Preparation And Application Of Mimic Enzymes Based On Multi-layer Core-Shell Nanocomposites

In biological systems,reactive oxygen radicals?ROS?are by-products resulting from incomplete oxidation during a series of biological and chemical oxidation processes.Hydrogen peroxide?H₂O₂?is one of the most stable ROS,which can cause the changes in cell membrane structure,DNA damage,intracellular damage and apoptosis through cytoplasmic peroxidation,so,high content reactive oxygen will be cause many various diseases,such as:Parkinson's,Alzheimer's disease,atherosclerosis,heart disease,and cancer.There is also much evidence shows that an appropriate level of ROS content plays an important role in the functioning of cells and signal transduction of cells,so its content can be used as an index for tracking the progress of biochemical reactions.Therefore,the determination of H₂O₂ content has very important significance in the fields of biomedicine and clinical diagnosis.Enzyme is a bio-macromolecule catalyst with catalytic function.Under mild reaction conditions,the enzymatic reaction generally has extremely high reaction efficiency and high efficiency selectivity.However,because of that natural enzymes are easily inactivated and difficult to prepare,they have severely limited the wide application of natural enzymes in actual production.Based on this issue,this article has prepared a series of nanocomposites with peroxidase catalytic activity as an artificial enzyme for the detection of H₂O₂ and glucose content.First,we used ethylene glycol as a solvent to prepare two different structures Fe₃O₄nanoparticles by one-step hydrothermal method,and characterized by XRD,TEM and other tests.At the same time,we build a Fe₃O₄/H₂O₂/TMB reaction system.In this reaction system,when we add appropriate concentrations of H₂O₂ and TMB solution to the buffer solution containing Fe₃O₄ NPs,the pH of the solution is adjusted to be weakly acidic,and the color changes are easy to observed.The color depth of the solution with the H₂O₂concentration also showed a regular change phenomenon.The linear detection range of solid structure Fe₃O₄ NPs for H₂O₂ was 0.81-1.8 mmol/L,and the detection limit was 0.81mmol/L.The linear detection range of hollow structure Fe₃O₄ NPs for H₂O₂ was 1.1-4.0 mmol/L,and the detection limit was 1.1 mmol/L.At the same time,we also measured the Michaelis constants and found that the K_m of the two structures was 2.46 mmol/L and 3.41mmol/L,respectively.Secondly,we use as-prepared Fe₃O₄@CeO₂ NCs with core-shell structure by hydrothermal method,and using hollow Fe₃O₄ NPs as the core.We using TMB,OPD and ABTS as the reaction substrates to determine its catalytic activity,we found that this materials has strong oxidase catalytic activity and can maintain high activity in strong acid environments.Its optimal working temperature is same like human body temperature,It's shown highly reusable structure.As a nanocomposite material,its Michaelis constant?1.13mmol/L?is lower than pure Fe₃O₄ NPs?3.75 mmol/L?and CeO₂ NPs?2.50 mmol/L?,and also confirms the synergy between this two materials.The effect of Fe₃O₄@CeO₂ NCs on the sensitivity of H₂O₂ and glucose is extremely strong.It can detect low levels of the reaction substrate and can be applied in real life.Finally,we used the as-prepared solid Fe₃O₄ NPs as the core,the layer of MnO₂ MCs was grown on the surface,and a Fe₃O₄@MnO₂/H₂O₂/TMB reaction system was constructed to detect the catalytic activity of H₂O₂ and glucose.The results showed that the detection limits of H₂O₂ and glucose were 0.5?mol/L and 1.1?mol/L,respectively.The experiment also studied its reproducibility and found that after many uses,it can still maintain high catalytic activity.And the detection limit of glucose in this system is lower than the glucose concentration in the body of diabetics?9 mmol/L?and normal people?3.9 mmol/L?,this shown that Fe₃O₄@MnO₂ MCM has great practical potentia in the field of catalyst.