Preparation and Properties of Electrochemical Glucose Biosensor Based on Highly Ordered Au Nanowire Arrays
|Tutor||WuYuCheng; Samuel Adeloju|
|School||Hefei University of Technology|
|Keywords||Au nanowire arrays physical adsorption cross-linking entrapment co-mediator FIA glucose electrochemical biosensor|
In this dissertation, glucose oxidase is immobilized onto the surface of gold nanowire arrays by different methods to construct highly sensitive electrochemical biosensors for glucose detection. On the one hand, well aligned gold nanowire arrays have been synthesized by direct electrodeposition in conjunction with AAO template based on appropriate AAO templates and electrolytes. The results indicate that the gold nanowire arrays fabricated in alkaline solution containing EDTA, Na2SO3, K2HPO4and HAuCl4are prone to obtain uniform morphology and rough surface compared with acidic solution composing of H3BO3and HAuCl4, which is in favour of immobilization of glucose oxidase and the stability of glucose electrochemical biosensors. On the other hand, the influences of different techniques for immobilization of glucose oxidase on the performance of glucose electrochemical biosensors are also investigated and the parameters for constructing glucose electrochemical biosensors with different techniques are optimized to improve the performance of as-prepared biosensors. In addition, FIA is employed to determine glucose concentration in combination with the as-prepared glucose nanobiosensors to improve the efficiency for glucose detection. Glucose bisensors based on different techniques are elucidated in detail as follows.Firstly, a great amount of glucose oxidase is adsorbed onto the surface of gold nanowire arrays due to their large specific surface area, a thin Nafion film is then cast onto the surface of GOx-modified gold nanowire arrays to improve the stability of as-prepard glucose biosensors in case of glucose oxidase loss during storage and measurements. SEM, TEM, XRD and CV are employed to characterize the morphology, microstructure and electroactive surface area of gold nanowire arrays, respectively. Furthermore, FTIR is also used to confirm whether GOx can be effectively immobilized or not by physical adsorption. Based on the optimum parameters such as immersing time, GOx solution concentration, Nafion solution concentration and electrodeposition time, as-prepared glucose biosensors achieve good performance with a high sensitivity of258.8μA·cm-2·mM-1for amperometric detection of glucose, while also achieving a low detection limit of0.2μM, and a wide linear range of10-3270μM, resulting from the accelaration of electron relay between active sites of GOx and gold nanowire arrays. Meanwhile, the presence of the two common interferants, uric acid (UA) and ascorbic acid (AA), has no effect on the performance of the glucose biosensor owing to selectivity of Nafion film. The apparent Michaelis-Menten constant Kmapp is calculated to be5.8mM, indicating an excellent affinity between immobilized GOx in the Nafion-GOx-AuNWA biosensor and the glucose in solution.Secondly, glucose oxidase is entrapped into polypyrrole (PPy) film formed on the surface of gold nanowire arrays in the process of electropolymerization to fabricate glucose electrochemical biosensors. SEM and TEM are employed to observe the morphology of gold nanowire arrays, potential-time curves of polymerization and EIS are utilized to analyze glucose oxidase loading in the polypyrrole film. It is apparent that the potential-time curves for pyrrole only and pyrrole/GOx mixture solution are completely different, implying that GOx is successfully entrapped into polypyrrole film in the process of electropolymerization, and this is further confirmed by EIS for different electrodes. PPy-GOx-AuNWA-based glucose biosensors also obtain good performance towards glucose detection with optimized parameters. The sensitivity of as-prepared glucose biosensor is183.3μA·cm-2·mM-1, linear range and detection limit achieve10-6140μM and0.5μM, respectively.Thirdly, cross-linking method is one of the most popular techniques to construct biosensors. In this thesis, two different kinds of glucose biosensors, the GLA-BSA-GOx-AuNWAs glucose biosensor and GLA-BSA-GOx-Med-AuNWAs glucose biosensor based on cross-linking method are introduced. As for GLA-BSA-GOx-AuNWAs glucose biosensor, the morphology of gold nanowire arrays is characterized by SEM and TEM, respectively. The mass transport through the biofilm made of GLA and BSA and interface features of GLA-BSA-GOx-AuNWAs glucose biosensor are also investigated by CV and EIS, respectively. Effect of parameters, such as GLA concentration, BSA concentration, GOx concentration etc. on the performance of GLA-BSA-GOx-AuNWAs glucose biosensors are studied and these parameters are optimized. GLA-BSA-GOx-AuNWAs glucose biosensors are sensitive to glucose, the sensitivity is as high as379.0μA·cm-2·mM-1. Linear range and limit of detection of GLA-BSA-GOx-AuNWAs glucose biosensors are5-5000μM and0.05μM, respectively. Furthermore,90%of the original amperometric response of the GLA-BSA-GOx-AuNWAs glucose biosensors can be maintained for over1month, indicating an excellent stability, due mainly to the3D nanostructure and the rough surface of the AuNWs.In order to overcome the disadvantage of O2-dependence for aforementioned glucose biosensors, and to improve the amperometric responses and enhance stability of glucose biosensors, mediator K3Fe(CN)6is incorporated into GLA-BSA-GOx-AuNWAs glucose biosensors. Functions of K3Fe(CN)6are investigated by amperometric measurement under O2and N2atomsphere. The results indicate that both K3Fe(CN)6and O2accept electrons from FADH2due to the synergisitic effect so that amperometric responses of GLA-BSA-GOx-Med-AuNWAs glucose biosensors increase drastically. In addition, K3Fe(CN)6also accepts electrons from FADH2instead of O2once the dissolved O2is insufficient. Parameters for constructing GLA-BSA-GOx-Med-AuNWAs glucose biosensors change greatly due to the change of microenvironment for GOx on the surface of AuNWAs after incorporation of K3Fe(CN)6. Compared with GLA-BSA-GOx-AuNWAs glucose biosensors, the performance of the GLA-BSA-GOx-Med-AuNWAs glucose biosensors is much improved because of the incorporation of K3Fe(CN)6after parameter re-optimization. GLA-BSA-GOx-Med-AuNWAs glucose biosensors achieve a high sensitivity as much as548.1"1, a wider linear range between2.5and5400μM and a lower detection limit of0.04 μM. The apparent Michaelis-Menten constant Kmapp is calculated to be5.6mM, indicating an excellent affinity between immobilized GOx in the GLA-BSA-GOx-Med-AuNWAs biosensors and the glucose in solution and good enzymatic dynamic response.Finally, glucose nanobiosensors in combination with FIA technique are employed to determine glucose concentration. On the one hand, glucose nanobiosensors have excellent performance with high sensitivity, wide linear range and low detection limit. On the other hand, FIA technique can analyze the concentration of analyte quickly and continuously. By combining the advantages of glucose nanobiosensors and FIA technique, efficiency for glucose detection is improved significantly. Experimental results indicate that Au-based glucose biosensors show high recovery rate and low relative stardard deviation (RSD), suggesting good reliability of as-prepard glucose biosensors towards glucose detection.