Study On Bacteriostatic Action Of Artificially Produced Antibiotic Peptides In Vitro

With the abuse of antibiotics, resistant strains have evolved,as they will almost inevitably do given sufficient time of exposure. A viable solution to this acquired resistance problemmay reside in new antibiotics,dissimilar to those that have lost their effectiveness. Antimicrobial peptide,one of the most important constituent components of host denfence system,is a kind of small peptides produced by inducing in organism.They are produced by all organisms,from plants and insects to human beings,as a major part of their immediately effective,nonspecific defences against infections.Although many demonstrate direct antimicrobial activity against bacteria,fungi,eukaryotic parasites and/or viruses,it has been established that antimicrobial peptides have a key modulatory role in the interface of innate and adaptive immunity. Anti-bacterial peptide is a kind of short peptide which characterized by high antimicrobial activities,wide antimicrobial sectrum and no side effects on animals.As an ideal substitute for antibioties,it has aroused people's more interests.Hybrid antimicrobial peptides are a new kind of defence active peptides,which charaeterized by higher antimicrobial activities and shorter peptide chain,composed by peptide chain of two or more anti-bacterial peptides.The design of anti-bacterial peptide is a hotspot which designed by acid replacement or sequence or combination with other peptides.The new designed anti-bacterial peptides are more excellent than the original. Exploring and designing new hybridantimierobial peptides in order to improve the activity and reduce the hemolytic effeets is very important and worthy,as the high level expression in pratical application.Objective: According with the relationship between the structure and function of APMs,we use peptide V13K as a template,choose amino acid sequence that is essential to its antibacterial activity, analysis this sequence and replace the amino acids in the place of 24.Then,we can get two types of new artificial hybrid antimicrobial peptides,and detect their hydrophobicity and antibacterial effects. Through the transformation of pre-and post changes in MIC and hydrophobicity,we can evaluat the modified peptide and provide a theoretical basis for peptide modifing. Methods: Spectrophotometric method is used to detect bacteria in the correlation between concentration and OD value,and thus to quantify bacteria more accurate; we use the method of chemical synthesis of the amino acid to synthesize the designed amino acid sequence.we use broth microdilution method to determine the minimum inhibitory concentration of the peptides.Results:We determined the concentrationship between OD value and the concentration of 12 bacterias respectively,and drawed the standard curve. We successfully synthesized pre-designed peptides,which names are: L-V13K,D-V13K,L-I24L,D-I24L,L-I24V,D-I24V. After purifing the peptides,we get 4mg for each sample for the MIC experiment. We determined peak time for V13K, I24L, I24V at 25℃, respectively, are 37.968min, 38.609min, 36.428min.We use broth microdilution method and get MIC values of the six peptides on 12 bacterial.At last we compared the MHB and LB medium on the antibacterial peptide effects.Conclusion: Both V-13K and the peptide with replacement of amino acid have inhibitory effect on the gram-negative and positive bacterias, effect on the gram-negative bacteria is more strong.L-V13K and the peptides with replacement of amino acid have more effects on bacteria than the D-type peptides. At a certain range, the V-13K and the peptide with replacement of amino acid will have better antibacterial ability with an increase in hydrophobicity.