Study On Producing Antimicrobial Peptides By Fermenting Soybean Meal

As an ideal alternative to antibiotics, antimicrobial peptides have the broad-spectrum antibacterial activity against Bacteria, Fungi, Protozoa, Tumor cell and Virus. In order to study on producing antimicrobial peptides by fermenting soybean meal, the crude peptides which had the antibacterial activity was extracted, the antimicrobial peptides were purified and its molecular weight was determined by mass spectrometry. The main research contents and results were as follows:(1) The result of antibacterial activity experiment showed that the Bacillus KN-BL-1 and KN-BS could product the antimicrobial substances in LB medium, liquid-state and solid-state fermentation of soybeanmeal. And the fermentation liquor of them were effectively inhibit on S.aureus, Micrococcus tetragenus, Microbacterium sp.(B009), Brevibacterium sp.(B021) and other Gram positive bacteria, the max diameter of antibiotic circle was 22.5mm. And the Bacillus KN-BL-1 and KN-BS had been determined that they were Bacillus amyloliquefaciens and Bacillus methylotrophicus by strain identification.(2) The crude peptides were extracted by acid precipitation method, and had made sure that the antibacterial activity components of the fermentation liquor of KN-BL-1 and KN-BS were polypeptides. The crude peptides could effectively inhibit on S.aureus, the max diameter of antibiotic circle was 18.6mm. The correlation coefficient of the antibacterial circle diameter and the peptide contents was more than 0.99, it means that the higher of the peptide contents, the better of the antibacterial activity.(3) The crude peptides which extracted from the fermentation liquor of KN-BL-1 and KN-BS were separated by DEAE-Sepharose ion exchange chromatography. Some of the peak samples had inhibitory effect on S.aureus, the max diameter of antibiotic circle was 23.8mm.(4) The samples which had antibacterial activity separated by DEAE-Sepharose ion exchange chromatography were purified again by Sephadex G-25 gel filtration chromatography. There were five antimicrobial peptides samples had been collected: LB 2-③, SM 2-⑤, SM 0.2-1-②, BS-LM 0.2-3-② and BS-SM 2-④. All of them could effectively inhibit on S.aureus, the antibacterial circle diameter of SM 2-⑤ and BS-SM 2-④ were both 19.8mm. The molecular weight of SM 2-⑤ was between 2000Da- 8000 Da, which was determined by mass spectrometry.