几种生物活性分子荧光探针的设计合成和生物成像

发布时间：2018-07-03 07:35

本文选题：荧光探针 + 喹啉　；参考：《中国科学技术大学》2017年博士论文

【摘要】：生物体内活性硫和活性氮物种是两类重要的生物活性分子。三种生物硫醇和硫化氢属于活性硫物种;一氧化氮是极具代表性的活性氮物种。大多数细胞中含有生物硫醇,浓度约在毫摩尔/升。硒代半胱氨酸(Sec)作为半胱氨酸(Cys)类似物,是构成硒蛋白(SePs)的基本成分。硫化氢(H_2S)和一氧化氮(NO)是细胞内重要的信号分子,因此,这些生物活性分子在细胞中的浓度的在线检测对生理和病理研究的意义重大。本论文以这几种生物活性分子为检测对象,开展了荧光探针的设计合成,传感性质研究以及生物体系中荧光成像等研究工作。具体为:.1.设计合成了两类取代的喹啉-2-甲撑丙二酸(QMAs)及其乙酯型(QMEs)的硫醇荧光探针。QMAs和QMEs与硫醇的迈克尔加成反应有着截然不同的pH效应:QMAs不同于QMEs以及大多数微碱性条件下的迈克尔型探针,能够在酸性环境(pHpKa3)中活化,与硫醇发生反应。通过对不同取代基的QMAs筛选,两种QMAs(取代基:NEt2,OH)能够定位于溶酶体内,与硫醇反应得到溶酶体的荧光成像,是首例反应型的溶酶体探针。2.以叠氮基修饰QME,设计合成了硫醇和硫化氢双输入型探针QME-N_3。该探针有较大的生物亲和性,能够分别响应硫醇和硫化氢这两种化学性质相近的活性硫物种,且识别产物是具有双光子性质的荧光产物、可进行双光子成像。与单一检测物的识别相比,双输入型荧光探针能够在更复杂的生物体系中参与生理和病理研究,是更具价值的生物传感器。3.设计合成了一种新的红色荧光发色团(B-BODIPY),以此构建了高灵敏、高选择性的硒醇探针Sel-p1和Sel-p2。实验表明,探针Sel-p2能够检测细胞中外源性Sec和亚硒酸盐诱导的内源性Sec,以及进行小鼠体内Sec的荧光成像,有望应用于探测体内硒化合物代谢,为研究硒化合物在癌症治疗中的作用提供了一条途径。4.设计合成了两种NO荧光探针NO-a5和NO-a6,都具有响应快、选择性高的特点。NO-a5与NO发生识别后呈现荧光打开的响应模式、具有双光子吸收性质;NO-a6与检测物NO反应前后,荧光呈比率变化,这一性质在这类荧光探针中属首次发现。探针NO-a5和NO-a6都能实现在活细胞中外源和内源性NO的检测。
[Abstract]:Active sulfur and nitrogen species are two important bioactive molecules. Three biological mercaptan and hydrogen sulfide belong to active sulfur species and nitric oxide is a representative active nitrogen species. Most cells contain biological mercaptan at concentrations of about ml / l. Selenocysteine (Sec), as a cysteine (Cys) analogue, is the basic component of selenium protein (SePs). Hydrogen sulfide (H _ 2S) and nitric oxide (no) are important signaling molecules in cells. Therefore, on-line detection of the concentrations of these bioactive molecules in cells is of great significance to physiological and pathological studies. In this paper, the design and synthesis of fluorescent probe, the study of sensing properties and the study of fluorescence imaging in biological system were carried out. Specifically:. 1. Two kinds of substituted quinoline-2-methylene malonic acid (QMAs) and their ethyl ester type (QMEs) mercaptan fluorescence probe. QMAs and QMEs have been designed and synthesized. The Michael addition reaction between QMEs and mercaptan has a different pH effect: QMAs is different from QMEs and most microbasic strips. The Michael probe under the device, It can be activated in acidic environment (pH pKa3) and react with mercaptan. Through the screening of QMAs with different substituents, two kinds of QMAs can be located in lysosomes and react with mercaptan to obtain the fluorescence imaging of lysosomes. These two QMAs are the first reaction type lysosomal probes. The double input probe QME-N _ 3 was designed and synthesized with azide modified QME-N _ 3. The probe has great biological affinity and can respond to two kinds of active sulfur species with similar chemical properties, mercaptan and hydrogen sulfide, respectively, and the recognition product is a fluorescence product with two-photon properties, which can be used for two-photon imaging. Compared with single detection, dual-input fluorescent probes can participate in physiological and pathological studies in more complex biological systems and are more valuable biosensors. A new red fluorescent chromophore (B-BODIPY) was designed and synthesized to construct selenol probes Sel-p1 and Sel-p2 with high sensitivity and selectivity. The results showed that the probe Sel-p2 could detect the endogenous Secs induced by exogenous Sec and selenite, as well as the fluorescence imaging of SECs in mice, which could be used to detect the metabolism of selenium compounds in vivo. It provides a way to study the role of selenium compounds in cancer treatment. 4. 4. Two kinds of no fluorescence probes, NO-a5 and NO-a6, were designed and synthesized. Both have the characteristics of fast response and high selectivity. The fluorescence of NO-a6 and NO-a6 showed a ratio change before and after the reaction between NO-a6 and no. This property is first found in this kind of fluorescent probe. The probes NO-a5 and NO-a6 can detect both exogenous and endogenous no in living cells.
【学位授予单位】：中国科学技术大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：O657.3

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