THE EFFECT OF BIOREACTOR CONDITIONS ON ATP SUPPLY AND DEMAND, ELECTRON FLOW, AND PRODUCT FORMATION IN THE ACETONE/BUTANOL FERMENTATION
MEYER, CHARLES LEE
Doctor of Philosophy
The availability and demand of biosynthetic energy (ATP) and reducing electrons (NADH, reduced ferredoxin) have been found to be crucial in the regulation of solvent production in steady state continuous cultures of Clostridium acetobutylicum. Production of acetic and butyric acids, which results in more efficient production of ATP, predominates when the availability of ATP is low, such as under conditions of glucose limitation. When the ATP supply is plentiful, solvent formation occurs together with a reduced rate of acid production. The intracellular level of ATP is a good measure of the energy supply of the cell and is itself one of the key factors regulating relative rates of acid and solvent formation. The intracellular ATP level was five to eight times as high during non-glucose limited steady states as during glucose limited steady states. When the ATP supply is not limiting, a lower growth rate imposed by biomass recycle leads to a reduced demand for ATP and substantially higher acetone and butanol yields. The solvent yield and the ATP ratio, a measure of the efficiency of ATP production, both exhibit a linear dependence on $\mu$ during non-glucose limited steady states. When the culture is glucose limited, however, biomass recycle results in lower solvent yields and higher acid yields. CO gassing of a culture of C. acetobutylicum can completely inhibit H$\sb 2$ production. An increased availability of reduction energy caused by inhibition of H$\sb 2$ formation triggers and enhances butanol, ethanol, and lactate production in batch and continuous cultures. Acetone production, however, which does not require NADH, is inhibited along with acid production. The intracellular NADH level increased more than four-fold during CO sparging and is implicated as the key factor leading to increased alcohol formation. ATP levels also increase substantially during CO gassing, but do not appear to be as important as the high levels of NADH. NADH and ATP levels were relatively high at all times during batch cultures compared to levels in continuous cultures. Although the levels of these nucleotides show systematic trends during normal batch fermentations, their role in the initation of solventogenesis in batch cultures is not clear.