Biomolecule Functionalized Nanoenzymes Mimic Enzyme Properties And Visualized Biosensing Applications

Nanomaterial-based artificial enzymes（nanozymes）have been attracting increasing attention in the past few years,because they exhibit several advantages such as low cost,stability against denaturing,and tunability in catalytic activities.In addition to the composition,crystallinity,and structure of nanozyme core,the catalytic performance of nanozymes is closely associated with the intrinsic properties of the surface modifier.Rational selection of modifiers can increase binding affinity and accelerate the catalyzed reaction by bringing substrates into proximity with the nanozyme’s active sites.Proteins and amino acids have been extensively explored to not only synthesize nanocrystals with small size and excellent monodispersity but also significantly increase the stabilization and biocompatibility of the system.More importantly,it introduces highly specific or multiple functionalities onto these hybrid nanoparticles based nanozyme for further multifunction application.In the present work,amino acid and protein modified nanozymes,and the enzyme activity and the application of the obtained nanozymes are studied.1.Amino acid-mediated ’turn-off/turn-on’ nanozyme activity of gold nanoclusters for sensitive and selective detection of copper ions and histidineHerein,we presented a facile strategy for highly sensitive and selective detection of both Cu²⁺and histidine（His）by combining the peroxidase-like nanozyme activity of gold nanoclusters with amino acid’s ambidentate nature.The peroxidase-like catalytic ability of histidine-Au nanoclusters（His-AuNCs）can be inhibited by the addition of Cu²⁺.The sensitivity of this probe to Cu²⁺ is significant with a linear range of 1-100 nM,and a low detection limit of 0.1 nM.More interestingly,His-AuNC/Cu²⁺ undergoes recovery of the activity upon exposure to free His,because His/Cu²⁺ complex is more stable due to the participation of the imidazole ring of His.The method displays a good selectivity toward histidine over all the other amino acids,with a wide linear relationship in the range of 20 nM-2μM,and a low detection limit of 20 nM.The feasibility of the probe for the rapid analysis of copper ion and His in human serum has been demonstrated with satisfactory results.With the merits of high sensitivity and selectivity,simplification,low cost,and visual readout with the naked eye,this novel ’turn-off/turn-on’sensing approach based on the amino acid’s ambidentate nature is potentially applicable to metal ions,amino acids and peptides in biological and environmental areas.2.Enzyme activity of histidine modified Fe₃O₄ nanoparticles and its application in ascorbic acid determinationThe surface modification of nanoparticles can not only effectively prevent the aggregation of Fe₃O₄ magnetic nanoparticles,but also affect its own mimetic enzyme activity.We synthesized histidine stabilized Fe₃O₄ nanoparticles through coprecipitation method.The products were characterized by TG,TEM,SEM,FT-IR,XRD and XPS measurements.The results show that the histidine plays an important role in the stability of the Fe₃O₄ nanoparticles.The composites as peroxidase mimetics were used to catalyze the oxidation of an enzyme substrate TMB by H2O2 to the oxidized blue color product.Based on the fact that His-Fe₃O₄ composite particles have efficient mimic enzyme activity,we have constructed a fast and sensitive method for ascorbic acid determination.The detection limit is 1 nM（s/n=3）,and the linear range is 1 nM-10μM.This method will have potential application in environmental chemistry,medical analysis and other fields,and can be applied to on-site and real-time detection.3.The properties of the simulated enzyme of magnetic particles functionalized by casein and the visual detection of alcoholIn this chapter,Fe₃O₄ nanoparticles coated with casein were prepared by hydrothermal method.The products were characterized by TEM,SEM,FT-IR,XRD and XPS measurements.The results showed that casein played an important role in the stability of Fe₃O₄ nanoparticles.The further study shows that the protein molecules in the Casein-Fe₃O₄ particles have an enhanced effect on the peroxidase activity of the Fe₃O₄ nanoparticles,and the affinity of the composite particles to the substrate TMB and the H2O2 is significantly higher than that of the unmodified Fe₃O₄ nanoparticles.We constructed a fast and sensitive method for the determination of H2O2.The detection limit is 50 nM（s/n=3）,and the linear range is 50 nM-10μM.AOx/Casein-Fe₃O₄ composite particles were prepared on the basis of Casein-Fe₃O₄ composite particles,and a fast and simple method for the determination of ethanol was established.The detection limit is 10μM and the linear range is 10μM-10 mM.The preparation of Casein-Fe₃O₄ composite nanoparticles is simple,easy to store,good biocompatibility,and easy to carry out magnetic separation.This method will be of potential application in the fields of environmental chemistry,medical analysis and other fields,and can meet field and real-time detection.