Development Of Composite Antibacterial Agents Based On Antimicrobial Peptides And Their Application In Milk

Harmful bacteria in food is the main cause of food spoilage,deterioration and a threat to consumer safety,which has been receiving increasing attention.Traditional food preservatives have drawbacks such as poor sterilization effectiveness,induce of resistance,and overuse.Therefore,it is of great significance to research and develop new,safe and efficient antibacterial agents for harmful bacteria in food,in order to improve food safety.The main research content and results of this article are as follows:1.Design of novel antibacterial short peptides and study on their antibacterial effects.Based on the antibacterial peptide killing principle and existing antimicrobial peptide databases,three new antimicrobial peptides were designed by sequence alignment(with amino acid sequences "FRLHF","FRWWR",and "FRLKFHF").Circular dichroism(CD)was used to measure the peptide structure,and the results shows that peptides "FRLHF" and "FRLKFHF" contained both α-helical and β-folded structures,while peptide "FRWWR" only had a β-folded structure.Minimal inhibitory concentration(MIC)experiments showed that peptide "FRWWR" had the best antibacterial effect against Listeria monocytogenes,Staphylococcus aureus,Escherichia coli,and Bacillus subtilis,with MIC values of 1 mg/m L,8 mg/m L,4 mg/m L,and 0.25 mg/m L,respectively.Laser confocal microscopy was used to observe the effect of peptide "FRWWR" on bacterial biofilms,and the results showed that the biofilm area of L.monocytogenes,S.aureus,E.coli,and B.subtilis was reduced by 84%,94.04%,89.87%,and 83.46%,respectively,after treatment with peptide "FRWWR".2.Research on antibacterial effect of Nisin.In order to investigate the antibacterial effect of Nisin,comprehensive evaluations were carried out through experiments including minimal inhibitory concentration,minimal bactericidal concentrations,minimum biofilm inhibitory concentration,and minimum biofilm eliminate concentration.The results shows that Nisin had different degrees of antibacterial effect on four strains,and its inhibition effect on L.monocytogenes and B.subtilis was significant.Furthermore,Nisin was able to effectively reduce the formation and growth of bacterial biofilms.3.The research on the synergistic antibacterial effect of the short peptide "FRWWR" and Nisin.The combined use of antibacterial agents can enhance the antibacterial effect and achieve broad-spectrum antibacterial activity.The synergistic effect of different concentrations of the short peptide "FRWWR" and Nisin on L.monocytogenes,S.aureus,E.coli and B.subtilis was studied.The results shows that the composite antibacterial peptide had a synergistic effect on the inhibition of L.monocytogenes and B.subtilis,an additive effect on the inhibition of S.aureus and E.coli,and a synergistic effect on the killing of L.monocytogenes,and an additive effect on the killing of S.aureus,E.coli,and B.subtilis.The composite antibacterial peptide had a synergistic effect on inhibiting the formation of biofilms by L.monocytogenes and S.aureus,an additive effect on inhibiting the formation of biofilms by E.coli and B.subtilis,and a synergistic effect on killing the biofilms formed by L.monocytogenes,and an additive effect on killing the biofilms formed by S.aureus,E.coli,and B.subtilis.The study results shows that the composite antibacterial peptide can enhance the antibacterial and bactericidal effects,and can kill more types of bacteria at lower concentrations.The combined use of the composite peptide has a better effect on the formation and elimination of biofilms.4.The application of composite antimicrobial peptides in milk.The application of composite antimicrobial peptides in milk has been shown to effectively kill harmful bacteria in milk,and compared with the control group,it can consistently maintain a sterile state.Toxicology experiments have been conducted to test the safety of composite antimicrobial peptides,and the results shows that the hemolysis rate of Nisin with a concentration of 1.2 mg/m L was the lowest,with a hemolysis rate of only 0.8% at 180 min.The hemolysis rate of the 4 mg/m L short peptide "FRWWR" reached the highest at60 min,at 2.06%.The hemolysis rate of the composite antimicrobial peptides formed by the short peptide "FRWWR" and Nisin reached the highest at 45 min,at 2.55%.The results of the study shows that whether the short peptide "FRWWR" and Nisin are used alone or in combination,the hemolysis rate is below 5%,which meets safety requirements.The application of composite antimicrobial peptides in milk has the advantages of low toxicity,good sterilization effect,maintaining milk quality,and extending the shelf life.In summary,harmful bacteria can reduce food quality and harm human health.The application of traditional preservatives in food has defects.New composite antimicrobial peptides have excellent sterilization effects,do not produce resistance,and have broad-spectrum antibacterial activity.They have a wide range of applications in the field of food preservation.