Ethanol production at elevated temperatures requires high potential thermotolerant ethanol-producing yeast. In this study, nine isolates of thermotolerant yeasts capable of growth and ethanol production at high temperatures were successfully isolated. Among these isolates, the newly isolated thermotolerant yeast strain, which was designated as Saccharomyces cerevisiae DBKKU Y-53, exhibited great potential for ethanol production from sweet sorghum juice (SSJ) at high temperatures. The maximum ethanol concentrations produced by this newly isolated thermotolerant yeast at 37 °C and 40 °C under the optimum cultural condition were 106.82 g·L⁻¹ and 85.01 g·L⁻¹, respectively, which are greater than values reported in the literatures. It should be noted from this study with SSJ at a sugar concentration of 250 g·L⁻¹ and an initial pH of 5.5 without nitrogen supplementation can be used directly as substrate for ethanol production at high temperatures by thermotolerant yeast S. cerevisiae DBKKU Y-53. Gene expression analysis using real-time RT-PCR clearly indicated that growth and ethanol fermentation activities of the thermotolerant yeast S. cerevisiae DBKKU Y-53 at a high temperature (40 °C) were not only restricted to the expression of genes involved in the heat-shock response, but also to those genes involved in ATP production, trehalose and glycogen metabolism, and protein degradation processes were also involved.

The optimum fermentation conditions for ethanol production from sweet sorghum juice (SSJ) by the thermotolerant yeast Saccharomyces cerevisiae DBKKUY-53 were determined using a statistical experimental design. Based on the Plackett–Burman design (PBD), yeast cell concentration, sugar concentration, and yeast extract were the significant independent fermentation factors affecting the ethanol production from SSJ at 37 °C by S. cerevisiae DBKKUY-53. These significant factors were optimized using response surface methodology (RSM) based on a central composite design (CCD). The result revealed that the optimum conditions for ethanol fermentation were 7.85 × 10⁷ cells/mL yeast cell concentration, 247 g/L sugar concentration, and 9.99 g/L yeast extract. Verification of the ethanol production using the optimum conditions revealed that the maximum ethanol concentration of 99.75 g/L and the productivity of 2.77 g/L/h were achieved. When the ethanol production was carried out in a 2 L fermentor under optimum conditions, the ethanol concentration was 101.81 g/L and the productivity was 2.83 g/L/h. This finding suggested that the thermotolerant yeast S. cerevisiae DBKKUY-53 has excellent potential for commercial ethanol production at high temperatures.