This conclusion was reinforced by m

This conclusion was reinforced by measuring the settling rate of yeast cells. A 150 mL broth sample was with- drawn from the reactor and placed in a mess cylinder. The height of the cell-supernatant boundary was recorded at intervals, and lower heights of the cell-supernatant boundary indicated higher settling rates. As shown in Fig. 1c, the settling rate of yeast cells was accelerated by adding Ca2+, especially with addition of 2.35 g/L of CaCl2·2H2O, which corresponded to a Ca2+ concentra- tion of 1000 mg/L. Retaining the number of yeast at high level in the reactor is essential for fermentation at a high dilution rate. Gratifyingly, the number of yeast reached 19 × 108 cells/mL at a Ca2+ concentration of 1000 mg/L without decreasing the ethanol yield (Fig. 1a and b). Thus, the feedstock used for con- tinuous ethanol fermentation was supplemented with CaCl2·2H2O to the Ca2+ concentration of 1000 mg/L in the following stud- ies.