Specific Determination of Glutathione using Micellar Elektrokinetic Chromatography and On-Column Reaction with 2,2-dipyridyldisulphide
| Authors | |
|---|---|
| Year of publication | 1999 |
| Type | Article in Proceedings |
| Conference | Book of Abstract, 1st International Symposium on Separations in the Biosciences, Amsterdam, Netherlands |
| MU Faculty or unit | |
| Citation | |
| Field | Biochemistry |
| Keywords | Micellar Elektrokinetic Chromatography; On-Column Derivatization; Thiols; Glutathione |
| Description | The on-column reaction of glutathione with the reactive disulphide 2,2-dipyridyldisulphide was used for the specific detection of glutathione during micellar electrokinetic chromatography: In this set-up the front end of the capillary is used as the reaction chamber. 2,2-dipyridyldisulphide and a sample of glutathione are injected consecutively in the capillary as two discrete plugs separated with short plug of background electrolyte. Due to the differences in the mobilities of the 2,2-dipyridyldisulphide and glutathione on column mixing and reaction occur. This reaction is rapid and quantitative, because thiol form is stabilised by thiol-thione tautomerism. This fact also causes a large shift of the absorption maxima toward longer wavelength. Glutathione is thus quantitatively transformed into a mixed disulphide concomitantly with formation of equimolar amounts of the 2-thiopyridone that is further separated by micellar electrokinetic chromatography and determined spectrophotometrically at 343 nm. |
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