Study On Fermentation Process Of1,3-propanediol By Klebsiella Pneumoniae

As an important chemical raw material,1,3-propanediol (1,3-PD) has been widelyused in the field of printing and dyeing, paint, ink, drugs, antifreeze, lubricants.Moreover, it has been attracted much attention as a monomer in constituting PTT(polyethylene terephthalate). There are two methods in produce1,3-PD-chemicalsynthesis and microbial fermentation. Nowadays, due to the shortage of petrochemicalresources, microbial fermentation becomes a hot research at home and abroad. Fromthe perspective of industrial production, the optimum fermentation conditions of batchand fed-batch fermentation process in5L fermentor and the feasibility of immobilizedfermentation of1,3-PD by klebsiella pneumoniae in shake flasks were studied andoptimized and then the1,3-PD separation and purification from the fermentation brothwas investigated, this study provided guidance for1,3-PD industrial production at last.Firstly, the optimal conditions of klebsiella pneumonia fermentation conditionswere studied. And then, the optimum conditions by fermenter single factor analysis andorthogonal experimentwere obtained: initial glycerol concentration50g/L,(NH4)2SO4concentration6g/L, temperature37°C, pH7.0, inoculation volume10%. Under suchcondition, the cell growth rate and1,3-propanediol formation rate rise fast and themaximum concentration of the1,3-PD was24.79g/L, the1,3-propanediol productivitywas2.479g/(L h) and the conversion rate was54%(g/g).Secondly, on the basis of batch fermentation optimization conditions, the differentfeeding start time, different way of fed-batch fermentation(constant rate feeding,intermittent steady feeding, glycerol feedback feeding) in klebsiella pneumonia2-1fed-batch fermentation process were investigated. After many experiments, theoptimum conditions were obtained: in the mid-logarithmic growth with the way ofglycerol feedback flow plus, maintained glycerol concentration at about20g/L,24hlater, the absorption of light value in650nm was10.38, the concentration of1,3-PD is80.45g/L, the1,3-propanediol productivity was3.35g/(L h) and the quality of theconversion rate of glycerol to1,3-PD was62.8%, which significantly better than otherways of feeding fermentation results.Thirdly, we have conducted the feasibility study of immobilized fermentation of1,3-PD by klebsiella pneumoniae. The conclusion revealed that calcium alginate showsgreat advantages in the microcapsules preparation and1,3-propanediol productivity compared with other four kinds of common embedding materials. The optimumconditions of calcium alginate encapsulation were that: sodium alginate concentration6%(w/v), calcium chloride4%(w/v), crosslinking time4h. The1,3-propanediolconcentration in the broth increased by7.4%compared with free fermentation, andproductivity reached to1.04g/(L h). In addition, the immobilized particles mechanicalstrength and fermentation level is not significantly reduced by6times fermentation.Finally, the removal process of bacteria, protein and salt in1,3-PD fermentationbroth were preliminary explorationed. The experiments show that chitosan is a goodkind of organic macromolecular flocculant. The best flocculation conditions were: finalconcentration of chitosan0.6g/L,37°C, pH5.0, both of stirring and standing time20min. Under this condition, the removal rate can reach to99.87%. Furthermore, theprotein remove rate reached to84.7%by added calcium chloride and boiled thefermentation broth. Owing to the alcohol precipitation-ultrafiltration desalination, thesalt removal rate can reached to79.2%, and obtained1,3-PD crude extracts with apurity of85.68%.