【作者】 刘丽；

【导师】 肖玲；

【作者基本信息】 武汉大学 ， 环境科学， 2013， 博士

【摘要】 本论文旨在构建低毒磁性近红外荧光双功能复合纳米材料,研究复合纳米材料与生物大分子之间相互作用,重点讨论紫外光辐射条件下复合纳米材料对生物大分子的毒性效应,为复合纳米材料的进一步体内生物应用提供理论和实验基础。具体研究内容及结论如下：1、巯基乙酸修饰ZnSe量子点的制备及表征采用巯基乙酸(TGA)为稳定剂直接在水相中以紫外光辐射辅助水热法制备了ZnSe量子点。系统地研究了紫外光辐射、反应pH值、前驱体配比和反应时间等实验条件对量子点荧光性能的影响。确定最佳合成条件为：反应pH值为9.5,Zn/NaHSe摩尔比为10：1,Zn/TGA摩尔比为1：3,100℃回流反应1h后,紫外辐射处理19h。采用透射电子显微镜(TEM)、粉体X射线衍射(XRD)、紫外-可见吸收光谱(UV-vis)、荧光发射光谱(FL)和傅里叶变换红外光谱(FT-IR)对量子点的晶体结构、表面结构和光学性能进行表征。制得的量子点为ZnSe/ZnS核壳型立方相闪锌矿结构,平均粒径4nm,荧光量子产率20%。2、CS-Fe3O4@ZnSe:Mn/ZnS磁性荧光复合纳米粒子的制备及表征采用化学共沉淀法制备了磁性壳聚糖CS-Fe304纳米粒子,采用水热法以巯基丙酸MPA为稳定剂通过成核掺杂模式制备了Znse:Mn/ZnS量子点,然后将制得的两种纳米粒子通过静电结合作用复合,制备得到CS-Fe3O4@ZnSe:Mn/ZnS磁性荧光复合纳米粒子。系统地讨论了成核温度、Zn/NaHSe摩尔比例、Mn2+离子掺杂量、稳定剂MPA加入量、溶液pH值、回流时间、ZnSe:Mn/ZnS量子点与CS-Fe3O4的摩尔比等实验条件的影响。确定最佳条件为：成核温度为4℃,Zn/NaHSe摩尔比为5：3,Mn2+离子掺杂量为4%,Zn/MPA摩尔比为1：24,溶液pH值为11,回流时间为7h,ZnSe:Mn/ZnS量子点与CS-Fe3O4的摩尔比为2.5：1。采用TEM.XRD.UV-vis、FL.VSM和FT-IR对其进行表征,结果表明,制得的CS-Fe3O4@ZnSe:Mn/ZnS具有很好的荧光性能和磁响应性能,在紫外灯照射下显示橙红色荧光,室温下饱和磁化强度为17.8emu/g。3.CS-Fe3O4@ZnS:Mn磁性荧光复合纳米粒子的制备及其与蛋白质的相互作用研究采用层层自组装原理,以磁性壳聚糖CS-Fe3O4纳米粒子为基体,通过表面壳聚糖与Zn2+和Mn2+离子的螯合,在CS-Fe3O4磁性纳米粒子上原位生长ZnS:Mn纳米晶,制备得到CS-Fe3O4@ZnS:Mn磁性荧光复合纳米粒子。研究了壳聚糖浓度、前驱体配比、反应温度等参数的影响。确定最佳合成条件为：壳聚糖浓度为1.0％,CS-Fe3O4磁性纳米粒子浓度为0.2mo1·L-1,前驱物Zn2+和S2-离子浓度为0.1mol.L-1,Mn2+离子掺杂量为50%,反应pH值为7,反应温度为100℃。以牛血清白蛋白BSA为模型蛋白质,采用UV-vis和FL等手段研究了没有紫外辐射和紫外辐射条件下CS-Fe3O4@ZnS:Mn与BSA的相互作用。实验结果表明,CS-Fe3O4@ZnS:Mn对BSA具有荧光猝灭作用,其荧光猝灭是CS-Fe3O4@ZnS:Mn与BSA通过静电作用形成了某种特定结构的配合物而引起的静态猝灭,且这是一个自发的反应过程。紫外辐射条件下CS-Fe3O4@ZnS:Mn对BSA的荧光猝灭作用更强。通过对反应过程中活性氧自由基的测定,探讨CS-Fe3O4@ZnS:Mn与BSA相互作用机理,认为CS-Fe3O4@ZnS:Mn与BSA之间的相互作用是荧光共振能量转移和光催化反应共同作用的一个过程。4、CS-Fe3O4@ZnS:Mn/ZnS磁性荧光复合纳米粒子的制备及其与蛋白质的相互作用研究采用外延生长法,制备得到CS-Fe3O4@ZnS:Mn/ZnS磁性荧光复合纳米粒子。系统地研究了核壳比和Mn2+离子掺杂量等参数对CS-Fe3O4@ZnS:Mn/ZnS性能的影响。确定最佳合成条件为：核壳比为6：1,Mn2+离子掺杂量为31%。ZnS壳层有效的钝化了CS-Fe3O4@ZnS:Mn的表面,使荧光强度提高了3.5倍。CS-Fe3O4@ZnS:Mn/ZnS与BSA相互作用研究结果表明,CS-Fe3O4@ZnS:Mn/ZnS对BSA具有荧光猝灭作用,猝灭方式为静态猝灭,相互作用力为静电作用力；紫外光辐射条件下CS-Fe3O4@ZnS:Mn/ZnS对BSA的猝灭作用进一步加强。5、复合体系中CS-Fe3O4@ZnS:Mn/ZnS磁性荧光复合纳米粒子与蛋白质的相互作用研究采用荧光光谱法分别讨论了共存离子Na+、K+、Fe2+和Fe3+离子,以及模拟体液对CS-Fe3O4@ZnS:Mn/ZnS与BSA相互作用的影响。结果发现,Na+或K+离子对BSA、 CS-Fe3O4@ZnS:Mn/ZnS以及CS-Fe304@ZnS:Mn/ZnS-BSA体系的荧光强度均没有明显影响。 Fe2+或Fe3+离子对BSA、CS-Fe3O4@ZnS:Mn/ZnS以及CS-Fe3O4@ZnS:Mn/ZnS-BSA体系具有荧光猝灭作用：Fe2+或Fe3+离子对BSA的猝火是由碰撞引起的动态猝灭；Fe2+或Fe3+离子对CS-Fe3O4@ZnS:Mn/znS MFNPs的猝灭为静态猝灭；Fe2+或Fe3+离子与CS-Fe304@ZnS:Mn/ZnS MFNPs对BSA的荧光猝灭作用具有协同效应,导致BSA荧光猝灭更剧烈。模拟体液使CS-Fe3O4@ZnS:Mn/ZnS-BSA体系荧光强度有所增加。更多还原

【Abstract】 This paper aimed to construct low toxic magnetic-fluorescent nanoparticles (MFNPs), the interaction and damage of MFNPs to serum albumin with or without UV illumination was also investigated. This work may be useful for further development of MFNPs in vivo biological applications. The main works and conclusions in the present study are as follows:1、Synthesis and characterizations of thioglycolic acid-capped ZnSe quantum dotsZnSe quantum dots (ZnSe QDs) were prepared in aqueous media and characterized by ultraviolet-visible absorption, fluorescence emission spectroscopy, X-ray powder diffraction, transmission electron microscopy, energy-dispersive spectroscopy, and inductively coupled plasma atomic emission spectroscopy. The influences of various experimental variables on the photoluminescence properties of ZnSe QDs were investigated. The results showed that the average diameter of these nanocrystals was4nm with narrow size distribution. The optimized conditions were as follows:pH value of9.5, Zn/NaHSe molar ratio of10:1, Zn/TGA molar ratio of1:3, heating time of1h, and irradiation time of19h. Under optimized conditions, the photo luminesce nce quantum yield of ZnSe QDs in aqueous solution reached20%.2、Synthesis and characterizations of CS-Fe3O4@ZnSe:Mn/ZnS MFNPsCS-Fe3O4@ZnSeMn/ZnS MFNPs were synthesized by incorporating chitosan coated Fe3O4magnetic nanoparticles CS-Fe3O4and ZnSe:Mn/ZnS QDs via electrostatic interaction. Effects of experimental variables, such as nucleation temperature, Zn/NaHSe molar ratio, the amount of Mn2+, the kind of stabilizer, Zn/MPA molar ratio, pH value, reaction time and QDs/CS-Fe3O4molar ratio on the fluorescence performance of CS-Fe3O4@ZnSe:Mn/ZnS MFNPs were discussed. The optimized synthetic conditions were found as follows:nucleation temperature as4℃, Zn/NaHSe=5:3, the amount of Mn2+was4%, Zn/MPA=1:24, reaction pH=11, growth duration as7h, QDs/CS-Fe3O4=2.5:1. The synthesized nanocrystals were characterized and analyzed with TEM, XRD, UV-vis, FL spectra, VSM and FT-IR. The obtained CS-Fe3O4@ZnSe:Mn/ZnS MFNPs exhibit orange fluorescent and superparamagnetic behavior, the saturation magnetization was17.8emu/g.3、Novel magnetic-fluorescent CS-Fe3O4@ZnS:Mn nanoparticles:preparation and interaction with serum albuminCS-Fe3O4@ZnS:Mn MFNPs were synthesized in aqueous media under ambient pressure and characterized by X-ray diffraction, UV-vis absorption, fluorescence emission, transmission electron microscope and energy dispersion spectrum. The effects of experimental conditions on the fluorescence properties of CS-Fe3O4@ZnS:Mn were studied. The optimal conditions were that:the amount of CS was1.0%,[CS-Fe3O4]=0.2mol-L-1, [Zn2+]=[S2-]=0.1mol·L-1, Mn2+was50%, reaction pH=7, reaction temperature was100℃In addition, the interaction of CS-Fe3O4@ZnS:Mn with bovine serum albumin (BSA) was studied by means of ultraviolet-visible and fluorescence spectra. The results show that the dominating fluorescence quenching mechanism of CS-Fe3O4@ZnS:Mn with BSA belongs to static quenching. The thermodynamic parameters indicated that electrostatic interaction play major roles in CS-Fe3O4@ZnSMn-BSA, while binding processes exist spontaneously. The damage of BSA is prone to happen in the presence of CS-Fe3O4@ZnS:Mn under UV illumination. The fluorescence quenching of BSA by CS-Fe3O4@ZnS:Mn MFNPs under UV illumination is mainly a result of a combination of FRET and photo-induced free radical procedure.4、Novel magnetic-fluorescent CS-Fe3O4@ZnSMn/ZnS nanoparticles:preparation and interaction with serum albuminNovel magnetic-fluorescent CS-Fe3O4@ZnS:Mn/ZnS nanoparticles were prepared and characterized. The optimum synthetic conditions were the molar ratio of C/S was6:1, the amount of Mn2+was31%. Introduction of ZnS shell significantly enhanced the photo luminescence intensity by3.5times. The interaction of CS-Fe3O4@ZnS:Mn/ZnS with BSA was investigated by ultraviolet-visible and fluorescence spectra. The results show that electrostatic interaction is the major force for the binding processes of BSA to the surface of CS-Fe3O4@ZnS:Mn/ZnS. The damage of BSA is prone to happen in the presence of CS-Fe3O4@ZnS:Mn/ZnS under UV illumination.5、 Effect of some ions on interaction of CS-Fe3O4@ZnS:Mn/ZnS MFNPs with serum albuminThe effect of Na+、K+、Fe2+、Fe3+and simulated body fluid (SBF) on the interaction of CS-Fe3O4@ZnS:Mn/ZnS MFNPs with BSA was studied by fluorescence spectra. The fluorescence intensity of CS-Fe3O4@ZnS:Mn/ZnS MFNPs, BSA, and CS-Fe3O4@ZnS:Mn/ZnS-BSA system is not influenced by the presence of Na+and K+. But the fluorescence intensity of CS-Fe3O4@ZnS:Mn/ZnS MFNPs, BSA, and CS-Fe3O4@ZnS:Mn/ZnS-BSA system is quenched by Fe2+or Fe3+. The dominating fluorescence quenching mechanism of Fe2+or Fe3+with CS-Fe3O4@ZnS:Mn/ZnS MFNPs and BSA belongs to static quenching and dynamic quenching, respectively. There is synergic effect of Fe2+or Fe3+and CS-Fe3O4@ZnS:Mn/ZnS MFNPs on the damage of BSA. SBF slightly increases the fluorescence intensity of CS-Fe3O4@ZnS:Mn/ZnS-BSA system.更多还原