高发光性能的贵金属纳米簇制备、生物学应用及相关分子机理研究

发布时间：2018-03-31 00:06

本文选题：单磷酸腺苷　切入点：金属纳米簇　出处：《吉林大学》2017年博士论文

【摘要】：金属纳米簇(metal nanocluster,NCs)也称为量子簇,由于其具有强而稳定的荧光发射性、低毒性及良好的生物相容性等优点以及其结构上具有粒径小(?2nm)、表面易被功能化等特点,因而成为近几年来的研究热点并被广泛地应用于纳米医学领域(包括生物传感、生物成像和生物治疗等)。金属纳米簇由几个到多于100个原子(通常为金(Au)或者银(Ag)原子)组成,与纳米颗粒不同的是其具有类分子的性质、在近红外区域发长寿命荧光以及具有较宽的双光子激发范围,这使得它们能够广泛地应用于体内和体外成像研究。由于金属纳米簇的表面活化能高易于聚合生长成不发光的大颗粒,因而需要使用合适的配体分子进行保护提高其稳定性。目前常用的配体分子包括:聚合物、有机小分子、生物分子等。尽管金属纳米簇相关研究已经取得了极大的进展,但仍存在一些问题,如荧光量子产率低、生物学应用中的局限性。基于此,我们利用新方法合成了一系列的金属纳米簇以及双金属纳米簇,进而提高其荧光量子产率并拓展其在生物学中的应用;并对其发光机理和结合模式进行了深入的研究。主要分为三个部分:一、通过加热搅拌法成功地制备了由单磷酸腺苷(Adenosine monophosphate,AMP)保护的金纳米簇(AuNCs@AMP)。对其反应条件如时间、温度及浓度比例进行了优化,最后确定1 h、80?C及AMP:Au=10:1为最佳合成条件。基于以上条件制备的AuNCs@AMP具有强而稳定的荧光,且荧光量子产率(QY%)高达14.52%。然后,我们通过红外吸收光谱、核磁氢谱磷谱及质谱对其结合模式及金和AMP的个数比列进行了深入的研究。结果表明,在配体AMP中,O、N及-NH2均为富电子原子或基团,都能够促进AuNCs@AMP的荧光发射;但是保护配体AMP上的嘌呤环和部分磷酸基团在金核表面的取向是决定AuNCs@AMP荧光量子产率高的主要因素。本章节通过对高亮发光AuNCs@AMP的制备机理的研究,希望可以激励更多的关于AuNCs的合成及实际应用方面的研究。二、乳酸脱氢酶(lactatedehydrogenase,ldh)是动物、植物及原核生物中普遍表达的一种酶,其主要生物学功能是作为一种质子转移酶催化乳酸盐(lactate)和丙酮酸盐(pyruvate)之间的转化。它是常见疾病(甲状腺功能减退、贫血、脑膜炎及急性胰腺炎)重要的生物靶标。鉴于荧光检测方法的灵敏度高,我们进一步以已制备的auncs@amp为新型的荧光探针对ldh进行定量检测。ldh能使auncs@amp的荧光猝灭,并且在ldh较宽的浓度范围(50-1000nm)内,呈线性猝灭,其涵盖了临床诊断的浓度范围。特别是,这种检测比已报道的检测手段更敏感且具有极低的检测限0.2nm(26pg/μl,0.8u/l)。对比研究表明auncs@amp对具有不同等电点的其它蛋白质无荧光猝灭响应,说明其对ldh的检测具有高选择性。另外,我们通过荧光、紫外光谱详细地研究了其响应机制,结果表明该猝灭响应本质是auncs@amp与ldh表面游离的巯基相互作用形成具有强结合能力的s-au键。因此,本章研究提供了一种低成本、快速简单检测ldh的方法,且此方法有高选择性和灵敏性,这为将来的临床诊断提供了一种可能的检测方法。三、我们进一步制备了amp保护的新型金银合金纳米簇(au/agncs@amp)。该工作是首次通过水热合成法制备粒径较均一的金属纳米簇。对其合成条件如温度、时间及浓度比例进行了优化,最终确定120?c、30min及amp:ag:au=5:1:0.2的最佳制备条件,此方法制备的au/agncs@amp具有较高的qy(8.46%)。通过荧光和紫外吸收光谱,观察到au/agncs@amp具有光敏性,使用时需要避光保存。通过对其制备过程的探索,au/agncs@amp形成了一种以金原子为核心银原子分布在其表面的结构。进一步我们利用阳离子型聚合物聚乙烯亚胺(pei)与该纳米簇结合构筑au/ag-amp-pei组装体系,进一步提高其荧光量子产率。基于pei的树枝状结构及pei中的-nh及-nh2基团与金属原子的强结合能力,使au/agncs@amp单分散地固定在pei结构中起到了很好的排水效应,进而使qy增强3.5倍左右(26.21%)。本章节,为小分子保护的金属纳米簇提供了一种新的合成方法,以及希望通过构筑纳米簇和大分子的组装体系为解决ncs低荧光量子产率的问题提供一个可行性方案。
[Abstract]:Metal nanoclusters (metal nanocluster, NCs) also known as quantum cluster, because of its stable and strong fluorescence emission, low toxicity and good biocompatibility and the structure with small particle size (? 2nm), the surface easy to be functional, and thus become a hot research topic in recent years it is widely used in the field of nanomedicine (including Bio sensing, biological imaging and biological treatment etc.). Metal nanoclusters by several to more than 100 atoms (usually gold or silver (Au) (Ag), different atoms) and nanometer particles is its molecular properties in the past the infrared region long life fluorescence and two-photon excitation has a wide range, which makes them can be widely used in the study of in vitro and in vivo imaging. The surface of the metal nanoclusters with high activation energy and easy aggregation into large particles with no light, and therefore need to use Suitable ligands to protect and improve its stability. The ligands commonly used include: polymer, small organic molecules, biological molecules. Although the research of the metal nanoclusters have made great progress, but there are still some problems, such as low fluorescence quantum yield, the application limits of the students. Based on this, we use the new method of a series of metal nanoclusters and double metal nanoclusters were synthesized, and then improve the fluorescence quantum yield and expand its application in biology; and the luminescent mechanism and binding mode were studied in depth. Mainly divided into three parts: one was prepared successfully by a single amp by heating method (Adenosine monophosphate AMP) gold nanoclusters protection (AuNCs@AMP). The reaction conditions were optimized such as time, temperature and concentration ratio, determine the final 1 h, 80 C and AMP:Au=10:1 is the best? The synthesis conditions. The above conditions were prepared based on AuNCs@AMP with stable and strong fluorescence, and the fluorescence quantum yield (QY%) up to 14.52%. and then we by infrared absorption spectroscopy, the number of NMR spectra and mass spectrometry combined with phosphorus pattern and gold and the AMP ratio is studied. The results showed that in N ligands AMP, O, and -NH2 are electron rich atom or group, could enhance the fluorescence of AuNCs@AMP; but the protection of ligand AMP on the purine ring and phosphate groups in the orientation of gold nuclear surface are the main factors that determine the AuNCs@AMP fluorescence quantum yield. This chapter through the research on the mechanism of the preparation high brightness AuNCs@AMP, hope that the study can inspire the synthesis and application of AuNCs more. Two, lactate dehydrogenase (LactateDehydrogenase, LDH) is a kind of animal, plant and enzyme widely expressed in prokaryotes, the main To biological functions as a proton transfer enzyme (lactate) lactate and pyruvate (pyruvate) between the transformation. It is a common disease (hypothyroidism, anemia, meningitis and acute pancreatitis). In view of the important biological target fluorescence detection method with high sensitivity, we further to auncs@amp has been prepared for fluorescence a new probe for quantitative detection of.Ldh LDH can quench the fluorescence of auncs@amp, and in a wide concentration range of LDH (50-1000nm), a linear quenching, which covers the range of clinical diagnosis. In particular, the detection of detection ratio detection means has been reported more sensitive and has a very low limit of 0.2nm (26pg/ L, 0.8u/l). The comparative study shows that the auncs@amp of other proteins with different isoelectric points without fluorescence quenching response, indicating that the detection of LDH with high selectivity. In addition, we through fluorescence, UV A detailed study of the spectral response mechanism, the results show that the quenching response is the essence with the formation of auncs@amp and LDH in surface free sulfhydryl interactions with strong S-Au bond ability. Therefore, provides a lower cost of this chapter, quick and simple method for the detection of LDH, and this method has high selectivity and sensitivity, this provides a possible method for clinical diagnosis in the future. Three, we further new gold alloy nanoclusters were synthesized AMP (au/agncs@amp). The work is first prepared by particle size homogeneous metal nanoclusters hydrothermal synthesis method. The synthesis conditions such as temperature, time were optimized and the concentration ratio, the final 120? C, 30min and amp:ag:au=5:1:0.2 the best preparation conditions, the preparation method of au/agncs@amp with higher QY (8.46%). By fluorescence and UV absorption spectrum, observed au/agncs@amp Photosensitive, use need to avoid light preservation. Through the exploration of the preparation process, au/agncs@amp formed the core of the silver atom distribution in the structure of the surface with a gold atom. Further we use cationic polymer polyethyleneimine (PEI) combined with the nanoclusters build au/ag-amp-pei assembly system, to further improve the the fluorescence quantum yield. The binding ability of dendritic structure and Pei Pei in -nh and -nh2 groups and the metal atom based on au/agncs@amp monodisperse fixed plays a very good drainage effect in the Pei structure, and the QY increased about 3.5 times (26.21%). This chapter provides a new synthetic method for metal nanoclusters of small molecule protection, and hope that through the assembly system to build the large molecules and nano clusters provide a feasible scheme for solving NCS low fluorescence quantum yield.

【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】：TB383.1;O657.3

【相似文献】