TABLE 1.

Protocol for standardization of fermentation trials in synthetic must carried out by Saccharomyces cerevisiae EC1118 and S. cerevisiae AWRI796.

Step	Procedures
Yeast strain supply	The active dry yeast (ADY) belonging to the same lot should be used
Synthetic must preparation	The synthetic must composition is reported in Table 1 of the resolution OIV-OENO 370-2012 (Anonymous, 2012)
Synthetic must distribution	500-ml Erlenmeyer flasks containing 350 ml of synthetic must and equipped with Muller valves are used. The trials are carried out in triplicates (three independent experiments)
Yeast rehydration	According to the resolution OIV/ENO 329/2009 (Anonymous, 2009), each ADY strain is rehydrated as follows: • weigh 1 g of ADY under aseptic conditions; • add 100 ml of 5% sucrose solution in water at 36–40°C under sterile conditions; • homogenize slowly using a rod or a magnetic stirrer for 5 min; • stop stirring and allow to stand for 20 min at a temperature between 36 and 40°C; • homogenize again at room temperature for 5 min; • take 10 ml under sterile conditions and then proceed to count viable yeast cells by Thoma counting chamber using 0.1% (w/v) methylene blue solution.
Yeast strain inoculum	Inoculate the rehydrated yeast in the synthetic must in order to get 2 × 106 cells/ml
Fermentation trial conditions	Incubate the Erlenmeyer flasks closed with Muller valves (containing sulfuric acid) at 25 ± 2°C in static conditions for 15 days
Fermentation monitoring	Check the weight loss daily after shaking each Erlenmeyer flask by hand for 1 min
Sample arrangement for analyses	At the end of the fermentation, centrifuge at 3,000 × g for 5 min at room temperature and separate the cells from the supernatant
Chemical analyses	Resulting wines should be analyzed at certified laboratory by official OIV methods.