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建立并优化了一种简单、灵敏、高选择性的高效液相色谱荧光检测法,可同时定量检测大鼠不同脑亚区中的单胺类神经递质及其代谢产物(左旋多巴,去甲肾上腺素,肾上腺素,多巴胺,5-羟色胺,二羟基苯乙酸,高香草酸和5-羟基吲哚乙酸)。在这种新方法中,所有待测物均采用未经过衍生化步骤而直接提取,并在20分钟内达到良好分离。荧光检测的激发波长280 nm,发射波长330 nm。色谱柱:Agilent Eclipse Plus C18柱(美国Agilent公司,4.6 mm×150 mm,5.0μm),保护柱:Agilent XDB-C18(美国Agilent公司,4.6 mm×12.5 mm,5.0μm),柱温:35 oC。流动相:柠檬酸–乙酸钠–辛烷磺酸钠缓冲体系(50 mmol/L柠檬酸,50 mmol/L无水乙酸钠,0.5 mmol/L辛烷磺酸钠,0.5 mmol/L Na2-EDTA,5 mmol/L三乙胺)–甲醇(90:10,v/v,p H 3.8),等度洗脱,流速:1.0 mL/min。所有待测物的检出限为0.9–23 nM,进样体积为50μL。该方法重现性良好,日内精密度在0.08%–1.85%之间(n=5)。对这些物质的同时测定可加深我们对不同成瘾性药物(甲基苯丙胺,海洛因及其二者的混合物)在大鼠中枢神经系统中神经生化作用机制的理解。
A simple, sensitive and highly selective high performance liquid chromatography (HPLC) fluorescence detection method was established and optimized to simultaneously detect monoamine neurotransmitters and their metabolites (levodopa, go Epinephrine, epinephrine, dopamine, serotonin, dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindoleacetic acid). In this new method, all analytes were extracted directly without derivatization and resolved well in 20 minutes. Fluorescence detection excitation wavelength 280 nm, emission wavelength 330 nm. Column: Agilent Eclipse Plus C18 column (4.6 mm × 150 mm, 5.0 μm, Agilent, USA), guard column: Agilent XDB-C18 . Mobile phase: sodium citrate-sodium acetate-octane sulfonate buffer system (50 mmol / L citric acid, 50 mmol / L anhydrous sodium acetate, 0.5 mmol / L sodium octane sulfonate, , 5 mmol / L triethylamine) -methanol (90:10, v / v, p H 3.8), isocratic elution, flow rate: 1.0 mL / min. The limit of detection for all analytes was 0.9-23 nM and the injection volume was 50 μL. The reproducibility of the method was good with intra-day precision of 0.08% -1.85% (n = 5). Simultaneous determination of these substances can deepen our understanding of the mechanisms of neurobiochemical effects of different addictive drugs (mixtures of methamphetamine, heroin and both) in the rat central nervous system.