Fermentation kinetics were monitored by CO2 release (Bely et al.,
1990 a, b). The amount of CO2 release (g/L) was determined by automatic
measurement of fermentor weight loss every 20 min. The CO2
production rate (g/L/h) was obtained by polynomial smoothing of the
last 11 CO2 measurements. Weight loss due to evaporation was under
2%.
Yeasts were pre-cultured in Erlenmeyer flasks filled with must at
24 °C for 24 or 48 h for S. cerevisiae and T. delbrueckii, respectively.
Fermentations were carried out at 24 °C with agitation in 1.2 L fermentors
locked to maintain anaerobiosis throughout alcoholic fermentation
(CO2was released through a sterile air outlet condenser). Four different
fermentations were conducted: two with pure cultures and two with
mixed cultures. Two types of mixed cultures were carried out:
simultaneous mixed modality where T. delbrueckii and S. cerevisiae
were inoculated at the same time and sequential mixed modality
where T. delbrueckiiwas inoculated 24 h before S. cerevisiae yeast. Single
and mixed cultures were inoculated with 1 × 107 viable cells/mL for
T. delbrueckii and 2 × 106 viable cells/mL for S. cerevisiae. All experiments
were performed in triplicate.