Integrated study of fermentation and downstream processing in a miniplant significantly improved the microbial 1,3-propanediol production from raw glycerol.

In this work, an integrated and optimized production process for 99 % pure 1,3-propanediol (PDO) from raw glycerol is presented. Glycerol fermentation is carried out applying a newly isolated strain Clostridium pasteurianum K1 under non-sterile conditions without any complex ingredients in the fermentation media. In this environment over 55 g/L PDO, yields of 0.52 g/g and space time yields over 2.3 g/(Lh) were achieved in up to 1 m(3) pilot scale. The downstream process for PDO purification consists of an ultrafiltration for biomass and protein separation, an evaporation step for concentration of PDO and a two-step rectification for final purification. For a proof of concept, process optimization and especially investigation of interactions of individual steps, the downstream process was performed in miniplant scale. A minimum salt input into the downstream process was shown to be important to overcome precipitation in evaporation as well as rectification. Thus, raw glycerol is desalinated before fermentation and the fermentation medium was minimized and complex nutrients, such as yeast extract, were avoided totally to prevent furthermore dark color formation. Furthermore, by titration of fermentation with ammonia instead of sodium hydroxide, the later separation of the major by-products, organic acids, in the evaporation step was significantly enhanced.

Influence of temperature on the production of an archaeal thermoactive alcohol dehydrogenase from Pyrococcus furiosus with recombinant Escherichia coli.

The heterologous production of a thermoactive alcohol dehydrogenase (AdhC) from Pyrococcus furiosus in Escherichia coli was investigated. E. coli was grown in a fed-batch bioreactor in minimal medium to high cell densities (cell dry weight 76 g/l, OD600 of 150). Different cultivation strategies were applied to optimize the production of active AdhC, such as lowering the cultivation temperature from 37 to 28 degrees C, heat shock of the culture from 37 to 42 degrees C and from 37 to 45 degrees C, and variation of time of induction (induction at an OD600 of 40, 80 and 120). In addition to the production of active intracellular protein, inclusion bodies were always observed. The maximal activity of 30 U/l (corresponding to 6 mg/l active protein) was obtained after a heat shock from 37 to 42 degrees C, and IPTG induction of the adhC expression at an OD600 of 120. Although no general rules can be provided, some of the here presented variations may be applicable for the optimization of the heterologous production of proteins in general, and of thermozymes in particular.