Bio-hydrogen production of Clostridium acetobutylicum DMS 792 and the pSOL1 deficient mutant Mu56 in deproteneized cheese whey

Anaerobic fermentation (dark fermentation) of whey derived from cheese manufacturing could represent an opportunity to increase the value of this dairy by-product which is generally poorly reused and requires onerous costs of disposal. Today over the 25% of the whey produced in the European Union is used for human consumption but after extraction and concentration of the protein fraction it becomes a by-product. Due to the reduced availability of fossil fuels, bio-hydrogen production can actually represent a future prospect of renewable energy. The ability of growing and producing hydrogen was demonstrated for the Clostridium acetobutylicum DSM 792 type strain and for its pSOL1 deficient mutant Mu56, using deproteinized whey derived from Grana Padano cheese production as substrate. The mutant Mu56 lacks the 210-Kb plasmid pSOL1 which carries the genes responsible for acetone and butanol production. This causes the interruption of the solventogenic pathway and the redirection of the electrons flow toward the production of hydrogen, with the consequent increasing in its yield. The loss of the pSOL1 megaplasmid in Mu56 was demonstrated by PCR and southern blot and it was confirmed by genome analysis. Genome-wide analyses also identified a number of mutations occurring during the 56 sub-culturing steps employed for plasmid curing. The higher yield achieved in deproteinized whey from the C. acetobutylicum Mu56, compared to the parent strain DSM 792, was demonstrated using two independent analysis: the BHP and the water displacement methods. The hydrogen production of the C. acetobutylicum mutant Mu56 in buffered whey was the 6% and the 19,59% more than the type strain DSM 792, as determined by the water displacement and the BHP analysis, respectively.