Chestnut fruit market is increasing worldwide, following the emergence of new chestnut based products (e.g.chestnut flour and purees). In Europe, Castanea sativa Mill is the main chestnut species commercialized. Portugal is one of the most important European producer countries, being Trás-os-Montes region the most important production area (85% of the national production/ total area of 35000 ha/ production of 19000 tons per year).Chestnut processing and consumption generate high amounts of by-products: shell, inner skin, outer skin and bur. This project is designed to extract active ingredients from Castanea sativa shells (CSS) using green technology (comparing conventional methodologies with supercritical fluids extraction). In fact, food by-products are an industrial burden requiring costly handling and waste management. Recent trends advocate added value of such materials as sources of compounds with biological properties. Through in vitro (Caco-2, HT29-MTX, Raji B, HaCaT and HFF-1 cells), ex-vivo (Franz cells) and in vivo assays (mouse and humans), the safety and efficacy of the final products developed (a functional food and a cosmetic product) will be assessed. The extracts from CSS will be characterized regarding chemical composition (HPLC/DAD/FL/MS) and scavenging activity against reactive oxygen and nitrogen species (ROS and RNS). Afterwards, in vitro tests with respect to the permeability (3D artificial model of intestine) and potential cytotoxicity and genotoxicity will be performed. Anti-inflammatory, antihypertensive and cholesterol-lowering ability will be also studied. Furthermore, different assays will be done to ensure the topical delivery (Franz cells and Raman Confocal Microscopy).In vivo tests will be performed to analyze the toxicity, efficacy and safety of the final products developed. Finally, the characterization of the final product, a cosmetic and a functional food containing the extract, will be performed by characterizing the textural/rheological properties and performing a sensory evaluation. 
In order to properly execute the tasks planned for this research project, the REQUIMTE-team has the facilities, equipment?s and knowhow gathered by scientific papers on functional food [1, 2], antioxidants [3, 4], cosmetics [5-9] and nutraceuticals [10, 11].The CESPU team has the facilities, equipment?s and knowhow by the performance of scientific papers on the evaluation of intestinal permeability and clinical tests on animal models to assess toxicity and efficacy [12-15].
With this research project is intended to:1)contribute to the agro-research with studies that will permit to use CSS in cosmetics and functional foods;2)optimize an efficient extraction of compounds with added value from CSS, using green technologies and low operational costs;3)valorize a food by-product, reducing waste and environmental impact;4)initiate a new era in Portuguese research, which has not considerable expression on food by-products re-use.