Environmental effects on growth and ethanol fermentation of immobilized Kluyveromyces fragilis

A trickle flow, short column reactor system using a natural sponge as a carrier was used to study the response of the immobilized yeast, Kluyveromyces fragilis NRRL 2415, to environmental and nutritional factors. The specific growth rate of adsorbed K. fragilis using a complete medium as feed during the start-up period was 0.0152 hr$\sp{-1}$ which was much lower than 0.60 hr$\sp{-1}$, the value for free cells. The transient responses of immobilized cells to changes in lactose concentration and temperature were quick, reaching new steady state values within half an hour. There was little or no detectable CO$\sb2$ evolved if the lactose concentration was below 3.8 g/l. The immobilized K. fragilis did not perform well at temperatures over 40$\sp\circ$C, although free cells maintained their viability at 40$\sp\circ$C. The inhibitory effect of added ethanol on immobilized K. fragilis was a nonlinear relationship. The thermotolerance of K. fragilis was significantly reduced by the presence of ethanol. A kinetic model was developed to determine the steady state productivity based upon growth and death rates as a function of temperature and ethanol concentration. Ergosterol was required for a stable performance of steady state immobilized K. fragilis using defined medium as feed. The cells could survive with a nitrogen-deficient medium and maintain a constant performance level without losing significant fermentative activity (ca. 26 to 30%) for 48 to 72 hours. Absence of vital nutrients for a period of up to three days did not cause cell washout or loss of viability. At least 0.375 g/l of yeast extract in a complex medium was required for stable fermentative activity. There was little difference between the presence and absence of ammonium sulfate if a suboptimal amount of yeast extract (0.375 g/l) was used. However, higher ammonium sulfate levels resulted in higher reactor productivity when higher amounts of yeast extract were used. Viability was not significantly influenced by ammonium sulfate.