On-line Monitoring Techniques for the Study of Yeast Physiology. Some Studies on the Yeasts Pichia stipitis and Saccharomyces cerevisiae

Doktorsavhandling, 1993

On-line monitoring techniques have been used with the goal of optimizing fermentation processes involving the yeasts Saccharomyces cerevisiae (Baker's yeast) and Pichia stipitis (a xylose fermenting yeast), and a highly equipped fermentor system has been developed for these studies. The fed-batch growth of S. cerevisiae on a synthetic glucose medium was controlled using a control strategy based on calorimetric measurements and an increase in cell yield by 10% compared to diauxic batch growth was obtained. Anaerobic chemostat growth of S. cerevisiae was studied under carbon limitation and nitrogen limitation. The ethanol yield was higher and the glycerol yield lower during nitrogen limited conditions. The nitrogen content of the cells was different in the two cases, with a lower level for the cells grown under nitrogen limitation. The ATP yield was also substantially lower during nitrogen limited conditions. This was probably due to either increased transport energy requirements or futile cycling. The xylose metabolism by the xylose fermenting yeast Pichia stipitis has been studied with in situ fluorescence measurements of the nucleotides NADH and NADPH. The fluorescence increase, caused by a step change from aerobic to anaerobic conditions, was much lower for P. stipitis than for C. utilis, which shows on a larger redox imbalance for the latter yeast during anaerobic conditions. A mathematical model for an open-ended fluorosensor has been developed and experimentally verified with measurements on a pure fluorophore solution. Oxygen programmed fermentation, a new method for the study of microaerobic fermentations based on a continuous ramp of oxygenation rates, is introduced and analyzed mathematically. It is shown that it may be possible to study metabolic transitions as a function of oxygenation rates at conditions which, normally, would lead to wash-out. The effect of carbon dioxide on the xylose fermentation rate by P. stipitis was also studied. In a carbon dioxide atmosphere, the specific fermentation rate was 45% lower than in a nitrogen sparged fermentor.