检测细胞内小分子巯基化合物的荧光探针合成及应用

发布时间：2018-02-21 22:08

本文关键词： 荧光探针近红外巯基化合物活细胞成像　出处：《兰州大学》2016年硕士论文　论文类型：学位论文

【摘要】：细胞内的小分子巯基化合物如半胱氨酸(Cys),同型半胱氨酸(Hcy),谷胱甘肽(GSH),在生物体内有着重要的生理功能。它们的含量水平是衡量生物体健康状况的重要标准。荧光探针具有高灵敏度,高选择性,操作便捷等优点,被广泛应用于检测各种离子和分子。因此,设计合成能够选择性检测细胞内的小分子巯基化合物的荧光探针具有十分重要的科学意义。本文设计合成了两个检测小分子巯基化合物的探针,并将其应用于活细胞成像。本文分为三章。第一章主要阐述了荧光的基础知识,荧光探针的主要机理,研究背景及小分子巯基化合物探针的研究进展。在第二章中,设计并合成了一个基于光诱导电子转移(PET)机理的选择性识别巯基化合物的荧光探针1。我们详细研究了探针1和不同的分析物作用后的紫外吸收和荧光发射性质。探针1在和巯基化合物作用后有一个很大的斯托克斯位移(~200 nm)并且可以在DMSO/PBS(9/1,v/v,10 mM,pH 3.5)中选择性检测Cys而不受GSH,Hcy,CN?和其他氨基酸的干扰,检测限为0.124μM。而在pH 5.5时,探针1对GSH,Hcy和Cys都响应,检测限分别为0.151μM,0.128μM和0.040μM。探针1的细胞毒性很低,被成功地应用于活细胞中巯基化合物的成像。在第三章中,合成了一个氯乙酰化的萘荧光素近红外(NIR)荧光探针2(NFC)。它可以选择性地识别Cys和Hcy而不受GSH和其它氨基酸(AAs)的影响。NFC对Cys和Hcy的检测限分别为0.30和0.42μM。萘荧光素(NF)作为荧光团,其荧光强度由分子内电荷转移(ICT)过程控制。该探针合成简单,自身荧光很弱,但和Cys或Hcy反应后,荧光明显增强。我们证明了荧光的关开过程是由于NFC与Cys和Hcy先发生了亲和取代反应,紧接着进行了分子内的环化生成了NF。该探针的毒性也比较低,被成功地应用于检测人血浆中的Cys和Hcy的总含量以及活细胞成像。
[Abstract]:Small molecule thiol compounds such as cysteine within the cell (Cys), homocysteine (Hcy), glutathione (GSH), have important physiological functions in organisms. Their content is an important standard to measure the organism's health. Fluorescent probe with high sensitivity, high selectivity, convenient operation and other advantages, is widely used in the detection of various ions and molecules. Therefore, the design and synthesis of which has very important scientific significance of small molecule thiol fluorescent probes for selective detection of cells. Two detection probe mercapto compounds were designed and synthesized, and its application in live cell imaging. This paper is divided into three chapters. The first chapter mainly introduces the basic knowledge of the main mechanism of fluorescence, fluorescence probe, probe the research progress of sulfhydryl compounds and small molecular background. In the second chapter, the design and synthesis based on a Photoinduced electron transfer (PET) fluorescence probe for selective recognition of sulfhydryl compounds mechanism 1. we studied in detail the probe and fluorescence emission properties of UV and 1 different analytes after absorption. In 1 probe and thiols. After a large Stokes shift (~200 nm) and the DMSO/PBS (9/1 10, v/v, mM, pH 3.5) in the selective detection of Cys but not by GSH, Hcy, CN and other amino acids? Interference, the detection limit is 0.124 M. and 5.5 in pH, 1 for Hcy and GSH probe, Cys response, detection limits were 0.151 M, 0.128 M cell toxicity and the 0.040 M. probe 1 is very low, imaging has been successfully applied to the sulfhydryl compounds in living cells. In the third chapter, the synthesis of a naphthalene fluorescein chloroacetylated near infrared fluorescent probe (NIR) 2 (NFC). It can selectively recognize Cys and Hcy but not by GSH and other the effects of amino acids (AAs). The NFC of Cys and the detection limit of Hcy were 0.30 and 0.42 M. naphthalene (NF) as fluorescein fluorophore, the fluorescence intensity of the intramolecular charge transfer (ICT) process control. The probe has simple synthesis, its fluorescence is very weak, but Cys or Hcy after the reaction, the fluorescence enhanced. We show that fluorescence the closing process is due to NFC and Cys and Hcy happened first nucleophilic substitution reaction, followed by intramolecular cyclization of the probe generated NF. toxicity is relatively low, the total content was successfully applied to detect Cys and Hcy in human plasma and live cell imaging.

【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：O657.3

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