Nineteen Tunisian onion genotypes were characterized based on phytochemical composition, sugar content, and antioxidant activity. The studied onions showed a great diversity on biochemical contents and composition. It seemed that there were differences between genotype categories (local landraces or local breeding lines) and within the same category. Quercetin and quinic acid were, respectively, the main flavonoid (flavonol) and organic acid (cyclitol) identified and quantified by HPLC. In all studied onions, quercetin was the predominant flavonoid, with the highest content in the local breeding line OP2-w (1142.19 mg 100 g^-1 DW). OP3-w showed the highest value of total phenolic as well as total flavonoid content at 12.52 mg GAE g^-1 DW and 48.28 RE g^-1 DW respectively. Fructose and sucrose were the most abundant sugars in all the genotypes. Clustering and PCA analysis showed a great dispersion of these genotypes which were classified into 3 major groups. The chemical and nutritional composition found highlights the great value of this onion germplasm which can be used for the sustainable conservation and management of Tunisian onion genetic resources.