Abstract
Repeated yeast pitching is a common practice in ale production, but yeast viability can decline across generations due to ethanol exposure, nutrient depletion, storage stress, and accumulated physiological damage. This study evaluates yeast viability changes during repeated pitching in controlled ale fermentation. Yeast slurry was harvested and repitched across successive fermentation cycles under standardized wort composition, pitching rate, oxygenation, and temperature conditions. Cell viability, membrane integrity, glycogen reserve, fermentation rate, attenuation, flocculation behavior, pH change, ethanol production, and off-flavour indicators were monitored. The results show that early repitching cycles maintained stable viability and fermentation performance, while later cycles showed slower attenuation, reduced vitality, and higher stress-related metabolite formation. Poor slurry storage and excessive trub carryover accelerated viability loss. The study demonstrates that repeated pitching can support consistent ale production when generation number, yeast handling, storage time, and physiological quality are carefully controlled before each reuse.