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Metabolic analysis of solventogenic clostridium saccharoperbutylacetonicum N1-4 (HMT).

Saunders, Elizabeth (2019) Metabolic analysis of solventogenic clostridium saccharoperbutylacetonicum N1-4 (HMT). Doctoral thesis, University of Surrey.

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The market for solvent production is predicted to reach $43.4 billion in 2018, with n-butanol having over 20% market share value where n-Butanol is the chemical precursor of several industrially important products, such as butyl-acetate, butyl-acrylate, glycol-ethers, and plasticisers. Butanol is currently produced from crude oil, and therefore in light of dwindling fossil fuel reserves, and more importantly, the need for green and clean production processes, synthesis of bio-butanol from biomass using Clostridia represents a viable and desirable alternative method.

This project focuses on the metabolic and physiologic characterisation of the acetone-butanol-ethanol (ABE) producing species Clostridium saccharoperbutylacetonicum (Csb). A minimal medium for Csb was defined based on literature data, modified by the addition of glutamate to support growth. Interestingly, batch cultures using this medium showed that Csb was able to grow and produce butanol under aerobic conditions, with titres of approximately 74% of those observed under anaerobic conditions. Steady state cultures in chemostats are essential to elucidate and characterise physiological features of microorganisms. Steady state cultures of Csb were used to determine the effect of acid production on solventogenesis, bacterial growth, and energy metabolism. Studies at different pH in the range 5.5 to 6.5 showed no correlation with the onset of solventogenesis. However, the pH and the growth rate seem to influence the productivity of butanol. In those experiments, significant increases in the production rate of butanol were observed when the dilution (growth) rate increased from 0.01 h-1 to 0.03 h-1 and the pH decreased from 6.5 to 5.5. Growth is potentially linked to production rate due to an increased demand for ATP and NADH recycling.

The use of genome scale metabolic models allows for the interpretation of metabolic and physiological changes upon changes in the culture conditions. A metabolic model of Csb was constructed based on the genome sequence of the microorganism and incorporating biomass synthesis equations specific for Csb which were constructed based on the analysis of the composition of the cells grown in the chemostat experiments, as opposed to current models that use biomass composition from related species (e.g. B. subtilis). The metabolic model was used to perform flux balance analysis to identify and interpret the changes in the distribution of metabolic fluxes that would explain the metabolic changes observed in Csb cultured under different conditions.

This work has demonstrated the basis for the presence of monophasic solventogenesis in C. saccharoperbutylacetonicum and provided important tools (defined media, GSMN equations) to improve industrial scale production of renewable sources of carbon-based feedstocks and thus reducing reliance on crude oil.

Item Type: Thesis (Doctoral)
Divisions : Theses
Authors : Saunders, Elizabeth
Date : 30 August 2019
Funders : Biotechnology and Biological Sciences Research Council (BBSRC)
DOI : 10.15126/thesis.00852363
Contributors :
ContributionNameEmailORCID Rossa,
Depositing User : Elizabeth Saunders
Date Deposited : 03 Sep 2019 15:16
Last Modified : 03 Sep 2019 15:17

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