本文選題：熒光分析法 + 有機熒光探針��； 參考：《南京大學》2017年碩士論文

【摘要】：隨著激光、納米材料等新技術和新材料的引入,熒光分析法得到了迅猛發(fā)展,并被廣泛應用于生物醫(yī)學領域。作為熒光分析法的重要工具,合適的熒光探針有利于提升熒光分析法的選擇性和靈敏度等性能,對于熒光分析法在生物檢測等方面的應用十分重要。以有機小分子為基礎的熒光探針具有合成簡單、成本低、易于操作等優(yōu)點。然而,有機熒光探針應用于生物檢測等時常常受到其復雜環(huán)境的影響。因此,合成靈敏度高、選擇性好、光物理性質(zhì)優(yōu)異的有機熒光探針對于熒光探針在生物醫(yī)學領域的應用尤為重要。本文針對目前一些有機熒光探針存在選擇性較差、靈敏度偏低、光穩(wěn)定性差等不足,設計合成了一些新型的有機熒光探針并用于生物檢測。具體內(nèi)容如下:1.通過Miyaura硼化反應合成一種以萘-苯并惡唑熒光團為報告基團,芳基硼酸酯為識別基團的比率型熒光探針R1。傳統(tǒng)的芳基硼酸酯探針對H2O2選擇性很好。令人驚訝的是,與傳統(tǒng)的芳基硼酸酯探針不同,R1探針對ClO-選擇性很好。詳細的機理研究表明,R1探針與ClO-作用經(jīng)歷了連續(xù)的氧化和氯化反應,氧化反應脫去硼酸酯基團而沒有使熒光發(fā)生明顯變化,氯化反應使得探針熒光發(fā)生紅移。而H2O2與探針R1只能發(fā)生氧化反應,熒光基本不變,其它ROS均不能與R1探針反應而使熒光發(fā)生變化。因此,R1探針對ClO-的檢測具有很好的選擇性。與此同時,R1探針對ClO-的檢測具有很好的靈敏度,檢測限達到6.4nM。芳基硼酸酯不僅作為探針的識別基團,還作為"lock"結構防止探針受到pH波動的影響。雖然R1與ClO-作用經(jīng)歷了多步反應,但它們的反應速度很快(只需2 min左右)。除了具有選擇性好、靈敏度高、對pH穩(wěn)定、反應時間較短等優(yōu)點,R1探針還具有很好的光穩(wěn)定性,這些優(yōu)良的性質(zhì)表明R1探針在生物檢測方面具有很好的應用前景;2.傳統(tǒng)的熒光探針在高濃度時會由于ACQ效應使熒光淬滅,因此這些熒光探針的光物理性質(zhì)容易受到濃度的影響,背景熒光較強、耐光漂白性較差,這些都限制了熒光探針在生物醫(yī)學領域的應用。我們以四苯乙烯(TPE)熒光團作為報告基團,胺作為酸堿反應位點合成了一種對pH敏感的聚集誘導發(fā)光(AIE)熒光探針TPE-Leu。不同于傳統(tǒng)的熒光探針,TPE-Leu探針具有AIE性質(zhì),在聚集狀態(tài)下熒光更強,這有效減小了背景熒光帶來的影響,對于探針在生物檢測方面的應用十分有利。探針TPE-Leu中含有芳香胺和脂肪胺兩種胺結構,偏堿性時,探針溶解性較差,熒光較強;在偏酸性條件下,由于脂肪胺的堿性比芳香胺強,更容易質(zhì)子化,使得探針溶解性增加,熒光減弱,利用探針的這種性質(zhì)實現(xiàn)對pH的檢測。此外,TPE-Leu檢測pH具有很好的選擇性,不易受到金屬離子及一些生物分子的影響。TPE-Leu探針還可以用于選擇性檢測乙酰膽堿酯酶(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.
【學位授予單位】：南京大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：O657.3
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本文編號：2010267