Study On The Structure-activity Relationship And Biomedical Applications Of Single-atom Nanozymes

Nanozymes are a class of nanomaterials with enzyme-like catalytic properties.Since the first discovery of the intrinsic peroxidase-like activity of Fe3O4 by Yan Xiyun’s group in2007,a series of different kinds of nanomaterials,such as metals,metal oxides,carbon-based nanomaterials and metal-organic framework(MOF),have been developed as nanozymes.At the same time,researchers have found that nanozymes can exhibit not only oxidoreductase activities,i.e.,peroxidase-like,catalase-like,oxidase-like and superoxide dismutase-like activities,but also hydrolase-like and decarboxylase-like activities.The development of these different nanozymes has greatly enriched the variety of artificial enzyme mimics.As an alternative to natural enzymes,nanozymes have many advantages over natural enzymes,such as low cost,high stability,adjustable activity,and scalable preparation.Therefore,nanozymes have been widely used in recent years in biomedical applications such as antibacterial therapy and antitumor therapy.However,the main challenges in the field of nanozyme research at this stage are how to achieve the controlled synthesis of nanozyme with high catalytic activity through the rational design,how to reveal the enzyme-like catalytic process and how to construct efficient and low toxic nanozyme-based catalytic therapeutic systems.These challenges require continuous exploration and understanding of the surface interface structure and properties of nanozymes,in-depth investigation of the structure-activity relationships of nanozymes,and ultimately the elucidation of the action mechanism of nanozymes in the complex in vivo microenvironment.In this thesis,based on the catalytic active structure and catalytic behavior of natural enzymes,we acheieved the bioinspired design of nanozymes.A variety of single-atom nanozymes(SAzymes)with well-defined active sites were developed in an in-depth and systematic manner from various aspects such as the modulation of metalloactive center,the selection of coordination atom,and the optimization of coordination number.The main contents of this thesis are as follows:Firstly,we developed a Zn-N-C single-atom catalyst derived from Zeolitic imidazolate framework-8(ZIF-8)as a research object for the first time,and developed a single-atom Zn nanozyme with excellent peroxidase-like activity,PMCS.PMCS has similar catalytic activity and M–Nx activity as natural enzyme sites.The experimental results showed that the high catalytic activity of PMCS can be attributed to the coordination-unsaturated Zn–N4active site.In addition,we revealed the catalytic mechanism of PMCS by density functional theory(DFT)calculations that Zn–N4 promotes the homolytic process of H2O2 and generates the free radical·OH.In an in vitro antibacterial assay,PMCS inhibited P.aeruginosa up to 99.87%.In a wound infection model,PMCS significantly promoted wound healing,which was close to healing by day 6,while showing no significant toxicity to various tissues and organs.To further modulate the metal active center and mimic the active structure of natural peroxidases,we developed a high-temperature pyrolysis activation strategy mediated by the nitrogen-rich mat erial melamine for the controlled synthesis of iron metal-centered single-atom nanozyme(Fe N5 SAzyme),which showed a five-coordinated active structure and catalytic behavior similar to that of natural enzymes.The Fe–N5 structure consisting of an axial fifth N ligand with Fe–N4 planes could effectively enhance the peroxidase-like activity,and the catalytic efficiency of Fe N5 SAzyme was 7.64 and 3.45×10~5 times higher than that of the single-atom iron nanozyme with four-coordinated structure(Fe N4 SAzyme)and Fe3O4,respectively.Experiments and DFT calculations confirmed that the Fe–N5 active structure exhibited higher affinity and better activation ability for H2O2,which could significantly reduce the energy barrier during the reaction rate-limiting step.By inducing the decomposition of H2O2 into toxic·OH,the Fe N5 SAzyme achieved specific killing of tumor cells and significantly prolonged the survival of 4T1 tumor-bearing mice.Finally,we constructed a sulfur-engineered single-atom copper nanozyme(Cu-N/S-C)by effectively introducing S element into carbon nanostructures using chemical vapor deposition,and determined that Cu-N/S-C possessed the active site of Cu–N1S2 by experimental characterizations.Compared with the non-S-engineered single-atom copper nanozyme Cu-N-C,Cu-N/S-C exhibited a higher peroxidase-like activity as well as an intrinsic catalase-like activity.During the catalytic reaction,the introduction of S plays an important role in the activation and decomposition of H2O2.In addition,due to its excellent absorption properties in the near-infrared region,Cu-N/S-C also exhibits photothermal and photodynamic properties,and its photodynamic properties can be significantly enhanced by its own catalase-like activity.Therapeutic experiments at the cellular and animal levels have shown that Cu-N/S-C with Cu–N1S2 sites can achieve specific damage to tumor cells,inhibit the proliferation process of tumor cells,and thus block the tumor process through the synergistic effect between nanozyme-based catalytic therapy and phototherapy,with the tumor inhibition rate of 85.9%.In summary,this thesis innovatively developed various metal-centered SAzymes via learning from the active structure and catalytic activity of natural enzymes.The homogeneous and dispersed metalloactive sites enable the SAzymes to maximize the atom utilization efficiency,and thus exhibit excellent enzyme-like activities.The well-defined active sites provide a convenient and advantageous way to reveal the structure-activity relationship of nanozymes.The unsaturated coordination metal sites of SAzymes play an important role in improving the affinity and activation ability of the substrate during the enzyme-like catalysis.The constructed SAzymes-based catalytic therapeutic system can achieve excellent antibacterial and antitumor therapeutic efficiency.These works will open up new prospects for the bioinspired design of nanozymes and the development of therapeutic agents with highly catalytic activity.