Design and synthesis of a hydrophilic fluorescent derivatization reagent for carboxylic acids, 4‐N‐(4‐N‐aminoethyl)piperazino‐7‐nitro‐2,1,3‐benzoxadiazole (NBD‐PZ‐NH₂), and its application to capillary electrophoresis with laser‐induced fluorescence detection

Abstract

A hydrophilic fluorescent derivatization reagent for fatty acids, 4-N-(4-N-aminoethyl)piperazino-7-nitro-2,1,3-benzoxadiazole (NBD-PZ-NH 2 ), was designed and synthesized. NBD-PZ-NH 2 possesses not only a fluorophore and a reacting group but also a positive charge group and, thus, was hydrophilic and suitable for application to capillary electrophoresis. NBD-PZ-NH 2 reacted with fatty acids in the presence of triphenylphosphine (TPP) and 2,2'-dipyridyl disulfide (DPDS) at room temperature within 10 min. The derivatives were strongly fluoresced and were positively charged at pH below 3. The derivatives of C4-C20 fatty acids were separated within 10 min in 50% acetonitrile in water containing 30 mM ammonium acetate and 1.0 M acetic acid by capillary electrophoresis with laser-induced fluorescence (CE-LIF) detection. The detection limits attained were 6.5 nM (signal-to-noise ratio of 3). It is proposed that NBD-PZ-NH 2 is a prominent derivatization reagent for fatty acids which is suitable for CE-LIF application.

Separation of Nile Blue‐labelled fatty acids by CE with absorbance detection using a red light‐emitting diode

“…Surprisingly, two reports on the use of indirect fluorescence detection using merocyanine 450 [12] and fluores-cein [13] have reported detection limits around 10 26 M, thereby offering only marginal improvement in sensitivity in comparison to indirect UV absorbance approaches. This is in stark contrast to reports where fatty acids have undergone precapillary derivatisation with an appropriate fluorophore, giving detection limits in the low nM (10 29 M) range, which is four orders of magnitude below those obtained by indirect detection methods [14][15][16][17]. However, the principal disadvantage of this approach is the expensive nature of the fluorescent reagents and equipment.…”

Separation of perfluorocarboxylic acids using capillary electrophoresis with UV detection

“…This has already been reported for CE determinations of carboxylic acids with MS [26,27] and LIF [28] detections. A high alkaline pH was used in these applications; hence, a BGE of 20 mM ammonium acetate, pH 9.5, was examined in this work.…”

“…It results in the decrease of EOF and alleviates the adsorption on the capillary walls. This has been applied in CE determinations of various carboxylic acids in phosphate BGE to which 15 [24] to 30% [21] acetonitrile was added, for the determination of dicarboxylic acids with 40% ethanol added [21], and for fatty acids with ammonium acetate buffer and 50% acetonitrile added [28]. Determinations of anionic surfactants required 50% methanol in BGE [23], whereas determinations of fatty acids with BGEs containing trimethoxybenzoic acid and dipentylamine with addition of 85% acetonitrile and 5% dioxane were reported [30].…”

Simple and practical derivatization procedure for enhanced detection of carboxylic acids in liquid chromatography–electrospray ionization-tandem mass spectrometry

“…The combination of DPDS and TPP has been conventionally used for the condensation of amines and carboxylic acids [14,15]. Based on this information, these agents were used to prepare the HPand PA-derivatives of the carboxylic acids in this study.…”