Cobalt Based Nanomaterials As Oxidase Mimics And Their Bioanalysis Applications

Enzymes are important substances that maintain the life movements.Compared with natural enzymes,nanoenzymes have significant advantages such as low cost,high stability,and easy regulation of activity.With the rapid development of nanotechnology,novel nanoenzymes have brought new vitality to the field of nanoenzymes,as well as new challenges.For nanoenzymes with multiple enzyme activities,taking oxidase-like activity and peroxidase-like activity for example.The oxidase-like activity will lead to the oxidation of the substrate which will cause color interference and affect the utilization of peroxidase.There are fewer studies on oxidase-like nanozymes than peroxidase-like nanozymes.Therefore,we hope to prepare novel nanoenzymes that only exihibit oxidase-like activity and no peroxidase-like activity.Cobalt-based nanomaterials have been widely used in the field of oxygen reduction reactions（ORR）due to their stable performance and high catalytic activity.The oxidase-like activity also includes oxygen reduction reactions,so cobalt-based nanomaterials have great potential to become novel oxidase-like nanozymes.In this paper,we focous on the construction of novel oxidase-like nanoenzyme based on cobalt-based nanomaterials,and application of biomolecules detection.The details are summarized as follows:（1）Rational design of N-doped carbon nanocages equipped with Co-N_x sites for oxidase mimicking and sensing applicationRational design of high-performance nanozyme is of great significance for sensing application.Here,the N-doped carbon nanocages-equipped Co-N_x active sites（CoN_x-NC）were fabricated by simple acid etching of the Co@NC,the carbonized product of the Co-Co Prussian blue analogues in N₂ atmosphere.Both the optimized Co@NC-650and CoN_x-NC-650 exhibit catalase-and oxidase-like properties.They can efficiently decompose H₂O₂into O₂ quickly,and oxidize colorless 3,3’,5,5’-tetramethylbenzidine（TMB）into blue product.This enables them no peroxidase-like activity.Impressively,the oxidase-like activity of CoN_x-NC-650 is 3.9-fold times that of un-etched Co@NC-650.Acid treatment induces the dissolution of Co nanoparticles generated during carbonization,resulting in the formation of extra mesoporous structure and exposure of more Co-N_x active species.Notably,the K_m value for CoN_x-NC-650 with TMB as substrate is 0.186 m M,which is 3.2-fold lower than that of classic oxidase-mimic CeO₂NPs,suggesting its higher affinity to TMB.Given its excellent oxidase-like activity,the CoN_x-NC-650 was used to sensitively and selectively determine acetylcholinesterase（AChE）activity based on the thiocholine（TCh）inhibition on TMB color reaction.TCh is produced by hydrolysis of acetylthiocholine catalyzed by AChE.The colorimetric biosensor has a linear response to AChE over 0.6-800 m U/L range and a detection limit（LOD,3σ）of 0.2 m U/L.The assay was successfully applied to determinate AChE activity in biological samples.（2）Hierarchical Co_0.85Se-NC nanozyme as oxidase-mimic for sensitive D-pen assayResearchers all over the world are on a perpetual search for nanozymes exploit.Transition metal selenides are widely used as catalysts in electrochemical and energy storage fields on account of ultra-conductivity.CoSe_x possesses unexpected oxidase-like activity.Herein,we report a sacrificed-template method to synthesize Co_0.85Se-NC with oxidase-like activity for sensitive D-pen detection.Notably,the K_m value for Co_0.85Se-NC with TMB as substrate is 0.13 m M,which is 3.2 fold lower than that of classic oxidase-mimic nanoceria suggesting the higher affinity to TMB.Under the optimum conditions,in the concentration range of 4-80μM,Co_0.85Se-NC showed a sensitive response to D-pen and the limit of detection（LOD,3σ/s）is 1.14μM.In addition,we also apply the proposed method to D-pen detection in pharmaceuticals and the results are satisfactory showing the good practical operability and clinical application potentialities.