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高效液相色谱-荧光法测定小鼠脑组织中单胺递质及其相关物质
引用本文:张璐璐, 冯芳, 叶小敏, 朱杰, 马冯飞. 高效液相色谱-荧光法测定小鼠脑组织中单胺递质及其相关物质[J]. 中国药科大学学报, 2010, 41(4): 367-371.
作者姓名:张璐璐  冯芳  叶小敏  朱杰  马冯飞
作者单位:1.中国药科大学药物分析教研室;2.药物质量与安全预警教育部重点实验室
基金项目:国家自然科学基金资助项目(No.30973858)
摘    要:建立高效液相色谱-荧光分析方法,研究小鼠脑皮层、海马、纹状体、丘脑等不同部位的单胺类神经递质及其相关物质。脑组织样本经高氯酸沉淀蛋白后进行色谱分析,采用Lichrospher C18柱(250 mm×4.6 mm,5 μm),流动相为pH 3.06缓冲液(含1 mmol/L庚烷磺酸钠;30 mmol/L磷酸二氢钠-磷酸溶液)-甲醇(85∶15);流速:1.0 mL/min;柱温:35 ℃。荧光检测:激发波长为280 nm,发射波长为315 nm。采用建立的方法,鉴别出小鼠脑中存在去甲肾上腺素(NE)、多巴胺(DA)、5-羟色胺(5-HT)、酪氨酸、色氨酸、3,4-二羟苯乙酸和5-羟吲哚乙酸等物质;对NE、DA和5-HT进行定量研究,确定它们的线性范围分别为2.4~471.0 ng/mL(r=0.999 6),2.6~519.5 ng/mL(r=0.999 4),2.4~469.3 ng/mL(r=0.999 1),日内、日间RSD均小于9.0%,准确度达到限度要求,提取回收率大于90%。本法专属性好,准确度高,可以反映不同鼠脑区域单胺类神经递质及相关物质含量上的差异。

关 键 词:高效液相色谱-荧光分析  单胺类神经递质  相关物质  脑组织

Determination of monoamine neurotransmitters and related substances in mouse brain tissue by HPLC-FLD
Determination of monoamine neurotransmitters and related substances in mouse brain tissue by HPLC-FLD[J]. Journal of China Pharmaceutical University, 2010, 41(4): 367-371.
Abstract:To establish an HPLC-FLD method for the investigation of monoamine neurotransmitters and related substances in mouse frontal cortex,hippocampus,striatum,and hypothalamus.Mouse brain samples were deproteinized by perchloric acid and analyzed.The separation was carried out on a Lichrospher C18 column(250 mm×4.6 mm,5 μm).The mobile phase was composed of pH 3.06 buffer(including 1 mmol/L heptanesulfonic acid sodium salt,30 mmol/L NaH2PO4-H3PO4 solution) and methanol in the ratio of 85∶15;the flow rate was 1.0 mL/min and the column temperature was 35 °C.Fluorescence detection:the excitation and emission wavelengths were set at λex=280 nm,λem=315 nm.Under the optimized conditions,norepinephrine(NE),dopamine(DA),5-hydroxytryptamine (5-HT),3,4-dihydroxy-phenyl acetic acid(DOPAC),5-hydroxyindoleacetic acid(5-HIAA),tyrosine(Tyr),and tryptophan(Trp) in mouse brain were detected.NE,DA,and 5-HT were quantified.The linearity ranges of NE,DA, and 5-HT were 2.4-471.0 ng/mL(r=0.999 6),2.6-519.5 ng/mL (r=0.999 4),and 2.4-469.3 ng/mL(r=0.999 1),respectively.The intra- and inter-day precision was always lower than 9.0%. The accuracy could attain the demands and the absolute recovery values were always higher than 90%. The proposed method has good specificity and accuracy.It could be used to analyze the content differences of monoamine neurotransmitters and related substances in different mouse brain areas.
Keywords:HPLC-FLD  monoamine neurotransmitter  related substances  brain tissue
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