【摘要】：分子印跡聚合物(MIPs)具有與目標分子大小、形狀及功能基團相匹配的印跡位點,可特異性識別模板分子,其化學及機械性能穩(wěn)定、選擇性強、可重復使用,廣泛應用于固相萃取、模擬酶催化及化學傳感器等領域。血紅蛋白(Hb)是生物體內重要的金屬蛋白質,在生命活動中有著重要作用,其濃度的變化會引起許多疾病、甚至死亡,準確檢測其含量具有實際意義。電化學分析方法快速、靈敏、成本低、易操作,因此,電化學方法檢測Hb一直是人們研究的焦點,也為電化學生物傳感器提供了研究基礎。本課題制備了血紅蛋白磁性分子印跡納米粒子Fe_3O_4@SiO_2 NPs(MMIPs NPs),并構建了血紅蛋白磁性分子印跡電化學傳感器,血紅蛋白磁性納米電極電催化檢測亞硝酸鹽(NO_2~-)。論文的主要研究內容及結果如下:1.選取Hb為模板分子,正硅酸乙酯為功能單體,Fe_3O_4@SiO_2 NPs作為載體,制備了MMIPs NPs。由TEM表征可知,MMIPs NPs形貌規(guī)整,具有核殼結構,單分散性良好,平均粒徑為80 nm(RSD=6.2%,n=100),印跡薄層大約12 nm。吸附實驗表明MMIPs NPs具有較薄的印跡層,且印跡位點在NPs的表面,加快了Hb的傳質速率,在1 h內就達到了吸附平衡。粒子的比表面積較大,具有較高的吸附容量107mg/g。實驗測得MMIPs NPs對Hb的印跡因子為α=1.75,Hb對牛血清白蛋白(BSA)、辣根過氧化物酶(HRP)、溶菌酶(Lyz)的分離因子分別是βHb/BSA=7.13,βHb/HRP=3.69,βHb/Lyz=2.76,表明MMIPs NPs能夠選擇性吸附Hb分子。2.在磁性玻碳電極(MGCE)的表面修飾MMIPs NPs,制備了快速、選擇性檢測Hb的磁性分子印跡電化學傳感器,實現(xiàn)了Hb的直接電化學檢測。使用磁性電極引入了磁場,其響應電流(Ipa)是無磁場時Ipa的2倍。由于Hb的印跡孔穴在NPs的表面,所以響應時間較短,在7 min時達到平衡。在最佳測試條件下,差分脈沖伏安法(DPV)測定表明,Hb在印跡傳感器上的Ipa是其在非印跡傳感器上的2.8倍。Hb對辣根過氧化物酶的分離因子是2.6。檢測Hb的線性范圍是0.005-0.1 mg/m L(r=0.998),檢出限是0.0010 mg/m L(S/N=3)。該傳感器成功用于分析檢測了血樣中的Hb,回收率在95.7%和105%之間。3.采用MMIPs NPs固定Hb,修飾在MGCE構建了電化學生物傳感器檢測NO2。在最佳測試條件下,DPV測定表明,使用磁性電極引入磁場,對NO_2~-的檢測有增敏的作用,NO_2~-在Hb/MMIPs NPs MGCE上的還原峰電流(Ip)比其在Hb/MMIPs NPs GCE的Ip增加18%。由于MMIPs NPs的表面有大量印跡孔穴,吸附較多的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%。線性范圍是10-100μmol/L,(r=0.998),檢出限是5μmol/L(S/N=3)。該傳感器具有較好的重復性和較長的使用壽命且在測定NO_2~-時不受常見的無機離子Na~+、K~+、Mg~(2+)、Ca~(2+)、F~-、Cl~-、SO_3~(2-)、NO_3~-、PO_4~(3-)、SO_4~(2-)的影響,可測定實際樣品中的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 偽 = 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.
【學位授予單位】：江南大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：O631.3;TP212.2

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