Metabolic selectivity and growth of Clostridium thermocellum in continuous culture under elevated hydrostatic pressure.

The continuous culture of Clostridium thermocellum, a thermophilic bacterium capable of producing ethanol from cellulosic material, is demonstrated at elevated hydrostatic pressure (7.0 MPa, 17.3 MPa) and compared with cultures at atmospheric pressure. A commercial limitation of ethanol production by C. thermocellum is low ethanol yield due to the formation of organic acids (acetate, lactate). At elevated hydrostatic pressure, ethanol:acetate (E/A) ratios increased >10(2) relative to atmospheric pressure. Cell growth was inhibited by approximately 40% and 60% for incubations at 7.0 MPa and 17.3 MPa, respectively, relative to continuous culture at atmospheric pressure. A decrease in the theoretical maximum growth yield and an increase in the maintenance coefficient indicated that more cellobiose and ATP are channeled towards maintaining cellular function in pressurized cultures. Shifts in product selectivity toward ethanol are consistent with previous observations of hydrostatic pressure effects in batch cultures. The results are partially attributed to the increasing concentration of dissolved product gases (H2, CO2) with increasing pressure; and they highlight the utility of continuous culture experiments for the quantification of the complex role of dissolved gas and pressure effects on metabolic activity.

Predicting vegetative inoculum performance to maximize phytase production in solid-state fermentation using response surface methodology.

Microbial phytase is used to reduce the environmental loading of phosphorus from animal production facilities. The limiting factors in the use of this enzyme in animal feeds can be overcome by solid-state fermentation (SSF), which is a promising technology for commercial enzyme production with lower production costs. Inoculum quality and the influence of inoculum quality on phytase production are important factors which need in-depth investigation before scaling-up of high-yielding fermentation process. A full factorial experimental design for 240 h with sampling at every 24 h was used to determine the effects of the treatments, inoculum age (plate and liquid culture), media composition and the duration of SSF on the production of fungal biomass and phytase in SSF systems using Aspergillus niger. The optimal treatment combination for maximal phytase production was determined by statistically comparing all treatments at each sampling time. Both 7- and 14-day plate cultures and M1 + medium composition with 72-h-old liquid inoculum treatments resulted in optimal phytase production at 144 h of SSF, which was the shortest duration observed for maximal phytase production. This resulted in maximal phytase production with a mean of 884 +/- 121 U/g substrate, while the maximal phytase production observed at 216 h of SSF (mean phytase activity of 1,008 +/- 121 U/g substrate), with the same treatment combinations, was not statistically significant from that at 144 h of SSF. Phytase production was strongly growth-associated with younger inocula. The significant treatment variables, age of liquid inoculum and the duration of SSF, were used to predict the system response for phytase production using response surface methodology. From the response surface model, the optimal response of the experiment was predicted and the reliability of the prediction was checked with the verification experiment.

Toxicity effects of compressed and supercritical solvents on thermophilic microbial metabolism.

Selection of biocompatible solvents is critical when designing bioprocessing applications for the in situ biphasic extraction of metabolic end-products. The prediction of the biocompatibility of supercritical and compressed solvents is more complicated than for liquid solvents, because their properties can change significantly with pressure and temperature. The activity of the anaerobic thermophilic bacterium, Clostridium thermocellum, was studied when the organism was incubated in the presence of compressed nitrogen, ethane, and propane at 333 K and multiple pressures. The metabolic activity of the organisms in contact with compressed solvents was analyzed using traditional indicators of solvent biocompatibility, such as log P, interfacial tension, and solvent density. The toxicity of the compressed solvents was compared with the phase and molecular toxicity effects measured in liquid alkanes at atmospheric pressure. Inactivation increased with time in the presence of the compressed solvents, but was constant in the presence of atmospheric liquid solvents. Knowledge of molecular and phase toxicity provides a framework for the interpretation of C. thermocellum metabolism in contact with atmospheric and compressed solvents.