Design And Synthesis Of Ag@AgCl-based Photoactive Antibacterial Materials And Study On Their Synergistic Action Against Drug-resistant Bacterial Infections

Infectious disease caused by antibiotic-resistant bacteria is considered as one of the major challenges of the public health worldwide.Bacterial resistance to antibiotics has been included in science list of the world’s 125 most cutting-edge scientific questions.Skin and soft tissue infections（SSTIs）are among the most common infections in outpatients.Drug-resistant bacteria are a principle factor which induced the failed treatment and increased mortality rate in patients with SSTIs.Local wound infections may lead to systemic complications,and coupled with the synergies of bacterial resistance,it makes treatment more difficult or even impossible to completely cure.Exploring new methods to promote wound healing while combating drug-resistant bacterial infections is a crucial challenge in the medical community.By virtue of the advantages of large specific surface area,adjustable material size,excellent optical responsiveness and flexible performance,photocatalytic nanostructures offer a more effective strategy to combat drug-resistant bacteria.Moreover,the synergistic mechanism of photodynamic and photothermal against drug-resistant bacteria can achieve the goal of high efficiency,non-toxic and no drug resistance,which possess the excellent practical application valueIn this project,with Ag@AgCl as the main research object,a series of novel Ag@AgCl based antibacterial materials with visible light catalysis and synergistic 808 nm near-infrared photothermal therapy have been constructed for the two major topics of drug-resistant bacteria and promoting healing of infected wounds.We designed and synthesized Ag@AgCl-based photoactive anti-drug-resistant functional materials which can be used to antibacterial from single-mode to multi-mode,and elucidated their role and effect in the antibacterial field,systematically investigated their antibacterial efficiency and antibacterial mechanism against drug-resistant strains of bacteria.The antibacterial and healing function of the prepared materials in the treatment of drug-resistant bacterial infection was systematically studied through animal model experiments,which laid the foundation for the development of new therapeutic strategies.The detailed research methods are as follows:（1）PVDF-Agmembranes were prepared by the phase conversion method.The PVDF-Agmembrane was used as a filtration membrane to remove residual chlorine by the cyclic filtration method,and the PVDF-Ag@AgCl antibacterial membrane with photocatalytic activity（PCT）was also prepared.The vitro antibacterial experiments demonstrated the efficient antibacterial activity against two drug-resistant bacteria icluding ampicillin-resistant Escherichia coli（AREC）and methicillin-resistant Staphylococcus aureus（MRSA）.The photocatalytic antibacterial mechanism was elucidated by the changes of bacterial morphological and bacterial cell permeability.Combined with the vitro hemolysis assay,cytotoxicity,vivo biochemical index assay and zebrafish acute toxicity assay,the good biocompatibility was strong demonstrated.In consideration that the residual chlorine easily induces the development of chlorine/drug-resistant bacteria,PVDF-Agmembranes were applied to filter domestic wastewater and medical wastewater,which not only effectively realized the chlorine recovery but also turns waste into treasure to prepare the PVDF-Ag@AgCl antibacterial membranes.The significant antibacterial effect of PVDF-Ag@AgCl membrane on the isolated strains of bacteria in chlorinated wastewater was further verified.Finally,the ability of the membrane to fight infection and promote healing in vivo was verified through wound infection healing treatments in mice.（2）AgCl-on-Ag NWs antimicrobial materials were synthesized by surface engineering design of N-halamine.In vitro antibacterial experiments demonstrated their efficient antibacterial activity against E.coli,S.aureus,AREC,and MRSA without developing drug resistance.The synergistic antibacterial mechanism of AgCl-on-Ag NWs with ROS（main）and Ag⁺release（supplementary）was revealed by bacterial morphological changes,bacterial cell membrane permeability,bacterial slicing,Ag⁺release,ESR and nucleic acid leakage.It was able to switch their antibacterial modes flexibly to common pathogenic and drug-resistant bacteria.The effect of AgCl-on-Ag NWs on the metabolic pathways of drug-resistant bacteria was further analyzed at the molecular level using transcriptomics.The pathways mainly affect bacterial protein composition by reducing the synthesis of essential compounds,disrupting membrane integrity and structure and disrupting energy metabolism.A wound model of MRSA infection was established to evaluate its ability to effectively eradicate drug-resistant bacteria and accelerate wound healing in vivo.The above experimental findings were also validated with theoretical calculations.（3）GDY-Ag@AgCl antibacterial nanoplatform with mechanisms of photothermal therapy（PTT）promoted PCT was designed and synthesized using in-situ oxidation method.The GDY-Ag@AgCl complexes were prepared by in situ oxidation of N-halamine,which select GDY nanosheets as substrates and prepare GDY-Agprecursors with controllable morphology and adjustable size by in situ reduction.Its therapeutic strategy for wound healing and keratitis was also investigated.Toxicological tests such as cell migration assays,hemolysis assays,biochemical index tests and histological staining of major organs were used to demonstrate its potential for clinical application.The effectiveness in eliminating drug-resistant bacterial infections and accelerating wound healing of GDY-Ag@AgCl under dual-light irradiation inclding visible light and an 808 nm near-infrared laser（VL+808 nm NIR）was evaluated by a wound model of drug-resistant bacterial infections and a keratitis model.The immunohistology test of GDY-Ag@AgCl nanoplatform demonstrated that the photothermal effect could promote the bacterial permeability and simultaneously increased the yield of ROS under the irradiation of both 808 nm NIR and VL dual-light.It can not only effectively kill AREC and MRSA but also promote fibroblast migration and vascular endothelial cell regeneration,thus effectively accelerate wound healing by MRSA infection as well as treat keratitis from MRSA infection.In addition,the conclusions above were also verified by transcriptomic perspectives.（4）Inspired by the morphology of sea urchin in nature,sea urchin-shaped Fe₃O₄@PDA@Ag@AgCl multifunctional composite microsphere with magnetic properties was designed and synthesized.The as prepared Fe₃O₄@PDA@Ag@AgCl multifunctional composite microsphere has the following functions:The sea urchin-like morphology gives it a rich specific surface area and a prominent geometric configuration,which enhances the adhesion to bacteria and the physical puncture ability;at the same time,the strong paramagnetic properties are fully demonstrated by the hysteresis line and the applied magnetic force,which can be applied to recycled for reuse;the biocompatibility is improved by PDA coating while imparting photothermal properties;most importantly,Ag@AgCl heterojunctions with visible light absorption was loaded in the surface,which enhanced the antibacterial activity against drug-resistant bacteria.Based on its excellent multifunctional structure,its physical and chemical properties are expanded,which provides the possibility for its clinical application.At last,its ability to accelerate wound healing was fully verified by establishing MRSA-infected mouse full-skin wound model experiments.