Comprehensive in vitro antimicrobial and antiviral evaluation, GC- MS profiling, in silico molecular docking and molecular dynamics simulations of Achillea millefolium essential oil

The rapid rise in antimicrobial drug resistance and recurrent viral outbreaks has necessitated the discovery of alternative multi- target anti-infective agents from natural resources. In this study, the essential oil of Achillea millefolium L. (AMEO), obtained by hydrodistillation of aerial parts collected from Saudi Arabia, was investigated using an integrated in vitro and in silico framework, including GC–MS profiling, antimicrobial and antiviral assays, molecular docking, molecular dynamics (MD) simulations, and MM- GBSA binding free energy analyses. Hydrodistillation yielded 0.55% (w/w) essential oil. GC–MS profiling identified 26 volatile constituents, representing 99.26% of the total oil composition, dominated by oxygenated monoterpenes and sesquiterpenes, with α-thujone (28.98%), β-thujone (14.70%), 1,8-cineole (13.39%), trans-sabinene hydrate (6.99%), trans-sabinyl acetate (5.73%), and spathulenol (4.02%) as the major constituents. AMEO exhibited concentration-dependent antimicrobial activity against methicillin- resistant Staphylococcus aureus, Escherichia coli, Candida albicans, and Aspergillus flavus, as well as notable antibiofilm effects. Antiviral evaluation showed measurable inhibition of influenza A virus (H1N1) and herpes simplex virus type 2 (HSV-2), whereas limited selectivity was observed against SARS-CoV-2 and adenovirus type 5. Extended 300-ns MD simulations coupled with MM-GBSA analysis provided insights into ligand–protein stability and the energetics of the interactions. Among the simulated complexes, 7B6M_spathulenol showed the most favorable binding free energy (−13.8 kcal/mol), whereas 8YVL_β-thujone displayed the weakest energetic profile (−0.5 kcal/mol), indicating a target-dependent binding behavior. The simulated complexes were mainly stabilized by van der Waals and hydrophobic interactions, with most systems maintaining structural stability during the MD simulations. Binding affinity estimates derived from MM-GBSA energetics showed variable predicted affinities, supporting strong target engagement for selected complexes while indicating weaker binding for others. Overall, the findings suggest that AMEO, particularly through selected constituents such as spathulenol, β-thujone, and trans-sabinyl acetate, possesses promising antimicrobial, antibiofilm, and selective antiviral potential, warranting further mechanistic and experimental validation as a natural anti-infective candidate.