Monitornig of Cysteine in Human Urine by On-column Derivatization during MEKC
| Authors | |
|---|---|
| Year of publication | 2002 |
| Type | Article in Proceedings |
| Conference | Book of abstracts of Developments in Chromatography and Electrophoresis & Chiranal 2002 |
| MU Faculty or unit | |
| Citation | |
| Field | Biochemistry |
| Keywords | derivatization; on-column; urine; MEKC |
| Description | Cysteine is a high sulphur containing amino acid synthesised by the liver. It is involved in a variety of important cellular functions, among others protein synthesis, detoxication and metabolisms. The biological significance of cysteine is well established but it is only recently that its level in physiological fluids such as plasma and urine has been established as an important indicator for a number of clinical disorders. Determination of urinary content of cysteine consequently may be taken as helpful non-invasive methods of diagnosis. This paper describes the development and validation of a micellar electrokinetic chromatography and on-column reaction with 2,2-dipyridyldisulphide method for the analysis of free and total cysteine in human urine. 2,2-dipyridyldisulphide and a sample of cysteine are injected consecutively into the capillary as two discrete plugs separated with a short plug of background electrolyte. Due to the differences in the mobilities of the 2,2-dipyridyldisulphide and cysteine, on-column mixing and reaction occur. Cysteine is in this reaction quantitatively transformed into a mixed disulphide concomitantly with formation of equimolar amount of the 2-thiopyridone that is further separated by micellar electrokinetic chromatography and determined spectrophotometrically at 343 nm. The concentration of cysteine is thus estimated indirectly from the result of 2-thiopyridone determination. The linear detection range for concentration versus peak area for the assay is from 0.05 to 5 mM (correlation coefficient 0.98) with a detection limit of 2.5 mM. The inter-day reproducibility of the peak area was 2.18 % and the inter-day reproducibility of the migration time 0.51 %. |
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