This paper studied bioethanol production at very high gravity (VHG) conditions using flocculent Saccharomyces cerevisiae, evaluating the response yield, ethanol concentration, productivity, and residual sugar through a central composite design (CCD). This CCD was evaluated at 12 and 24 h fermentation times. In the CCD evaluated for 12 h of fermentation, the best condition for alcoholic fermentation was 27 °C, 260 g/L substrate concentration and a 30% v/v cell concentration; a maximum overall desirability of 0.937 was achieved. For CCD at 24 h of fermentation, the best condition was 27 °C, 300 g/L substrate concentration, and a 26% v/v cell concentration. The desirability achieved was 0.811. These conditions allowed us to verify, experimentally, that the CCD models described the fermentation behavior well. VHG alcoholic fermentation in fed-batch with the reuse of cells without chemical treatment was performed using the optimum conditions obtained from the desirability function (27 °C, 300 g/L, 26% v/v). This resulted in favorable alcohol content 132.90 g/L in comparison to the conventional fermentation process.