A synthetic medium for ethanol production by recombinant xylose-fermenting Zymomonas mobilis was optimized using Plackett-Burman (PB) design and response surface methodology (RSM). The effects of 19 medium components were investigated by PB design. Eight of these components were determined to have significant effects on ethanol production. The statistical model was constructed via central composite design (CCD) using five selected variables, including xylose, trisodium citrate, choline chloride, pyridoxine, and thiamine. The validity of the developed model was verified. The ethanol concentration in the optimized medium was 6.7% higher than in the RM medium. The optimized medium was then simplified to give the final synthetic medium (S2), in which the ethanol concentration was 20.7% higher than in the RM medium. Xylose and trisodium citrate were the key medium components influencing ethanol production, while calcium pantothenate was the only growth factor required by recombinant Z. mobilis. Ethanol production and growth response were directly proportional to the logarithm of the concentration of trisodium citrate in the range of 0.1-1.6 g/L.recombinant Zymomonas mobilis, synthetic medium, ethanol production, response surface methodology, trisodium citrate, xylose Citation:Dong H N, Zhao X M, Ma Y Y, et al. Optimization of a synthetic medium for ethanol production by xylose-fermenting Zymomonas mobilis using response surface methodology.

Three xylose-fermenting recombinant Zymomonas mobilis strains containing different Peno-talB/tktA operon terminators were engineered. Each showed similar levels of foreign protein expression and xylose fermentation performance. Strain CP4-P2-1 was further used to compare the glucose/xylose co-fermentation under various cultivation environments to improve the efficiency of the process. Optimal co-fermentation was achieved at 30-34 °C and pH 5.5 using xylose-grown preculture cells giving 20.5 g ethanol/l, ethanol productivity of 0.43 g/l h and ethanol yield of 0.44 g/g at 48 h. Adverse culture conditions mainly influenced the efficiency of xylose fermentation but not glucose fermentation. The key factors affecting co-fermentation were also explored at the molecular level. This study provides valuable insights into the effective harnessing of biomass resources.

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