Herbal plants are known to contain a wide number of bioactive compounds that are considered as potential sources of natural biological activities. Therefore, this study aims to compare and analyze the chemical composition, the polyphenolic compounds, and the biological activities of ethanolic extracts of Saussurea costus (SCE) and Withania somnifera (WSE). The chemical composition and the polyphenolic compounds of the ethanolic SCE and WSE were elucidated. Each of their antioxidant activity, antifungal activity, and anti-aflatoxigenic activities were evaluated. The obtained data indicated that SC contained higher levels of carbohydrates and protein than WS, while higher levels of ash, moisture, and oil were detected in WS. Although SCE contained higher levels of total phenolic and flavonoid contents and antioxidant activity, the high content and diversity of some polyphenolic compounds such as Gallic acid, Coffeic acid, Syringic acid, Pyro catechol, Rutin, Vanillin, Kaempferol and Cinnamic acid detected in WSE making it more efficient as an antifungal and anti-aflatoxigenic agent than SCE. It can be concluded that both SCE and WSE were potential sources of polyphenolic compounds with antioxidants, antifungal and anti-aflatoxigenic properties.