Syntrophic acetate oxidizing bacteria convert acetate into hydrogen and carbon dioxide and through the mutualistic syntrophic partnership with methanogens the products are further converted to methane in biogas processes operating at high ammonia concentrations. There is very little known about SAOBs, only five have been characterized and had their genome analyzed. The aim of this project was to gain further knowledge about the methane producing pathway of SAOBs with a proteomic approach. Proteins were extracted from biogas sludge with a phenol-based approach and trypsin digestion and peptide recovery were performed using the Suspension Trapping method. Measurement of the peptide content was made with LC-MS/MS. The peptide profiles obtained were screened for the proteins expressed of the mesophilic SAOB Syntrophaceticus schinkii. The data supports earlier suggestions that it utilizes the Wood-Ljungdahl pathway for hydrogen production. Furthermore, the peptide profile revealed that enzymes for the glycine reductase complex and the glycine cleavage system were expressed during high ammonia concentration, indicating a potential role of these enzymes in the methane producing pathway. However, due to partial failure of the sample preparation for mass spectrometry measurements no quantification conclusions could be made. A discussion on how to further improve sample preparation methods as well as how to access the proteome to a large extent is presented.

Biofuel production via fermentation is produced primarily by fermentation of simple sugars. Besides the sugar fermentation route, there exists a promising alternative process that uses syngas (CO, H2, CO2) produced from biomass as building blocks for biofuels. Although syngas fermentation has many benefits, there are several challenges that still need to be addressed in order for syngas fermentation to become a viable process for producing biofuels on a large scale. One challenge is mass transfer limitations due to low solubilities of syngas species. The hollow fiber reactor (HFR) is one type of reactor that has the potential for achieving high mass transfer rates for biofuels production. However, a better understanding of mass transfer limitations in HFRs is still needed. In addition there have been relatively few studies performing actual fermentations in an HFR to assess whether high mass transfer rates equate to better fermentation results. Besides mass transfer, one other difficulty with syngas fermentation is understanding the role that CO and H2 play as electron donors and how different CO and H2 ratios effect syngas fermentation. In addition to electrons from CO and H2, electrodes can also be used to augment the supply of electrons or provide the only source of electrons for syngas fermentation. This work performed an in depth reactor comparison that compared mass transfer rates and fermentation abilities. The HFR achieved the highest oxygen mass transfer coefficient (1062 h-1) compared to other reactors. In fermentations, the HFR showed very high production rates (5.3 mMc/hr) and ethanol to acetic acid ratios (13) compared to other common reactors. This work also analyzed the use of electrons from H2 and CO by C. ragsdalei and to study the effects of these two different electron sources on product formation and cell growth. This study showed that cell growth is not largely effected by CO composition although there must be at least some minimum amount of CO present (between 5-20%). Interestingly, H2 composition has no effect on cell growth. Also, more electron equivalents will lead to higher product formation rates. Following Acetyl-CoA formation, H2 is only used for product formation but not cell growth. In addition to these studies on electrons from H2 and CO, this work also assessed the redox states of methyl viologen (MV) for use as an artificial electron carrier in applications such as syngas fermentation. A validated thermodynamic model was presented in order to illustrate the most likely redox state of MV depending on the system setup. Variable MV extinction coefficients and standard redox potentials reported in literature were assessed to provide recommended values for modeling and analysis. Model results showed that there are narrow potential ranges in which MV can change from one redox state to another, thus affecting the potential use as an artificial electron carrier.