Metabolite analyses over the first two hours of fermentation with regard to the production of ethanol, acetate, intra- and extracellular glycerol are shown in Fig. 3. Ethanol (Fig. 3a) increased in all strains after 5–10 min from inoculation into the must, growing constantly during all the experiments and at 120 min reached values between 3.4 ± 0.1 and 8.0 ± 0.2 mmole/OD. At this time point, ISE19 and ISE90 strains showed superior ethanol content, with respect to commercial strain BK1, and reached values which were 115–135% higher. Acetate production (Fig. 3b) showed increments starting from 30 min after inoculation; at 120 min strain BK1 was different with a lower production, by about 50%, compared with the other strains, ISE19 and ISE90, which showed a similar metabolite production. Analysis of the extracellular glycerol (Fig. 3c) showed a trend that is substantially similar to that of acetate with a rapid increase from 30 min. In this case, BK1 and ISE19 were different in comparison to strain ISE90 with a lower rate of accumulation, presenting a 25–30% lower concentration at 120 min. The accumulation of intracellular glycerol (Fig. 3d) showed relevant differences with respect to the metabolites previously described. The inoculation of strains ISE19 and ISE90 into the must induced the accumulation of glycerol in the cell with a greater increase between 10 and 30 min. After 30 min, the accumulation was interrupted and the concentration remained essentially unchanged until the end of the experiment. Once again, strain BK1 differed in its accumulation of intracellular glycerol: starting from inoculation, this strain accumulated glycerol at a lower rate with respect to the other two strains, but continuously up to 120 min and did not exhibit any stopping of the accumulation. At 120 min, the concentrations of the three strains didn't show any significant differences. Interestingly, cell growth measured as OD shows an increase starting from 30 min in strain ISE19 and 60 min in ISE90, while growth is not observed in the case of strain BK1