Preliminary Exploration Of The Enhangce-ment Of Copper Based Nanoenzyme Activity And The Detection Of The H₂O₂ And Glucose

Nanoenzyme is an analog enzyme with unique properties and catalytic functions.The catalytic activity of nanoenzyme can be regulated by changing the size and specific surface area,the structure of nanomaterials,and modifying the surface by groups and so on.Nanoenzyme is widely used in food,medicine,chemical industry,agriculture and other fields due to its high efficiency in catalysis,economy and large-scale preparation.And,it is often used in biochemical analysis in basic research.The amount of hydrogen peroxide and glucose in food affects the health of people,hydrogen peroxide and glucose in body fluids are important indicators for the diagnosis of diseases.Therefore,it is significant to develop highly sensitive hydrogen peroxide and glucose detection methods.In this paper,three methods for the detection of hydrogen peroxide and glucose were developed,starting from the enhancement of nano-analog enzyme activity and the establishment of high sensitivity detection methods.The specific work and results are as follows:1.The colorimetric detection of hydrogen peroxide and glucose was established based on Cl-enhanced CuO NPs mimic haloperoxidase.Firstly,the catalytic mechanism of CuO NPs under the presence of three substrates was investigated,and it was found that it conforms to the model of Michaelis-Menten and ping-pong mechanism,which is similar to the natural haloperoxidase.Then,the reactive chlorine species was confirmed by the halogenation experiments of signal molecules,and it was first proposed that CuO NPs has the activity of haloperoxide,and the reaction mechanism was summarized:CuO NPs bind and react with the first substrate(H2O2),releasing the activated peroxo-intermediate before reacting with the second substrate(Cl-)to yield hypochlorous acid.This compound further reacts with TMB or an additional equivalent of hydrogen peroxide to release singlet oxygen.Meanwhile,Cl-accelerates the CuO-Fenton reaction by facilitating the reduction of Cu2+ to form Cu+on the nanosurfaces.At last,the colorimetric detection method of hydrogen peroxide and glucose was established by using the system of CuO-H2O2-Cl-.The linear range and detection limit of hydrogen peroxide were 2.5 ?M to 200 ?M and 2.5?M,respectively.The detection limit was 40 times higher than that of CuO-H2O2.The linear range and detection limit of glucose were 10 ?to 300 ?M and 10 ?M,respectively.The detection limit was 20 times higher than that of CuO-H2O2.Signals from other saccharides were not enough to interfere with glucose measurements.The results were consistent with the standard method,and the deviation of the two methods was 1.5%.2.The fluorescence detection of hydrogen peroxide and glucose was established based on the high efficiency catalytic activity of the low-valent metal in CuFe02 nanoparticles.First,the properties of the mimic enzyme of CuFeO2 nanoparticles were investigated,and it was found that it could catalyze the oxidation of the TMB substrate,resulting in a blue colored product.The catalytic mechanism of CuFe02 nanoparticles conforms to the model of Michaelis-Menten and ping-pong mechanism.The catalytic activity of CuFeO2 was higher than that of Fe2O3,CuO,Fe3O4 and other nanoparticles.Then the reasons for the efficient catalytic activity of CuFeO2 were determined.Except the process of Cu+(?)Cu2+and Fe3+(?)Fe2+ to produce ·OH,there is synergies between Cu+ and Fe3+,that is Cu+ reduces Fe3+ to Fe2+,and the increase of Fe2+ lead to more hydroxyl radicals generated,thus accelerating the reaction rate and enhancing the catalytic activity.Finally,based on the High catalytic activity of CuFe02,a fluorescence detection method for hydrogen peroxide and glucose was established based on the principle of oxidation of terephthalic acid by hydroxyl radical to produce fluorescence substance.The linear range and detection limit of hydrogen peroxide were 0.05 ?M to 10 ?M and 32 nM,respectively.The linear range and detection limit of glucose were 1 ?M to 10 ?M and 0.24 ?M,respectively.The results were satisfactory for the determination of glucose in the actual serum samples,with a difference of 3.9%between this method ands tandard method.3.The mimicry enzyme activity of the magnetic CuFe2O4 nanoparticles was studied,the catalytic process conforms to the model of Michaelis-Menten and ping-pong mechanism.The combination of CuFe2O4 and MWCNTs with good conductivity into CuFe2O4-MWCNTs nanocomposite material not only maintains the inherent properties of the two materials,but also gives new properties to the materials.The catalytic activity of CuFe2O4-MWCNTs nanocomposite was higher than that of CuFe2O4 and MWCNTs alone.The intensity of chemiluminescence and the concentration of hydrogen peroxide under 0.02 mM?0.5 mM had good linearity and the detection limit was 0.5?M.