an organic modifier such as acetonitrile. The CL emission from the glassy carbon thin-layer flow cell used for the in situ studies was found to be dependent upon the concentration of Ru(bpy)(3)(2+) in the carrier buffer, the buffer flow-rate through the cell, and the applied potential at the working electrode. The detection scheme was applied to high performance ion-exchange chromatography (HPIEC) by utilizing a column packed with a polystyrene-divinylbenzene anion-exchange resin. The technique was found to provide a linear response over 3 orders of magnitude for leucine, a representative amino acid. Limits of detection (LOD) ranged from 100 fmol for proline to 22 pmol for serine at a S/N of 6. The CL-flow-injection analysis response of injected leucine was found to decay less than 8% over the course of 40 h of continuous electrolysis of Ru(bpy)(3)(2+) in the detection cell. The method is also shown to be applicable to the sensitive detection of PTH-glycine. Arguments toward the future miniaturization of the detection scheme in order to facilitate application to capillary electrophoresis and subsequent development of new protein sequencing methodologies will be presented.
- Analytica Chimica Acta
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- 1873-4324; 0003-2670