高效液相色谱荧光检测流速梯度法测定去卵巢小鼠脑内单胺类神经递质张雷pdf.docVIP

2005 年 1 月 色 谱 Vol .23 No.1 January 2005 Chinese Journal of Chromatography 76 ～ 78 高效液相色谱-荧光检测流速梯度法测定去卵巢小鼠脑内单胺类神经递质 张 雷, 卢英俊, 邓同乐, 郑筱祥 (浙江大学生物医学工程系, 浙江杭州 310027) 摘要:建立了同时检测去卵巢(OVX)小鼠脑内的去甲肾上腺素(NE)、多巴胺(DA)、 5-羟色胺(5-HT)3 种神经递质 的高效液相色谱方法。 采用的色谱条件为:C1 8柱分离, 荧光检测器检测, 流动相为 0.1 mol/L 的 KH2 PO4 缓冲液-甲醇(体积比为 9 1), 设置单泵流速梯度洗脱。 方法的检测限(S/ N =3)均在 10 nmol/ L以下, 绝对回收率均在 80 % 以上 , 样品日内测定峰面积的相对标准偏差在 3.03%以下。该方法具有检测灵敏度高、样品制备简便等优点, 对小鼠脑内的 3 种单胺类递质的检测具有实用价值。 关键词:高效液相色谱;流速梯度;去甲肾上腺素;多巴胺;5-羟色胺;去卵巢小鼠 中图分类号:O658 文献标识码:A 文章编号:1000-8713(2005)01-0076-03 Measurement of Monoamine Transmitters in the Brains of Ovariectomized Mice Using High Performance Liquid Chromatography with Fluorescence Detection and Gradient Flow Rate Elution ZHANG Lei, LU Yingjun, DENG Tongle , ZHENG Xiaoxiang (Department of Biomedical Engineering of Zhejiang University , Hangzhou 310027 , China) Abstract :A method for the determination of norepinephrine (NE), dopamine (DA)and 5-hydrox-ytryptamine (5-HT)in brains of ovariectomized mice was established by using high performance liquid chromatograph with a fluorescence detector , a C18 column and a gradient flow rate delivered by a single pump.The mobile phase was a mixture of 0.1 mol/L KH2PO4 and CH3OH (9/1 ,v/v). Detection limits of the method were determined to be lower than 10 nmol/L , absolute recoveries were above 80 %and the relative standard deviations (RSDs)of intra-day were all within 3.03 %. The method has the advantages of high sensitivity and of simple sample preparation .Short analysis time and better separation of samples were also obtained. Key words :high performance liquid chromatography ;gradient flow rate;norepinephrine; dopamine ;5-hydroxytryptamine;ovariectomized mice  单胺类神经递质在中枢神经系统中参与脑缺血、镇痛、学习记忆等多种生理过程 ,具有重要的生理意义。该类神经递质包括儿茶酚胺和吲哚胺等 , 前者主要指去甲肾上腺素(NE)和多巴胺(DA), 后者主要指 5-羟色胺(5-HT)[1] 。国内关于 NE ,DA 和 5-HT 3 种神经递质的检测多采用高效液相色谱-电化学分析法(HPLC-ECD)[2 ～ 6] , 但由于 ECD 对流动相变化较为敏感, 故较少采用梯度洗脱。梯度洗脱中以流动相的浓度变化最为常见 ,而以流动相的流速梯度的应用在文献中少有报道。本文采用高效液  相色谱-荧光检测法 ,尝试设置单