Study On The Targeted Antibacterial Activity Of GO/Apt/BBR And GO/Apt/AE Nanoparticles Against Staphylococcus Aureus

This paper took Staphylococcus aureus(S.aureus)as the main research object.S.aureus specific aptamers(Apt)were modified on the surface of graphene oxide(GO),and then the active components of traditional Chinese medicine berberine(BBR)and aloe-emodin(AE)were further loaded to produce traditional Chinese medicine targeted nanoparticles(GO/Apt/BBR and GO/Apt/AE).After preparation and characterization,the targeted antibacterial properties of two nanoparticles in vitro and vitro were studied.The research contents were as follows:1 Characterization and antibacterial properties study of GO/Apt/BBR and GO/Apt/AEIn this chapter,covalent modification of S.aureus specific Apt on GO surface was first carried out,followed by loading of the active components of traditional Chinese medicine(BBR or AE)with great antibacterial effect to synthesize GO/Apt/BBR and GO/Apt/AE.Then,the prepared nanoparticles were characterized by ultraviolet visible absorption spectrum(UV-vis),fourier transform infrared spectroscopy(FTIR),dynamic light scattering(DLS)and zeta potential.After that,the drug loading ability and release performance of nanoparticles were studied by UV-vis.The results showed that both nanoparticles had good drug loading ability and the release of BBR or AE in vitro was p H dependent.The results of standard plate counting method demonstrated that the antibacterial effect of GO/Apt/BBR and GO/Apt/AE against S.aureus significantly enhanced compared with BBR and AE in vitro.The above results showed that GO/Apt/BBR and GO/Apt/AE had been successfully prepared,and the antibacterial effect against S.aureus significantly enhanced compared with BBR and AE.2 Study on the targeted antibacterial activity of GO/Apt/BBR and GO/Apt/AE combined nanoparticles in vitroIn this chapter,the best concentration ratio of GO/Apt/BBR and GO/Apt/AE were studied by the micro broth dilution method.It showed that the best inhibition effect of GO/Apt/BBR and GO/Apt/AE was molar ratio 5:1.Fluorescence microscope and flow cytometry results showed that the fluorescence response of the combined nanoparticles to Escherichia coli was very weak,while a strong red fluorescence signal can be observed on the surface of S.aureus.The inhibition effect of the combined nanoparticles on S.aureus was further evaluated by standard plate counting method,dynamic antibacterial test and transmission electron microscope(TEM)for bacterial morphology observation.The results showed that the combined nanoparticles could destruct the S.aureus bacterial walls.Compared with single targeted nanoparticles,bacteria inhibition rate significantly increased corresponding with the increase of concentration.Crystal violet staining tests confirmed that combined nanoparticles could inhibit about 64.37% formation of S.aureus biological membrane.The results of fluorescence quantitative PCR determination confirmed that the combined nanoparticles could significantly down-regulate the expression level of hrt A,hrt B,rpm G3,deo D,sec E,sar A,agr A,hla,cid A,ica A genes(P<0.05).The above results showed that GO/Apt/BBR and GO/Apt/AE combined nanoparticles(5:1)possessed good targeted delivery property,significantly down-regulated the expression level of various genes related to bacterial nutrition metabolism,biofilm formation and toxin,and inhibited the growth and reproduction of S.aureus.3 Study on the safety evaluation and targeted antibacterial activity of GO/Apt/BBR and GO/Apt/AE combined nanoparticles in vivoIn this chapter,the safe concentration of the combined nanoparticles was evaluated by the toxicity test of bovine mammary epithelial cells,erythrocyte hemolysis test,zebra fish toxicity test experiment and mouse mammary perfusion toxicity test.The results showed that the combined nanoparticles had no obviously negative effects in the range of 0-200 μg/m L.Then,S.aureus induced mouse mastitis model was constructed to evaluate the targeted antibacterial activity of the combined nanoparticles against S.aureus in vivo.The results of three-dimensional imaging experiments in small animals showed that combined nanoparticles had better targeting performance for S.aureus,significantly reduced the number of S.aureus in the mammary gland,tissue congestion and neutrophil infiltration.The above results showed that 200 μg/m L of combined nanoparticles had good biosafety and S.aureus targeting,effectively inhibited the number of S.aureus in breast tissue in mice,reduced the damage and inflammatory symptoms caused by the bacteria.Two GO-based targeted nanoparticles of GO/Apt/BBR and GO/Apt/AE were successfully prepared.Compared to either targeted nanoparticles,the combined utilization of both nanopariticles with the ratio of 5:1 showed stronger antibacterial performance.Combined nanoparticles can fulfill the target of S.aureus perfused into mouse breast,and effectively alleviate tissue congestion and neutrophil infiltration,which provided new ideas for the application of active traditional Chinese medicine components(such as BBR and AE)in S.aureus infection-related diseases.