Screening Of ε-polylysine-producing Strain And Studies On The Fermentation Process

ε-poly-L-lysine (ε-PL), which usually consists of 25-30 residues of an essential amino acid L-lysine, is a homo-poly-amino acid characterized by the peptide bond between the carboxyl and ï¿¡-amino groups of L-lysine, and its molecular weight is approximately 3500 to 4500. ï¿¡-PL shows a wide range of antimicrobial activity against most gram-positive and gram-negative bacteria, fungi and also some viruses, and is stable and safe. Due to its antimicrobial activity, e-PL is widely used as a food preservative. Hitherto, its industrialized production of e-PL is only carried out in Japan.A simple and sensitive screening method to screen e-PL-producing strains from soil samples was developed in the text. 150mg/L K₂Cr₂O₇ was added to the SG medium to enrich actinomycetes. A basic dye, Methylene Blue, which could react with the secreted basic polymers by electrostatic interaction and form special zoon, was incorporated in the agar plate to detect alkali producers, and 219 alkali producers were isolated, and 46 alkaloid producers were screened by Dragendorff regent reaction then. And PL6-3, one of the four ε-PL producers which were obtained by TLC analysis, showed the highest productivity, and its fermentation capability was also very stable.Phylogenetic analysis of PL6-3 strain, including morphology, physiological and biochemical characters and chemotaxomy were performed and phylogenetic tree was constructed based on the 16S rDNA sequences. And the results indicated that PL6-3 strain is a member of Kitasatospora, so it was named Kitasatospora sp. PL6-3.The culture medium was optimized, and the optimal culture for the reciprocal flask culture is 50 glucose, 10 (NH₄)₂SO₄, 1.5 Na₂HPO₄, 1.5 KH₂PO₄, 0.5 MgSO₄·7H₂O, 0.03 ZnSO₄·7H₂O, 0.01 FeSO₄·7H₂O, 5 yeast extract per liter. Original pH is pH6.8, capacity is 80mL/500mL, 28℃, 72h.The effects of different stirred speeds on Kitasatospora sp. PL6-3 fermentation were inspected, and the optimum stirred speed is 350r/min. The effects of differentinitial glucose concentration on e-PL production were inspected under pH controlling pH4.0, too. When the initial glucose concentration was 3% with glucose feeding, 6.5g/L biomass was obtained, and ï¿¡-PL production was 1.59g/L, both were higher than that when the initial glucose concentration was 5%. And the biomass and the yield of e-PL were improved 10% and 65%, respectively, under the same consumption of glucose. Under the condition of 350r/min, controlling pH4.0 and the initial glucose concentration was 3% with glucose feeding, ï¿¡-PL productivity was as high as 6.65g/L, which was 10-fold higher than that of batch culture.