醛类、酸类化合物的毛细管电泳分离研究

【作者】 白新伟；

【导师】 尤进茂；

【作者基本信息】 曲阜师范大学 ， 分析化学， 2010， 硕士

【摘要】 高效毛细管电泳是离子或荷电离子以电场为驱动力,在毛细管中按其淌度和分配系数不同进行高效、快速分离的一种电泳新技术。它具备高效、快速、样品用量少,仪器的自动化程度高、操作简单、试剂消耗少、环境污染小等优点。正因为HPCE具备如此诱人的优点,使其在生命科学、生物技术、医学药物、环境科学、法医科学等领域都显示了极其重要的应用前景,在十几年的时间中,毛细管电泳技术发展迅速,特别是在最近几年,受到分离分析科学家的极大关注,成为生物化学和分析化学中最受瞩目,发展最快的一种分离分析新技术。有毒化学品优先控制名单上部分醛类位居前列,是公认的变态反应源,所以醛类的研究受到人们越来越多的关注。芳香族羧酸化合物在食品、药物等生产中有着很广泛的应用。其中苯甲酸钠常作为调味料中的食品防腐剂,但是用量过多会对人体有较大的毒害作用,因此测定食品中芳香族羧酸类化合物含量具有重要的意义。脂肪酸在自然界中分布广泛,是生物体内重要的营养代谢物质,对于调节某些代谢作用和生理功能起着重要作用。胆汁酸广泛存在于各种动物胆汁中,能够乳化水中的疏水分子利于消化吸收,平衡胆固醇,消化脂肪,防止胆结石的形成因此对两者含量的测定对进一步深入研究生理学和病理学以及临床诊断具有重要的意义。在醛类和酸类的研究领域,由于这些生物分子没有或者仅有微弱的紫外吸收,直接紫外检测满足不了低含量组分测定的要求,因此化学衍生化法应运而生,进行了脂肪醛、芳香酸、脂肪酸、胆汁酸的毛细管电泳分离,结果令人满意。论文分三章：第一章简单介绍了毛细管电泳的发展及其在药物分析、生物分子分离、基因工程研究、临床医学等各方面的应用,并对毛细管电泳分离被分析物的模式及其相关的基础知识进行了阐述。第二章以9,10-菲醌作柱前衍生试剂,采用非水毛细管电泳模式考察并优化了脂肪醛的分离条件。不加其他添加剂可实现七种脂肪醛高效基线分离,并在最优化条件下对实际样品进行了测定,结果令人满意。第三章7-氨基-1,3-萘二磺酸(ANDSA)作为柱前衍生试剂用于酸类物质的毛细管电泳分离研究,分为四部分。3.1以7-氨基-1,3-萘二磺酸作柱前衍生试剂,采用区带毛细管电泳模式,在15 mmol/Lβ-CD, pH9.5,30 mmol/L硼酸盐缓冲体系下,采用58.5 cmx50μm i.d毛细管(有效柱长50 cm),20℃,50 mbar,8 s进样,可实现10种苯甲酸及其一元取代物的快速高效基线分离。并在最优化条件下对生抽、老抽样品中苯甲酸进行定量检测,结果令人满意。3.2以7-氨基1,3-萘二磺酸作柱前衍生试剂,采用区带毛细管电泳模式考察并优化了C1～C10十种短链脂肪酸衍生化产物的分离条件。在pH为9.5的30 mmol/L硼酸盐缓冲体系下,20℃,50mbar压力进样,进样8 s。17分钟内可实现快速高效基线分离,检测限0.027-0.042线性范围0.07-5μmol/L,并在最优化条件下对葫芦藓、卷柏实际样品进行了定性定量检测,结果令人满意。3.3建立了一种用于测定苔藓类植物中长链脂肪酸的毛细管区带电泳-紫外检测方法(CZE-UV)。用硼砂缓冲溶液为背景电解质,经7-氨基-1,3-萘二磺酸柱前衍生的10种脂肪酸在18 min内可实现快速高效的基线分离和测定。在最优化条件下对葫芦藓实际样品进行的定性定量检测实验,结果令人满意。3.4以7-氨基-1,3-萘二磺酸作柱前衍生试剂,采用区带毛细管电泳光二极管阵列检测器分离测定6种胆汁酸。实验综合考察了缓冲液浓度、pH、温度及添加剂对胆汁酸衍生化产物分离效果的影响。在pH为9.5的25 mmol/L硼酸盐缓冲体系下,20℃,5×107Pa压力进样,进样时间8 s,不加其他添加剂情况下,13 min可实现6种胆汁酸的快速高效基线分离。更多还原

【Abstract】 High-performance capillary electrophoresis is a kind of new electrophoresis technology with high efficiency and rapid separation, which carries out separation by a ion or charged ions taking high voltage electricity for motive force and in accordance with the difference of mobility and distribution behaviors of all the components of samples. It has some advantages of high efficiency, fast analysis, little sample consumption, high automaticity, simple operation and little reagent dosage.Because of HPCE having so attractive advantages, it has shown extremely important application prospects in the life sciences, biotechnology, medical drugs, environmental science, forensic science and other fields. In last ten years, capillary electrophoresis technology develops rapidly, especially in recent years it has been one of the most-watched and most advanced technologies in biochemistry and analytical chemistry.Some aldehydes are recognized as the source of allergy. Hence the study of aldehydes is paid people’s more and more attention. Aromatic carboxylic acid compounds have a very wide range of application in foods, drugs and other productions. In these compounds, sodium benzoate is often used as food preservatives in flavoring compounds, but excessive dosage will have a great toxicity in the human body. As a result, determination of the content of aromatic acids has great significance. Fatty acid is widely distributed in nature, which is an important nutrient metabolism material in organism and plays an important role in regulating certain metabolic and physiological functions. Bile acid is widely found in a variety of animals’bile, which can emulsify hydrophobic molecules in water, improve digestion, balance cholesterol, digest fat, and prevent the formation of gallstones. Therefore the determination of them has an important significance in further study on physiology and pathology, and clinical diagnosis.As these biological molecules have no or weak ultraviolet absorption and the low contents cannot be detected directly by UV detection, chemical derivation has been developed in the separation of aliphatic aldehydes, aromatic carboxylic acid, fatty acids and bile acids. The results are satisfactory.The thesis consists of three chapters.Chapter OneThe developments of capillary electrophoresis (CE) and the its applications in pharmaceutical analysis, biomolecular separation, genetic engineering, clinical medicine and many other fields have been simply introduced. At the same time, the separation strategy of analyte by capillary electrophoresis and the related basic knowledge are reviewed.Chapter Two A simple and mild method for the separation of aldehydes based on a condensation reaction with 9,10-phenanthrenequinone as labeling reagent with nonaqueous capillary electrophoresis has been developed. The results indicate that seven aliphatic aldehyde derivatives and actualspecimen can be achieved baseline resolution under the proposed conditions.Chapter ThreeThe separation of acid based on a condensation reaction with 9,10-phenanthrenequinone as labeling reagent with capillary electrophoresis, The principal contents of this chapter consist of four parts.3.1 The benzoic acid and substituted benzoic acids can be well-separated with 7-amino-1,3-naphthalenedisulfonic acid as labeling reagent by capillary zone electrophoresis. The optimum conditions are as follows:30 mmol/L borate,15 mmol/Lβ-CD, pH 9.5. It has been applied to the determination of the content of benzoic acid in light soy sauce and dark soy sauce with satisfactory results.3.210 short chain fatty acids(C1～C10)can be well-separated with 7-amino-1,3 naphthalene disulfonic acid as labeling reagent with capillary zone electrophoresis. The optimum conditions are as follows:30 mmol/L borate buffer,15 mmol/Lβ-CD, pH 9.5, temperature 20℃, pressure 50 mbar, injection time 8 s, and the funaria hedw and selaginella samples are qualitatively and quantitatively detected under the optimum conditions with satisfactory results finally.3.3 A simple and mild method for the separation of 10 long fatty acids based on a condensation reaction with 7-amino-1,3-naphthalene disulfonic acid as labeling reagent with CZE-UV has been developed. Under the optimum conditions,10 fatty acids can be well-separated within 18 min. and the brachymenium nepalense sample is qualitatively and quantitatively detected under the optimum conditions with satisfactory results finally.3.46 bile acids can be well-separated based on a condensation reaction with 7-amino-1,3-naphthalenedisulfonic acid as labeling reagent with capillary zone electrophoresis. The optimum conditions are as follows:25 mmol/L borate, pH 9.5; column temperature,20℃. The method was rapid, simple and effective.更多还原