Comparative study of rosmarinic acid content in methanolic extracts of Bulgarian Thymus species, section Hyphodromi

Genus Thymus consists of around 250 species known for their therapeutic properties: antioxidant, anti-inflammatory, expectorant, and carminative. Twenty-one species occur naturally in Bulgaria, with eight belonging to the section Hyphodromi: Bulgarian endemics: T. jalasianus Stoyanov & Marinov. and T. perinicus (Velen.) Jalas; Balkan endemics: T. aznavourii Velen. T. compus Friv.; T. atticus Čelak (sub-endemic), as well as T. leucotrichus Halácsy T. striatus Vahl. and T. zygioides Griseb., all widespread. Due to the species’ limited distribution and the overall complexity of the genus, extensive studies are lacking. Rosmarinic acid is a phenolic compound with highly valued biological activities, including antioxidant, anti-inflammatory, and antimicrobial properties. Many thyme species are reported to be sources of this polyphenol, showcasing their potential beneficial properties. This study aimed to estimate the rosmarinic acid content by a quick, accurate, and readily available method such as HPTLC in the methanolic extract of the Thymus species section Hyphodromi distributed in Bulgaria. Plant materials were collected from the natural localities of the studied taxa. The quantification of rosmarinic content was achieved using HPTLC silica gel 60 F₂₅₄ plates. The mobile phase was chloroform/ethyl acetate/formic acid (50:40:10). The compounds were visualized by spraying with ‘Naturstoffreagenz A’ reagent. The fluorescence emission of rosmarinic acid was recorded under UV radiation at 336 nm, using a digital camera. The images were analyzed by QuantiScan 2.1 Biosoft software. The results shows that the content varied between 1.20 and 2.50 μg/g dry weight. Thymus atticus and T. jalasianus extracts were found to be the richest sources of rosmarinic acid, while those of T. comptus, T. aznavouiri, and T. striatus showed moderate levels. The obtained data showed Thymus atticus and T. jalasianus as promising for cultivation when plant material with a high rosmarinic acid content is needed.