基于荧光素的一氧化氮荧光探针的构建及其识别性能研究

发布时间：2018-03-12 14:03

本文选题：荧光探针　切入点：5-氨基荧光素　出处：《遵义医学院》2017年硕士论文　论文类型：学位论文

【摘要】：目的:构建基于荧光素的NO型荧光探针,并对其识别NO的性能进行研究。方法:以4-硝基邻二甲苯为起始原料,经氧化、脱水、加成及还原等反应制备5-氨基荧光素(L_1),再将其与4-(2-氨乙基)吗啉反应制备5-氨基荧光素吗啉衍生物(L_2)。采用~1H-NMR,~(13)C-NMR,HR-MS,FT-IR及单晶衍射等现代分析技术对L_1和L_2的组成和结构进行表征。在Tris-HCl缓冲溶液中利用紫外-可见吸收光谱法和荧光分光光度法考察L_1和L_2识别NO的性能和机理。结果:经~1H-NMR,~(13)C-NMR,HR-MS,FT-IR及单晶衍射确证L_1和L_2均为预期目标化合物。它们均有选择性识别NO的性能。L_1是以荧光增强脱氨基化对NO进行识别,其荧光强度与一氧化氮供体NOC-18在0-8.0×10~(-5) mol/L浓度范围内呈现良好的线性关系;研究范围内的活性氧和氮物种中Cl O-对L_1识别NO稍有干扰,而抗坏血酸和脱氢抗坏血酸对L_1识别NO有较大干扰;L_1识别NO机理经~1H-NMR与HRMS证实为脱氨基作用。细胞毒性实验发现:L_1对Raw 264.7细胞具有低毒性,并能在Raw 264.7细胞中实现对NO的荧光成像。L_2是以荧光猝灭脱氨基机理进行荧光猝灭识别NO。L_2自身在紫外灯下呈蓝色的荧光,在L_2与NO作用后呈现的是荧光猝灭的现象,并且其猝灭作用在研究范围内的活性氧和氮物种中受NO_3~-的影响。细胞毒性实验显示:L_2对Raw 264.7细胞具有低毒性。结论:构建的两个5-氨基荧光素衍生物L_1和L_2均通过脱氨基反应实现对NO的荧光探测,L_1为荧光增强型NO荧光探针,L_2为荧光猝灭型NO荧光探针。
[Abstract]:Objective: to construct no fluorescent probe based on fluorescein and study its ability to recognize no. Methods: 4-nitro-o-xylene was oxidized and dehydrated. 5-aminofluorescein L _ (1) was prepared by addition and reduction reactions, and then synthesized by reaction of 5-aminofluorescein morpholine with 4-( 2-aminoethyl) morpholine. The composition and structure of L1 and L2 were analyzed by modern analytical techniques such as 1H-NMR-13C-NMR-MSFT-IR, single crystal diffraction and other modern analytical techniques, such as the addition and reduction of 5-aminofluorescein and the synthesis of 5-aminofluorescein morpholine derivatives. The performance and mechanism of no recognition by L _ (1) and L _ (2) in Tris-HCl buffer solution were investigated by UV-Vis absorption spectrometry and fluorescence spectrophotometry. Results: 1H-NMR-13C-NMR-HR-MSFT-IR and single crystal diffraction confirmed that L _ 1 and L _ S _ 2 were expected target compounds. They all have the ability of selective recognition of no. LS-1 is used to recognize no by fluorescence enhanced deamination. The fluorescence intensity showed a good linear relationship with the concentration of nitric oxide donor NOC-18 in the range of 0-8.0 脳 10 ~ (-5) mol/L. However, ascorbic acid and dehydroascorbic acid have great interference on the recognition of no in L1. The mechanism of no recognition in L1 was confirmed by 1H-NMR and HRMS. The fluorescence imaging of no in Raw 264.7 cells. LST2 is a fluorescence quenching mechanism to recognize the blue fluorescence of NO.L_2 under ultraviolet lamp, and the phenomenon of fluorescence quenching after the interaction between L2 and no. The cytotoxicity test showed that the two 5-aminofluorescein derivatives L1 and L2 were of low toxicity to Raw 264.7 cells. Conclusion: both of the constructed 5-aminofluorescein derivatives L1 and L2 are common. Fluorescence detection of no by over deamination reaction: L1 is a fluorescence enhanced no fluorescence probe and L2 is a fluorescence quenching no fluorescence probe.
【学位授予单位】：遵义医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.3

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