Mechanisms of antibacterial action of three monoterpenes

Abstract

In the present paper, we report the antimicrobial efficacy of three monoterpenes [linalyl acetate, (+)menthol, and thymol] against the gram-positive bacterium Staphylococcus aureus and the gram-negative bacterium Escherichia coli. For a better understanding of their mechanisms of action, the capability of these three monoterpenes to damage biomembranes was evaluated by monitoring the release, following exposure to the compounds under study, of the water-soluble fluorescent marker carboxyfluorescein from unilamellar vesicles with different lipidic compositions (phosphatidylcholine, phosphatidylcholine/phosphatidylserine [9:1], phosphatidylcholine/stearylamine [9:1], and phosphatidylglycerol/cardiolipin [9:1]). Furthermore, the interaction of the terpenes tested with dimyristoylphosphatidylcholine multilamellar vesicles as model membranes was monitored by means of differential scanning calorimetry. Finally, the results were related to the relative lipophilicity and water solubility of the compounds examined. Taken together, our findings lead us to speculate that the antimicrobial effect of (+)menthol, thymol, and linalyl acetate may result, at least partially, from a perturbation of the lipid fraction of microorganism plasma membrane, resulting in alterations of membrane permeability and in leakage of intracellular materials. Besides being related to physicochemical characteristics of the drugs (such as lipophilicity and water solubility), this effect seems to be dependent on lipid composition and net surface charge of microbial membranes. Furthermore, the drugs might cross the cell membranes, penetrating into the interior of the cell and interacting with intracellular sites critical for antibacterial activity.

FIG. 1.

Effect of thymol challenge on CF release from CF-encapsulated large unilamellar vesicles with different lipid compositions.

FIG. 2.

Effect of (+)menthol challenge on CF release from CF-encapsulated large unilamellar vesicles with different lipid compositions.

FIG. 3.

Effect of linalyl acetate challenge on CF release from CF-encapsulated large unilamellar vesicles with different lipid compositions.

FIG. 4.

Effect of (+)menthol, thymol, and linalyl acetate, at different molar fractions, on the transitional temperature of dimyristoylphosphatidylcholine multilamellar liposomes.

FIG. 5.

Effect of (+)menthol, thymol, and linalyl acetate (X = 0.09) on T_m values of dimyristoylphosphatidylcholine MLVs evaluated by differential scanning calorimetry (full signs). Empty signs at t_∞ refer to samples prepared by direct interaction between the examined compounds and MLVs, at the same organic fraction, by organic solvent dissolution and MLV preparation.