Development and validation of a simple, rapid and sensitive LC-MS/MS method for the measurement of urinary neurotransmitters and their metabolites

Abstract

Neurotransmitters play crucial roles in physiological functions and their imbalances have demonstrated association in the pathology of several diseases. The measurement of neurotransmitters possesses a great potential as a significant clinical tool. This study presents the development and validation of an LC-MS/MS method for simultaneous quantification of multi-class neurotransmitters associated with dopamine, tryptophan and glutamate-γ-aminobutyric acid pathways. A total of ten neurotransmitters and their metabolites (dopamine, epinephrine, metanephrine, tryptophan, serotonin, kynurenic acid, kynurenine, anthranilic acid, GABA, glutamic acid) were determined based on a simple and rapid 'dilute and shoot' method using minimal urine volume. The chromatographic separation was achieved using a Poroshell 120 Bonus-RP LC Column in combination with a gradient elution within an 8.5-min time frame. The method exhibited good sensitivity as the limits of quantification ranged between 0.025 and 0.075 μg/mL with acceptable matrix effects (< ± 14.5%), no carryover and good linearity (r > 0.98). The accuracy and precision for all analytes were within tolerances, at < ± 9.9% mean relative error (MRE) and < 8.6% relative standard deviation (RSD), respectively. The method was successfully applied in measuring the neurotransmitter concentrations in urine of healthy donors. Furthermore, the undertaken stability experiments indicated that acidified urine specimens allowed the analytes to be stable for prolonged durations in comparison to those untreated. The study also reveals the performance of the method is unaffected by the absence of expensive deuterated reference standards under the experimental conditions employed which further simplifies the analytical procedures and provides a significant cost saving for running the assay. Graphical abstract The quantification of multi-class neurotransitters associated with the dopamine, tryptophan and GABA-glutamate pathways using a simple 'dilute and shoot' LC-MS/MS method.

Urinary Profile of Endogenous Gamma-Hydroxybutyric Acid and its Biomarker Metabolites in Healthy Korean Females: Determination of Age-Dependent and Intra-Individual Variability and Identification of Metabolites Correlated With Gamma-Hydroxybutyric Acid

“…Few studies have been previously conducted on the endogenous urinary concentrations of biomarker metabolites of GHB. In a previous study on determining 10 urinary neurotransmitters and their metabolites in urine samples of six males and four females, glutamic acid and GABA levels were measured, and their concentrations were 3.050 ± 0.150 and 0.543 ± 0.036 μg/ml, respectively ( Yan et al, 2017 ). Further, phase 1 metabolite and GHB levels were measured in urine samples of 77 males and 55 females with ranges as follows: GHB, not detected–1.94 μg/ml, glycolic acid, 1.30–400 μg/ml; succinic acid, 1.17–273 μg/ml; 2,4-OH-BA, 0.72–26.2 μg/ml; and 3,4-OH-BA, 1.88–122 μg/ml ( Jarsiah et al, 2020 ).…”

Correlation between monoamine neurotransmitter and cytokine levels and the occurrence of post-traumatic stress disorder among operating room nurses

“…Neurotransmitters play a crucial role in maintaining physiological function at both the cellular and organ levels, and their imbalances have demonstrated a correlation in the pathology of several diseases (8). Monoamine neurotransmitters, a category of neurotransmitters, can act as biomarkers for the diagnosis and prognosis of some neurological disorders, such as Parkinson's disease and Alzheimer's disease (9,10).…”

Ion Mobility–Mass Spectrometry for Bioanalysis

“…Due to the inherent complexity of biological matrices, samples are usually subjected to clean-up or preconcentration procedures to enrich the sample and to eliminate possible interferences that can hinder the determination of the compound(s) of interest. The complexity of such procedures ranges from simple dilution of the sample (dilute and shoot (DAS)) [17][18][19] or protein precipitation (PPT) [20,21] to intricate procedures dealing with solid-phase (SPE) [22][23][24] or liquid-liquid (LLE) [25] extractions [26,27]. It is clear that the sample treatment procedure is determined by the information available regarding the structural and the physicochemical properties of the analytes, the matrix, and the purpose of the analysis.…”