from indigenous fermented foods and human gut ... - Thapar University

3.7 Characterization of probiotic properties of Lactobacillus strains
3.7.1 Low pH and bile salt tolerance
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Chapter III Material and methods
The isolated Lactobacillus strains were tested for their ability to resist at low pH and
bile salt. The pH value of gastric acid varies in the range of about 1.5-4.5 in a period of 2 h,
depending on the entering time and the type of gastric contents. In the present study, pH 2
was used as a representative gastric pH value. 24 h old culture of each Lactobacillus (10 8
CFU/ml) was suspended in a MRS broth at pH 3 for 5 h at 37°C. The suspensions were then
centrifuged at 3000 rpm for 10 min at 4°C twice and washed in sterile saline solution to
remove the media. Cells were suspended in physiological solution and a series of 10-fold
dilution (10 -2 -10 -10 ) was prepared. A given amount of each dilution (50 µl) was plated on to
de Man Rogosa Sharpe (MRS) agar and incubated anaerobically at 37°C for 24 h. The
percentage of the viable bacteria was calculated. Tolerance to bile salts was verified by
inoculating 100 µl of bacterial suspension of each strain (10 8 CFU/ml) on to MRS agar
containing bile salt at 0.3% concentration. Survival of the Lactobacillus strains was examined
by counting the cells after 24 h of incubation at 37°C on to MRS agar.
3.7.2 Resistance to 0.4% phenol
Some aromatic amino acids derived from dietary or endogenously produced proteins
can be deaminated in the gut by bacteria, leading to the formation of phenols (Suskovic et al.,
1997). These compounds may exert a bacteriostatic effect against some Lactobacillus strains.
Thus, testing for the resistance to phenol may generate further information on the potential
for survival of Lactobacilli in gastrointestinal conditions (Xanthopoulos et al., 2000).
Therefore, the ability of Lactobacillus strains to grow in the presence of phenol by
inoculating cultures (1% of an overnight culture) in MRS broth with and without 0.4%
phenol was tested. Serial dilutions were spread plated (100 μl aliquots) onto MRS agar at