A study of the minimum inhibitory concentration and mode of action of oregano essential oil, thymol and carvacrol

TLDR

Knowledge of the inhibition extent and mode of plant‑derived compounds can aid the use of natural preservatives in foods, as carvacrol–thymol combinations achieve oregano essential oil’s potency with less flavor impact. The study aimed to determine the MIC of oregano essential oil, thymol, and carvacrol against *Pseudomonas aeruginosa* and *Staphylococcus aureus*, and to investigate their mechanisms of action. Using a 100‑well microtitre plate, the authors measured turbidimetric growth across a range of concentrations for each compound and their mixtures to generate inhibition profiles. The analysis showed that carvacrol–thymol mixtures act additively, with low concentrations increasing cell permeability, disrupting pH gradients, and causing ion leakage, thereby producing membrane damage that underlies oregano essential oil’s overall inhibition.

Abstract

Aims: The minimum inhibitory concentration (MIC) of oregano essential oil (OEO) and two of its principle components, i.e. thymol and carvacrol, against Pseudomonas aeruginosa and Staphylococcus aureus was assessed by using an innovative technique. The mechanism of action of the above substances was also investigated. Methods and Results: The applied technique uses 100‐well microtitre plate and collects turbidimetric growth data. To produce the inhibition profiles, a wide range of concentrations were tested for each of the three compounds, as well as for carvacrol–thymol mixtures. Following a specific mathematical analysis of the observed inhibition profiles from all compounds, it was suggested that mixtures of carvacrol and thymol gave an additive effect and that the overall inhibition by OEO can be attributed mainly to the additive antimicrobial action of these two compounds. Addition of low amounts of each additive: (a) increased permeability of cells to the nuclear stain EB, (b) dissipated pH gradients as indicated by the CFDA‐SE fluorescent probe irrespective of glucose availability and (c) caused leakage of inorganic ions. Conclusions: Mixing carvacrol and thymol at proper amounts may exert the total inhibition that is evident by oregano essential oil. Such inhibition is due to damage in membrane integrity, which further affects pH homeostasis and equilibrium of inorganic ions. Significance and Impact of the Study: The knowledge of extent and mode of inhibition of specific compounds, which are present in plant extracts, may contribute to the successful application of such natural preservatives in foods, since certain combinations of carvacrol–thymol provide as high inhibition as oregano essential oil with a smaller flavour impact.