Production Of 1,3-Propanediol By Encapsulated Klebsiella Pneumoniae In The NaCS/PDMDAAC Microcapsule

Microbial production of 1,3-propanediol (1,3-PD) has been among the most remarkable research focuses in the realm of biology. To extend the application of immobilization technique in this bioconversion, the sodium cellulose sulfate / poly-dimethyl-diallyl-ammonium chloride microcapsule encapsulated cells of Klebsiella pneumoniae for dissimilating glycerol to 1,3-PD. By virtue of advantages of encapsulated cells, this immobilization fermentation aimed at enhancing 1,3-PD concentration, yield and productivity, achieving cell reutilization and continuous fermentation, as a result, decreasing the cost of microbial method from materials and operation.Firstly, the free cell culture was carried out with a newly isolated K. pneumoniae strain and effects of pH, oxygen, glucose on the 1,3-PD formation were investigated and the fermentation information about the new strain was obtained. Under the condition of pH7.0 and no aeration, glycerol was consumed in 14 hours, obtaining a maximum biomass of 3.20g/L. The 1,3-PD final concentration was 16.35g/L with a yield of 0.50mol/mol and a productivity of 1.17g/L/h. Glucose as a co-substrate was priorly consumed to enhance the biomass to 4.60g/L.Secondly, a serious of shake flask cultures with encapsulated K. pneumoniae were performed under different preparation and cultivation conditions. It was determined that the optimal dilution multiplier of cell suspension for encapsulation, volume ratio between microcapsules and medium, initial pH and glycerol concentration of medium were 5, 1:2, 7.0, 40~60g/L, respectively. Then the cell growth, glycerol consumption and 1,3-PD production of encapsulated cells were investigated. Compared with free cell culture in the same condition, biomass of encapsulated cell culture was 2.6-fold higher than that of free cell culture;glycerol consumption period was decrease from 47 hours in free cell culture to 12 hours in encapsulated cell culture;1,3-PD final concentration reached 12.8g/L with a yield of 0.63mol/mol in encapsulated cell culture, 17g/L and 0.57mol/mol in free cell culture;also in encapsulated cell culture, glycerol concentration inside microcapsule remained 0.1~0.3g/L while 1,3-PD concentration accumulated to 10.6g/L and acetic acid was the main byproduct.Thirdly, the encapsulation fermentation was amplified to fixed-bed bioreactor. The fermentation mode included batch, fed-batch and continuous culture. Batch culture further reduced the glycerol consumption period to 5 hours and obtained a 1,3-PD concentration of 63.1g/L with a yield of 0.65mol/mol when initial glycerol concentration was 120g/L;fed-batch culture lasted 67 hours and its maximum 1,3-PD concentration was 51.86g/L with a yield of 0.39mol/mol and a productivity of 1.08g/L/h;continuous culture was carried out at three different dilution rates and reached the optimal result under 0.33h"], the productivity of which was prompted to 4.49g/L/h with a 1,3-PD concentration of 13.6g/L and a yield of 0.43mol/mol in the steady state.Fourthly, a two-stage fermentation process from glucose to 1,3-PD was designed and performed, in which glycerol formation in the air-lift bioreactor with encapsulated Candida krusei was combined with 1,3-PD formation in the fixed-bed bioreactor with encapsulated K. pneumoniae. The first stage obtained a glycerol concentration of ca. 52g/L and a yield of 0.59molGiyceroi/niolGiucose;the second stage achieved a maximum 1,3-PD concentration of 7.55g/L with a yield of 0.50molpD/molGiyCeroi and a total yield of 0.3 5molpi)/molGiucose-Finally, the mixed culture with free cells of C. krusei and K. pneumoniae was studied in a fermentation medium containing lOOg/L glucose and 80g/L glycerol. A 1,3-PD concentration of 21.11 g/L was obtained in the mixed culture while only 9.17g/L 1,3-PD was obtained only by K. pneumoniae under the same medium and cultivation conditions.