【作者】 穆建帅；

【导师】 王炎；

【作者基本信息】 哈尔滨工业大学 ， 化学工程与技术， 2014， 博士

【摘要】 无机纳米材料一般被认为是生物惰性物质，但最近研究者发现一些无机纳米材料如磁性氧化铁纳米粒子、氧化铈纳米材料和贵金属纳米材料等展现出酶的催化性能。无机纳米材料的这一新颖性能促使产生了一个全新的研究领域-纳米材料模拟酶。本文以Co3O4纳米材料为研究对象，构建了新的纳米材料模拟酶体系，研究了其模拟过氧化物酶及模拟过氧化氢酶的催化性质和机理，探讨了结构与性能之间的关系，并将其应用于H2O2、葡萄糖和钙离子等物质的分析检测。通过筛选研究发现Co3O4纳米粒子具有较强的模拟过氧化物酶催化活性。详细研究了各因素对Co3O4纳米粒子模拟过氧化物酶催化能力的影响，结果显示催化能力依赖于pH、温度和过氧化氢浓度。对模拟过氧化物酶的催化动力学过程进行了研究，米氏常数显示Co3O4纳米粒子对底物四甲基联苯胺的结合能力大于天然过氧化物酶，催化常数表明Co3O4纳米粒子的催化能力略小于天然过氧化物酶。通过荧光探针法和电化学法探讨了催化反应机理，分析出Co3O4纳米粒子模拟过氧化物酶的催化能力来源于它们在催化过程中的电子传递作用。基于Co3O4纳米粒子模拟过氧化物酶，建立了检测H2O2和葡萄糖的方法。检测H2O2的线性范围为：0.05-25mmol/L，检测限是10μmol/L。检测葡萄糖的线性范围是0.01-10mmol/L，检测限是5μmol/L。其它糖如乳糖、果糖和麦芽糖对测定葡萄糖的干扰很小，说明方法具有较好的选择性。研究了结构因素对Co3O4纳米材料模拟过氧化物酶催化性能的影响，包括形貌和表面官能团修饰。合成出不同形貌Co3O4纳米材料：纳米片、纳米棒和纳米立方体，高倍透射电镜分析结果显示它们暴露的主要晶面分别是{112}、{110}和{100}晶面。研究发现各形貌材料模拟过氧化物酶的催化能力大小顺序为：纳米片>>纳米棒>纳米立方体，其中纳米片的催化能力大于天然过氧化物酶。实验结果表明各形貌材料催化能力的差异或许与它们暴露的晶面有关，利用相关软件研究了不同晶面的原子排列方式，分析了晶面对催化能力的影响。将Co3O4纳米片的表面修饰上不同官能团，包括-NH2、-SH、-COOH和-OH，并利用红外光谱、热重分析和Zeta电位分析等手段进行了表征。对其催化性能的研究表明除羟基修饰的纳米片之外，其它修饰物的催化能力都大于未修饰的纳米片，催化能力的大小为：Co3O4（-NH2）> Co3O4（-SH）> Co3O4（-COOH）> Co3O4> Co3O4（-OH），这可能与不同官能团修饰的Co3O4纳米片带有的不同电荷有关。对结构与模拟过氧化物酶催化能力之间关系的研究，为构建高催化能力的模拟过氧化物酶提供了一种有效方法，同时表面修饰可增加其生物亲和性，促进了其潜在的应用能力。发现Co3O4纳米粒子还具有模拟过氧化氢酶性质。并研究了Co3O4纳米粒子模拟过氧化氢酶催化能力的影响因素、催化动力学和稳定性。结果表明：催化能力随pH和温度的上升而增加；通过阿伦尼乌斯公式计算得到Co3O4纳米粒子和天然过氧化氢酶的活化能分别是43.3KJ/mol和42.8KJ/mol，说明两者催化分解H2O2的难易程度相似；Co3O4纳米粒子和天然过氧化氢酶的Kcat和Kcat/Km数值非常接近，表明Co3O4纳米粒子具有很高的模拟过氧化氢酶催化能力；Co3O4纳米粒子的稳定性远高于天然过氧化氢酶。基于Co3O4纳米粒子模拟过氧化氢酶性质，制备了电化学传感器，应用于检测H2O2。电化学检测H2O2的线性范围是：10μmol/L-4mmol/L，检测限是3μmol/L。和其它天然过氧化氢酶相比，具有较宽的检测范围和较低的检测限。对不同形貌Co3O4纳米材料的模拟过氧化氢酶催化性能进行了研究。测定了各材料的催化动力学过程，结果显示活化能大小为：纳米片<纳米棒<纳米立方体，催化能力大小为：纳米片>纳米棒>纳米立方体。阐述了Co3O4纳米材料模拟过氧化氢酶的催化机理，即Co3O4纳米材料上的Co(Ⅲ)从一分子过氧化氢得到一个电子变成Co(Ⅱ)，并使过氧化氢生成氧气和水；其次Co3O4纳米材料上的Co(Ⅱ)将电子传递给另外一分子的过氧化氢变至Co(Ⅲ)，使过氧化氢还原成氢氧根离子。通过催化机理解释了各材料催化能力的差异是来源于它们具有不同的电子传递能力。研究发现钙离子能急剧增加Co3O4纳米材料模拟过氧化氢酶的催化能力。基于钙离子的促进作用，构建了一种新型的钙离子传感器。检测钙离子的线性范围是：0.1-1mmol/L，检测限为4μmol/L。传感器具有良好的选择性和重复性，应用于牛奶样中钙离子的检测，测定值和ICP-AES测定值相吻合，说明构建的传感器具有较高的准确性。更多还原

【Abstract】 Inorganic nanomaterials are generally considered as biologically inert substances. In recent years, it is surprising that some nanomaterials exhibit the catalytic properties of enzymes, such as magnetic iron oxide nanoparticles, cerium oxide nanomaterials, precious metals nanomaterials and so on. The novel properties of nanomaterials promote the establishment of a new field─nanomaterial-based artificial enzymes. We studied Co3O4nanomaterials and constructed the new nanomaterial-based artificial enzymes. The catalytic properties, kinetics and mechanisms of their mimetic peroxidase and mimetic catalase were studied in detail. And Co3O4nanomaterials were applied in the detection of some substances.Based on the systematic research, Co3O4nanoparticles were found to show the peroxidase mimetic activity. The effect of various factors on the catalytic activity of Co3O4nanoparticles was studied, the results showed that the catalytic activity was dependent on pH, temperature and H2O2concentration. The catalytic kinetics of Co3O4nanoparticles were investaged with the Michaelis-Menten equation. Michaelis constants showed that Co3O4nanoparticles had a higher affinity for TMB than that of natural peroxidase. The catalytic constants indicated that the catalytic activity of Co3O4nanoparticles was lower than natural peroxidase. The reaction mechanism was investigated with fluorescence probe and electrochemical methods. The catalytic activity of Co3O4nanoparticles was due to the electron transfer in the catalytic process. Co3O4nanoparticles were applied in the detection of H2O2and glucose with its mimetic peroxidase activity. The linear range for H2O2was from0.05to25mmol/L and the detection limit was10μmol/L. The linear range for glucose was from0.01to10mmol/L and the detection limit was5μmol/L. The demonstrated biosensing system was highly selective for glucose detection.The impact of structure of Co3O4nanomaterials on their peroxidase mimic activity was studied, including morphology and surface modification with functional groups. The Co3O4nanomaterials of different morphology including nanoplates, nanorods and nanocubes were synthesized, and HRTEM results showed that their mainly exposed planes were {112},{110} and {100} planes, respectively. The catalytic activities of Co3O4nanomaterials with different morphology followed the order of nanoplates>> nanorods> nanocubes. The results indicated that the difference in catalytic activity might be related to their explosed planes. The surface atomic configurations in different crystal planes were stuided by relative software, indicating that planes were the important factor on the catalytic activity. Finally, the surfaces of Co3O4nanosheets were modified by different functional groups, including-NH2,-SH,-COOH and-OH, which were verified using infrared spectroscopy, thermal analysis instruments and Zeta potential analysis. Other than the complex modified by hydroxy, the catalytic activities of the complexs were higher than Co3O4nanosheets. The order of their catalytic activities was Co3O4(-NH2)> Co3O4(-SH)> Co3O4(-COOH)> Co3O4> Co3O4(-OH), which might be related with the different electric charges on the surface of the complexs. Studies on the relationship between the structure and the catalytic ability of peroxidase mimic provided an effective method for constructing the peroxidase mimic with high catalytic activity. Surface modification can also increased their biological affinity and promoted their potential applications.The catalytic properties of Co3O4nanoparticles as catalase mimic was firstly studied. The external factors, catalytic kinetics and stability were studied. The results showed that: the catalytic activity increased as pH and temperature; the activation energy of Co3O4nanoparticles and catalase were obtained by Arrhenius formula to be43.3KJ/mol and42.8KJ/mol, indicating close catalytic decomposition barriers; the close Kcat and Kcat/Km values of Co3O4nanoparticles and catlase showed Co3O4nanoparticles had high catalase mimic activity; the stability of Co3O4nanoparticles was higher than that of catalase. Based on the mimetic catalase activity of Co3O4nanoparticles, they were used as the amperometric sensor for the detection of H2O2. The linear range of the H2O2sensor was from10μmol/L to4mmol/L, with the detection limit of3μmol/L. Compared with some other catalases, it had wide range and low detedction.The catalytic properties of Co3O4nanomaterials as catalase mimic with different morphology are studied. The catalytic kinetics of Co3O4nanomaterials as catalase mimic were studied, the activation energies followed the order of nanoplates <nanorods <nanocubes, and the order of their catalytic activity was: nanoplates> nanorods> nanocubes. The catalytic mechanism of Co3O4nanomaterials as catalase mimic was proposed: firstly, the Co(III) of Co3O4nanomaterials obtained electrons easily from H2O2and turned into Co(II), oxidizing H2O2into oxygen and water; secondly, Co(II) in Co3O4nanomaterials transferred electrons to H2O2and changed back to Co(III), reducing H2O2to OH. The difference of catalase mimic activities was due to the different electron transfer ability of Co3O4nanomaterials. Calcium ion was found to dramatically increase the catalase mimic activity of Co3O4nanomaterials. Based on the stimulation of calcium ion, an amperometric biosensor with Co3O4nanoplates was developed to detect Ca2+. The linear range of for detecing Ca2+was0.1and1mmol/L with a detection limit of4μmol/L. It showed high selectivity against other metal ions and good reproducibility. The proposed method was successfully applied for the determination of calcium in milk samples, showing high accuracy.更多还原