Development and validation of versatile cellular models for investigating the absorption and bioactivity of natural extracts

Objectives In order to explain the health benefits provided by high quality extra virgin olive oil (EVOO), the objectives of the work are the following: a) Assessing whether EVOO polyphenols are absorbed at intestinal level and their antioxidant capacity on human intestinal Caco-2 and hepatic HepG2 cells. b) Evaluating their effects on cholesterol metabolism in Caco2 and HepG2 cells and on ox-LDLs and lipid peroxidation in HepG2 cells. Methods For the absorption experiments, differentiated human intestinal Caco-2 cells are incubated in the apical side with the tested samples and absorbed species are detected and quantified by HPLC-chip/ESI/MS/MS. The antioxidant effect is evaluated in vitro and at cellular level by DPPH assay and analysing the reactive oxygen species (ROS) production. The effects on cholesterol metabolism are assessed by measuring the OxLDL production and lipid peroxidation. Results All methodologies were optimized working on plant protein hydrolysates as testing samples. Using the HPLC-chip/ESI/MS/MS approach, absorbable lupin peptides were identified and strategies developed to analyse further their biological activities. Methods to evaluate the antioxidant activity at in vitro and cellular level were optimized on soybean protein hydrolysates. Their ability to reduce the ROS and nitric oxide (NO) production after induced oxidative stressed on HepG2 cells were assessed. The same approaches are currently applied to study the EVO polyphenols bioactivity.