Studies Of The Third-Generation Enzyme Biosensors Based On Nano-Materials

Enzymes are biological macromolecules, its electroactive groups or redox center was usually deeply buried within the polypeptide chain, difficult to contact with the electrode surface. So it was hard to directly transfer electron between the electrode and enymes. However, because of nanomaterials' unique electrical, electro-catalytic properties and good biocompatibility, it has the very high activity and selectivity as modified material. When nano-materials are used as the modifier on the electrode, the specific physical and chemical properties of the nanomaterial with the large electrode surface area and good adsorption properties can result in the increase of the current response with low detection limit. In this paper, the ChOx/CS-GR/GCE cholesterol enzyme biosensor, GCE/GR-ZnO-Ag-CS/GOx/nafion and nafion/GOx/CS-g-C3N4/GCE glucose enzyme biosensor were prepared with GR, GR-ZnO-Ag and g-C3N4 nanomaterials as modification materials, respectively, studying the direct electrochemistry and electrical catalytic behavior of enzymes. The prepared sensors have wide detection range, low detection limit, good reproducibility and stability, etc., which can be used in the detection of real samples.(1) Graphene oxide (GO) was prepared by the microwave method with natural graphite as raw materials, and chitosan-graphene oxide (CS-GO) was reducted for chitosan-graphene (CS-GR) by in situ reduction method. The ChOx/CS-GR/GCE electrode based on direct electrochemistry of cholesterol oxidase was fabricated by immobilizing cholesterol oxidase (ChOx) on Glassy carbon electrode (GCE) functionalized by chitosan-graphene (CS-GR) composites. The product and the electrode have been characterized by transmission electron microscope (TEM), scanning electron microscopy (SEM), fourier transform infrared spectroscopy (FI-IR), cyclic voltammetry (CV), AC impedance (EIS), differential pulse voltammetry (DPV) and other characterization methods, studying the conductivity, catalytic performance, stability and reproducibility of the electrode. The results showed that, the fabricated electrode exhibited a wide linear response to cholesterol (0.005-1.0 mM), a low detection limit (0.715?M), the low Michaelis-Menten constant (17.39?M) and the high rate constant ks(2.69 s-1). In addition, the biosensor also exhibited excellent reproducibility, stability and very high specificity to cholesterol with complete elimination of interference from UA, AA, DA and glucose. It can be used to test cholesterol in human serum.(2) The partially reducted graphene oxide-zinc oxide-sliver (PRGO-ZnO-Ag) nanocomposites were prepared by the microwave method with natural graphite, silver nitrate (AgNO3), citric acid (C6H8O67) and two zinc acetate hydrate (zinc (CH3COO)2 2H2O) as raw materials, and PRGO-ZnO-Ag-CS was reducted for GR-ZnO-Ag-CS by in situ reduction method. The GCE/GR-ZnO-Ag-CS/GOx/nafion electrode based on direct electrochemistry of glucose oxidase(GOx) was fabricated by immobilizing GOx on Glassy carbon electrode (GCE) functionalized by GR-ZnO-Ag-CS composites. The product and the electrode have been characterized by X-ray diffraction(XRD), Raman spectrum(RS), scanning electron microscopy (SEM),transmission electron microscope (TEM), cyclic voltammetry (CV), differential pulse voltammetry (DPV) and other characterization methods, studying the conductivity, catalytic performance, stability and reproducibility of the electrode. The results showed that, the fabricated electrode exhibited a wide linear response to glucose (0.1-12.0 mM), a low detection limit (10.6?M), the low Michaelis-Menten constant (0.25mM) and the high rate constant ks (2.85 s-1). In addition, the biosensor also exhibited excellent reproducibility, stability and very high specificity to glucose with complete elimination of interference from UA, AA and DA. It can be used to test glucose in human serum.(3) g-C3N4 nanomaterial was successfully prepared with tripolycyanamide as raw materials. The nafion/GOx/CS-g-C3N4/GCE electrode based on direct electrochemistry of glucose oxidase(GOx) was fabricated by immobilizing GOx on Glassy carbon electrode (GCE) functionalized by g-C3N4-CS composites. The product and the electrode have been characterized by X-ray diffraction(XRD), transmission electron microscope (TEM), cyclic voltammetry (CV), differential pulse voltammetry (DPV) and other characterization methods, studying the conductivity, catalytic performance, stability and reproducibility of the electrode. The results showed that, the fabricated electrode exhibited a wide linear response to glucose (0.1-8.0 mM), a low detection limit (46.7?M) and the low Michaelis-Menten constant (0.42mM). In addition, the biosensor also exhibited excellent reproducibility, stability and very high specificity to glucose with complete elimination of interference from UA, AA and DA. It can be used to test glucose in human serum.