2.4. Simulated in vitro gastrointestinal digestion
The digestions were performed with simulated gastric fluid and
simulated intestinal fluid, both prepared according to procedures
implemented by Moura and Canniatti-Brazaca (2006). The simulated
gastrointestinal digestion was performed with pepsin solubilised
with 0.1 mol L1 HCl during the gastric phase and pancreatin–
bile salts solubilised with 0.1 mol L1 NaHCO3 in the intestinal
phase. Twenty grams of each sample was added to 100 mL of
0.01 mol L1 HCl and adjusted to pH 2 with 2 mol L1 HCl solution.
After the pH adjustment, 3.2 mL of pepsin was added, and the sample
was stirred in a thermostat at 37 C/2 h to simulate the digestion
of the food in the stomach. After that, titration with
0.5 mol L1 NaOH was performed until pH 7.5 to simulate the pH
found in the intestine of an individual. Dialysis was performed
for two hours in dialysis membranes (33 21 mm, molecular
weight: 12.000–16.000, porosity: 25 Angstrons – INLAB, Brazil)
containing 0.1 mol L1 NaHCO3 equivalent to titratable acidity.
After the pH adjustment, the dialysis membranes were added
and stirred in a water bath thermostated at 37 C/30 min, then
5.0 mL of pancreatin solution and bile salts were added and stirred
in a bath at 37 C/2 h. This step simulates the digestion of food in
the intestine. At the end of this step the contents of the membrane
(dialysate) were removed and samples were stored at 20 C until
the time of analysis.