2.7. Microbiological analysis
The FDA’s standard method of Bacteriological Analytical Manual
[24] was used to determine the microbial population of juice samples.
Pour plate method was used to determine the total plate
counts. Proper serial dilutions were prepared by mixing sterilized
distilled water followed by further decimal dilutions of samples
(up to 105). These dilutions were poured into sterile petri dishes
with the help of a pipette. Molten agar (15 mL) was added to each
petri dish. Mixing of sample dilutions with agar medium was
immediately done by moving each petri dish 5 times in vertical,
clockwise and horizontal, anticlockwise directions. The petri dishes
were held at room temperature (25 ± 1 C) for 30 min followed by
incubation at 37 C for 48 h in an incubator (GSP-9080 MBE, Shanghai
Boxun Industry & Commerce Co., Ltd., China). The bacterial colonies
were counted by multiplying with reciprocal in the samples
and the results were expressed as log colony-forming units (CFU)/
mL of juice.
Yeast and mold counts were determined by pour plate method
using PDA. A known amount of PDA powder (31.2 g) was
dissolved in distilled water (800 mL) to prepare media. Tartaric
acid (10%, w/v) was added in PDA media to protect it from the
growth of microbes by cross contamination. All the media plates
were put in an incubator at 32 ± 1 C. After 48 h of incubation,
yeast and mold were counted in each plate and the results were
expressed as CFU/mL of juice. All the analysis was carried out in
triplicate.