A high-performance liquid chromatography with fluorescence detection method for the simultaneous quantitation of monoamine neurotransmitters and their metabolites in subregions of rat brain

doi:10.5246/jcps.2015.07.059

Abstract

Abstract:

In the present study, wesimultaneously quantified the levels of monoamine neurotransmitters (MANTs) and their metabolites (levodopa, norepinephrine, epinephrine, dopamine, 5-HT, 3,4-dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindole-3-acetic acid) in different brain subregions of rats using a newly developed simple, sensitive and selective high-performance liquid chromatography with fluorescence detection (HPLC-FLD) method. In this new HPLC-FLD method, analytes were directly extracted and separated without deriveatization step within 20 min. The FLD wavelength was set at 280 nm and 330 nm for excitation and emission, respectively. The analytes were separated on an Agilent Eclipse Plus C₁₈ column (4.6 mm×150 mm, 5.0 μm) equipped with an Agilent XDB-C₁₈ security guard column (4.6 mm×12.5 mm, 5.0 μm), and the column temperature was maintained at 35 ºC. The mobile phase for elution was isocratic. The mobile phase consisted of citric acid buffer (50 mmol/L citric acid, 50 mmol/L sodium acetate, 0.5 mmol/L octane sulfonic acid sodium salt, 0.5 mmol/L Na₂EDTA and 5 mmol/L triethylamine, pH 3.8) and methanol (90:10, v/v) at a flow rate of 1.0 mL/min. The detection limit (DL) was 0.9–23 nM for all the MANTs and their metabolites with a sample volume of 50 μL. The method was shown to be highly reproducible in terms of peak area (intraday, 0.08%–1.85% RSD, n = 5). The simultaneous measurement of these MANTs and their metabolites improved our understanding of the neurochemistry in the central nervous system (CNS) in relation to different addictive drugs (methamphetamine, heroin and their mixture) in drug-addicted rat models.

Key words: High performance liquid chromatography, Fluorescence detection, Monoamine neurotransmitters, Addictive drug, Brain subregions, Corticolimbic system

CLC Number:

R917

Supporting:

Peng Xu, Yanping Bai, Haisong Yang, Jing Li, Wei Lu, Xiaomei Ling. A high-performance liquid chromatography with fluorescence detection method for the simultaneous quantitation of monoamine neurotransmitters and their metabolites in subregions of rat brain[J]. Journal of Chinese Pharmaceutical Sciences, 2015, 24(7): 458-466.

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