Construction Of Novel Biosensor To Detect Environmental Pollutants

Based on the detection of generation or consumption of electroactive material inenzymatic reaction, electrochemical enzyme sensor was one of the mostly usedbiosensor. The key to construct electrochemical enzyme sensor succcessfully wasimmobilizing enzyme onto the surface of the matrix and promote the elctron transfer.The thesis prepared diffrent enzyme electrode based on different material andimmobilization method and applied in detecting environmental pollution in order toestablish simple, fast, stable and sensitive electrochemical methods. The main points ofour thesis was shown as follows.1Hollow nano-nickel oxide microsphere （NiO） was fabricated by microwavehydrothermal process and characterized by SEM. The prepared NiO was used toimmobilize hemoglobin （Hb） on the carbon paste electrode with1-butyl-3-methylimidazolium tetrafluoroborate （BMIMBF4）. The structure of Hb in thecomposite film was still maintained native-like demonstrated by FTIR spectra andUV-visible spectra. A pair of stable and quasi-reversible redox peaks was observed inphosphate buffer solution which indicating that heme-proteins could achieve its directelectron transfer effectively. The modified electrode displayed an excellent and rapidelectrocatalytic response to the oxidation of NaNO₂. The proposed biosensor exhibited agood linear response to the concentration of NaNO₂in the range from10to170μM,with a detection limit of4.6μM （S/N=3）. The apparent Michaelis-Menten constantwas estimated to be2.4mM. Furthermore, the biosensor possesses satisfactory stability and good reproducibility.2Environmental estrogen attracted extensive attention and research because theycan disturb the human body metabolism, it’s necessary to detect their content inenvironment. Bi₂WO₆nano/microstructures were prepared by hydrothermal method andcharacterized by scanning electron microscopy （SEM）. The synthesized Bi₂WO₆nano/microstructures were then used to fabricate modified carbon paste electrode（Bi₂WO₆-CPE） by a simple rubbing method. Electrochemical impedance spectroscopy（EIS） of the sensor indicated electron transfer resistance of the Bi₂WO₆-CPE reducedvastly compared with CPE, while cyclic voltammetry （CV） demonstrated the Bi₂WO₆nano/microstructures could accelerate the rate of electron transfer of diethylstilbestrol（DES）. Moreover, the sensor exhibited good selectivity toward simultaneousdetermination of DES and bisphenol A （BPA）. By using DPV method, low detectionlimits of0.009μM （S/N=3） and0.01μM （S/N=3） for DES and BPA were obtained, withthe linear calibration curves over the concentration range0.02-2.3μM and0.03^-1.3μM,respectively. In addition, the sensor was applied for the determination of analytesextracted from milk powder for middle and old age with a recovery of91%⁹9%and88%⁹8%, respectively.3Hot research of the third generation biosensor focuses on the immobilization ofenzyme or protein onto an electrode through suitable immobilized method whileretaining their biology catalytic activity at the same time. Amino-functionalizedhydrogel （PNIPAM-NH₂） was prepared by emulsifier-free emulsion polymerization andthen applied to covalent-embed hemoglobin （Hb） to achieve a novel trichloroacetic acid（TCA） biosensor （Hb/PNN/CPE）. The Hb immobilized in the hydrogel membrane canretain its bioactivity partially with an apparent activity concentration of4.09×10-9Mcm^-1.The proposed biosensor exhibited a good linear response to the concentration ofTCA in the range from20to485μmoL/L, with a detection limit of11μmoL/L （S/N=3）, the apparent Michaelis-Menten constant was estimated to be0.18mmoL/L.