HPLC-fluorescence detection and MEKC-LIF detection for the study of amino acids and catecholamines labelled with naphthalene-2,3-dicarboxyaldehyde

Abstract

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.

Study of the naphthalene-2,3-dicarboxaldehyde pre-column derivatization of biogenic mono- and diamines in mixture and fluorescence−HPLC determination

“…Compared with the products formed by derivatization with OPA, the NDA/CN derivatives exhibit improved chemical stability, excitation maxima (420 and 440 nm) in the visible region, efficient chemiluminescence by the peroxyoxalate method and enhanced fluorescence quantum efficiency [41]. NDA has been used for the derivatization of amino acids and peptides [41][42][43][44][45], catecholamines [46][47][48][49], aliphatic amines [50], heterocyclic amines [51,52], aliphatic diamines [53], drugs [54][55][56] and lipids [57]. The main features of these methods are summarized in Electronic supplementary material (ESM) Table S1.…”

The use of naphthalene‐2,3‐dicarboxaldehyde for the analysis of primary amines using high‐performance liquid chromatography and capillary electrophoresis

“…Recently a study was presented to show the greater sensitivity of CE/LIF relative to conventional HPLC/ fluorescence detection in the analysis of microdialysates (Siri et al, 2006). This study also shows the benefits of using lithium dodecylsulfate to separate derivatized amino acids in MEKC instead of conventional SDS: the lithium salt results in a better separation with shorter migration times.…”

Determination of amino acids by micellar EKC: Recent advances in method development and novel applications to different matrices

“…An interesting method is that recently published by Siri et al [50] for the separation of: (i) few amino acids (Gly, Glu, Asp) and of the catecholamines dopamine and noradrenaline and (ii) a mixture of 20 standard amino acids and catecholamines. They used 22.5 mM lithium tetraborate buffer, pH 9.2, containing 20 mM lithium dodecyl sulfate.…”

“…When applied to microdialysates collected from the region of the suprachiasmatic nucleus of male Syrian hamsters, the approach was able to measure daily variations of GABA levels. The method developed by Siri et al [50], already presented in Section 2.1, was also applied to microdialysates of rat striatum. Glu, GABA, Asp, Gly, NOR, and DOPA could be easily detected with recoveries ranging between 88 and 102% for amino acids and between 86 and 117% for NOR and DOPA.…”