本文選題：熒光探針　切入點：雙光子熒光　出處：《蘭州大學》2017年碩士論文

【摘要】：雙光子吸收技術自問世以來一直受到了廣泛的關注。與單光子吸收材料相比,雙光子吸收材料在分辨率、穿透深度具有顯著的優(yōu)勢,可以用于顯微成像、微紡織技術、三維數(shù)據(jù)存儲、光限幅、上轉換發(fā)光、光動力學治療以及藥物靶向釋放等諸多領域。特別是雙光子顯微技術,以近紅外的激光為光源對生物樣品進行成像,具有穿透性強,空間分辨率高,背景熒光干擾小,以及對生物樣品的光損傷較小等優(yōu)點,在生物醫(yī)學領域具有廣闊的應用前景。然而,傳統(tǒng)的雙光子材料常常具有大共軛結構,水溶性差、細胞穿透能力差、生物毒性也較大,并不適用于生物成像。因此,設計合成具有較高雙光子吸收截面的有機小分子用于生物體內(nèi)細胞、血管、組織成像,具有重要的研究價值。本文設計合成了兩種具有雙光子吸收特性的熒光小分子,對其發(fā)光性能進行了系統(tǒng)的研究,探索它們在生物成像中的應用。具體的研究內(nèi)容包括:1、設計合成了一類以寡聚苯乙烯為骨架的雙光子次氯酸熒光探針OPV-HOCl,并將其應用于活細胞及組織內(nèi)的雙光子成像。在寡聚苯乙烯骨架上引入次氯酸識別基團——氧硫雜環(huán)戊烷,通過1H-NMR、13C-NMR、HRMS對其結構進行了表征,并通過紫外光譜、熒光光譜等進一步研究了該探針對次氯酸的響應性能,測定了其雙光子吸收截面。加入次氯酸以后,探針分子末端的氧硫雜環(huán)戊烷基團被氧化,并生成醛基。由于分子內(nèi)強烈的電荷轉移導致產(chǎn)物的雙光子吸收截面提高了近15倍(從78.9 GM提高到1131.5 GM),因此OPV-HOCl可以作為一個雙光子“turn-on”型次氯酸熒光探針。此外,該探針還具有反應速度快、選擇性好、pH適用范圍寬等優(yōu)點。MTT實驗表明該探針具有較小的細胞毒性。由于該探針優(yōu)異的次氯酸響應性能和較小的生物毒性,我們成功地將其用于小鼠膠質瘤細胞BV-2中次氯酸的檢測,研究表明該探針可以透過細胞膜,并對細胞中外源性次氯酸和脂多糖誘導產(chǎn)生的內(nèi)源性次氯酸具有高選擇性的快速響應。2、合成了具有雙光子吸收性質的吲哚四方酸染料SQA-Br,并將其負載于微米級的聚苯乙烯微球中,用于腦中風區(qū)域的檢測研究。研究表明,聚苯乙烯微球可以為染料提供相對惰性的環(huán)境,避免了聚集誘導熒光猝滅,使得染料在水溶液中仍具有良好的發(fā)光性能,并且染料在微球中的熒光壽命和量子產(chǎn)率與溶液中相比均有一定程度的提高,因此在生物成像中具有潛在的應用價值。我們將負載四方酸的聚苯乙烯微球注射進入中風小鼠體內(nèi),發(fā)現(xiàn)微球從中風損傷區(qū)域擴散出去并在周圍區(qū)域富集,可以用于中風損傷區(qū)域的初步識別。
[Abstract]:Two-photon absorption technology has been paid more and more attention since it came into being. Compared with single-photon absorption material, two-photon absorption material has obvious advantages in resolution and penetration depth, so it can be used in micro-imaging and micro-textile technology. Three dimensional data storage, optical limiting, up-conversion luminescence, photodynamic therapy and drug targeted release, etc., especially two-photon microscopy, imaging biological samples with near-infrared laser as light source, has strong penetrability. Because of its advantages of high spatial resolution, low background fluorescence interference and low light damage to biological samples, it has broad application prospects in biomedical fields. However, traditional two-photon materials often have large conjugate structures and poor water solubility. Therefore, organic small molecules with high two-photon absorption cross sections are designed and synthesized for biological imaging of cells, blood vessels and tissues. In this paper, two kinds of fluorescent small molecules with two-photon absorption characteristics are designed and synthesized, and their luminescence properties are systematically studied. The specific research contents include: 1, a two-photon hypochlorite fluorescence probe OPV-HOCl with oligopolystyrene as skeleton was designed and synthesized, and applied to two-photon formation in living cells and tissues. Like. Introduction of hypochlorite recognition group-oxo-thiocyclopentane on the framework of oligopolystyrene, The structure of the probe was characterized by 1H-NMR-13C-NMR-HRMS. The response of the probe to hypochloric acid was further studied by UV and fluorescence spectra. The two-photon absorption cross section of the probe was determined after the addition of hypochloric acid. The oxygen-thiocyclopentane group at the end of the probe was oxidized. Due to the strong intramolecular charge transfer, the two-photon absorption cross section of the product was increased by nearly 15 times (from 78.9 GM to 1131.5 GMN), so OPV-HOCl can be used as a two-photon "turn-on" type hypochloric acid fluorescence probe. MTT assay showed that the probe had less cytotoxicity because of its excellent hypochloric acid response and low biotoxicity. We have successfully applied it to the detection of hypochloric acid in mouse glioma cell line BV-2. The results show that the probe can penetrate the cell membrane. And the endogenous hypochloric acid induced by exogenous hypochloric acid and lipopolysaccharide was highly selective and rapid response. SQA-Br2, a two-photon absorption dye, was synthesized and loaded into micrometer polystyrene microspheres. The results show that polystyrene microspheres can provide a relatively inert environment for dyes, avoid agglomeration induced fluorescence quenching, and make the dyes still have good luminescence properties in aqueous solution. Moreover, the fluorescence lifetime and quantum yield of dyes in the microspheres were improved to a certain extent compared with those in the solution. We injected polystyrene microspheres loaded with tetragonal acid into apoplectic mice and found that the microspheres spread out from the wind-damaged area and enriched in the surrounding area. It can be used to identify the area of stroke injury.
【學位授予單位】：蘭州大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：O657.3

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