| ObjectiveGraphene oxide is an antibacterial material,but its antibacterial effect is relatively simple with high biotoxicity.Based on the negative charge on the surface of bacterial membrane and the acidic microenvironment of infected inflammatory sites,cationic nanoparticles are promising on better targeting to bacterial and bactericidal effect.In this study,new antimicrobial nanomaterials with positive charge were prepared from GO and cationic polymer for the purpose of killing bacterial after targeting bacterial membrane and the antimicrobial resistance of bacteria is reverse,which provides new research schemes and ideas for the design of new antimicrobial materials.Methods(1)The improved Hummer method was used to prepare GO nanosheets,and the azithromycin bond onto GO through esterification reaction to construct the antibacterial nanosheets AZI@GO.(2)The cationic polymer PEI-CA was constructed by poly(polyethyleneimine)and limonin anhydride to increase the positive charge density of acid responsiveness.(3)The acid-responsive antibacterial nanoparticles AZI@GO-PEI-CA were prepared by coupling the cationic polymer with the antibacterial nanosheets.(4)The properties of AZI@GO,PEI-CA,and AZI@GO-PEI-CA were characterized by a series of methods.(5)The biosafety of antibacterial nanomaterials was evaluated by MTT and hemolysis tests.(6)The size,potential and morphology of AZI@GO-PEI-CA were monitored under acidic conditions,and the density change of surface charge and drug release process of the new antibacterial nanomaterials were verified at different p H condition.(7)AZI@GO-PEI-CA was incubated with Escherichia coli and Staph aureus,and the antibacterial effect of AZI@GO-PEI-CA was studied by dilution plate counting method.The targeted adsorption of AZI@GO-PEI-CA a to bacteria and the destruction of membrane integrity were evaluated by fluorescence microscopy and scanning electron microscopy.The synergistic antibacterial effect of graphene antibacterial nanomaterials was systematically studied.(8)We have established an animal model of bacterial infection,and systematically studied the anti-infection therapeutic effect of new antimicrobial nanomaterials by detecting the number of bacteria in the infected site,the level of inflammatory factors,and the repair of tissue damage.ResultsThe antibacterial graphene oxide nanosheets AZI@GO were successfully prepared by graphene oxide nanosheets loaded with azithromycin.A novel acid sensitive antibacterial nano-drug AZI@GO-PEI-CA was further synthesized by modified with cationic polymer.(2)Under acidic conditions,the release efficiency of antibiotics from AZI@GO is as high as 75% within 6h,much higher than 20% under neutral conditions,which demonstrated acid-responsive release of antibiotics.(3)The change of surface potential of antibacterial nanomaterials under acidic conditions verified the increase of positive charge density of acid responsiveness.(4)The results of light microscopy and scanning electron microscopy revealed that the effective antibacterial activity of AZI@GO-PEI-CA under acidic conditions was achieved through the dual effect of targeted adsorption of bacteria and destruction of membrane integrity.(5)In neutral and acidic conditions,the antibacterial nano drugs were incubated with bacteria,and the results showed that AZI@GO and AZI@GO-PEI-CA had excellent antibacterial effect compared with AZI and GO-PEI-CA,and the antibacterial effect of AZI@GO-PEI-CA was the best under acidic conditions,which indicated that AZI-GO had synergistic antibacterial effect,and AZI@GO-PEI-CA played an acid responsive and efficient antibacterial effect.The experimental results on animal level show that this novel antimicrobial nanomaterial has excellent anti-infection therapeutic effect.ConclusionGO combined antibiotics with cationic polymers to form antimicrobial nanomedicines improved the targeted adsorption on the surface of bacterial membrane,realizing the synergistic antimicrobial function and overcoming bacterial drug resistance. |
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