Development of a bioprocess for enhanced butanol production: Integrating laboratory experiments with in-silico predictions

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Increasing global population and energy intensive lifestyle has resulted in increased global energy demand. Butanol, produced by clostridial fermentation, is a promising transportation alternative as a drop-in fuel due to its physiochemical properties. The present study aims to address pressing bottlenecks and develop a suitable bioprocess for butanol production using Clostridium sp. Suitable strain for fermentation, C. acetobutylicum MTCC 11274, was selected from procured cultures based on a maximum butanol production capability of 11.5 g L-1. The maximum butanol titer obtained under screening experiments restricted by butanol toxicity. Hence, for media optimization, maximization of biomass productivity and maximization of butanol productivity were selected as objective functions. Based on batch experiments conducted using the media obtained for maximization of biomass productivity and maximization of butanol productivity, a two-stage fed-batch fermentation strategy was developed. The process involved sequential use of biomass productivity media followed by butanol productivity media, where butanol induction served as the transition point from one stage to the second stage. The strategy resulted in a 77 % improvement in the butanol productivity (0.55 g L-1 h -1) when compared to batch fermentation using the optimized medium for maximization of butanol productivity. Further, studies on limitation and starvation of nutrients in stage one yielded magnesium limitation as the most suitable factor and addition of 5 g L-1 calcium carbonate in stage two was found to be the best among supplementation experiments. Based on these results, a combinatorial approach was designed which improved butanol titer and butanol productivity to 16.5 g L-1 and 0.59 g L-1 h -1, respectively, one of the highest values reported in literature.
Supervisor: Debasish Das