Antifungal activity and synergistic potential of Achillea santolina ethanolic extract and its bioactive constituents against trichophyton mentagrophytes

Chemical analysis results of the alcoholic extract of Achillea santolina revealed the presence of 33 bioactive compounds. The most prominent of these compounds responsible for the extract's biological activity were phytol (17.31%), benzophenone (14.23%), scopa one (12.78%), the 3-methyl-3-buten-1-ol derivative (13.29%), and trithiocyanuric acid (11.74%). MIC and MFC tests demonstrated the extract's effectiveness against Trichophyton mentagrophytes, with an MIC of 8 ± 0 mg/mL and an MFC of 9 ± 0 mg/mL, compared to the standard flucytoconazole (MIC = 16 µg/mL and MFC = 32 µg/mL). The results showed that the ethyl fraction (Teac) recorded the highest antifungal activity, with an MIC of 2.0 ± 0.1 mg/milk MFC = 3.0 ± 0.3 mg/mL when the extract was separated into organic fractions, while the aqueous fractions were less effective (MIC = 10 mg/mL, MFC >10 mg/mL). Stronger activity was observed for the isolated compounds scopa one and phytol at lower concentrations, with a synergistic effect when mixed in a 1:1 ratio (MIC = 150 ± 15 µg/mL, MFC = 300 ± 15 µg/mL). The results of the synergistic experiment between luliconazole and the crude extract indicated a significant decrease in both MIC values ​​(from 7 mg/mL to 3 mg/mL for the extract and 16 µg/mL to 6 µg/mL for the drug), with an FIC index of 0.50, indicating a synergistic effect between them. The results of Time-kill assay showed that the ethyl fractions at 2×MIC reduced the microbial density to less than 1.0 log CFU/mL after 24 hours, confirming a fungicidal effect, while the crude extract recorded a decrease to 3.8 log CFU/mL. Cytotoxicity assay showed that the isolated compounds had lower toxicity and greater selectivity than the crude extracts and semi-pure fractions. This makes them safer treatment options. These results confirm the strong potential of Achillea santolina as a natural source of antifungals. It could be developed into additional treatment options, especially when used alongside traditional drugs.