Bilayer-Modifying Potential of Limonene and its Metabolites

Abstract

Membrane protein function depends on lipid bilayer properties, which can be modified by small molecules. This may account for some of the undesired clinical effects of amphiphatic drugs that modify bilayer properties. In this study, we assessed whether D-limonene and its metabolites (perillyl alcohol, perillaldehyde, and perillic acid) alter lipid bilayer properties, as sensed by bilayer-spanning gramicidin (gA) channels, using a fluorescence assay and single-channel electrophysiology. D-limonene and its metabolites are terpenes that are found in a variety of foods, particularly citrus oils. They have been shown as potential cancer prevention and treatment agents and as antimicrobials. These terpenes are also used as solvents and flavor and fragrance agents. Given the wide range of biological functions of these hydrophobic/amphiphilic compounds, it becomes important to determine to what extent they alter bilayer properties. Using the fluorescence assay, we find that at micromolar concentrations (nominal concentrations): D-limonene decreases gA channel activity, perillyl alcohol and perillaldehyde increase activity, and perillic acid has no apparent effect. When examined using single-channel electrophysiology, each terpene increased gA channel lifetime and appearance rate. In the case of perillaldehyde and perillyl alcohol there is agreement between the two assays but not for D-limonene and perillic acid. We are exploring the basis for these differences. The changes in gA channel function (bilayer properties) were observed at low membrane concentrations (mole fraction ∼0.02-0.03), indicating that these terpenes are potent bilayer modifiers.