This study investigates the antimicrobial mechanisms of three monoterpenes—linalyl acetate, (+)menthol, and thymol—against Gram-positive *Staphylococcus aureus* and Gram-negative *Escherichia coli*. The research focuses on how these compounds damage bacterial membranes, which is crucial for their antibacterial activity. The study evaluates the ability of these monoterpenes to disrupt biomembranes by monitoring the release of the fluorescent marker carboxyfluorescein (CF) from large unilamellar vesicles (LUVs) with different lipid compositions. The results indicate that the antimicrobial effect of these compounds is largely due to membrane disruption, leading to increased permeability and leakage of intracellular materials. The study also examines the interaction of these monoterpenes with model membranes using differential scanning calorimetry (DSC), revealing that they can lower the transition temperature of phosphatidylcholine (PC) membranes and alter their thermotropic behavior. These findings suggest that the monoterpenes interact with lipid membranes, possibly by introducing lipophilic molecules that disrupt the ordered structure of the lipid bilayer. The antimicrobial activity of the monoterpenes is also related to their physicochemical properties, such as lipophilicity and water solubility. Thymol and (+)menthol showed stronger antimicrobial activity against *S. aureus* compared to linalyl acetate, likely due to their higher lipophilicity and water solubility, which enable them to penetrate the aqueous extracellular medium and interact with lipid membranes. Additionally, the lipid composition and net surface charge of bacterial membranes influence the susceptibility of the microorganisms to these compounds. The study concludes that the antimicrobial effect of these monoterpenes is at least partially due to their ability to penetrate lipid membranes and disrupt the lipid fraction of the plasma membrane, leading to changes in membrane permeability and leakage of intracellular materials. The findings highlight the importance of lipid composition and physicochemical properties in determining the antimicrobial activity of monoterpenes.This study investigates the antimicrobial mechanisms of three monoterpenes—linalyl acetate, (+)menthol, and thymol—against Gram-positive *Staphylococcus aureus* and Gram-negative *Escherichia coli*. The research focuses on how these compounds damage bacterial membranes, which is crucial for their antibacterial activity. The study evaluates the ability of these monoterpenes to disrupt biomembranes by monitoring the release of the fluorescent marker carboxyfluorescein (CF) from large unilamellar vesicles (LUVs) with different lipid compositions. The results indicate that the antimicrobial effect of these compounds is largely due to membrane disruption, leading to increased permeability and leakage of intracellular materials. The study also examines the interaction of these monoterpenes with model membranes using differential scanning calorimetry (DSC), revealing that they can lower the transition temperature of phosphatidylcholine (PC) membranes and alter their thermotropic behavior. These findings suggest that the monoterpenes interact with lipid membranes, possibly by introducing lipophilic molecules that disrupt the ordered structure of the lipid bilayer. The antimicrobial activity of the monoterpenes is also related to their physicochemical properties, such as lipophilicity and water solubility. Thymol and (+)menthol showed stronger antimicrobial activity against *S. aureus* compared to linalyl acetate, likely due to their higher lipophilicity and water solubility, which enable them to penetrate the aqueous extracellular medium and interact with lipid membranes. Additionally, the lipid composition and net surface charge of bacterial membranes influence the susceptibility of the microorganisms to these compounds. The study concludes that the antimicrobial effect of these monoterpenes is at least partially due to their ability to penetrate lipid membranes and disrupt the lipid fraction of the plasma membrane, leading to changes in membrane permeability and leakage of intracellular materials. The findings highlight the importance of lipid composition and physicochemical properties in determining the antimicrobial activity of monoterpenes.