Four Portuguese chestnut cultivars from the "Castanha da Terra Fria" protected designation of origin were selected: Aveleira, Boa Ventura, Judia and Longal. The nutritional parameters (moisture, fat, protein, carbohydrates, ash and energy) as well as fibers (neutral detergent fiber, acid detergent fiber, acid detergent lignin and cellulose) were characterized. Moisture was the major component followed by carbohydrates, protein and fat, resulting in an energetic value lower than 195 kcal/100 g of fresh fruit. In order to find significant differences among cultivars, the lipidic fraction was studied in detail. Fatty acids (FA) were determined by gas-liquid chromatography with flame ionization detection, revealing a clear prevalence of C18:1 and C18:2, two FA very well-known due to their beneficial effects on human health, e.g., in the prevention of cardiovascular diseases. A triacylglycerols (TAG) profile was obtained by reversed-phase high-performance liquid chromatography with evaporative light-scattering detection. TAG analysis is very important because it furnishes highly specific information due to genetic control of the stereospecific distribution of FA on the glycerol molecule. OLL, PLL, OOL and POL were the major compounds. As far as we know this is the first complete characterization of TAG in chestnut. The obtained data were screened through an analysis of variance (to evaluate the accuracy of the method as well as the uniformity of results for each cultivar) and a discriminant analysis (DA), which gave good results, once that, in some cases, the four cultivars were clustered in four individual groups, obtained through the definition of two DA dimensions.