The Effects Of Luminescent CdSe Quantum Dot-Functionalized Antimicrobial Peptides Nanoparticles On Antibacterial Activity And Molecular Mechanism

The problem of bacterial resistance to antibiotics has seriously threatened human health in the world.Due to the irregular use of antibiotics,the threat of bacterial resistance is worsening.Therefore,the development of new antimicrobial drugs is imminent.Antimicrobial peptides are highly effective and broad-spectrum antimicrobial agents.They are used for pathogens through different action mechanism and have potential antibacterial activity.Therefore,antimicrobial peptides are expected to become a new type of drug for the treatment of drug-resistant bacteria.Through experiments,this subject was artificially synthesized according to the amino acid sequence of the polypeptide Perplanetasin-4（Leu-His-LysVal-Tyr-Cys-Val-Leu-Gly-Pro-NH₂）extracted from the American cockroach.Based on the unique optical properties of quantum dots,through experiments,the exterior of the quantum dots is modified with antimicrobial peptides,which greatly improves its stability,and it is found that the antibacterial activity of antimicrobial peptides is also improved.Therefore,the combination of antimicrobial peptides and quantum dots can be improved the effectiveness of the two groups compounds.This subject mainly carried out the following research:（1）By using 2-Cl resin as the amino acids binding carrier,the crude peptide was synthesized by Fmoc solid phase synthesis method,and the crude peptide was analyzed by mass spectrometry to confirm that the crude peptide contained the target peptide,and then separated by high performance liquid chromatography.Purify to obtain the obtain the target product with a purity of more than 90%.The obtained target product was subjected to infrared spectroscopy and mass spectrometry to confirm than the separated peptide was the antimicrobial peptide periplanetasin-4,which provided a material basis for the next experiments.（2）Under the protection of nitrogen,the prepare Se precursor solution was poured into a three-necked flask containing the prepared Cd precursor solution to obtain Cd Se NPs after reaction.Then prepare AP-Cd Se NPs.The antimicrobial peptides,CdSe and AP-CdSe NPs are characterized by ultraviolet spectrophotometer and fluorescence spectrophotometer.By analyzing the ultraviolet spectrum,infrared spectrum and fluorescence spectrum,it is known that the antimicrobial peptide has been successfully modified on the CdSe quantum dots and has fluorescence characteristics.Scanning electron microscopy revealed that AP-CdSe NPs are uniformly distributed,the particle size of CdSe is about 9 nm,and the particle size of AP-CdSe NPs is about 14 nm.（3）In the vivo and vitro bacteriostatic experiments of AP-CdSe NPs,Staphylococcus aureus,Escherichia coli,and MDR Escherichia coli and Staphylococcus aureus were used to for antibacterial experiments verification.After the antimicrobial peptides were successfully modified on the surface of CdSe quantum dots,in vitro tests were performed on the individual antimicrobial peptides,CdSe NPs and AP-CdSe NPs.AP-CdSe NPs had a low MIC value against common bacteria in vitro experiment.For example,AP-Cd Se NPs still showed strong antibacterial activity against E.coli and S.aureus at the concentration of 8.0μg/mL.Through the CFU test,it is found the survival rates of MDR E.coli and S.aureus were 10.3%and 3.8%respectively when the AP-Cd Se NPs concentration was 40μg/mL.These data indicate that AP-CdSe NPs have higher antibacterial properties compared with individual AP and Cd Se nanoparticles.The mechanism of cells integrity is studied by flow cytometry.The enrichment analysis of GO and KEGG showed that AP-Cd Se NPs affected the growth of E.coli due to its influence on bacterial metabolic pathways.These all confirmed that AP-Cd Se NPs has high antibacterial properties in vitro.In vivo experiments,through the establishment of a bacteremia model of S.aureus infection in mice,it is further concluded that AP-Cd Se NPs also has a higher antibacterial activity in vivo,and AP-Cd Se NPs is also proved to have no side effects on mice through safety evaluation.This subject experiment has laid a certain experimental foundation for further research on antimicrobial peptide drug preparation in the future.