After fermentation, the ethanol was concentrated to 99.5 wt% through a purification process. The mash column was operated for purifying the fermented broth and Table 5 lists the results. In the initial operation, the temperature at the top of the column was 94–95 °C, and the ethanol concentration was 6–8 wt%. When the ethanol was collected in the mash column reflux tank, ethanol was transferred to the top portion of the column. The temperature of the top portion was gradually decreased and the ethanol concentration increased with increasing reflux ratio. The mash column could concentrate ethanol to 73.8 wt% (approximately 80 vol%) when column conditions were the following: 1/8 reflux ratio, 82–83 °C top portion temperature, and 100.6 °C bottom portion temperature.