Selective and sensitive liquid chromatographic determination method of 5-hydroxyindoles with fluorous and fluorogenic derivatization

Abstract

A liquid chromatographic (LC) method with improved selectivity for the simultaneous determination of 5-hydroxyindoles (5-HIs; 5-hydroxytryptophan, 5-hydroxytryptamine, N-acetyl-5-hydroxytryptamine, 5-hydroxyindole-3-acetic acid, and 5-hydroxytryptophol) is described. This method involves precolumn derivatization with 4-(3',3',4',4',5',5',6',6',7',7',8',8',9',9',10',10',10'-heptadecafluorodecyl)benzylamine (HFBA) and separation of the derivatives using a fluorous LC column. In this study, stable benzoxazole derivatives of 5-HIs with HFBA have been obtained by a simple derivatization procedure; their fluorescent properties enabled highly sensitive detection. In addition, only the HFBA derivatives of 5-HIs has been selectively retained on the fluorous LC column via fluorous interaction whereby perfluoroalkyl compounds show affinities with each other, while the non-fluorous compounds did not. The HFBA derivatives were separated within 30 min and the detection limits for 5-HIs in a 20-μL injection volume were 1.2-14 fmol (S/N=3). Furthermore, this method was applied to the analysis of 5-HIs in the human plasma from healthy subjects.

Recent Trends in the Quantification of Biogenic Amines in Biofluids as Biomarkers of Various Disorders: A Review

“…The data summarizing chromatographic methods for BA determination confirmed that gradient elution of BAs was mostly applied, based on the controlled modification of the composition of the mobile phase during the run-time of the analysis, although the isocratic mode with a constant mobile phase composition during chromatographic separation was also reported [29,38,42,44,51,70,71]. Gradient elution allows the separation of the compounds whose peak resolution under isocratic conditions is unsuccessful.…”

“…Sakaguchi et al applied sample derivatization with 2-(perfluorooctyl)ethyl isocyanate (PFOEI) for the determination of L-DOPA, DA, NE, E, M, L-Tryp, 5-HTrp and 5-HT in 10 µL of urine samples, reaching LODs from 0.21 to 4.2 nmol/mL and with recovery results from 96.9 to 103.3% [37]. In a different study concerning plasma samples, Sakaguchi et al used tetrahydrofuran (THF) and 4-(heptadecafluorodecyl)benzylamine (HFBA) to deproteinize and derivatize 5-HT, 5-HIAA and three other 5-hydroxyindoles in 70 µL of plasma samples [38]. This method provides LODs of 4.3 pmol/mL for 5-HT and 1.5 pmol/mL for 5-HIAA but the extraction efficiencies were worse and equalled 84.2% for 5-HT and 72.2% for 5-HIAA.…”

“…Another derivatizing agent, such as HFBA, was applied before the LC-FL analysis of 5-HT and 5-HIAA in plasma, based on a fluorous phase column [38]. Moreover, the derivatization of the abovementioned analytes with benzylamine and potassium hexacyanoferrate (III) before the LC-FL assay based on RP18 was also reported [49].…”

“…An interesting approach is also the application of the MRM 3 mode, which can improve the reliability of the obtained results [78]. Therefore, the LC-MS/MS technique was applied for the identification and the quantification of BAs in serum [29,31,35,39,43,49,58,61,72,75], urine [7,22,23,24,29,31,32,34,37,41,44,46,48,50,60,63,65,68,69], plasma [7,16,25,26,27,30,32,38,52,53,57,69,71], whole blood [51] and other human samples [3,16,31,42,59,76] (Table 2). A method based only on ESI-MS/MS for the determination of 13 amino acids in human plasma, including L-Tyr, has also been reported [56].…”

Fluorous and Fluorogenic Derivatization for Selective Liquid Chromatographic Analysis of Cyanide in Human Plasma

“…[17][18][19][20] In addition, its combination with fluorescentbased detection approaches was also utilized as a selective analysis method to focus on some biogenic compounds. [21][22][23] The aim of this study is to develop a fluorous and fluorogenic derivatization method for the highly selective and sensitive analysis of cyanide with LC-FL detection. To this end, NDA and 4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11-heptadecafluorounde cylamine (HFUA) were used as derivatization reagents (Fig.…”