Automated mass spectrometric analysis of urinary and plasma serotonin

Abstract

Serotonin emerges as crucial neurotransmitter and hormone in a growing number of different physiologic processes. Besides extensive serotonin production previously noted in patients with metastatic carcinoid tumors, serotonin now is implicated in liver cell regeneration and bone formation. The aim was to develop a rapid, sensitive, and highly selective automated on-line solid-phase extraction method coupled to high-performance liquid chromatography–tandem mass spectrometry (XLC-MS/MS) to quantify low serotonin concentrations in matrices such as platelet-poor plasma and urine. Fifty microliters plasma or 2.5 μL urine equivalent were pre-purified by automated on-line solid-phase extraction, using weak cation exchange. Chromatography of serotonin and its deuterated internal standard was performed with hydrophilic interaction chromatography. Mass spectrometric detection was operated in multiple reaction monitoring mode using a quadrupole tandem mass spectrometer with positive electrospray ionization. Serotonin concentrations were determined in platelet-poor plasma of metastatic carcinoid patients (n = 23) and healthy controls (n = 22). Urinary reference intervals were set by analyzing 24-h urine collections of 120 healthy subjects. Total run-time was 6 min. Intra- and inter-assay analytical variation were <10%. Linearity in the 0–7300 μmol/L calibration range was excellent (R2 > 0.99). Quantification limits were 30 and 0.9 nmol/L in urine and plasma, respectively. Platelet-poor serotonin concentrations in metastatic carcinoid patients were significantly higher than in controls. The urinary reference interval was 10–78 μmol/mol creatinine. Serotonin analysis with sensitive and specific XLC-MS/MS overcomes limitations of conventional HPLC. This enables accurate quantification of serotonin for both routine diagnostic procedures and research in serotonin-related disorders.

Development of an LC-MS/MS method for the analysis of serotonin and related compounds in urine and the identification of a potential biomarker for attention deficit hyperactivity/hyperkinetic disorder

“…This method has a low sensitivity (LOD and LOQ of 0.10 and 0.13 μmol/L) and a marginally reduced recovery (81-98%) in comparison to the method reported here. de Jong et al [27] have also recently described an automated LC-MS method for serotonin in urinary and plasma samples. The LOQ for serotonin (30 nmol/L) in urine is matched by this study (36 nmol/L) and our recoveries are in the range of 88-135%; with absolute recoveries of 83-85%.…”

“…A variety of methods have been developed to measure serotonin including spectrophotometry [21,22], LCfluorometry [23,24], enzyme immunoassay, and LCelectrochemical detection [25,26]. LC-mass spectrometric methods is however the method of choice as tandem LC-MS methods provide increased selectivity while reducing the need for dependence on chromatographic separation [27][28][29][30][31][32][33][34][35]. While these methods report good results for the analytes under investigation, the methods do not facilitate multi-target analysis which was the aim of this study.…”

Principles and Applications of LC-MS/MS for the Quantitative Bioanalysis of Analytes in Various Biological Samples

“…Platelet serotonin is specifically measured for the detection of carcinoid tumors that secrete little serotonin. With LC-MS/MS, it is possible to measure accurate and reproducible serotonin both in platelet-rich and plasma-poor plasma (de Jong et al, 2009b;Monaghan et al, 2009;de Jong et al, 2010b). For this purpose, protein precipitation, using acetonitrile, combined with chromatography based on strong cation exchange and reversed-phase interaction, was used with a total run time of 6 min (Monaghan et al, 2009) and a detection limit of 5 nmol/L.…”

Applicability of 2D gel electrophoresis and liquid chromatography in proteomic analysis of urine using mass spectrometry MALDI-TOF

“…This procedure linked with the use of analytical column with the C-18 phase of the 25 cm length enables better separation than the comparable gel 2D electrophoresis system. However, liquid chromatography used as a method of preparation of a sample for analysis has also its own limitations (Erni 1982, Adachi et al 2006, De Jong et al 2010. One of the main assumptions in sample preparation for analysis is adjustment of the number of theoretical shelves to applied mobile phase.…”