Optimization Of Fermentation Factors In ε-Poly-L-Lysine Production By Streptomyces Albulus GS114

ε-poly-L-lysine(ε-PL) is an L-lysine homopolymer, which is synthesised with 25-35 L-lysine by nonribosomal peptide synthetases(NRPSs). Because of its wide spectrum of microorganism, thermal stability, wide used range of pH and safety, it is mainly used as a food preservative. In our preliminary study, we obtained a high ε-PL producing strain, named Streptomyces albulus GS114. The yield of ε-PL in the shaking flask with S. albulus GS114 is 1.7-fold greater than that of S. albulus M-Z18. In order to excavate the potential of S. albulus GS114, we optimized its fermentation medium and pH control strategy to improve the domestic ferment level. The main results as follows:(1) We optimized the fermentation medium in shaking flask and 5 L fermentor by single factor experiment and Plackett-Burman degsin. Based on the medium named M3 G, we investigated the effect of nutrition component on ε-PL synthesis by single factor experiment. Then we screened out the remarkable factors by Plackett-Burman design, which contained glucose, yeast powder and K2HPO4. After that, we optimized the concentration of three remarkable factors in 5 L fermentor. The composition of optimized fermentation medium was established as follows: 90 g·L-1 glucose, 10 g·L-1(NH4)2SO4, 10 g·L-1 yeast powder, 4 g·L-1K2HPO4, 0.50 g·L-1, MgSO4·7H2O, 0.04 g·L-1 ZnSO4·7H2O, 0.03 g·L-1 FeSO4·7H2O. With the optimized fermentation medium, ε-PL production and DCW achieved 37 g·L-1 and 32.6g·L-1 respectively, which increased by 43.9% and 32.5% compared with the control.(2) We selected glucose as the fermentation carbon source by comparing the fermentation parameters, the degree of polymerization of ε-PL and the antibacterial activity in different carbon sources with S. albulus GS114. The concentration of ε-PL was 43.4 g·L-1 and45.7 g·L-1 when carbon source was glucose and glycerol respectively in fed-batch fermentation, while the total mass of ε-PL was 172 g with glucose as carbon source, which was 25.5% higher than that of glycerol. Furthermore, The degree of polymerization of ε-PL with glucose as carbon source ranged from 20 to 34, and the peak of degree of polymerization was 30, which was more centralized and higher than that of glycerol. Moreover, the antibacterial activity of ε-PL with glucose as carbon source was also higher than that of glycerol.(3) The pH control strategy was established with glucose as carbon source by optimizing pH in the process of fermentation. Based on the effect of pH on cell growth and ε-PL synthesis, we proposed a two-stage pH(3.8-4.0) control strategy. With this strategy, the concentration of ε-PL and DCW reached 38.0 g·L-1 and 46.0 g·L-1 respectively. To increase the ferment level of ε-PL further, we investigated the strategy of pH(3.5-4.0) and pH(3.3-4.0). The result illustrated that pH(3.3-4.0) control strategy was the best choice. With this strategy, the concentration of ε-PL and DCW reached 43.0 g·L-1 and 35.0 g·L-1. Finally, we optimized the inhibition pH, inhibition time and recover pH based on the two-stage pH(3.3-4.0) control strategy. The result showed that when the inhibition pH, inhibition time and the recover p H were 3.3, 4 hour and 4.0 respectively, we can achieve the highest ferment level. With the optimal condition, the concentration of ε-PL and DCW were 47.3 g·L-1 and 29.5 g·L-1.