2.1 Cultures and Media
C. acetobutylicum ATCC55025 was cultured in modified P2 medium containing (per liter, at pH 5.5): 0.5 g KH2PO4, 0.5 g K2HPO4, 1 g yeast extract, 0.5 g tryptone, 0.2 g MgSO4∙7H2O, 0.01 g MnSO4∙H2O, 0.01 g NaCl, and 0.01 g FeSO4∙7H2O, 0.1 mg p -aminobenzoic acid, 0.1 mg thiamine, 1 µg biotin, and 50 g glucose as the carbon source, unless otherwise noted (Xu et al., 2015). Concentrated mineral (MgSO4∙7H2O, MnSO4∙H2O, NaCl, FeSO4∙7H2O) and vitamin (p -aminobenzoic acid, thiamine, biotin) solutions were prepared separately, filter-sterilized with 0.2 µm pore size filter and stored in a refrigerator until use. The medium without glucose was sterilized by autoclaving at 121 ºC for 30 minutes. The main carbon source, glucose, was sterilized separately in a concentrated solution and added aseptically to the medium to a final concentration of ~50 g/L or as specified. Butyric acid at various amounts (up to 10 g/L) was also added to the medium to study its effects on cell growth and fermentation kinetics. The medium was adjusted to pH 5.5 with NH4OH and purged with sterile N2 for ~30 min to reach anaerobiosis before use.