Preparation Of MSN@Ce6@MnO₂-CS/Ag Nanocomposites And Their Synergistic Antibacterial Properties

BACKGROUND:Bacterial infections are one of the biggest public health problems in the world,and various complications caused by wound infections can be life-threatening.With the increase of antibiotic resistance,many scholars have turned their research attention to antimicrobial nanomaterials,but the existing nanomaterials generally have the disadvantages of poor stability,low water solubility,low biocompatibility,high preparation cost and high toxicity.In recent years,phototherapy,which converts light energy into heat or generates reactive oxygen species（ROS）to produce bactericidal effects,has received much attention due to its excellent performance,high tissue penetration ability,and low side effects.PURPOSE:In this study,Chlorine e6（Ce6）was grafted onto mesoporous silica nanoparticle（MSN）to obtain MSN@Ce6,thus improving the dispersion of Ce6 and its photodynamic efficacy.Then,Mn O₂ nanosheets and chitosan（CS）modified Nano Silver（Ag NPs）were used for surface coating to obtain MSN@Ce6@Mn O₂-CS/Ag（MCMA）nanostructures,which can be used for photodynamic therapy（PDT）and photothermal therapy（PTT）and Ag NPs were used to verify the excellent antibacterial performance and biosafety under the synergistic effect of multiple mechanisms.In addition,the combination of hydrogel performance to form antibacterial dressing can be used in clinical treatment of skin wound infections,providing new ideas and solutions for chronic wound infections in clinical settings.METHODS:The main results are as follows:1)MSN was synthesized by template method and MSN was aminated,activated Ce6 was grafted onto MSN surface by amide condensation reaction to get MC.Mn O₂ was synthesized by in-situ reduction under ultrasonic condition to get MSN@Ce6@Mn O₂（MCM）,which was connected with CS modified Ag NPs by electrostatic interaction to get MCMA.The morphology of the material was studied by transmissionelectronmicroscope（TEM）,the surface functional groups were characterized by Fouriertransforminfrared（FTIR）,and the light absorption was evaluated by ultraviolet-visible absorption photometer Ultraviolet-visible Spectrophotometer（UV-vis）and fluorescence spectrum;2)the cytotoxicity and hemolysis of MCMA on mouse fibroblasts L929 were detected by CCK8;3)Co-culture Ce6 and bacteria to determine the best irradiation time of PDT;4)determine the photothermal source and photothermal characteristics of the nanocomposite system by 808nm laser;5)detect the antibacterial property of MCMA and the ability of scavenging biofilm by bacterial coating plate experiment and biofilm scavenging experiment;6)explore its antibacterial mechanism by zeta potential,ROS production,peroxidase-like（POD）,high temperature and oxidized glutathione（GSH）;7)on the basis of the above,MCMA is presented in the form of dressing and acts on the body.The antibacterial property and in vivo biosafety of the dressing were verified by the mouse wound infection model.RESULTS:The UV-vis shows that MC has absorption peaks at 410 nm and 660nm,which proves the successful grafting of Ce6 onto MSN,and the spectrum of MCM shows a new absorption peak at 300-400 nm,which proves the successful cladding of Mn O₂.XPS showed that MCMA contained C,O,N,Si,Mn and Ag elements,which proved the success of MCMA synthesis.The fluorescence spectra showed that the absorption peaks of Ce6,MC,MCM and MCMA appeared at 660 nm at the same time,which proved that the synthesized MCMA still had the photodynamic properties of Ce6.TEM images showed that MCMA was irregularly shaped;uniform particle size;no obvious agglomeration;positive surface charge（+16.49 m V）;and the size was about 321 nm.The hemolysis assay,CCK8 assay and in vivo toxicity test showed that the nanotherapeutic system has excellent biocompatibility.The results of antibacterial experiments showed that the antibacterial rates of MSN,MC,MCM,and MCMA increased sequentially under the same treatment conditions,and the final MCMA obtained had the best antibacterial ability.MCMA at a concentration of 250μg/m L,after simultaneous 808 nm and 660 nm laser irradiation on Escherichiacoli（E.coli）and Staphylococcus aureus（S.aureus）after laser irradiation at concentrations of 250μg/m L and 808 nm and 660 nm,respectively,and achieved 100%and 99.6%antibacterial rates,and 58%and 55.8%clearance of biofilm formation,respectively.The main reasons for its excellent antibacterial properties may be due to its positive surface charge,induction of ROS production,POD-like properties,ability to oxidize GSH,and high temperature effect.The in vivo antibacterial results showed the best healing of infected wounds in the MCMA+808 nm+660 nm group on day 7 of administration on a mouse infection model,and tissue sections showed the most fiber production in this group.CONCLUSION:In this study,we successfully synthesized composite nanomaterials with good biocompatibility and PDT/PTT/CDT synergistic antibacterial effect by using the photothermal material Mn O₂ integrated photosensitizer and metal nanomaterials,which have good prospects for clinical applications.