二氧化钛纳米粒子与铜离子联合毒性研究

发布时间：2018-06-28 06:57

本文选题：二氧化钛纳米粒子 + 铜离子　；参考：《山西医科大学》2017年硕士论文

【摘要】：目的:研究二氧化钛纳米粒子(TiO_2-NPs)与Cu~(2+)联合,对牛血红蛋白(Bovine Hemoglobin,BHb)、人支气管上皮细胞(Human bronchial epithelial cells,16HBE)、斑马鱼胚胎的毒性作用,从分子、细胞和动物整体水平评价TiO_2-NPs与Cu~(2+)联合产生的毒性效应。方法:采用透射电镜等表征TiO_2-NPs及其与Cu~(2+)混合物的表面形貌、粒径及Zeta电位。以BHb为模型蛋白,采用紫外-可见吸收光谱法、荧光光谱法、同步荧光光谱法、时间分辨荧光光谱法及圆二色光谱法结合,探讨TiO_2-NPs、Cu~(2+)及二者联合与BHb的相互作用,并评价它们对BHb的毒性作用。以16HBE细胞为研究对象,采用噻唑蓝(MTT)比色法测定TiO_2-NPs、Cu~(2+)及二者联合对16HBE细胞的毒性作用,利用吖啶橙(AO)/溴化乙锭(EB)荧光双染色法检测16HBE细胞凋亡。选用斑马鱼胚胎为动物模型,以斑马鱼胚胎的孵化率、死亡率、畸形率及鱼鳔正常发育率为指标,构建体内评价TiO_2-NPs、Cu~(2+)及二者联合的毒性体系。结果:TiO_2-NPs粒径约20 nm,Zeta电位为0.0606 mV,在水溶液中的平均粒径为76.44nm。光谱学结果表明,TiO_2-NPs、Cu~(2+)及二者混合物与BHb均发生相互作用,均使BHb的荧光发生猝灭,其猝灭机制主要为静态猝灭;TiO_2-NPs、Cu~(2+)及二者联合均使BHb的构象及二级结构发生变化,螺旋结构变得更为松散,三者对BHb均产生毒性效应。TiO_2-NPs、Cu~(2+)及二者联合对16HBE细胞的毒性作用均呈剂量效应、时间效应;AO/EB结果证明TiO_2-NPs、Cu~(2+)及二者联合均可以诱导16HBE细胞凋亡;0-800mg·L-1的TiO_2-NPs联合10 mg·L-1的Cu~(2+)对16HBE细胞的毒性作用强于TiO_2-NPs。当TiO_2-NPs浓度介于0-200 mg·L-1时,斑马鱼胚胎死亡率均小于15%,畸形率、孵化率及鱼鳔正常发育率与对照组均无显著性差异;当Cu~(2+)浓度大于0.125 mg·L-1时,斑马鱼胚胎72 h孵化率(26%)和染毒终点时鱼鳔正常发育率(27%)较低、染毒终点时的死亡率(8%)和畸形率(25%)较高,且存在剂量依赖关系;0-1mg·L-1 TiO_2-NPs与0.125 mg·L-1 Cu~(2+)联合对斑马鱼胚胎的毒性作用与单独Cu~(2+)相近,当TiO_2-NPs的浓度为5-200 mg·L-1时,二者联合后斑马鱼胚胎的孵化率和鱼鳔正常发育率均增高,死亡率和畸形率均降低。结论:TiO_2-NPs、Cu~(2+)及二者联合均对BHb具有一定的毒性作用。体外0-800 mg·L-1的TiO_2-NPs联合10 mg·L-1 Cu~(2+)对16HBE细胞呈增毒效应。体内5-200 mg·L-1的TiO_2-NPs与0.125 mg·L-1的Cu~(2+)联合对斑马鱼胚胎呈减毒效应。
[Abstract]:Objective: to study the toxic effects of TIO _ 2 nanoparticles (TIO _ 2-NPs) combined with Cu2 on bovine hemoglobinine (BHB), human bronchial epithelial cells (16HBE) and zebrafish embryos. The toxicity of TiO- 2-NPs combined with Cu2 was evaluated at the cellular and animal levels. Methods: the surface morphology, particle size and Zeta potential of TiO2-NPs and their mixtures with Cu2 were characterized by transmission electron microscopy. Using BHB as model protein, the interaction of TiO2-NPsCu2 and its combination with BHB was investigated by UV-Vis absorption spectrum, fluorescence spectrum, synchronous fluorescence spectrum, time-resolved fluorescence spectrum and circular dichroism spectroscopy. Their toxicity to BHB was evaluated. The cytotoxicity of TiO2-NPsCu2 and its combination on 16HBE cells was determined by MTT colorimetry. The apoptosis of 16HBE cells was detected by acridine orange (AO) / ethidium bromide (EB) fluorescence double staining. Using zebrafish embryos as animal model, the toxicity system for evaluating TiO2-NPsCu ~ (2) and its combination in vivo was constructed by using the hatching rate, mortality rate, deformity rate and normal development rate of swim bladder as the indexes of zebrafish embryos. Results the Zeta potential of 20 nmPs was 0.0606 MV and the average particle size in aqueous solution was 76.44 nm. The spectroscopic results show that TiO2-NPsCu ~ (2) and their mixtures interact with BHB, and both of them cause fluorescence quenching of BHB. The quenching mechanism is mainly static quenching of TiO2-NPsOCu2 and the combination of both of them to change the conformation and secondary structure of BHB. The spiral structure became looser, and the toxic effects of the three on BHB. The toxicity of TiO- 2-NPsCu ~ (2) and their combination on 16HBE cells were dose-dependent. The results of AOP / EB showed that TiO2-NPsCu ~ (2) and its combination could induce apoptosis of 16HBE cells. The toxicity of TiO2-NPs combined with Cu2 (10 mg L-1) on 16HBE cells was stronger than that of TiO2-NPs. When tio _ 2-NPs concentration was between 0-200 mg 路L ~ (-1), the embryonic mortality of zebrafish was lower than 15 mg 路L ~ (-1), and there was no significant difference in abnormal rate, hatching rate and normal development rate of swim bladder between the two groups, and when the concentration of Cu2 was more than 0.125 mg 路L ~ (-1), there was no significant difference between the two groups. The hatching rate of zebrafish embryos at 72 h (26%) and the normal development rate of swim bladder at the end of exposure (27%) were lower, the mortality rate (8%) and abnormal rate (25%) were higher at the end point of exposure to zebrafish. The toxicity of TiO2-NPs and 0.125 mg L-1 Cu2 to zebrafish embryos was similar to that of Cu2 alone. When the concentration of TiO2-NPs was 5-200mg L-1, the hatching rate of zebrafish embryos and the normal development rate of swim bladder increased after the combination of TiO2-NPs and TiO2-NPs. Mortality and deformity rate decreased. Conclusion\% TiO2-NPsCu ~ (2) and their combination have some toxic effects on BHB. In vitro, 0 800 mg L -1 TiOs 2 NPs combined with 10 mg L -1 Cu2 showed a toxic effect on 16 HBE cells. The combination of TiOs 2-NPs 5-200mg L-1 and Cu2 (0.125 mg L-1) showed a attenuated effect on zebrafish embryos.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R99

【参考文献】