血红蛋白磁性分子印迹电化学传感器的研制

发布时间：2018-11-15 21:57

【摘要】：分子印迹聚合物(MIPs)具有与目标分子大小、形状及功能基团相匹配的印迹位点,可特异性识别模板分子,其化学及机械性能稳定、选择性强、可重复使用,广泛应用于固相萃取、模拟酶催化及化学传感器等领域。血红蛋白(Hb)是生物体内重要的金属蛋白质,在生命活动中有着重要作用,其浓度的变化会引起许多疾病、甚至死亡,准确检测其含量具有实际意义。电化学分析方法快速、灵敏、成本低、易操作,因此,电化学方法检测Hb一直是人们研究的焦点,也为电化学生物传感器提供了研究基础。本课题制备了血红蛋白磁性分子印迹纳米粒子Fe_3O_4@SiO_2 NPs(MMIPs NPs),并构建了血红蛋白磁性分子印迹电化学传感器,血红蛋白磁性纳米电极电催化检测亚硝酸盐(NO_2~-)。论文的主要研究内容及结果如下:1.选取Hb为模板分子,正硅酸乙酯为功能单体,Fe_3O_4@SiO_2 NPs作为载体,制备了MMIPs NPs。由TEM表征可知,MMIPs NPs形貌规整,具有核壳结构,单分散性良好,平均粒径为80 nm(RSD=6.2%,n=100),印迹薄层大约12 nm。吸附实验表明MMIPs NPs具有较薄的印迹层,且印迹位点在NPs的表面,加快了Hb的传质速率,在1 h内就达到了吸附平衡。粒子的比表面积较大,具有较高的吸附容量107mg/g。实验测得MMIPs NPs对Hb的印迹因子为α=1.75,Hb对牛血清白蛋白(BSA)、辣根过氧化物酶(HRP)、溶菌酶(Lyz)的分离因子分别是βHb/BSA=7.13,βHb/HRP=3.69,βHb/Lyz=2.76,表明MMIPs NPs能够选择性吸附Hb分子。2.在磁性玻碳电极(MGCE)的表面修饰MMIPs NPs,制备了快速、选择性检测Hb的磁性分子印迹电化学传感器,实现了Hb的直接电化学检测。使用磁性电极引入了磁场,其响应电流(Ipa)是无磁场时Ipa的2倍。由于Hb的印迹孔穴在NPs的表面,所以响应时间较短,在7 min时达到平衡。在最佳测试条件下,差分脉冲伏安法(DPV)测定表明,Hb在印迹传感器上的Ipa是其在非印迹传感器上的2.8倍。Hb对辣根过氧化物酶的分离因子是2.6。检测Hb的线性范围是0.005-0.1 mg/m L(r=0.998),检出限是0.0010 mg/m L(S/N=3)。该传感器成功用于分析检测了血样中的Hb,回收率在95.7%和105%之间。3.采用MMIPs NPs固定Hb,修饰在MGCE构建了电化学生物传感器检测NO2。在最佳测试条件下,DPV测定表明,使用磁性电极引入磁场,对NO_2~-的检测有增敏的作用,NO_2~-在Hb/MMIPs NPs MGCE上的还原峰电流(Ip)比其在Hb/MMIPs NPs GCE的Ip增加18%。由于MMIPs NPs的表面有大量印迹孔穴,吸附较多的Hb,所以,NO_2~-在Hb/MMIPs NPs MGCE上的Ip比其在Hb/MNIPs NPs MGCE的Ip增加24%。线性范围是10-100μmol/L,(r=0.998),检出限是5μmol/L(S/N=3)。该传感器具有较好的重复性和较长的使用寿命且在测定NO_2~-时不受常见的无机离子Na~+、K~+、Mg~(2+)、Ca~(2+)、F~-、Cl~-、SO_3~(2-)、NO_3~-、PO_4~(3-)、SO_4~(2-)的影响,可测定实际样品中的NO_2~-。
[Abstract]:Molecularly imprinted polymer (MIPs) has imprinted sites that match the size, shape and functional groups of the target molecule, and can specifically identify template molecules, with stable chemical and mechanical properties, strong selectivity, and reusable. It is widely used in solid phase extraction, mimic enzyme catalysis and chemical sensor. Hemoglobin (Hb) is an important metal protein in organism and plays an important role in life activities. The change of hemoglobin (Hb) concentration can cause many diseases and even death. It is of practical significance to accurately detect the content of hemoglobin (Hb). Electrochemical analysis is rapid, sensitive, low-cost and easy to operate. Therefore, electrochemical detection of Hb has always been the focus of research, and also provides a research basis for electrochemical biosensor. In this paper, hemoglobin magnetic molecularly imprinted nanoparticles (Fe_3O_4@SiO_2 NPs (MMIPs NPs),) were prepared and the electrochemical sensor of hemoglobin magnetic molecular imprinting was constructed. Electrocatalytic determination of nitrite (NO_2~-) with hemoglobin magnetic nano electrode. The main research contents and results are as follows: 1. MMIPs NPs. was prepared by using Hb as template molecule, ethyl orthosilicate as functional monomer and Fe_3O_4@SiO_2 NPs as carrier. TEM analysis shows that, MMIPs NPs has a regular morphology, a core-shell structure, a good monodispersity, an average particle size of 80 nm (RSD=6.2%,n=100), and an imprinted thin layer of about 12 nm.. The adsorption experiments showed that MMIPs NPs had a thin imprinted layer, and the imprinting site was on the surface of NPs, which accelerated the mass transfer rate of Hb and reached the adsorption equilibrium within 1 h. The particle has a large specific surface area and a high adsorption capacity of 107 mg / g. The imprinting factors of MMIPs NPs to Hb were determined as 伪 = 1.75 Hb for bovine serum albumin (BSA), horseradish peroxidase (HRP), lysozyme (Lyz) and 尾 Hb/BSA=7.13, 尾 Hb/HRP=3.69, 尾 Hb/Lyz=2.76, for bovine serum albumin (BSA), horseradish peroxidase (HRP), lysozyme, respectively. The results show that MMIPs NPs can selectively adsorb Hb molecule. 2. A fast and selective magnetic molecularly imprinted electrochemical sensor for the detection of (MGCE) was prepared by modified MMIPs NPs, on the surface of the magnetic glassy carbon electrode. The direct electrochemical detection of Hb was realized. The magnetic field is introduced into the magnetic electrode, and the response current (Ipa) is twice as high as that of the Ipa without magnetic field. Because the imprinted pores of Hb are on the surface of NPs, the response time is shorter and equilibrium is achieved at 7 min. Under the optimum conditions, the differential pulse voltammetry (DPV) method showed that the Ipa of Hb on the imprinting sensor was 2.8 times higher than that on the non-imprinted sensor, and the separation factor of horseradish peroxidase by Hb was 2.6. The linear range of Hb was 0.005-0.1 mg/m / L (r = 0.998) and the detection limit was 0.0010 mg/m / L (S/N=3). The sensor was successfully used to analyze and detect the recovery of Hb, in blood samples in the range of 95.7% and 105%. Electrochemical biosensor for NO2. detection was constructed by using MMIPs NPs fixed Hb, modification in MGCE. Under the optimum test conditions, DPV measurement shows that the magnetic field introduced into the magnetic electrode can enhance the sensitivity of NO_2~- detection, and the reduction peak current (Ip) on Hb/MMIPs NPs MGCE is increased by 18% than that on Hb/MMIPs NPs GCE Ip. Because of the large number of imprinted holes on the surface of MMIPs NPs and the adsorption of more Hb, the Ip of NO_2~- on Hb/MMIPs NPs MGCE is 24% higher than that of Ip on Hb/MNIPs NPs MGCE. The linear range is 10 ~ 100 渭 mol/L, (r ~ 0. 998) and the detection limit is 5 渭 mol/L (S/N=3). The sensor has good repeatability and long service life and is not subject to the common inorganic ions Na~, K ~, Mg~ (2), Ca~ (2), FGC-CCl-Con so _ 3 ~ (2-), NO_3~-, in the determination of NO_2~-. The effect of PO_4~ (3-) and SO_4~ (2-) on the determination of NO_2~-. in real samples can be obtained.
【学位授予单位】：江南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O631.3;TP212.2

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