The Peroxidase-like Activity Based On Composites Of Ferrocene Derivatives And Their Analytical Application

Natural enzymes occupy a very important position in many areas because of the specificity and efficient catalytic activity etc.However,some of the inherent shortcomings of natural enzymes,such as easy deactivation and difficult purification,limit its wide use.Therefore,it is particularly important to seek a mimetic enzyme material with natural enzyme activity.Compared with natural enzymes,mimic enzyme has the advantages of easy preparation and storage,good stability and easy regulation of activity.Among them,the study of peroxide mimetics is the most prevalent,because hydrogen peroxide is an important active oxygen form in the living body.Since 2007,the ferric oxide magnetic nanoparticles have been reported to have the catalytic activity of peroxide mimetic enzyme,the synthetic materials have been developed rapidly in the field of peroxide mimetic enzyme,and are widely used in electrochemical sensors,environmental pollutants degradation and biological macromolecule detection and so on.In this paper,we have constructed a series of new methods of materials based on ferrocene derivatives and their applications.Firstly,cobalt-ferrocene carboxylic acid?Fc-COOH-Co?was synthesized by the coordination of carboxyl groups with cobalt ions in ferrocenecarboxylic acid.The successful synthesis of the complex was confirmed by infrared characterization.We have found that Fc-COOH-Co has a peroxidase-like activity,that is to say,Fc-COOH-Co can catalyze the oxidation of TMB by H₂O₂,while the oxidized TMB is ablue product.Based on this,we constructed a simple colorimetric method to detect H₂O₂.Under the optimum conditions,the method had a wide linear range?0.01 ?M-70?M?and a lower detection limit?0.01 ?M?.Steady-state kinetic experiments showed that relative to the substrate H₂O₂,the Michaelis-Menten constant of Fc-COOH-Co was0.63 mM,which was significantly lower than HRP?3.70 m M?,indicating that Fc-COOH-Co had a good affinity with H₂O₂.By combining glucose oxidase to catalyze the reaction of glucose and oxygen to produce hydrogen peroxide,the method can be used to detect glucose,with a linear range of 0.01 ?M to 70 ?M and a detection limit of0.01 ?M.This method can also be successfully used for the determination of glucose in serum.In addition,Fc-COOH-Co had excellent pH?2-11?and temperature?10-80??tolerance relative to HRP.Secondly,ferrocenyl-L-histidine Schiff base derivative?L-Fc-CHO?was prepared by hydrothermal method and its peroxidase mimetic enzyme activity was studied.L-Fc-CHO exhibited a synergistic catalytic effect relative to L-histidine and ferrocenecarboxaldehyde.Based on this,we constructed an ultra-sensitive method to detect H₂O₂ and glucose.The detection limit of this method was as low as 8 nM.Steady-state kinetic experiments showed that the Michaelis-Menten constant of L-Fc-CHO relative to the substrate H₂O₂ was 0.22 mM,which is nearly 17 times lower than that of HRP,indicating that the high catalytic activity of L-Fc-CHO was derived from the H₂O₂ excellent affinity.Fluorescence experiments based on the free radical trapping agent-terephthalic acid showed that ·OH was produced in the catalytic reaction and then it further oxidized the substrate.Finally,this method can be used to accurately determine serum glucose.Finally,ferrocene and its derivatives were used as precursors to synthesize magnetic carbon materials containing no functional groups and different oxygencontaining functional groups?hydroxyl,aldehyde and carboxyl groups?by solvothermal method.The characterization of XPS and FT-IR indicated that the oxygen-containing functional groups in the raw material remained in the magnetic carbon.We have found that these four magnetic carbon materials have different peroxidase mimetic enzymecatalytic activity?aldehyde carbon ? carboxyl carbon> carbon> hydroxyl carbon?.The difference in their catalytic activity was mainly due to their different oxygen-containing functional groups,because the other properties of the four kinds of magnetic carbon,such as surface Fe content,morphology and size and the specific surface area were basically no difference.Steady-state kinetic experiments show that the aldehyde-based carbon has a higher maximum reaction rate than the magnetic carbon with non-functional group,the carboxy carbon has a lower Michaelis-Menten,that is,better substrate affinity,while the hydroxyl group has lower maximum reaction rate and higher Michaelis constant.This indicates that the different functional groups in the carbon material act as a catalytic reaction:-C=O groups are as catalytic site,-COOH groups as affinity site,and-OH groups as inhibitor.Through the study of this paper,we can achieve the purpose of regulating the activity of magnetic carbon by different oxygen-containing functional groups.