The first Task will be to collect and identify wild mushrooms, with the help of an expert in this area and the second Task will consist in the isolation and in vitro production of mushrooms mycelia. The third Task will consist in the preparation of the extracts for further studies. The
extracts will be further characterized in LMW (phenolic compounds will be analysed by HPLC-DAD/MS, tocopherols by HPLCfluorescence,ascorbic acid and carotenoids by spectrophotometer techniques) and HMW (polysaccharides will be characterized by HPSEC, oligosaccharides by FACE, and their composition in monosaccharides by HPLC-RI). The fourth Task will initially consist in carrying out a screen of the growth inhibitory properties of the various extracts/fractions, in four cancer cell lines. Furthermore,
further characterizing the effects in cellular proliferation and apoptosis, selecting for this study one cell line only and the extracts or fractions (obtained after HPSEC separation) which showed more potent growth inhibition effect. Finally, in the fifth Task we intend to elucidate the mechanism of action of some of the bioactive compounds identified in the extracts (either antioxidants or polysaccharides). QSAR (quantitative structure-activity relationship) studies will be performed using the PLS (partial least squares)statistical method implemented in SIMCA-P software, in order to correlate the identified bioactive compounds in the extracts with their antiproliferative and apoptosis inducing properties. Several proteins involved on antiproliferative and apoptotic activity have been
identified as targets for modulation with compounds isolated from functional foods. Many of these protein targets have their 3D structure experimentally determined by X-Ray crystallization or NMR studies and are readily available at the Protein Data Bank (PDB).
We will select targets with known 3D structures and perform protein-ligand molecular docking studies with the identified compounds as ligands, using Autodock4 software.