The isolation and purification of EPS from fermented skim milk
samples were performed according to a protocol described previously
(Prasanna et al., 2011). The fermented milk sample was mixed with
12% (v/v) of 80% (w/v) trichloroacetic acid (TCA) and stored at 4 °C
for 24 h, for the precipitation of proteins. The samples were centrifuged
at 25,000×g for 30 min at 4 °C. The supernatant was mixed
with two volumes of chilled absolute ethanol, and stored at 4 °C for
24 h for the precipitation of EPS. The precipitate was separated by
centrifugation at 25000×g for 30 min, at 4 °C. The pellet containing
the EPS was dissolved in ultrapure water and dialysed against distilled
water at 4 °C for 2 days, with two water changes per day,
using a dialysis membrane tube (molecular mass cut-off value
12,400 Da, Sigma-Aldrich, UK). The EPS-containing solution was
then freeze dried (IEC Lyoprep - 3000). The freeze dried samples
were stored at −20 °C for further analysis.