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A reinvestigation of cellulose degradation by Clostridium cellulolyticum in a bioreactor with pH control of the batch culture and using a defined medium was performed. Depending on cellulose concentration, the carbon flow distribution was affected, showing the high flexibility of the metabolism. With less than 6.7 g of cellulose liter ؊1 , acetate, ethanol, H 2 , and CO 2 were the main end products of the fermentation and cellulose degradation reached more than 85% in 5 days. The electron flow from the glycolysis was balanced by the production of H 2 and ethanol, the latter increasing with increasing initial cellulose concentration. From 6.7 to 29.1 g of cellulose liter؊1 , the percentage of cellulose degradation declined; most of the cellulase activity remained on the cellulose fibers, the maximum cell density leveled off, and the carbon flow was reoriented from ethanol to acetate. In addition to that of previously indicated end products, lactate production rose, and, surprisingly enough, pyruvate overflow occurred. Concomitantly the molar growth yield and the energetic yield of the biomass decreased. Growth arrest may be linked to sufficiently high carbon flow, leading to the accumulation of an intracellular inhibitory compound(s), as observed on cellobiose (E. Guedon, M. Desvaux, S. Payot, and H. Petitdemange, Microbiology 145:1831-1838, 1999). These results indicated that bacterial metabolism exhibited on cellobiose was distorted compared to that exhibited on a substrate more closely related to the natural ecosystem of C. cellulolyticum. To overcome growth arrest and to improve degradation at high cellulose concentrations (29.1 g liter ؊1 ), a reinoculation mode was evaluated. This procedure resulted in an increase in the maximum dry weight of cells (2,175 mg liter ؊1 ), cellulose solubilization (95%), and end product concentrations compared to a classical batch fermentation with a final dry weight of cells of 580 mg liter ؊1 and 45% cellulose degradation within 18 days.Cellulolytic clostridia play a major role in cellulose decomposition, which is a key process in carbon cycling (29). Clostridium cellulolyticum is a nonruminal cellulolytic mesophilic bacterium isolated from decayed grass and capable of degrading crystalline cellulose (36). The biotechnological exploitation of this microorganism as well as the understanding of the role it plays in its own ecosystem requires knowledge of its metabolism and of its behavior when developed on cellulose.C. cellulolyticum is a low-GϩC gram-positive anaerobe belonging to clostridial group III (39, 40); it is also placed in family 4, genus 2, in a new proposed-hierarchical structure for clostridia (7). Recent metabolic investigations with this bacterium indicated that (i) compared to a complex medium previously used, mineral salt medium clearly produced a different regulatory response and permitted better control of the carbon flow (19, 34), (ii) early growth inhibition was associated with a carbon excess (18), and (iii) carbon-limited and carbon-saturated chemostats...

The effect of the component concentrations of a synthetic medium on acetone and butanol fermentation by Clostridium acetobutylicum ATCC 824 was investigated. Cell growth was dependent on the presence of Mg, Fe, and K in the medium. Mg and Mn had deleterious effects when in excess. Ammonium acetate in excess caused acid fermentation. The metabolism was composed of two phases: an acid phase and a solvent one. Low concentrations of glucose allowed the first phase only. The theoretical ratio of the conversion of glucose to solvents, which was 28 to 33%, was obtained with the following medium: MgSO 4 , 50 to 200 mg/liter; MnSO 4 , 0 to 20 mg/liter; KCl, 0.015 to 8 g/liter (an equivalent concentration of K + was supplied in the form of KH 2 PO 4 and K 2 HPO 4 ); FeSO 4 , 1 to 50 mg/liter; ammonium acetate, 1.1 to 2.2 g/liter; para -aminobenzoic acid, 1 mg/liter; biotin, 0.01 mg/liter; glucose, 20 to 60 g/liter.