*3.5. Total Acidity and Antibacterial Activity of Kombucha Tea*

The antibacterial activities of the kombucha tea that was fermented under different conditions were investigated against pathogenic enteric bacteria. The fermented tea was tested at 15 days in terms of acidity and then after neutralization (pH 7.0). Moreover, the samples were heated to analyze the thermostability of the active components. Kombucha tea prepared from green, oolong, and black tea showed antibacterial activity against all tested enteric bacteria; *Escherichia coli*, *E. coli* O157: H7 DMST 12743, *Shigella dysenteriae* DMST 1511, *Salmonella* Typhi DMST 22842, and *Vibrio cholerae*. The diameter of the inhibition zones of the kombucha prepared from green tea ranged from 20.0 ± 0.0–24.7 ± 0.6 mm. The diameter of the inhibition zones of kombucha prepared from oolong tea ranged from 19.3 ± 0.6–24.7 ± 0.6 mm and the diameter of the inhibition zones of black tea ranged from 20.0 ± 0.0–21.3 ± 0.6 mm. The antibacterial activity of each type of kombucha tea was similar to that of the acetic acid at the same total acidity content of each type of kombucha tea. In this study, gentamicin (1 mg/mL) was used as a positive control and presented the diameters of the inhibition zones ranged from 19.3 ± 0.6–23.3 ± 0.6 mm. after testing with *E. coli*, *E. coli* O157:H7, *S. dysenteriae*, *S.* Typhi, and *V. cholera* (Table 2).


**Table 2.** Antibacterial activity of kombucha prepared from green tea, oolong tea, and black tea against pathogenic enteric bacteria.

<sup>a</sup> Inhibition zone diameter is presented as mean ± SD of three independent experiments. <sup>b</sup> Fermented tea: kombucha tea fermented at 15 days without any treatment. <sup>c</sup> Unfermented tea: broth culture contained only 1% of tea leaves. <sup>d</sup> Neutralized kombucha: kombucha tea neutralized with NaOH (1 M) at pH 7.0. <sup>e</sup> Heat-denatured kombucha M1: kombucha tea treated at 100 ◦C for 20 min. <sup>f</sup> Heat-denatured kombucha M2: kombucha tea treated at 121.5 ◦C for 15 min. <sup>g</sup> Acetic acid: acid prepared at the same total acidity content of kombucha for each type of tea leaf.

On the other hand, the antibacterial activity of kombucha after neutralization to pH 7.0 did not reveal any inhibition of all of the tested enteric bacteria. Kombucha tea was also treated by being boiled (100 ◦C for 20 min) and autoclaved (121.5 ◦C for 15 min) to determine the activity of the thermostability of its components. The total acidity of the kombucha tea prepared from green, oolong, and black teas were titrated after thermal treatments, while the total acidity of kombucha tea prepared from green, oolong, and black teas after treatment by boiling was 10.93, 11.61, and 15.67 g/L, respectively. In addition, the total acidity of kombucha tea from green, oolong and black teas after treatment by being autoclaved was 8.12, 9.16, and 12.16 g/L, respectively. Thus, the total acidity level of kombucha tea after treatment by boiling was maintained since the total acidity was significantly higher than that of the kombucha tea that was treated by being autoclaved. The results indicated that the antibacterial activity of kombucha tea after treatment by boiling revealed significantly higher inhibitory activity on the tested enteric bacteria than in the kombucha tea that had been treated by being autoclaved (Table 2).
