Catecholamines, analysis/separation using

Excess of the reagent hydrolyses to a non-fluorescent residue and the reagent itself does not fluoresce. The optimum wavelength of the excitation light is 390 nm and that of the emitted light 475 nm. This regent is, however, less sensitive than Fluoropa and the derivative is unstable consequently, it must be injected onto the column immediately after formation if used in pre-column derivatization. It has been used successfully in the separation and analysis of polyamines (32), catecholamines (33) and amino acids (34). 

Tornkvist A, Sjoberg PJ, Markides KE, Bergquist J. 2004. Analysis of catecholamines and related substances using porous graphite carbon as separation media in liquid chromatography tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 801 323. 

Faure et al. [22] described nanoelectrochromatography on poly (dimethyl)-siloxane microchips using organic monolithic stationary phases for analysis of derivatized catecholamines. Surface modification of the PDMS material was carried out by UV-mediated graft polymerization. The efficiency of the unit was ascertained by measuring theoretical plates, which were 200,000 per meter. Furthermore, the authors optimized the separation by using pinched and electrokinetic modes at different applied potential of l.OkV/cm (Fig. 7.6) and 30kV/cm (Fig. 7.7) for the pinched and electrokinetic... 

Reversed-phase chromatography is the most popular mode for the separation of low molecular weight (<3000), neutral species that are soluble in water or other polar solvents. It is widely used in the pharmaceutical industry for separation of species such as steroids, vitamins, and /3-blockers. It is also used in other areas for example, in clinical laboratories for analysis of catecholamines, in the chemical industry for analysis of polymer additives, in the environmental arena for analysis of pesticides and herbicides, and in the food and beverage industry for analysis of carbohydrates, sweeteners, and food additives. 

As with the catecholamines, fluorescence methods have also been reported for urinary metanephrine analysis. Fluorescent derivatization of the metanephrines NM and MN by chemical oxidation was based on modification of the trihydroxyindole reaction used for catecholamines. The individual metanephrines were measured following chromatographic separation and fluorescent derivatization or through the formation of differential fluorescent compounds by oxidation at different pH levelsSince the stability of the fluorescent products was variable, with some products decomposing within 10 min,this method has limited application in current practice. Other early methods for analysis of NM and MN included electrophoresis and paper and thin-layer chromatography. These assays were technically complex and had poor analytical sensitivity. 

Catecholamines and amines of the histamine and tryptophan group are highly polar compounds and hence have low volatility. It is preferable, as is the case with amino acids, q.v., therefore, to form derivatives to obtain satisfactory gas-liquid chromatograms. The main problem in the analysis of urine for these amines lies not in the gas chromatographic aspect of the procedure, but rather in their isolation from the urine prior to GLC, where there may be as little as 1 /tg of amine per 100 ml of urine. It appears that this problem of extracting the amines into a sufficiently small volume of suitable solvent prior to GLC has not yet been solved satisfactorily. The fact that GLC can be used successfully to separate catecholamines through their derivatives when starting with pure substances should provide a stimulus for increased research activity in this field. 

Acetone, butanone, cyclobutanone and similar ketones have been used to prepare Schiff bases from long-chain amines, aromatic amines, diamines, anriino-phenols and catecholamines in biological media [29-34], and from hydrazines in air [35], prior to separation by GC. When electron capture detection was used, penta-fluorobenzaldehyde was the chosen reagent. This procedure was applied to the analysis of traces of amines in water samples [36] and biological extracts [37,38], as well as hydrazine in tobacco smoke, technical maleic hydrazide and pyrolysis products [39]. 

Plasma catecholamines The catecholamines are present in human blood plasma in the low nanomolar range, hence some preconcentration, extraction, and sample cleanup of endogenous electrophores is required prior to analysis. The catecholamines from 1 to 3 ml of plasma are adsorbed onto alumina and then eluted into 100 pi of acid to give a 10-30-fold increase in concentration. Preparation of trihydro-xyindole derivatives provides sufficient sensitivity for fluorescent detection. Reversed-phase (Cl 8) columns with addition of an anionic ion-pair reagent or specific ion-exchange columns are suitable, producing separation of all three catecholamines within 15-20 min with isocratic elution. Electrochemical detection provides acceptable specificity with an oxidation potential of -1-0.75 V when using ampero-metry. 

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