Synergistic bactericidal mechanism of carvacrol and ε-polylysine against Staphylococcus aureus: dysregulation of cellular homeostasis and metabolism

Abstract

This study aims to explore the synergistic bactericidal interaction of carvacrol (CAR) combined with ε-polylysine (ε-PL) against Staphylococcus aureus and its underlying mechanism. The combination of CAR (12.50 µg mL−1) and ε-PL (7.66 µg mL−1) exhibited synergistic bactericidal and antibiofilm activities, primarily attributed to combined destruction of the bacterial cell membrane, as evidenced by increased membrane permeability, time-dependent depolarization, and reduced cell surface hydrophobicity after 3 h of treatment. Membrane damage facilitated intracellular accumulation of CAR, which rapidly triggered energy depletion and early metabolic disturbances. Using metabolomics, the combination after 0.5 h was found to primarily perturb three key pathways, including tryptophan metabolism, cofactor biosynthesis (thiamine, biotin, nicotinamide), and nucleotide metabolism. These early metabolic changes, along with ATP decline, occurred prior to full membrane depolarization, oxidative stress, and pH imbalance at 3 h. Ultimately, the synergy caused irreversible energy depletion, membrane disruption, and severe oxidative damage. Furthermore, the combination reduced S. aureus counts by approximately 1.0 log10 CFU g−1 in raw beef under refrigeration, supporting its practical potential. This study for the first time constructs a complete temporal mechanism chain from initial membrane damage and rapid metabolic disorder to downstream cellular dysfunction for the CAR-ε-PL synergy, providing a novel theoretical basis for developing natural antimicrobial strategies in food preservation.