Application of flow cytometry to monitor the evolution of Clostridium acetobutylicum during extractive ABE fermentation
Tuesday, April 29, 2014: 1:05 PM
Grand Ballroom F-G, lobby level (Hilton Clearwater Beach)
Butanol obtained by Acetone-butanol-ethanol fermentation is a promising petroleum
substitute. In order to make its production
economically viable, it is necessary to maximize the yield of alcohol in the bioreactor, and thus
to overcome microorganism’s inhibition at product threshold concentration. The
liquid-liquid extractive fermentation is an in situ removal approach that can be used to minimize
the product inhibition and the high energy demand for downstream. The effectiveness of the
process will depend on the extraction performance and the ability to keep high microbial cells
viability.
The solventogenic Clostridia go through a complex cell-cycle
during butanol production. The role of the different cell types coexisting in the culture is still
under discussion. Flow cytometry allows to analyze the bacterial morphology evolution at the
single cell level (size and internal granulometry). Combining this technique with specific
fluorescent stains, it is also possible to discriminate cells by their physiological state and
metabolic activity.
In this work, two solvents have been tested on batch extractive
fermentations, in which butanol production increased up to 60 % related to conventional
fermentation. The morphological changes of different cell types
were quantified for both control and extractive fermentation, and compared to the
production rate of the metabolic products. Results gave us evidence that butanol production can
be achieved by vegetatives cells, with low endospore/spore participation. The percent of
membrane permeable cells are systematically higher in the case of conventional fermentation,
while the viability is preserved to a certain extent in the extractive fermentation with vegetable
oil. Considering these results, flow cytometry appears as a useful potential application for
metabolic pathways comprehension, and further bioprocess control and optimization at
industrial scale.
substitute. In order to make its production
economically viable, it is necessary to maximize the yield of alcohol in the bioreactor, and thus
to overcome microorganism’s inhibition at product threshold concentration. The
liquid-liquid extractive fermentation is an in situ removal approach that can be used to minimize
the product inhibition and the high energy demand for downstream. The effectiveness of the
process will depend on the extraction performance and the ability to keep high microbial cells
viability.
The solventogenic Clostridia go through a complex cell-cycle
during butanol production. The role of the different cell types coexisting in the culture is still
under discussion. Flow cytometry allows to analyze the bacterial morphology evolution at the
single cell level (size and internal granulometry). Combining this technique with specific
fluorescent stains, it is also possible to discriminate cells by their physiological state and
metabolic activity.
In this work, two solvents have been tested on batch extractive
fermentations, in which butanol production increased up to 60 % related to conventional
fermentation. The morphological changes of different cell types
were quantified for both control and extractive fermentation, and compared to the
production rate of the metabolic products. Results gave us evidence that butanol production can
be achieved by vegetatives cells, with low endospore/spore participation. The percent of
membrane permeable cells are systematically higher in the case of conventional fermentation,
while the viability is preserved to a certain extent in the extractive fermentation with vegetable
oil. Considering these results, flow cytometry appears as a useful potential application for
metabolic pathways comprehension, and further bioprocess control and optimization at
industrial scale.