Unveiling the phytochemical landscape of Phlomis pungens willd.: LC-MS/MS insights into antioxidant and multi-enzyme inhibitory potential

Phlomis pungens Willd. (syn. Phlomis herba-venti subsp. pungens (Willd.) Maire ex DeFilipps) is a medicinal and aromatic species that has been studied mainly for its essential oil composition, whereas its non-volatile phytochemical constituents have remained largely unexplored. In this study, the ethanolic extract of P. pungens was  systematically characterized for the first time using a targeted LC–MS/MS method. In parallel, total phenolic and flavonoid contents, antioxidant potential, and multi-enzyme inhibitory activities were evaluated. LC–MS/MS profiling revealed a phenolic acid–rich composition, with quinic acid (14.146 mg/g extract) and chlorogenic acid (13.187 mg/g) identified as the predominant compounds.These were followed by cynaroside (1.541 mg/g), vanillic acid (1.453 mg/g), and cosmosiin (0.830 mg/g). The extract demonstrated considerable levels of total phenolics and flavonoids (37.18 ± 1.97 μg PEs/mg and 18.19 ± 0.90 μg QEs/mg extract, respectively). Antioxidant evaluation indicated moderate activity, particularly in the ABTS (IC50 = 32.05 ± 0.28 μg/mL) and CUPRAC (A0.5 = 20.83 ± 0.37 μg/mL) assays. Enzyme inhibition studies revealed selective inhibitory effects against collagenase (24.73 ± 0.32%), tyrosinase (15.04 ± 0.11%), and butyrylcholinesterase (13.62 ± 0.12%) at 50 μg/mL, while no inhibition was detected toward acetylcholinesterase, urease, or elastase.These findings suggest that the biological activities of P. pungens may be associated with its non-volatile phenolic constituents.This work provides novel phytochemical data and highlights the potential of the species for future pharmaceutical and dermocosmetic applications.