HPLC-fluorescence detection and MEKC-LIF detection for the study of amino acids and catecholamines labelled with naphthalene-2,3-dicarboxyaldehyde.
Naphthalene-2,3-dicarboxyaldehyde (NDA) is commonly used for detection of primary amines in conjunction with their separation with HPLC and CE. The fluorescence of the derivatives can be measured by a conventional fluorometer or via LIF. NDA is a reactive dye, which can replace o-phthaldehyde (OPA) and provides for derivatives which are considerably more stable than OPA derivatives. In addition, NDA can be used to derivatize primary amines at concentrations as low as 100 pM. In this work, HPLC/fluorescence and MEKC/LIF experiments were performed to separate/detect six neuroactive compounds, the amino acids, Gly, Glu, Asp, gamma-aminobutyric acid (GABA) and the catecholamines, dopamine and noradrenaline. The two methods were compared in terms of performance of separation. The amino acids can be separated in HPLC in less than 30 min and an identical separation is obtained in CE using MEKC and lithium salts with greater resolution (the number of theoretical plates was approximately 5000 for HPLC and 200 000 for MEKC). The lowest detected concentration was in the range of 0.1 nM for CE/LIF. The presence of a high salt concentration does not affect the separation of the samples. Examples of the analysis of microdialysate samples as well as amino acids in Ringer's solution are presented.
Siri
Nathalie
N
Picometrics, Toulouse, France.
Lacroix
Marlene
M
Garrigues
Jean-Christophe
JC
Poinsot
Véréna
V
Couderc
François
F
D000596:Amino Acids / Q000302:isolation & purification*; D002395:Catecholamines / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D020374:Chromatography, Micellar Electrokinetic Capillary; D019075:Electrophoresis, Capillary; D005470:Fluorometry / Q000379:methods*; D009281:Naphthalenes; D012680:Sensitivity and Specificity
D000596:Amino Acids; D002395:Catecholamines; D009281:Naphthalenes; C027717:2,3-naphthalenedicarboxaldehyde