3.2.8.1. Evaluation of the Impact of Digested Formulations on Intestinal Epithelium Viability

To evaluate the impact of digested formulations on intestinal epithelium viability, digested formulations were serially diluted in digestive fluids (from 1:2 up to 1:32 dilution) and added to the apical side of the in vitro intestinal epithelia, while HBSS buffer was placed in the basolateral compartment. Digestive fluids (without formulations) were added to the apical side of the in vitro intestinal epithelia as a negative control. After 3 h incubation, monolayers were washed twice with pre-warmed HBSS and viability of intestinal epithelia was evaluated with MTS assay, according to manufacturer's instructions. This assay is based on MTS tetrazolium compound reduction by viable cells to generate a colored formazan product that can be quantified by measuring the absorbance at 490 nm. The color intensity at 490 nm was determined with a microplate reader (Synergy4, Biotek, Colonia Santo Domingo Azcapotzalco Distrito Federal, México). Cell viability (%) was expressed as the ratio of the color intensity in the treated groups to that in the control (untreated) group. Absorption rate experiments were performed using non-toxic concentrations determined by dose-response curves.
