ADMET analysis and evaluation of the effect of ortho-eugenol on the QacA/B efflux pump of Staphylococcus aureus

Antimicrobial resistance (AMR) is a significant global problem, with Staphylococcus aureus being a prominent multidrug-resistant bacterium, mainly due to mechanisms such as β-lactamase production and the QacA/B efflux pump system. These mechanisms impede effective treatment of the infection, necessitating the development of innovative therapeutic strategies. In this context, natural products such as ortho-eugenol, a synthetic isomer of eugenol, have attracted attention for their potential to modulate bacterial resistance. This study evaluated the antimicrobial activity of ortho-eugenol against S. aureus K4414, a strain harboring the QacA/B efflux pump. Despite having a minimum inhibitory concentration (MIC) of ≥ 1024 μg/mL, indicating weak antibacterial activity, its combination with penicillin resulted in a fivefold reduction in the MIC of the antibiotic (from 512 μg/mL to 101.6 μg/mL). This potentiating effect was not observed with the standard efflux pump inhibitor chlorpromazine, suggesting that the mechanism is not related to QacA/B inhibition but likely involves β-lactamase inhibition, as confirmed by the combination of ampicillin with sulbactam. In addition, ortho-eugenol in combination with ethidium bromide (EtBr) increased the MIC of EtBr, suggesting an antagonistic interaction. In contrast, chlorpromazine with EtBr decreased its MIC, confirming the functionality of the QacA/B efflux pump. Biophysical and predictive analyses revealed that ortho-eugenol has lipophilic properties and moderate metabolic stability, making it suitable for oral bioavailability with low central nervous system toxicity. However, a structural vulnerability related to the isolated alkene group was identified, indicating potential epoxidation during phase I metabolism, warranting controlled dosing. These findings highlight the potential of ortho-eugenol as a β-lactamase inhibitor.