Selection Of High Yield Pediocin Strains Of Pediococcus Acidilatici By Chemical Mutagenesis

Bacteriocins produced by lactic acid bacterias are metabolited products of food-grade microorganisms, and nowadays some of them are used for food preservation. Pediocin is the most likely to put into production since nisin. However, it's negative to study about pediocin because the bacterias producing pediocin are wild stains which have low yields that can't meet the requirement of industrial production. The strain Pediococcus acidilactici PA003 was isolated from fermented vegetable food in the laboratory. A high yield pediocin strain was expected to be obtained during the compand chemical mutagenesis, and it might be very useful for laboratory researches and industrial productions.Three methods were established to survey the titer of pediocin fast and accuratedly. One was the quality weighing method used to determine the production of pediocin in scale production processes. The second was the agar diffusion method which had accurate results, and it used for the selection of mutants. The third was the spectrophotometry which had a short test period, and it used to find the optimal production conditions of the high yield pediocin strain.DES and LiCl were used as compand chemical mutagents to the strain Pediococcus acidilactici PA003. Optimum mutagenic conditions of DES were determined by orthogonal experiments and the results were: the bacterial concentration 107cfu/mL, the concentration of DES 0.8%, the mutagenic time 30min and the mutagenic temperature 37℃. By DES mutagenesis, 20 mutants of high yield of pediocin were obtained. Of the 20 mutants, 5 with higher pediocin production were selected for mutagenic test by LiCl. Afer LiCl mutagenesis, additional 5 mutants were chosen. One of the mutant strains P2 was 24.15 times of the stain Pediococcus acidilactici PA003 in the same culture conditions.Related elements on the pediocin production of the mutant strain P2 were explored. The single-factor experiments demonstrated that medium beginning pH, cultivate temperature, cultivate time and inoculum were the most influential factors. A 4-factor, 3-level Box-Behnken experimental design was applied by Design Expert 7.1.6, and the interactions of each factor on the titer of pediocin were also investigated. The results showed that the optimal producing conditons of the mutant strain P2 were: medium beginning pH 6.5, inoculum 0.5%, cultivate temperature 37℃and cultivate time 17.6h. Under these conditions, the titer of the mutant stain P2 was 1.12 times of that of the srain in common conditions.The pediocin production seemed to be stable during the continuous 20 slope passages'experiments of the mutant strain P2. The production declined about 2.3% in generation 20.The pediocin produced by Pediococcus acidilactici was safe and efficient, and its antibacterial activity had a favorable potential application in food preservation. The high yield pediocin strain P2 obtained in this work was expected to be used for industrial production, and it might benefit the economy and the society.