Species-Specific Metabolite Profiles and Biological Activities of Bulgarian Thymus Species from Section Hyphodromi

This study investigates the phytochemical composition and biological activity of eight Thymus species distributed in Bulgaria, with a focus on taxa from section Hyphodromi. High-performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC–MS) were used to characterize the methanolic (MeOH) and diethyl ether (Et2O) extracts, respectively. MeOH extracts revealed high concentrations of phenolic acids, particularly rosmarinic acid, salicylic acid, and flavonoid glycosides, with T. atticus, T. jalasianus, and T. leucotrichus showing the most diverse profiles. Et2O extracts were dominated by triterpenic acids (ursolic and oleanolic) and monoterpenes such as thymol and carvacrol, especially in T. zygioides and T. leucotrichus. All MeOH extracts exhibited significant antioxidant activity in the DPPH assay (IC50 < 50 µg/mL), with T. jalasianus and T. atticus demonstrating the strongest effects. Radical scavenging potential generally followed the trend of total phenolic content. Moderate acetylcholinesterase-inhibitory activity was observed only in T. zygioides and T. leucotrichus. The study reports for the first time data on the biological activity and metabolic composition of extracts from endemic and rare species and from the species of Bulgarian origin. The results provide new data on the phytochemical composition and in vitro antioxidant and acetylcholinesterase-inhibitory activities of selected Thymus species, contributing to the characterization of their overall in vitro biochemical profiles.