【作者】 王清江；

【导师】 方禹之； 何品刚；

【作者基本信息】 华东师范大学 ， 分析化学， 2004， 博士

【摘要】 毛细管电泳（Capillary Electrophoresis，CE）是近二十年来发展最快的分离分析技术之一，它将分离柱效提高到上百万理论塔板数，进样量也从微升水平进入纳升水平，其研究和应用涉及环境分析、药物分析、生化分析等几乎所有的分析化学领域。但毛细管电泳较小的进样量和极细的毛细管通道内径也给检测也带来了困难。紫外、激光诱导荧光、质谱等各种检测方法都具有一定的优点，但往往存在仪器比较昂贵、衍生化步骤烦琐等缺点。电化学检测中的安培检测技术，具有比紫外检测更高的灵敏度，且仪器简单、价格成本低、线性范围宽、操作简便，因而其与毛细管电泳联用后在分析化学领域得到了广泛的研究和应用。 同高效液相色谱分离相比，毛细管电泳在分离效率和仪器等方面有明显的优势。但由于毛细管电泳进样量远远小于液相色谱，利用紫外或荧光检测时光程很短，导致浓度检测灵敏度较低，不能满足有些痕量分析的要求。利用毛细管电泳本身的一些特殊性质，通过改变其进样和分离时的一些条件，如采用电堆集、场放大进样和等速电泳等简单易行的方法，可以显著提高毛细管电泳检测灵敏度。 本论文的目的之一是研究毛细管电泳-安培检测联用技术（CE-AD）中提高灵敏度的方法，包括场放大进样和等速电泳等。论文也探讨了毛细管电泳-安培检测联用技术在药物分析、生化分析、食品分析中的一些应用。主要内容共分为六章： 第一章为绪论。在该章中综述了毛细管电泳的特点、分离模式和理论基础、联用的检测器、提高灵敏度的方法、毛细管电泳-安培检测的理论研究和应用现状等，并简单介绍了本论文的目的意义。 第二章共分为两节。在第一节中探讨了毛细管电泳-安培检测法测定复方降压中药珍菊降压片中两个含量较高的有效成分：芦丁和氢氯噻嗪。在最佳条件下，这两种分析物在12min内能够被很好的检测出，在碳电极上也有较大的电流响应，检测灵敏度分别是7.02×10⁴ nA·L／mol和2.17×10⁵ nA·L／mol，检测限分别是5.0×10^-7molL^-1和2.0×10^-7molL^-1（S／N=3）。应用该联用技术成功测定了这两种有效成分体内代谢后在尿液中的含量。 在第二节中探讨了使用不同稀释剂时毛细管电泳-安培检测中的场放大进样摘要现象。我们通过研究用不同稀释溶剂对在水溶液中分别带正电荷、中性电荷、负电荷的盐酸可乐定、氢氯唆嗓和芦丁三种组分稀释后电流响应的变化，探索了在毛细管电泳电动进样时稀释溶剂对场放大进样的影响规律。从实验中发现:a.改变样品溶液的酸度，可以实现带正电荷、负电荷的离子同时进行场放大进样，该法比以往改变电极极性的方法简单易行;b.使用更高的缓冲溶液稀释样品，也可以使带负电荷的离子达到场放大进样; c.通过选择合适的稀释溶剂，可以调节同一样品中不同成分的检测灵敏度，使其电流响应可以达到同一数量级水平。如在珍菊降压片中，盐酸可乐定、氢氯唆嗦和芦丁三种有效成分的相对含量相差上百倍，若用缓冲溶液稀释样品，其电流响应难以在同一张电泳图上反映出，但用水做稀释剂，它们的电流响应可以在同一张电泳图上直观地反映出来。 第三章包括两节内容。在第一节中讨论了毛细管区带电泳一安培检测法间接测定Fent。n反应中的经基自由基的条件。在3ommol·L一’N处B4o7一Hel（pH 7.4）缓冲溶液中，实现了水杨酸、2，3一DHBA和2，5一DHBA的完全分离，检测限达到了10一smol/1数量级。并且通过对当归多糖对轻基自由基的清除能力的研究，证明该方法可用来鉴定不同药物对轻自由基的清除能力。 在第二节中探讨毛细管电泳一安培检测中的等速电泳富集现象。由于人体内新陈代谢产生的自由基的浓度极低，如在尿样中轻自由基捕获剂的浓度在10月mol/1数量级，所以毛细管电泳一安培检测法尚不能直接进行生化样品中轻自由基的测定。我们尝试了用毛细管瞬时等速电泳的方法对2，3一DHBA进行了管内富集，选用对甲基苯甲酸为前导电解质，运行缓冲溶液为终止电解质，使2，3一DHBA的检测限降低了20倍，在灵敏度上达到了进行生化样品中轻自由基分析的要求。本文对适合于毛细管电泳一电化学检测特性的等速电泳条件也作了初步的探讨。 第四章利用胶束电动毛细管电泳一安培检测联用同时测定了食品中的活性胺及其前体氨基酸。我们以碳圆盘电极为工作电极，用胶束电动毛细管电泳一安培检测法（MEcc一AD）同时分离和检测食品中生物活性胺及其前体氨基酸，即色胺、酪胺、色氨酸和酪氨酸。所建立的方法简单、可靠、重现性较好。分离和检测的最佳条件是:o.oZmol·L一‘硼酸钠（NaZBZo7）一氢氧化钠（NaoH）缓冲溶液（PH10.35）作为运行缓冲液，其中含0.03 mol.L一，十二烷基硫酸钠（SDS）;分离电压为华东师范大学2004年申请博士学位论文摘要20kV;检测电位为+0 .80V（VS.SCE）。在优化条件下，色胺、酪胺、色氨酸和酪氨酸得到完全分离，在三个数量级范围内，峰电流与待测物呈良好的线性关系，检测限（S/N=3）达到1.0xl0一7mol.L一‘。将建立的方法用于实际样品米酒的测定，测定结果令人满意。 第五章应用毛细管电泳一安培检测测定中药杂多糖?更多还原

【Abstract】 Capillary Electrophoresis (CE) is one of the most important separation techniques in recent twenty years. Because the theoretical plate number is great developed to tens of thousands and even millions and the volume of sample injection reaches nano milliliter level in CE analysis, it is studied and applied to most analytical areas. However, the small sample injection volume and very thin capillary bring about difficulties to detections. The common used detection methods are ultra-violet (UV), laser induced fluorescence (LIF), mass spectra (MS) and electrochemical detection (ED). Every detection method performs some advantages and suffers from some shortages too. For example, LIF and MS are very sensitive but the instrumentations are very expensive, and some complicated derivation procedures are needed. Amperometric detection (AD), one kind of electrochemical detections, is more sensitive than the common used UV detection, and have many merits when coupled with CE, such as simple instrumentation and operation, low cost, wide linear range, etc. The combination of CE with AD is extensively studied and applied in most analytical fields for its above advantages.The advantages of CE in separation efficiency and instrumentation are obvious when compared to HPLC. However, the concentration detection sensitivity of CE is much lower than HPLC owing to its smaller sampling volume and the short light distance when coupled with UV or FI detectors. The detection sensitivity of CE could be great improved by some special characters itself such as field-amplification-injection and isotachophoresis.One goal of this dissertation is to explore the methods of improving the sensitivity of CE-AD. Another one is applying this CE-AD technique to resolve some actual analytical problems in pharmaceutical analysis, biochemical analysis and food analysis. The contents of this dissertation include six chapters:In the first chapter, the characteristics of CE, the separation models and basictheories of CE, the detectors, the sensitivity-enhanced methods, the studies and applications of CE-AD simply is introduced. The goal and significance of this dissertation are introduced too.Chapter 2 includes two sections. In the first section, capillary zone electrophoresis (CZE) with amperometric detection (AD) was firstly applied to the simultaneous determination of rutin (RT) and hydrochlorothiazide (HCT) in compound Chinese herb medicines and human urine samples. The two analytes could be perfectly analyzed within 12 min and showed significant current responses at carbon electrode under the optimum conditions and their detection limits were 5.0x10-7 molL-1 and 2.0x10-7 molL-1 respectively (S/N=3).In the second section of this chapter, we explored the effects of diluents during electro-kinetic sampling on field amplification injection by studying the changes of current responses of clonidine hydrochloride, hydrochlorothiazide and rutin which are originally positive, neutral, negative respectively in water solution when diluted in different solvents. It was found that the simultaneous field amplification injection for positive and negative particles could be carried out by changing the acidity of samples. This method is much easier than converting electricity polarity. The effect for negative particles could be also reached by using buffer solution with higher concentration as diluents. The detection sensitivity of different ingredients existed in one sample could be adjusted and then their current responses could be reach the same magnitude level by using suitable diluents. For example, the current responses of clonidine hydrochloride, hydrochlorothiazide and rutin are difficult to compare clearly in the same electropherogram if using the running buffer as diluent because their contents are different in one hundred times. But, their current responses could be compared clearly in one electropherogram if water is used as the diluent.Chapter 3 includes two sections too. In the first section, the indirect determination of hydroxyl radical (OH) b更多还原