Development of novel clostridial fusants for enhanced biobutanol production

Biobutanol, a fuel with higher energy content than ethanol was produced in the present work through Simultaneous Saccharification and Fermentation (SSF) of wheat straw (WS) that traditionally produces acetone, butanol and ethanol solvents (ABE). Novel Clostridium strains were developed and tested for enhanced biobutanol production. Thermal stability was imparted to two mesophilic clostridial wild strains through protoplast fusion with that of a corresponding thermophilic clostridial strain. This novel development eliminated the need to add external enzymes during the SSF process, allowed for SSF to be conducted at an elevated temperature of 45°C, enhanced the activity of internally produced enzymes and tolerated butanol toxicity to new limits. In order to determine the stability of the genetically modified strains, the parent fusants were passed on to five successive growth cycles. The fused strains of each growth cycle were inoculated with substrate and examined for solvent production and compared to the parent generation. Polymerase Chain Reaction Technique (PCR) followed by Agarose Gel Electrophoresis demonstrated reproducible genetic stability of the fused strains. Highest biobutanol production of 13.8 g/L (almost twice the concentration reported by other laboratory scale batch SSF research studies) was produced by one of the novel fusants. Generally, fused strain that achieved the highest biobutanol production exhibited relatively constant butanol concentration over the five growth cycles. Optimistic results obtained from batch SSF at lab scale demonstrate a clear potential of these novel strains to improve productivity and yield of biobutanol at a large-scale facility.