Determining Neurotransmitters in Spinal Cords with UHPLC

To innovate current sample pretreatment and detection techniques in the management of neurological disorders, researchers at Jilin University combined ultrasound-assisted magnetic ionic liquid dispersion liquid-liquid microextraction (UA-MIL-DLLME) with ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-QqQ/MS²) for the extraction and quantification of neurotransmitters (NTs). This could help improve clinical diagnosis and management of these conditions. A paper based on this work was published in Scientific Reports (1).

NTs, which are essential chemicals in the body, facilitate communication between neurons and target cells through chemical synapses, with their main responsibility being the facilitation of information exchange between the brain and the rest of the body, ensuring a normal flow of life activities (2). In addition, NTs also play a crucial role in the regulation of a variety ofneural activities such as stress responses, motor coordination, and cognitive functions (3.4). Categorized into monoamines (dopamine, adrenaline, norepinephrine, serotonin) and amino acids (glutamic acid, glycine, gamma-aminobutyric acid),the importance of NTs in the central and peripheral nervous systems has made them the subject of extensive research in the fields of biomedicine, medical diagnosis, clinical chemistry, and pharmaceuticals fields (5-7).

The performance of the UA-MIL-DLLME method was assessed by the researchers under optimized conditions through the evaluation of a variety of metrics, including linearity, enrichment factor (EF), detection limit (LOD), limit of quantitation (LOQ), recovery rate, precision, and robustness. The team analyzed standard and labeled sample solutions to determine calibration curves; peak areas were correlated to analyte concentrations. The concentration capacity of the method, measured by the enrichment factor (EF), ranged from 7.0 to 9.8 (EF = C_s/C₀). The quantification of sensitivity was determined using the LOD (S/N = 3) and LOQ (S/N = 10), which indicated a high sensitivity regarding NT detection.

NT concentration was accurately added to the sample matrix accurately, and the experiment was conducted five times by the researchers to confirm the trustworthiness of the results. Recovery experiments with added NTs showed stable recovery rates between 83.9% and 117.9%, with precision (RSD) less than 7.5%, which demonstrated both high recovery and reliable accuracy. The analytical performance of this method, compared with that of other literatures for NTs analysis, revealed that the technique proposed holds multiple advantages, including the number of NTs able to be detected (1).

The authors of the study believe that their use of an external magnet for phase separation simplifies past extraction processes as well as sidesteps avoiding complex centrifugation steps. The method, under optimized conditions, presents, in their opinion, a broad linear detection range, very low detection limits, and excellent recovery performance, thus offering a technique is efficient, effective, and economically friendly, and exhibits strong potential for clinical analysis applications (1).

Spinal cord. © sorin - stock.adobe.com

REFERENCES

1. Fan, Z.; Yu, W.; Liu, Z. Ultra Performance Liquid Chromatography with Ultrasound Assisted Magnetic Ionic Liquid Dispersive Liquid Liquid Microextraction for Determination of 20 Neurotransmitters in Spinal Cords. Sci. Rep. 2025, 15 (1), 5151. DOI: 10.1038/s41598-025-89692-9

2. Olesti, E.; Rodríguez-Morató, J.; Gomez-Gomez, A,; Ramaekers, J. G.; de la Torre, R.; Pozo, O. J. Quantification of Endogenous Neurotransmitters and Related Compounds by Liquid Chromatography Coupled to Tandem Mass Spectrometry. Talanta 2019,192,93-102. DOI: 10.1016/j.talanta.2018.09.034

3. Zhou, W.; Zhu, B.; Liu, F.; Lyu, C.; Zhang, S.; Yan, C.; Cheng, Y.; Wei, H. A Rapid and Simple Method for the Simultaneous Determination of Four Endogenous Monoamine Neurotransmitters in Rat Brain Using Hydrophilic Interaction Liquid Chromatography Coupled with Atmospheric-Pressure Chemical Ionization Tandem Mass Spectrometry. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 2015, 1002, 379-386. DOI: 10.1016/j.jchromb.2015.08.042

4. Ye, H.; Wang, J.; Greer, T.; Strupat, K.; Li, L. Visualizing Neurotransmitters and Metabolites in the Central Nervous System by High Resolution and High Accuracy Mass Spectrometric Imaging. ACS Chem. Neurosci. 2013, 4 (7), 1049-1056. DOI: 10.1021/cn400065k

5. Matys, J.; Gieroba, B.; Jóźwiak. K. Recent Developments of Bioanalytical Methods in Determination of Neurotransmitters in vivo. J. Pharm. Biomed. Anal. 2020, 180, 113079. DOI: 10.1016/j.jpba.2019.113079

6. Salvadore, G.; van der Veen, J. W.; Zhang, Y.; Marenco, S.; Machado-Vieira, R.; Baumann, J.; Ibrahim, L. A.; Luckenbaugh, D. A.; Shen, J.; Drevets, W. C.; Zarate, C. A. Jr. An Investigation of Amino-Acid Neurotransmitters as Potential Predictors of Clinical Improvement to Ketamine in Depression. Int. J. Neuropsychopharmacol. 2012, 15 (8), 1063-1072. DOI: 10.1017/S1461145711001593

7. Marc, D. T.; Ailts, J. W.; Campeau, D. C.; Bull, M. J.; Olson, K. L. Neurotransmitters Excreted in the Urine as Biomarkers of Nervous System Activity: Validity and Clinical Applicability. Neurosci. Biobehav. Rev. 2011, 35 (3):635-644. DOI: 10.1016/j.neubiorev.2010.07.007