| Plasma treatment is proved to be safe and high efficiency for its transient existence and immediate disappearance after dealing with samples. It is widely used in the field of breeding and surface sterilization owing to the special merit. Most researches concentrated on sterilization mechanism, but the biological effect of plasma under different operate conditions is still unknown. Meanwhile, when cells were treated by plasma, the permeability of cell membrane would be changed, so what will the effect change when ions added is also very interesting.1,3-Propanediol (1,3-PD) is an important chemical, due to the presence of two hydroxyl groups at1and3positions, it is wildly used in the production of polymers. In microbial production of1,3-PD, pure glycerol is used as substrate, but the high cost of glycerol limits the commercialization of microbial method. It is necessary to replace it by crude and cheap glycerol. However, strains which can tolerate rude glycerol are very few. In order to solve problems above, this paper focuses on the below topices:Firstly, Response Surface Methodology was employed to evaluate the impact of plasma on1,3-PD production by Klebsiella pneumoniae. Results showed that the production was increased under the conditions:voltage24～26kV, electric current1.7～1.8mA, treating time120～160s and voltage30～32kV, electric current1.8-1.9mA, treating time20～60s, where K. pneumoniae was preserved more than3months in refrigeratory at-70℃. The effect of liquid volumes was furthur tested, and it showed no difference to the1,3-PD production. With the results received, the optimization experiment was conducted, under the optimum conditions as follows:voltage24kV, electric current1.8mA, treating time140s, resulting in enhanced production of1,3-PD by21.7%. Meanwhile, the intracellular concentration glutathione (GSH) and total antioxidant capability (T-AOC) were tested, and they both increased after the inducement of plasma under the optimisum conditions.Secondly, the complex treatment of plasma and ions was investigated by the experiment. The results showed that the function of Mg2+, Li+, K+except for Ba2+with plasma was benefit to K. pneumoniae for1,3-PD production. The seed handled by plasma and Mg2+was further used to produce71.5g/L of1,3-PD at23h in fed-batch fermentation, increased by18.8%comparing to the control, and the yield was also increased slightly.Finally, based on the complex treatment of plasma and LiCl, a mutant strain of K. pneumoniae was isolated by using the medium containing FeCl3and selective pressure. The mutant strain named KpM30improved1,3-PD production from biodiesel-derived glycerol through producing less organic acid than the wild strain. The fed-batch fermentation in a5L bioreactor showed that the concentration of1,3-PD reached70.2g/L, the yield of1,3-PD on glycerol was0.576(mol/mol), which were increased by24.9%and17.7%respectively compared with the wild strain. Meanwhile, the production of lactic acid was decreased by47.0%. According to the metabolic analysis, the production of ATP and NADH by KpM30were1.6-fold and1.3-fold of the wild strain at the beginning of the fermentation. At the end of the fermentation, the carbon flux flowed from glycerol to1,3-PD was increased by22.6%. On the other hand, KpM30was also proved a better utilization of pure glycerol than wild strain. |
PDF Full Text Request