Chestnut (Castanea sativa) is an important basic food in rural diets and a major starch crop used in a similar way to potatoes. Chestnuts are a fundamental economic resource in the "chestnut regions" not only for the fruit but also for the chestnut wood. Chestnuts have become increasingly important with respect to human health, for example, as an alternative gluten-free flour source. Chestnuts are also a rich source of other beneficial compounds, but there have been few studies on the composition during processing. In this study, we analyzed the chemical composition of three Portuguese cultivars at different stages of industrial processing. The chestnut cultivars were Longal, Judia, and Martaínha. All three cultivars had high moisture contents but were low in ash, crude fat, and crude protein contents, with high starch and low fiber contents. The free amino acid contents, including various essential amino acids, varied depending on the cultivar. All three cultivars also had a significant content of polyphenolics with gallic acid; ellagic acid was predominant among hydrolyzable and condensed tannins. Many of these compounds are known to exert significant positive effects on human health. The one-way analysis of variance for fresh chestnut shows significant differences among the three cultivars for most of the studied parameters. The same statistical analysis applied to each one of the two cultivars (Judia and Longal) sampled for the four processing steps analyzed indicates a significant effect of this factor in practically all of the constituents. On the other hand, the two-way analysis of variance shows that, besides the residual, the processing step and the interaction cultivar x processing step were the factors that more contributed for the total variation observed in the constituents analyzed, while the contribution of cultivar was much less significant.