Application Of Cascade Reaction In Food System Based On Nanozyme And Natural Enzyme Composites

Nanozymes are defined as nanomaterials with enzymatic activity,aiming to solve the limitations of natural enzymes and traditional artificial enzymes.Peroxidase is a type of enzyme that catalyzes the oxidation of substrates by peroxide,which can be found in all organisms.Peroxidase nanozymes have been extensively studied among the development.In the analysis of food components,people use natural oxidase that catalyzes the production of hydrogen peroxide and nanozymes that mimic peroxidase for cascade catalysis to detect oxidase substrates,such as glucose,with high selectivity.However,the low diffusion efficiency of the cascade reaction and unstable intermediate limit the catalytic performance.In addition,due to the mismatch between the working pH of natural oxidase and nanozyme,it is always necessary to perform two incubation processes in different buffers,which not only complicates the reaction system,but also prolongs the detection time.Metal organic framework(MOF)can be used as an independent nanozyme catalyst with peroxidase-like activity due to its abundant active sites and active functional groups.At the same time,due to its good stability,biocompatibility and high modifiability,it has received more and more attention as a type of solid support for immobilized enzymes.In this thesis,based on the copper-based MOF,three natural oxidase/nanozyme composite materials were successfully designed,prepared,and used in the cascade reaction to construct a new method for the detection of the corresponding analytes,and they were successfully applied in different types of food systems.Among the natural oxidase/nanozyme composite materials,the copper-based MOF material exhibits a dual role.On the one hand,it exhibits stable mimic peroxidase activity under neutral conditions.On the other hand,due to its reliable stability,it is used as a solid carrier for complex natural enzymes.The specific work is as follows:1.Preparation of copper-based MOF materials: by the self-assembly method,4,4'-bipyridine with a rigid planar conjugated structure and copper(?)were coordinately combined to form a peroxide at room temperature MOF nanozyme with enzymatic activity,and its morphology and spectrum were characterized.At the same time,the catalytic mechanism of MOF nanozymes under neutral conditions(pH=7)was explored.2.A new method for glucose detection based on composite enzyme materials:under mild conditions,glucose oxidase(GOx)and Cu-MOF were compounded to form GOx@MOF.In the composite system,glucose is oxidized by GOx to gluconic acid and hydrogen peroxide in the presence of oxygen.Under the catalysis of MOF nanozymes,hydrogen peroxide acting as an intermediate product is immediately converted to ·OH,which rapidly oxidizes the chromogenic substrate to generate a colorimetric signal,thereby realizing quantitative glucose detection with a detection limit of 2.67 ?M and successful used in beverages.3.A new method for choline detection based on composite enzyme materials: CuMOF was combined with choline oxidase(Ch Ox)to form a double enzyme complex(Ch Ox@MOF).In the presence of oxygen,the composite enzyme catalyzes choline,accelerating the chromogenic substrate producing signal changes,thereby realizing the quantitative detection of choline,with a detection limit of 2 ?M and successfully applied to the deter mination of choline in milk samples of different brands.4.A new method for galactose detection based on composite enzyme materials:Cu-MOF was combined with galactose oxidase(GAOx)to form GAOx@MOF.In the presence of oxygen,the composites catalyze galactose,accelerating the chromogenic substrate producing signal changes,thereby realizing the quantitative detection of galactose,with a detection limit of 6.67 ?M and successfully applied in milk.