有机荧光探针的设计、合成及其生物应用

发布时间：2018-06-12 16:13

  本文选题：荧光分析法 + 有机荧光探针　； 参考：《南京大学》2017年硕士论文

【摘要】：随着激光、纳米材料等新技术和新材料的引入,荧光分析法得到了迅猛发展,并被广泛应用于生物医学领域。作为荧光分析法的重要工具,合适的荧光探针有利于提升荧光分析法的选择性和灵敏度等性能,对于荧光分析法在生物检测等方面的应用十分重要。以有机小分子为基础的荧光探针具有合成简单、成本低、易于操作等优点。然而,有机荧光探针应用于生物检测等时常常受到其复杂环境的影响。因此,合成灵敏度高、选择性好、光物理性质优异的有机荧光探针对于荧光探针在生物医学领域的应用尤为重要。本文针对目前一些有机荧光探针存在选择性较差、灵敏度偏低、光稳定性差等不足,设计合成了一些新型的有机荧光探针并用于生物检测。具体内容如下:1.通过Miyaura硼化反应合成一种以萘-苯并恶唑荧光团为报告基团,芳基硼酸酯为识别基团的比率型荧光探针R1。传统的芳基硼酸酯探针对H2O2选择性很好。令人惊讶的是,与传统的芳基硼酸酯探针不同,R1探针对ClO-选择性很好。详细的机理研究表明,R1探针与ClO-作用经历了连续的氧化和氯化反应,氧化反应脱去硼酸酯基团而没有使荧光发生明显变化,氯化反应使得探针荧光发生红移。而H2O2与探针R1只能发生氧化反应,荧光基本不变,其它ROS均不能与R1探针反应而使荧光发生变化。因此,R1探针对ClO-的检测具有很好的选择性。与此同时,R1探针对ClO-的检测具有很好的灵敏度,检测限达到6.4nM。芳基硼酸酯不仅作为探针的识别基团,还作为"lock"结构防止探针受到pH波动的影响。虽然R1与ClO-作用经历了多步反应,但它们的反应速度很快(只需2 min左右)。除了具有选择性好、灵敏度高、对pH稳定、反应时间较短等优点,R1探针还具有很好的光稳定性,这些优良的性质表明R1探针在生物检测方面具有很好的应用前景;2.传统的荧光探针在高浓度时会由于ACQ效应使荧光淬灭,因此这些荧光探针的光物理性质容易受到浓度的影响,背景荧光较强、耐光漂白性较差,这些都限制了荧光探针在生物医学领域的应用。我们以四苯乙烯(TPE)荧光团作为报告基团,胺作为酸碱反应位点合成了一种对pH敏感的聚集诱导发光(AIE)荧光探针TPE-Leu。不同于传统的荧光探针,TPE-Leu探针具有AIE性质,在聚集状态下荧光更强,这有效减小了背景荧光带来的影响,对于探针在生物检测方面的应用十分有利。探针TPE-Leu中含有芳香胺和脂肪胺两种胺结构,偏碱性时,探针溶解性较差,荧光较强;在偏酸性条件下,由于脂肪胺的碱性比芳香胺强,更容易质子化,使得探针溶解性增加,荧光减弱,利用探针的这种性质实现对pH的检测。此外,TPE-Leu检测pH具有很好的选择性,不易受到金属离子及一些生物分子的影响。TPE-Leu探针还可以用于选择性检测乙酰胆碱酯酶(AChE)以及脲酶(Urease)。
[Abstract]:With the introduction of laser, nanomaterials and other new technologies and materials, fluorescence analysis has been rapidly developed and widely used in biomedical fields. As an important tool of fluorescence analysis, suitable fluorescence probe is helpful to improve the selectivity and sensitivity of fluorescence analysis, and is very important for the application of fluorescence analysis in biological detection. Fluorescent probes based on organic small molecules have the advantages of simple synthesis, low cost and easy operation. However, organic fluorescent probes are often affected by their complex environment when they are used in biological detection. Therefore, organic fluorescent probes with high sensitivity, good selectivity and excellent photophysical properties are particularly important for the application of fluorescent probes in biomedical fields. In this paper, some novel organic fluorescent probes have been designed and synthesized and used for biological detection, aiming at the disadvantages of poor selectivity, low sensitivity and poor photostability of some organic fluorescent probes. The details are as follows: 1. A ratio fluorescent probe R1 with naphthalene benzoxazole fluorescence group as report group and aryl borate as recognition group was synthesized by Miyaura boride reaction. The traditional aryl borate probe has good selectivity for H _ 2O _ 2. Surprisingly, different from the traditional aryl borate probe, the R1 probe has good CLO-selectivity. The detailed mechanism studies show that the reaction between the probe and ClO- has undergone continuous oxidation and chlorination reaction, the oxidation reaction deborate group without obvious changes in fluorescence, chlorination reaction makes the probe fluorescence red shift. However, the H _ 2O _ 2 can only react with the probe R _ 1, but the fluorescence is almost unchanged, and no other Ros can react with the probe to make the fluorescence change. Therefore, the detection of ClO- by the R1 probe has a good selectivity. At the same time, the detection limit of ClO- was 6.4 nm. Aryl borate not only serves as the recognition group of the probe, but also acts as a "lock" structure to prevent the probe from being affected by pH fluctuations. Although the interaction between R1 and ClO- has undergone multi-step reaction, the reaction rate of R1 and ClO- is very fast (only about 2 min). In addition to the advantages of good selectivity, high sensitivity, stable pH and short reaction time, the R1 probe has good photostability. These excellent properties indicate that the R1 probe has a good application prospect in biological detection. Because of the high concentration of ACQ, the photophysical properties of the traditional fluorescent probes are easily affected by the concentration, the background fluorescence is strong, and the photobleaching resistance is poor. All these limit the application of fluorescent probe in biomedical field. A pH sensitive fluorescent probe TPE-Leu was synthesized by using tetrastyrene (TPE) fluorescence group as the reporting group and amine as the acid-base reaction site. Different from the traditional fluorescent probe, TPE-Leu probe has the properties of AIE and is stronger in aggregation state, which effectively reduces the influence of background fluorescence, and is very beneficial to the application of the probe in biological detection. The probe TPE-Leu contains two kinds of amines, aromatic amine and aliphatic amine. When the probe is alkaline, the solubility of the probe is poor and the fluorescence is strong. In the condition of partial acidity, the solubility of the probe increases because the alkalinity of the fatty amine is stronger than that of the aromatic amine. The fluorescence was weakened and pH was detected by using this property of the probe. In addition, TPE-Leu can be used to detect acetylcholinesterase (ache) and urease as well as urease Ureaseau, which is not easily influenced by metal ions and some biomolecules.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.3
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本文编号：2010267