Ecological Control Strategies for Biobutanol Production

As the world transitions towards a bio-based economy, less dependent on current liquid fossil fuels, it becomes important to develop and implement sustainable alternatives to the many derivatives of oil refining. With the transportation sector accounting for more than half of the world’s consumption of liquid fossil fuels, liquid energy sources like gasoline and diesel are the primary targets of current research into green energy alternatives.

Due to their many similarities, butanol can already be used as direct replacement for gasoline. Butanol is also a key chemical in the solvent industry, and is used as bulk chemical in industrial sectors like plastics, polymers and even in the pharmaceutical industry, making it an all-around chemical of high interest. However, biological production of butanol still relies on expensive pure substrates (e.g. glucose) or feedstocks that compete with the agro-food sector, and on
pure culture fermentation operated in batch mode.

The goal of this thesis was therefore to develop a working microbiome, capable of continuous production of butanol using butyrate as a low-value carbon source. The substrate choice aimed at providing a route for resource valorization in the biotechnology industry, where butyrate can many times be found in wastewater. Taking from the successful example of wastewater remediation, where degradation of organic matter allows the recovery of resources and energy,and from an extensive literature review on their advantages and weaknesses, an anaerobic open microbiome was the inoculum of choice.

Through ecological selection principles, the metabolic flux of the open microbiome was tailored towards butanol production from butyrate and hydrogen as electron donor. Process control in a CSTR significantly increased butanol titers and volumetric productivities, over serum bottle batch fermentation, and minimized by-product formation from butyrate. A thermodynamic evaluation of catabolic reactions showed measured by-product formation to be unfeasible from butyrate conversion, but generally energetically favorable from residual carbon dioxide present in the inoculum. A better understanding of the structure of the open microbiome, capable of producing butanol from butyrate and hydrogen, was gained through DNA sequencing combined with 16S rRNA gene amplicon analysis. Results of the microbial community analysis also support the formation of by-products from carbon sources other than butyrate.

Building on the findings of butanol production solely from butyrate and hydrogen under well-controlled conditions during batch fermentation, the microbial community was further enriched for butanol-forming species through continuous fermentation. Two independent continuous fermentations, operated under steady conditions for up to three retention times, hint at the possibility of continuous butanol production from butyrate and hydrogen without the need for glucose addition. To benchmark different production platforms, butanol production with an open microbiome was successfully achieved in a UASB type reactor. Results were however inferior to those of a CSTR, due to operational constraints. Further experiments can help to assess the best reactor configuration for biobutanol production and the results obtained in both studied reactor configurations were benchmarked to proposed production platforms in literature.