光交联四苯基乙烯类分子的设计合成及生物成像应用研究

发布时间：2018-01-19 05:26

本文关键词： 氧杂环丁烷四苯基乙烯顺反异构体光交联纳米粒子特异性标记超分辨荧光成像　出处：《吉林大学》2017年博士论文　论文类型：学位论文

【摘要】：作为典型的聚集诱导发光分子,四苯基乙烯自发现以来,由于结构简单、合成方便、易于修饰等优点而得到了广泛的研究和应用。但是相关领域仍然存许多问题需要进一步研究和解决,例如四苯基乙烯顺反异构体难以分离的问题和小尺寸聚集诱导发光纳米粒子制备和特异性标记问题,本文将围绕这两个问题进行研究解决。为了解决四苯基乙烯顺反异构体分离困难的问题,我们在第三章中设计合成了氧杂环丁烷取代的四苯基乙烯分子,通过简单的柱层析分离得到了单一构型的异构体,具有较高的分离产率。而且通过质谱、红外和核磁等测试方法,证明这两个分离的产物的确为异构体。借助二维核磁,实现了对两个分子构型的确定。这两个异构体在适当的光/热刺激下会发生异构化,相互转化。重要的是与反式异构体相比,顺式异构体在固体状态下具有更加红移的发射和更高的量子效率,顺式异构体量子效率是反式异构体的五倍。通过理论计算可以发现,两种异构体最大发射波长和量子效率的差别来自于两种异构体不同的发光机理,其中反式异构体呈现的是局域激发态发射,而顺式异构体呈现的是电荷转移态发射。这两个异构体都是优良的力刺激变色材料,因为在力刺激下它们不仅会发生荧光颜色的变化,而且存在明显的力刺激荧光亮度增加的现象,因而在力刺激下具有较高的对比度,通过一定的处理又可以恢复到初始状态。此外,反式异构体呈现出一定的自修复能力,其可以自发的从研磨后的无定型状态回到初始的结晶态,伴随着荧光发射的恢复。在溶液中,顺式异构体聚集体仍然具有较高的量子效率,利用两种异构体制备的纳米粒子也存在明显的区别。通过细胞标记实验可以发现,相比于反式异构体,顺式异构体制备的纳米粒子具有更高的细胞标记亮度。因此,通过简单反应我们实现了四苯基乙烯顺反异构体的宏观分离,进一步加深了我们对四苯基乙烯顺反异构体的理解,对进一步设计合成不同功能的四苯基乙烯分子具有一定的指导意义。在第四章中,我们通过合理的结构调整,设计合成了三个分别具有蓝色、绿色、红色发射的氧杂环丁烷修饰的四苯基乙烯衍生物分子,最大发射波长分别为479 nm、503 nm和612 nm。这三个分子呈现典型的聚集诱导发光性质,都具有较高的荧光量子效率,固体状态下量子效率分别为69%、63%和57%。利用氧杂环丁烷修饰的聚苯乙烯衍生物作为基质,通过纳米再沉淀法制备了三种小尺寸(≈16 nm)、稳定的、高亮度的荧光纳米粒子。随后,利用氧杂环丁烷的光交联特性,将纳米粒子进行交联。光交联后的纳米粒子中会产生新的共价网络,因此纳米粒子的稳定性得到明显的改善,在多种生理条件甚至是极端有机溶剂条件下都具有一定的稳定性。通过控制交联条件,不仅可以明显改善纳米粒子的稳定性,而且可以维持纳米粒子的小尺寸。得益于高亮度、小尺寸、超稳定等优势,这三种纳米粒子水溶液可以直接作为荧光墨水应用于喷墨打印机中,实现荧光图案化和荧光防伪。在生物成像方面,首先利用酰胺化反应进行纳米粒子的生物功能化,得到链霉亲合素修饰的三种纳米粒子。利用抗原抗体相互作用,实现了对细胞膜表面蛋白的特异性标记。首次实现了聚集诱导发光纳米粒子对亚细胞结构微管的标记,具有较高的分辨率。更重要的是实现了聚集诱导发光纳米粒子在受激辐射损耗超分辨荧光成像中的应用,成像分辨率得到明显提高,定量分析表明半峰宽由190 nm缩减至95 nm,。通过除氧系统的引入,进一步提高了纳米粒子在成像中的稳定性,为进一步的长期荧光定位追踪奠定了基础。总之,我们设计合成了一系列氧杂环丁烷修饰的四苯基乙烯类分子,利用氧杂环丁烷的结构特点,我们实现了四苯基乙烯顺反异构体的宏观分离。通过详细的对比研究,进一步揭示了顺反异构对分子性质的影响,对未来分子的设计合成具有重要的指导意义。利用氧杂环丁烷的光交联特性,我们发展了一种新的制备纳米粒子的方法,制备了小尺寸、高亮度、超稳定的荧光纳米粒子。通过生物功能化,真正实现了聚集诱导发光纳米粒子的特异性标记,实现了对细胞膜表面蛋白和亚细胞结构微管的特异性标记。利用超分辨荧光成像技术,进一步提高了成像的分辨率。
[Abstract]:As a typical molecular aggregation induced emission, four phenyl vinyl since the discovery, because of its simple structure, easy synthesis, easy modification and other advantages and has been widely studied and applied in related fields. But still many problems need to be further studied and solved, such as the four benzene vinyl CIS trans isomers difficult separation problems and small size the AIE nanoparticles preparation and specific markers, this paper will study and solve on the two issues. In order to solve the difficult problem of four phenyl ethylene CIS trans isomer separation, we design four phenyl ethylene oxetane substituted were synthesized in the third chapter, the single isomer configuration obtained by column chromatography is simple, the separation efficiency is higher. And by mass spectrometry, IR and NMR measurements, proved that the product of these two separate indeed isomers. With the help of 2D NMR, the determination of the two molecular configurations. The two isomers in the proper light / heat stimulation occurs under isomerization, into each other. The important thing is compared with the trans isomer, the cis isomer is more red shifted emission and higher quantum efficiency in solid state the cis isomer, the quantum efficiency is five times of the trans isomer. Through theoretical calculations can be found, two isomers of the maximum emission wavelength and quantum efficiency differences from the two isomers of different luminescence mechanism, the trans isomer is the locally excited state emission, and cis isomers show the charge transfer state emission. The two isomers are excellent photochromic materials stress stimulation, because of changes in force under the stimulation of them will not only bring the fluorescent color, and there are obvious stimulation fluorescence to increase the brightness of the phenomenon, so in force thorn High contrast with shock, through certain processing and can be restored to the initial state. In addition, the trans isomer showed a certain self-healing ability, it can spontaneously from non crystalline state back to the initial setting after grinding, with the recovery of fluorescence emission. In the solution, with quantum efficiency isomer aggregates are still high, the use of two kinds of heterogeneous nanoparticles preparation has obvious difference. Through cell labeling experiments can be found, compared to the trans isomers, cis isomers with nanoparticles prepared cells were labeled with higher brightness. Therefore, the macro separated by simple reaction we achieved four phenyl ethylene CIS trans isomers, and further enhance our CIS trans isomers of four phenyl vinyl understanding, has certain guidance for further design and synthesis of four phenyl ethylene molecules with different functions Meaning. In the fourth chapter, we through reasonable structural adjustment, designed and synthesized three respectively with blue, green, four phenyl ethylene derivatives oxetane theemission modification, the maximum emission wavelength were 479 nm, 503 nm and 612 nm. three molecules that are typical of aggregation induced emission properties and have high fluorescence quantum efficiency, quantum efficiency of solid state were 69%, 63% and 57%. using polystyrene derivatives oxetane modified as matrix, three kinds of small size were prepared by nano re precipitation (about 16 nm), stable, high brightness fluorescent nanoparticles. Then, using light crosslinking characteristics oxetane, the nanoparticles were cross-linked nanoparticles. Light after crosslinking will produce new covalent network, so the stability of nanoparticles have been significantly improved in various physiological conditions or even extremely The end has a certain stability in organic solvent conditions. By controlling the cross-linking conditions, not only can improve the stability of nanoparticles, and the nanoparticles can maintain small size. Due to the high brightness, small size, high stability and other advantages, these three kinds of nanoparticles aqueous solution can be directly used as fluorescent ink used in inkjet printers, realize fluorescence and fluorescence. Patterned in biological imaging, biological function, first by amidation reaction of nanoparticles, obtained the streptavidin affinity of three kinds of nanoparticles was modified. The anti body interaction with antigen, the specific marker of cell surface proteins. For the first time labeled AIE nanoparticles on subcellular structure microtubules, with high resolution. The more important is to realize aggregation induced emission of nanoparticles in the stimulated radiation loss of super resolution The application of fluorescence imaging, the imaging resolution was improved and the quantitative analysis showed that the half peak width from 190 nm down to 95 nm. Through the introduction of deaerator system, further improve the stability of nanoparticles in imaging, which laid the foundation for long-term follow-up fluorescence localization. In short, we designed and synthesized a series of oxygen oxetane modified four phenyl ethylene molecules, based on the structural characteristics of the oxetane, macro separation we achieved four phenyl ethylene CIS trans isomers. Through detailed comparative study, further reveals the CIS trans isomerization effect on molecular properties, has an important guiding significance for the future design of synthetic molecules. Using light crosslinking characteristics of oxetane, we developed a new method for preparation of nanoparticles, preparation of small size, high brightness, ultra stable fluorescent nanoparticles by biological function. The specific markers of aggregated induced luminescent nanoparticles were realized, and the specific markers for the surface proteins and subcellular microtubules of cell membrane were achieved. The resolution of imaging was further improved by super-resolution fluorescence imaging technology.

【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：O48

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