Development and Full Validation of a Novel Liquid Chromatography Electrochemical Detection Method for Simultaneous Determination of Nine Catecholamines in Rat Brain

IF 1.8 Q3 PHARMACOLOGY & PHARMACY Turkish Journal of Pharmaceutical Sciences Pub Date : 2022-12-28 DOI:10.4274/tjps.galenos.2022.06606
Saniye Özcan, Murat Kozanlı, Aysun Geven, Nafiz Öncü Can
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Abstract

INTRODUCTION: Chemical neurotransmission, managed by neurotransmitters, has a crucial role in brain processes such as fear, memory, learning, and pain, or neuropathologies such as schizophrenia, epilepsy, anxiety/depression, and Parkinson’s disease. The measurement of these compounds is to elucidate the disease mechanisms and evaluate the outcomes of therapeutic interventions. However, this can be quite difficult due to various matrix effects and the problems of chromatographic separation of analytes. In the current study; for the first time, an optimized and fully validated fully according to FDA and EMA Bioanalytical Validation Guidance HPLC-EC method was developed for the simultaneous analysis of nine neurotransmitter compounds which are dopamine (DA), homovanilic acid (HVA), vanilmandelic acid (VA) serotonin (SER), 5-hydroxyindole-3-acetic acid (5-HIAA), 4-hydroxy-3-methoxyphenylglycol (MHPG), norepinephrine (NE), 3,4 dihydroxyphenylacetic acid (DOPAC) and 3-methoxytyramine (3-MT) and simultaneously determined in rat brain samples. METHODS: The separation was achieved with 150 mm × 4.6 mm, 2.6 μm F5 Kinetex (Phenomenex, USA) column isocratically, and analysis was carried out HPLC equipped with DECADE II electrochemical detector. RESULTS: The method exhibited good selectivity and correlation coefficient values for each analyte’s calibration curves were >0.99. The detection and quantification limits ranged from 0.01 to 0.03 ng/mL and 3.04 to 9.13 ng/mL, respectively. The stability of the analytes and method robustness were also examined in detail in the study, and the obtained results are also presented statistically. DISCUSSION AND CONCLUSION: The developed fully validated method has been successfully applied to real rat brain samples and important results have been obtained. In the rat brain sample analysis, the least amount of SER and the highest amount of NA were found.
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一种同时测定大鼠脑中9种儿茶酚胺的新型液相色谱-电化学检测方法的建立和充分验证
导读:由神经递质控制的化学神经传递在诸如恐惧、记忆、学习和疼痛等大脑过程或精神分裂症、癫痫、焦虑/抑郁和帕金森病等神经病理中起着至关重要的作用。这些化合物的测量是为了阐明疾病机制和评估治疗干预的结果。然而,由于各种基质效应和分析物的色谱分离问题,这可能相当困难。在目前的研究中;根据FDA和EMA生物分析验证指南,首次建立了一种优化的HPLC-EC方法,用于同时分析多巴胺(DA)、香草酸(HVA)、香草酸(VA)、血清素(SER)、5-羟基吲哚-3-乙酸(5-HIAA)、4-羟基-3-甲氧基苯基乙二醇(MHPG)、去甲肾上腺素(NE)、3,4二羟基苯乙酸(DOPAC)和3-甲氧基酪胺(3- mt),同时测定大鼠脑样品。方法:采用150 mm × 4.6 mm, 2.6 μm F5 Kinetex (Phenomenex, USA)柱等柱分离,配备DECADE II型电化学检测器进行HPLC分析。结果:该方法具有良好的选择性,各分析物的标度曲线相关系数均为0.99。检测限为0.01 ~ 0.03 ng/mL,定量限为3.04 ~ 9.13 ng/mL。研究中还对分析物的稳定性和方法的稳健性进行了详细的检验,并对所得结果进行了统计分析。讨论与结论:本方法已成功应用于实际大鼠脑样品,并获得重要结果。在大鼠脑样品分析中,发现SER含量最少,NA含量最高。
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CiteScore
3.60
自引率
5.90%
发文量
79
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