金—高分子杂化纳米材料的制备及应用研究

【作者】 朱钦舒；

【导师】 沈健； 毛春；

【作者基本信息】 南京师范大学 ， 生物物理化学， 2014， 博士

【摘要】 运用微尺度或分子设计和分子工程理念进行无机-有机功能纳米材料的组装、杂化以及加强其特殊功能性的研究,已成为材料科学中最为热门和前沿的领域。本文通过研究金-高分子杂化纳米材料的合成、组装,以及材料的结构与功能之间的相关性,探索无机-有机杂化纳米材料的生长机理和决定无机-有机杂化纳米产物形貌结构及性质等的关键作用,并且努力构建金-高分子杂化纳米材料的生物安全性评价体系,为此类无机-有机杂化纳米材料在可控制备以及生物医学、光电和催化等方面的应用提供理论与实验基础。1.基于离子凝聚技术制备了草莓型Au-TPL@CS杂化纳米粒子,表征了杂化纳米粒子的形貌、组成、大小以及表面带电性。并通过凝血常规检测、血浆复钙时间、补体激活和溶血试验及红细胞形态变化,评价了草莓型Au-TPL@CS杂化纳米粒子的血液相容性。试验结果表明,草莓型Au-TPL@CS杂化纳米粒子具有良好的血液相容性,无明显的红细胞毒性。将草莓型Au-TPL@CS杂化纳米粒子用于电极表面的修饰,并用循环伏安法测量了 Hb/Au-TPL@CS/MWCNTs/GCE的直接电子转移性能,该生物传感器对还原H2O2表现出良好的电催化活性。2.利用两亲性嵌段共聚物F127的还原性、模板作用以及掺杂分散能力,通过不同合成工艺,制备、表征了三种不同形貌结构的Au-F127杂化纳米粒子,并探讨了它们各自的形成机理。红毛丹型Au-F127杂化纳米粒子的存在能够有效促进Hb和电极表面之间的电子传导,并为Hb提供了具有良好的生物相容性的微环境,使其能够很好保持自身原本的生理活性。因此,红毛丹型Au-F127杂化纳米粒子有望在生物传感领域得到应用。而核-壳型Au@F127杂化纳米粒子由于外部壳层F127胶束本身具有优异的生物相容性,因此具有一定的抗凝血性能,且无明显的细胞毒性。3.通过凝血常规检测、补体激活和血小板激活、溶血和MTT试验,证实了红毛丹型Au-F127杂化纳米粒子具有一定的抗凝血性能,且无明显的细胞毒性。将红毛丹型Au-F127杂化纳米粒子修饰到GCE表面,再固定上GOx从而制备出一种新颖的GOx/Au-F127/GCE葡萄糖生物传感器,并对这种生物传感器的电化学性能进行研究。进一步将该生物传感器应用到全血中对血液中的葡萄糖进行检测,检测结果表明该生物传感器具有抗生物污垢性能,检测线性范围宽,检测灵敏度良好,同时具有很好的抗干扰能力、重现性,在最低检测限方面具有一定的优越性。在全血中的测得的数据与同源血清样品中使用生化分析仪测得的数据结果非常接近。基于我们的检测原理,该检测数据在理论上比医院血清样品检测所得数据更符合血液的真实状况。4.制备了一种类肝素化的磺酸化超支化聚酯纳米粒子HBPE-S03H,然后利用自组装技术构建了 GOx-(HBPE-S03H)/Au/APTES/GCE生物传感器,将其直接应用于全血中葡萄糖的电化学检测,实验结果显示金-磺酸化超支化杂化纳米粒子在电极表面的修饰能够起到抗生物污垢的作用,使得传感器可以在全血中直接检测,检测结果可信度高,并且表现出抗干扰能力强、重现性好,在最低检测限、检测范围和线性相关系数方面都具有一定的优越性。5.通过绿色化学和自组装技术制备一类表面携带负电荷的金-聚吡咯杂化纳米粒子,并通过凝血常规检测、补体激活和血小板激活等,证实了该杂化纳米粒子具有一定的抗凝血性能。将其修饰到电极表面,并进一步和凝血酶适配体进行结合构建了一种新型的电化学适体传感器,然后应用于全血中凝血酶的检测。试验结果显示该生物传感器在凝血酶浓度为1 fM-500 nM的检测范围内存在良好的线性关系,检测限为0.29fM(S/N=3),表明该适配体传感器具有较高的灵敏度,还具有良好的特异性选择,抗干扰能力强,稳定性和重现性表现也很好。在全血中的测得的数据与同源血清样品中使用荧光法测得的数据结果非常接近。由此表明我们所制备的纳米电化学生物传感器在全血中的检测具有高可信度。更多还原

【Abstract】 Base on the concepts of microscale or molecular design and molecular engineering, the assembly or hybrid preparation of inorganic-organic functional nanomaterials has become one of the most popular fields of materials science. In this paper, the synthesis, assembly methods and the correlations between the structure and function of some novel gold-polymer hybrid nanomaterials were investigated for explore the growth mechanism of inorganic-organic hybrid nanomaterials, and the decision key of morphologies,structure and nature of inorganic-organic hybrid nanomaterials. Moreover, the biological safety evaluation systems for these gold-polymer hybrid nanomaterials were established.The results will provide the theoretical and experimental basis for the applications of inorganic-organic hybrid nanomaterials in future.The main results can be concluded as follows:1. A novel kind of raspberry-type hybrid nanoparticles was prepared by hybridization of chitosan (CS) with thioethyl pendant ligand (TPL) modified Au nanoparticles(Au-TPL@CS NPs). Such method was based on ionic gelation using sodium tripolyphosphate as a counterion. The blood compatibility of Au-TPL@CS NPs was characterized by coagulation tests, plasma recalcification time,hemolysis assay,morphological changes of red blood cells (RBCs) and complement activation in vitro.The results showed that Au-TPL@CS NPs exhibited good blood compatibility. The possible underlying mechanism was also present. Finally, the direct electron transfer reactivity of the Hemoglobin/Au-TPL@CS NPs/multi-walled carbon nanotubes/glassy carbon electrode was investigated with cyclic voltammetry measurements.The biosensor exhibited a good electrocatalytic activity to the reduction of H2O2. Such new type of Au-TPL@CS NPs provides a promising platform of biological system for early illness detection and treatment in future.2. Three kinds of novel facile fabrication routes to prepare Au-F127 nanocolloids with different morphologies including rambutan-like Au-F127 hybrid nanospheres, Au@F127 core-shell nanostructure, and multiple shapes of Au nanosheets are reported. The different Au-F127 nanocolloids are characterised by transmission electron microscopy,electron diffraction patterns, and UV-vis spectroscopy. The different formation mechanisms of the Au-F127 nanocolloids with different morphologies are discussed. The intrinsic properties and application potential of Au-F127 nanocolloids are determined by their size,shape,composition,and structure, A biosensor made of rambutan-like Au-F127 hybrid nanospheres exhibited a good electrocatalytic activity for the reduction of hydrogen peroxide. The Au@F127 core-shell nanoparticles exhibited good blood compatibility attributable to the surface hydrophilicity of the F127 layer. These strategies open up the exciting possibility of using F127 in synthesis of gold nanomaterials of variable morphology, and provide great potential for biomedical applications.3. The good blood compatibility and no-cytotoxicity of the rambutan-like Au-F127 hybrid nanospheres were confirmed by coagulation tests, complement and platelet activation, hemolysis assay, and MTT test in vitro. Further, we established an amperometric glucose biosensor applied in whole blood directly,which was based on the direct electron transfer of glucose oxidase (GOx) entrapped onto the Au-F127 nanospheres. Here, the Au-F127 nanospheres could provide a blood compatible surface with antifouling property for determination of glucose in whole blood. The cyclic voltammetric results indicated that GOx immobilized on the Au-F 127 nanospheres exhibited direct electron transfer reaction, and the cyclic voltammogram (CV) displayed a pair of well-defined and nearly symmetric redox peaks with a formal potential of 93 mV.The biosensor had good electrocatalytic activity toward glucose with a low detection limit 3.15 pM. The glucose biosensor did not respond to ascorbic acid (AA) and uric acid(UA) at their high concentration encountered in blood. In this method, the biosensor was used for quantification of the concentration of glucose in whole blood samples. The data obtained from the biosensor showed good agreement with those from a biochemical analyzer for homologous serum in hospital.4. Acknowledging the benefits of hyperbranched polymers and their nanoparticles,herein we report the design and synthesis of sulfonic acid group functionalized hydroxyl-terminated hyperbranched polyester (HBPE-SO3H) nanoparticles and their biomedical application. The HBPE-SO3H nanoparticles were characterized by transmission electron microscopy (TEM), Fourier transform infrared (FTIR)spectroscopy and proton nuclear magnetic resonance spectroscopy (1H NMR). The good hemocompatibility of HBPE-SO3H nanoparticles was also investigated by coagulation tests, complement activation and platelet activation. The novel glucose biosensor was fabricated by immobilizing the positively charged Au nanoparticles, HBPE-SO3H nanoparticles and GOx onto the surface of glassy carbon electrode (GCE). It can be applied in whole blood directly, which was based on the good hemocompatibility and antibiofouling property of HBPE-SO3H nanoparticles. The biosensor had good electrocatalytic activity toward glucose with a wide linear range (0.2-20 mM), a low detection limit 1.2 × 10-4M in whole blood and good anti-interference property. The development of materials science will offer a novel platform for application to substance detection in whole blood.5. Based on the green chemistry and self-assembly technology, a novel kind of gold-polypyrrole (Au-PPy) hybrid nanoparticles with surface negative charge was synthesized, and its good blood compatibility,no-cytotoxicity, conducting and antibiofouling property were proved. Further, we developed a label-free electrochemical aptamer biosensor using thrombin-binding aptamer (TBA) as receptor for the measurement of thrombin in whole blood. Binding of the thrombin at the modified GCE electrode surface greatly restrained access of electrons for a redox probe of [Fe(CN)6]3-/4-.Moreover, the aptamer biosensor could be used for detection of thrombin in whole blood,a wide detection range (1 fM-500 nM) and a detection limit on the order of 0.29 fM(S/N=3) were demonstrated. Control experiments were also carried out by using bull serum albumin (BSA) and lysozyme in the absence of thrombin. The good stability and repeatability of this aptamer biosensor were also proved. The data obtained from the biosensor showed good agreement with those from a fluorescence analyzer for homologous serum in hospital.更多还原