Ethanol and acetate production from synthesis gas using microbial catalysts

by 1982- Cotter, Jacqueline Louise

COTTER, JACQUELINE LOUISE. Ethanol and Acetate Production from Synthesis Gas using Microbial Catalysts. (Under direction of Mari S. Chinn.) The hybrid technology of gasification and fermentation has the potential to serve as a viable approach for ethanol production from plant biomass. Autotrophic bacteria can use gaseous product streams from a gasifier, CO and CO2 and H2, to produce ethanol and acetate. The work presented investigates the potential of resting cells to enhance ethanol production by Clostridium ljungdahlii and Clostridium autoethanogenum as well as the processing parameters significant to synthesis gas fermentation. Resting cells can offer improved cell stability under harsh environmental conditions and enhance the production of secondary metabolites (i.e. ethanol). The objectives of the resting cell culture experiments were to develop methods to induce the resting state in key autotrophic bacteria and evaluate performance based on culture stability over time, ethanol and acetate production, and culture viability. In effort to increase ethanol formation by resting cultures, the effects of benzyl viologen and medium pH were also examined. Studies describing microbial performance of growing C. ljungdahlii and C. autoethanogenum cultures supplied continuous synthesis gas streams have been limited. The objectives assessing synthesis gas processing conditions included: examining the overall culture metabolism of C. autoethanogenum grown on bottled synthesis gas supplied at different flow rate; and the effects of pH and bottled synthesis gas flow rate on substrate utilization and metabolic end product formation by C. ljungdahlii in liquid-batch, continuous-gas fermentation. Variations of nitrogen limited media were tested for function in creating non-growing cells, while maintaining cell viability and density. C. ljungdahlii was able to maintain a stable cell density when transferred to basal medium supplemented with vitamins and trace elements and all major nitrogen removed. Despite the culture’s viability, these resting cells did not produce ethanol and acetate in large quantities. Cultures at pH 4.5 and 6.8 produce maximum ethanol concentrations of 0.52 mM and 1 mM over 144 hours, respectively. Cultures at pH 5.5 did not produce any ethanol. Poor performance at the lower pH levels may also be related to viability, where less than 50% viability was observed. The addition of benzyl viologen negatively affected culture viability and resulted in little ethanol and acetate production. C. autoethanogenum was sensitive to the amount and source of nitrogen in the different media formulations. Ammonia chloride was necessary for minimizing culture density loss, however non-growing cells did not produce significant quantities of ethanol and acetate. Considering viability of the cells, ethanol seems to be a primary metabolite for these autotrophic bacteria on sugar substrates. Growth of C. ljungdahlii and C. autoethanogenum on continuous synthesis gas substrates was slower with longer lag phases than what was observed for growth on sugars. For C. ljungdahlii higher cell densities were achieved at a pH of 6.8 (579 mg/L) compared to pH 5.5 (378 mg/L) after 48 hours. In addition, the ethanol concentration at pH 6.8 (3.8 mM) was 110% greater than that at pH 5.5. Acetate concentrations were not statistically affected by pH level. For ethanol formation and ethanol to acetate ratios, flow rate was not a significant factor. Unlike C. ljungdahlii, C. autoethanogenum was significantly influenced by gaseous flow rates. More dense cultures were achieved at 10 ml/min compared to 5 ml/min gas flow, 136 mg/L and 109 mg/L, respectively. Although ethanol concentrations were less than 0.5 mM after 60 hours, the 10 ml/min flow rate resulted in a 38% increase in ethanol compared top the 5ml /min treatment. Overall, ethanol was not observed as a secondary metabolite on sugar and synthesis gas substrates. More acidic initial medium pH levels do not promote ethanol production over acetate and reduced microbial growth potential. Synthesis gas flow rate will have a greater impact on cell culture densities and growth than regulation on end product formation.