Screening Of Antibacterial Peptides Against Pet Skin Pathogens And The Preparation Of Nanoemulsion

In recent years,the number of pets kept growing and pet diseases have followed.In particular,Canine cat bacteria,fungal and mixed skin diseases account for a large proportion of veterinary clinical diseases.Therefore,a large number of clinically resistant strains resistant to multiple antibiotics.Antimicrobial peptides have become an alternative to antibiotics in recent years.They have an efficient,broad-spectrum activity against bacteria,fungi,viruses and some protozoa,and have short amino acid sequences and simple synthesis.Clinical use of drugs to optimize the role of antimicrobial peptides,increase stability and safety.We screened short sequence antimicrobial peptides for use in nano-drug delivery systems to increase their transdermal absorbability,and studied their stability,safety,and in vitro antibacterial properties.In this experiment,Detection of antibacterial activity of various antimicrobial peptides synthesized by chemical solid-phase synthesis by disk diffusion method drug susceptibility test.Taking Escherichia coli and Staphylococcus aureus as representative strains,the antibacterial peptides RR-9 and RI-12 were preliminarily screened to have ideal antibacterial activity.Further verification by(MIC)detection,bactericidal concentration detection,bactericidal curve determination,scanning electron microscopy,and transmission electron microscopy.The results showed that both RR-9 and RI-12 antibacterial peptides could inhibit the growth of standard strains of common pathogenic bacteria in pet skin at the same time.The MIC values for S.aureus ATCC 29213 were 16 ?g/mL and 6.6 ?g/m L,respectively.The MIC value of RR-9 against clinically resistant S.aureus was 64 ?g/mL.When the concentration of RR-9 reaches 4 times the MIC,S.aureus can be completely killed within 20 minutes.Electron microscopic results show that the antibacterial peptide RR-9 can quickly destroy the complete structure of bacterial cell membrane,by disrupting the cell membrane and allowing the outflow of the content material,bacterial growth is inhibited and the purpose of sterilization is achieved.This experiment adopted the reported formula: isopropyl myristate(IPM)as the oil phase,caprylic acid phthalic acid polyethylene glycol glyceride(EP)as the surfactant,polyglyceryl fatty acid esters as cosurfactants.On this basis,the oleic acid and azone is selected to the best ratio increases nanoemulsion stability and skin permeability.The antibacterial peptide nanoemulsion containing RR-9 was prepared by this formula,and its quality was evaluated.The determination of antimicrobial peptides in nanoemulsion was established by UV spectrophotometry.The experimental results show that the optimal ratio of oleic acid and azone is 1% and 2%.The particle size analyzer detects that the average particle size of nano-emulsion is95.3 nm,and the size is uniform and evenly distributed;The established content detection method is simple and accurate.In this study,rat and rabbit skin irritation test,acute toxicity test and HaCat cytotoxicity test were used to evaluate the safety of nano-emulsion containing RR-9,and the in vitro efficacy of antimicrobial peptide nanoemulsion was evaluated.The results showed that the antibacterial peptide nanoemulsion formulation did not show acute toxicity,irritation,or allergic reaction after external application to the skin.The cell proliferation RGR was 61.54% and the toxicity was grade 2.There was no cytotoxicity.In vitro efficacy experiments showed that the MIC value of the antibacterial peptide nanoemulsion against Staphylococcus aureus ATCC29213 and Candida albicans ATCC10231 was 3.9 ?g/m L,and the MIC value of resistant Staphylococcus aureus resistant to antibiotics was 7.8 ?g./mL.It can be seen that the antibacterial activity of nanometer milk containing RR-9 is superior to the antibacterial peptide water solution.