Changes in fermentation characteristics of commercial Kombucha determined by time of inoculation of a mixed culture of bacteria and yeast—Gluconobacter oxydans SFT-27, Acetobacter pasteurianus SFT-18, Leuconostoc mesenteroides SFT-45, and Saccharomyces cerevisiae SFT-71
Main Article Content
Keywords
inoculation time; Kombucha; metabolite; mixed culture fermentation; process optimization
Abstract
Our study aimed to investigate the effects of varied inoculation times of a mixed culture of four microor-
ganisms on the fermentation characteristics of kombucha to standardize the process. We selected strains
through prior phenotypic and genotypic screening to establish a controlled, mixed microbial fermentation
system for kombucha production. A combination of Gluconobacter oxydans SFT-27, Acetobacter pasteur-
ianus SFT-18, Leuconostoc mesenteroides SFT-45, and Saccharomyces cerevisiae SFT-71was used for inoc-
ulation at different times (Days 1–5) to assess the effects on fermentation characteristics. Green tea extract
was prepared and supplemented with sucrose, glucose, and appropriate nitrogen sources. The extract was
inoculated with a primary inoculation of acetic acid bacteria (G. oxydans SFT-27, A. pasteurianus SFT-18).
There were subsequent inoculations with yeast (S. cerevisiae SFT-71) and lactic acid bacteria (L. mesenteroi-
des SFT-45) from Day 1–5. We analyzed key fermentation indices, including pH, titratable acidity, ethanol
content, °Brix, reducing sugar content, viable microbial counts, and gluconic acid levels over 25 days. Results
demonstrated that pH decreased sharply after the third day of inoculation, with titratable acidity peaking
at 4.64%, surpassing Korean food code standards. Ethanol peaked on Day 5 and decreased thereafter. There
was a steady increase in reducing sugars, particularly in the samples inoculated later, which correlated with
enzymatic sucrose hydrolysis and cellulose degradation. Microbial counts indicated stable populations of
acetic acid bacteria and yeast when the samples were inoculated on Days 3 and 5. Gluconic acid production
was the highest in the sample inoculated on Day 5, exceeding levels reported in symbiotic culture of bacteria
and yeast (SCOBY)-based fermentations. These findings suggest that inoculation with yeast and lactic acid
bacteria, following prefermentation with acetic acid bacteria, yields superior fermentation performance in
terms of acidity, metabolite production, and microbial stability.
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