Electronic Thesis and Dissertation Repository

Thesis Format



Master of Engineering Science


Chemical and Biochemical Engineering


Rehmann, Lars


Butanol, a next-generation biofuel, can be produced by fermenting glycerol using Clostridium pasteurianum. To address product inhibition, an integrated system that combined a fed-batch process with pervaporation was assessed against conventional batch and fed-batch fermentations. This study showed that with the novel process configuration, the productive fermentation time could be extended, translating to a 2.4-fold and 1.9-fold increase in butanol production relative to baseline fed-batch and batch operation, respectively. Further, it was demonstrated that butanol concentrations were able to be maintained below inhibitory levels throughout the fermentation. Despite this outcome, metabolic oscillations were revealed, indicating instability in the process. The introduction of secondary sugar substrates into the modified system improved the butanol selectivity and did not result in fluctuations of the product profile. Overall, these findings provide strong evidence of the advantages of in-situ product recovery in glycerol fermentation which can be aided by the addition of secondary carbon sources.

Summary for Lay Audience

Efforts to improve the sustainability of biofuel production practices have redirected attention to new generation biofuels, such as biobutanol. Butanol is an ideal biofuel candidate that can be easily integrated into current infrastructure and provides a higher carbon content relative to other biofuels such as ethanol. Several avenues exist to produce biobutanol, yet an attractive route is via fermentation with Clostridium pasteurianum. This anaerobic process is unique in that it can exclusively use crude glycerol, a ‘waste’ product from the biodiesel industry as a low-cost feedstock. However, current limitations of low productivity render the process economically unfeasible on an industrial level. One factor that restricts the efficiency of the fermentation in batch operation is butanol toxicity. A solution to minimize the accumulation of butanol in the reactor is to incorporate in-situ product removal.

The focus of this project is to develop a new process configuration for glycerol fermentation that addresses the accumulation of butanol in the reactor. To accomplish this goal, a membrane-assisted fed-batch fermentation was performed using pervaporation, a selective method of butanol recovery. In this system, fermentation broth is pumped out from the bioreactor and across a membrane, where butanol diffuses through the surface while the remaining cell-rich broth is returned to the fermenter.

When compared to a baseline fed-batch control without pervaporation beginning with the same initial glycerol concentration, the modified set-up resulted in a 2.4-fold increase in butanol production and extended the fermentation time by 19.5 hours. No evidence of butanol inhibition was observed as concentrations were maintained below inhibitory levels. Despite this improvement, the process was variable, with fluctuations observed in gas production and glycerol consumption, indicative of metabolic shifts of the culture. To overcome this oscillatory behaviour, additional simple sugar substrates were introduced into the modified system. Under co-substrate fermentation, the system performed more consistently, without observed oscillations but an imbalance between removal capacity and production resulted in toxic butanol levels, prematurely ending the fermentation. This outcome indicates the need for more refinement of process conditions but overall highlights the value of in-situ product removal in glycerol fermentation.

Available for download on Sunday, April 30, 2023