Quantitative determination of monoamine neurotransmitters in rat brain homogenates using HPLC-MS/MS

Relevance. Evaluation of the effect of drugs on neurotransmitter processes is an important component of pharmacodynamic studies. The quantitative determination of monoamine neurotransmitters in the brain structures of laboratory animals is an urgent task of pharmacology and physiology.Purpose of the study. Development of a method for the quantitative determination of serotonin, dopamine, norepinephrine, histamine and epinephrine in rat brain homogenates using HPLC-MS/MS.Methods. The isolation of neurotransmitters from the brain of rats was carried out by homogenizing the biomaterial with acetonitrile and hydrochloric acid. The extraction was purified by liquid-liquid extraction with chloroform and isopropanol. Monoamines were detected using an AB Sciex QTrap 3200MD mass spectrometer, chromatography was performed using an Agilent Technologies 1260 Infinity II HPLC. Methanol and deionized water were used as eluent.Results. Sample preparation consisted of centrifugation of the resulting homogenate, drying of the supernatant in a stream of nitrogen, dissolution of the precipitate in the mobile phase, and purification of the solution using a mixture of chloroform and isopropanol. An Agilent InfinityLab Poroshell 120 EC-C18 4.6×100 mm, 2.7 μm analytical column was used to separate monoamine neurotransmitters. The total time of the chromatographic analysis was 12 minutes, the retention time of norepinephrine, epinephrine, dopamine, serotonin, histamine was 2.8; 3.2; 5.4; 7.9; and 2.2 minutes, respectively. The analytical range of the technique was 25.0–5000.0 ng/g for epinephrine, histamine, and dopamine; 5.0–5000.0 ng/g for serotonin and 50.0–5000.0 for norepinephrine. To test the technique, we analyzed monoamine neurotransmitters in the striatum of intact Wistar rats.Conclusion. The developed bioanalytical HPLC-MS/MS method for the quantitative determination of monoamine neurotransmitters in the rat brain fully complies with the validation requirements. The metrological characteristics of the technique make it possible to estimate the content of norepinephrine, epinephrine, dopamine, serotonin, and histamine in the brain structures of rats with high accuracy.

[1]  A. Starkweather,et al.  Associations of neurotransmitters and the gut microbiome with emotional distress in mixed type of irritable bowel syndrome , 2022, Scientific Reports.

[2]  Peter A. Johnson,et al.  Maladaptive or misunderstood? Dopamine fasting as a potential intervention for behavioral addiction , 2021, Lifestyle Medicine.

[3]  Chung S. Yang,et al.  An Unrecognized Fundamental Relationship between Neurotransmitters: Glutamate Protects against Catecholamine Oxidation , 2021, Antioxidants.

[4]  Di Chen,et al.  A simultaneous extraction/derivatization strategy coupled with liquid chromatography-tandem mass spectrometry for the determination of free catecholamines in biological fluids. , 2021, Journal of chromatography. A.

[5]  Zheng-Wei Zhang,et al.  Determination and Application of Nineteen Monoamines in the Gut Microbiota Targeting Phenylalanine, Tryptophan, and Glutamic Acid Metabolic Pathways , 2021, Molecules.

[6]  N. Conejo,et al.  Sex-Specific Effects of Early Life Stress on Brain Mitochondrial Function, Monoamine Levels and Neuroinflammation , 2020, Brain sciences.

[7]  V. Narkevich,et al.  The Levels of Monoamines and Their Metabolites in the Brain Structures of Rats Subjected to Two- and Three-Month-Long Social Isolation , 2020, Bulletin of Experimental Biology and Medicine.

[8]  S. Jung-Klawitter,et al.  Analysis of Catecholamines and Pterins in Inborn Errors of Monoamine Neurotransmitter Metabolism—From Past to Future , 2019, Cells.

[9]  P. Lorkiewicz,et al.  Comprehensive, robust, and sensitive UPLC-MS/MS analysis of free biogenic monoamines and their metabolites in urine. , 2018, Journal of chromatography. B, Analytical technologies in the biomedical and life sciences.

[10]  M. Akhtar,et al.  Histamine H3 receptor antagonists display antischizophrenic activities in rats treated with MK-801 , 2016, Journal of basic and clinical physiology and pharmacology.

[11]  D. Vohora,et al.  A validated HPLC-UV method and optimization of sample preparation technique for norepinephrine and serotonin in mouse brain , 2015, Pharmaceutical biology.

[12]  N. Zilka,et al.  Liquid chromatography-tandem mass spectrometry method for determination of panel of neurotransmitters in cerebrospinal fluid from the rat model for tauopathy. , 2014, Talanta.

[13]  H. Stark,et al.  Microdialysate analysis of monoamine neurotransmitters--a versatile and sensitive LC-MS/MS method. , 2013, Analytica chimica acta.

[14]  M. Zoli,et al.  Simultaneous measurement of adenosine, dopamine, acetylcholine and 5-hydroxytryptamine in cerebral mice microdialysis samples by LC-ESI-MS/MS. , 2012, Journal of pharmaceutical and biomedical analysis.

[15]  R. Šlamberová,et al.  Monitoring of dopamine and its metabolites in brain microdialysates: method combining freeze-drying with liquid chromatography-tandem mass spectrometry. , 2011, Journal of chromatography. A.

[16]  Dan-ni Zhu,et al.  HPLC/MS/MS for quantification of two types of neurotransmitters in rat brain and application: myocardial ischemia and protection of Sheng-Mai-San. , 2011, Journal of pharmaceutical and biomedical analysis.

[17]  C. Hoppel,et al.  Quantification of carnitine and acylcarnitines in biological matrices by HPLC electrospray ionization-mass spectrometry. , 2008, Clinical chemistry.

[18]  R. Kostiainen,et al.  Analysis of acetylcholine and choline in microdialysis samples by liquid chromatography/tandem mass spectrometry. , 2005, Rapid communications in mass spectrometry : RCM.

[19]  P J Kontur,et al.  Brain regional manganese levels and monoamine metabolism in manganese-treated neonatal rats. , 1988, Neurotoxicology and teratology.

[20]  T. Greve,et al.  An automated direct-injection HPLC-method for the electrochemical/fluorimetric quantitation of monoamines and related compounds optimized for the screening of large numbers of animals. , 1986, Biomedical chromatography : BMC.