AbstractThe genus Stachys L., one of the largest genera of the Lamiaceae family, is highly represented in Turkey. This study was conducted to determine the bio‐pharmaceutical potential and phenolic contents of six different extracts from aerial parts of Stachys tundjeliensis. The obtained results showed that the ethanol extract exhibited the highest antioxidant activity in the antioxidant assays. Meanwhile, the ethanol extract displayed strong inhibitory activity against α‐tyrosinase, the dichloromethane extract exhibited potent inhibition against butyrylcholinesterase, and the n‐hexane extract against α‐amylase. Based on ultra‐high performance liquid chromatography coupled to high‐resolution mass spectrometry analysis, more than 90 secondary metabolites, including hydroxybenzoic acid, hydroxycinnamic acid, and their glycosides, acylquinic acids, phenylethanoid glycosides, and various flavonoids were identified or tentatively annotated in the studied S. tundjeliensis extracts. It was observed that the application of S. tundjeliensis eliminated H2O2‐induced oxidative stress. It was determined that protein levels of phospho‐nuclear factor kappa B (NF‐κB), receptor for advanced glycation endproducts, and activator protein‐1, which are activated in the nucleus, decreased, and the synthesis of matrix metalloproteinase (MMP)‐2 and MMP‐9 also decreased to basal levels. Overall, these findings suggest that S. tundjeliensis contains diverse bioactive compounds for the development of nutraceuticals or functional foods with potent biological properties.