Influence of aeration on cytoplasmic pH of yeast in an NMR airlift bioreactor

Nuclear magnetic resonance spectroscopy has been increasingly pursued as a tool for noninvasive, real-time studies of metabolic processes of cell suspensions in bioreactors. One acute challenge in NMR bioreactor design has been supplying enough oxygen for cell respiration in a suspension that contains sufficient cells for NMR signal detection. The use of cytoplasmic pH as an intracellular marker of adequate oxygenation was evaluated from P-31 NMR spectra of the yeast Saccharomyces cerevisiae at several cell densities, ranging from low (0.9% (v/v)) to very high (45% (v/v)) cell densities, in an airlift bioreactor. P-31 NMR spectra were obtained for derepressed yeast cells prior to, and during, glycolysis under nongrowth conditions. During endogenous respiration, pH(cyt) can be used as an intracellular marker for aeration for cell densities up to 18% (v/v) based on two criteria: a value of pH(cyt) at least 0.2 pH units higher under aerobic than anaerobic conditions and an absolute pH(cyt) value of 7.1-7.2. These results were more conservative than values of the maximum cell density obtained from calculations using k(L)a and respiration rate estimates and highlight the utility of intracellular measurements in conjunction with engineering design calculations. During glycolysis, pH(cyt) values were similar under aerobic and anaerobic conditions and hence pH(cyt) cannot be used as a marker under these conditions. Carbon dioxide in the influent gas was observed to aid cells in maintaining physiological pH(cyt) at high cell densities.