毛细管电泳分析尿神经递质：场放大样品进样和分子印迹聚合物固相萃取提高灵敏度。
Analysis of urinary neurotransmitters by capillary electrophoresis: sensitivity enhancement using field-amplified sample injection and molecular imprinted polymer solid phase extraction.
机构信息
Institut de Chimie Organique et Analytique, CNRS FR UMR, Université d'Orléans, France.
出版信息
Anal Chim Acta. 2011 Aug 12;699(2):242-8. doi: 10.1016/j.aca.2011.05.014. Epub 2011 May 17.
Capillary electrophoresis (CE) has been investigated for the analysis of some neurotransmitters, dopamine (DA), 3-methoxytyramine (3-MT) and serotonin (5-hydroxytryptamine, 5-HT) at nanomolar concentrations in urine. Field-amplified sample injection (FASI) has been used to improve the sensitivity through the online pre-concentration samples. The cationic analytes were stacked at the capillary inlet between a zone of low conductivity - sample and pre-injection plug - and a zone of high conductivity - running buffer. Several FASI parameters have been optimized (ionic strength of the running buffer, concentration of the sample protonation agent, composition of the sample solvent and nature of the pre-injection plug). Best results were obtained using H(3)PO(4)-LiOH (pH 4, ionic strength of 80 mmol L(-1)) as running buffer, 100 μmol L(-1) of H(3)PO(4) in methanol-water 90/10 (v/v) as sample solvent and 100 μmol L(-1) of H(3)PO(4) in water for the pre-injection plug. In these conditions, the linearity was verified in the 50-300 nmol L(-1) concentration range for DA, 3-MT and 5-HT with a determination coefficient (r(2)) higher than 0.99. The limits of quantification (10 nmol L(-1) for DA and 3-MT, 5.9 nmol L(-1) for 5-HT) were 500 times lower than those obtained with hydrodynamic injection. However, if this method is applied to the analysis of neurotransmitters in urine, the presence of salts in the matrix greatly reduces the sensitivity of the FASI/CE-UV method.Therefore, a solid phase extraction (SPE) on a dedicated imprinted polymer (MIP) was developed to extract specific neurotransmitters, catecholamines, metanephrines and indolamines, from urine. Matrix salts were thus discarded after sample extraction on AFFINIMIP™ Catecholamine & Metanephrine (100mg) cartridge. Therefore, lower limits of quantification were determined in artificial urine (46 nmol L(-1) for DA, 11 nmol L(-1) for 3-MT and 6 nmol L(-1) for 5-HT).The application of this protocol MIP-SPE/FASI-CE-UV analysis of neurotransmitters in human urine gave rise to electropherograms with a very good base line and signal to noise ratios above 15.
毛细管电泳（CE）已被用于分析一些神经递质，如多巴胺（DA）、3-甲氧基酪胺（3-MT）和 5-羟色胺（5-HT），其浓度低至纳摩尔水平。场放大样品进样（FASI）已被用于通过在线预浓缩样品来提高灵敏度。阳离子分析物在毛细管入口处堆积，位于低电导率区-样品和预进样塞-和高电导率区-运行缓冲液之间。已经优化了几个 FASI 参数（运行缓冲液的离子强度、样品质子化剂的浓度、样品溶剂的组成和预进样塞的性质）。使用 H 3 PO 4 -LiOH（pH 4，离子强度 80mmol L -1 ）作为运行缓冲液、甲醇-水 90/10（v/v）中 100μmol L -1 的 H 3 PO 4 作为样品溶剂和水 100μmol L -1 的 H 3 PO 4 作为预进样塞时，可获得最佳结果。在这些条件下，DA、3-MT 和 5-HT 的线性验证范围为 50-300nmol L -1 ，测定系数（r 2 ）高于 0.99。DA 和 3-MT 的定量限（10nmol L -1 ）和 5-HT（5.9nmol L -1 ）比动态注射法低 500 倍。然而，如果将该方法应用于尿液中神经递质的分析，基质中的盐会大大降低 FASI/CE-UV 方法的灵敏度。因此，开发了一种基于专用印迹聚合物（MIP）的固相萃取（SPE），从尿液中提取特定的神经递质，儿茶酚胺、代谢物和吲哚胺。因此，在 AFFINIMIP™儿茶酚胺和代谢物（100mg）萃取小柱上进行样品萃取后，基质盐被丢弃。因此，在人工尿液中确定了较低的定量限（DA 为 46nmol L -1 ，3-MT 为 11nmol L -1 ，5-HT 为 6nmol L -1 ）。应用该方案，通过 MIP-SPE/FASI-CE-UV 分析人尿中的神经递质，得到了基线非常好、信噪比大于 15 的电泳图谱。
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