The essential oil of Melaleuca alternifolia (tea tree) exhibits broad-spectrum antimicrobial activity. Its mode of action against the Gram-negative bacterium Escherichia coli AG100, the Gram-positive bacterium Staphylococcus aureus NCTC 8325, and the yeast Candida albicans has been investigated using a range of methods. We report that exposing these organisms to minimum inhibitory and minimum bactericidal/fungicidal concentrations of tea tree oil inhibited respiration and increased the permeability of bacterial cytoplasmic and yeast plasma membranes as indicated by uptake of propidium iodide. In the case of E. coli and Staph. aureus, tea tree oil also caused potassium ion leakage. Differences in the susceptibility of the test organisms to tea tree oil were also observed and these are interpreted in terms of variations in the rate of monoterpene penetration through cell wall and cell membrane structures. The ability of tea tree oil to disrupt the permeability barrier of cell membrane structures and the accompanying loss of chemiosmotic control is the most likely source of its lethal action at minimum inhibitory levels.

C .M . M A NN AN D J. L. M AR KH A M. 1998. A new microdilution method has been developed for determining the minimum inhibitory concentration (MIC) of oil-based compounds. The redox dye resazurin was used to determine the MIC of a sample of the essential oil of Melaleuca alternifolia (tea tree) for a range of Gram-positive and -negative bacteria. Use of 0·15% (w/v) agar as a stabilizer overcame the problem of adequate contact between the oil and the test bacteria and obviated the need to employ a chemical emulsifier. A rapid version of the assay was also developed for use as a screening method. A comparison of visual and photometric reading of the microtitre plates showed that results could be assessed without instrumentation; moreover, if the rapid assay format was used, rigorous asepsis was not necessary. Accuracy of the resazurin method was confirmed by plate counting from microwells and MIC values were compared with results obtained using an agar dilution assay. The MIC results obtained by the resazurin method were slightly lower than those obtained by agar dilution.

Research into the mode of action of the essential oil of Melaleuca alternifolia (Tea tree oil) is briefly reviewed. Its mode of action is interpreted in terms of the membrane-toxicity of its monoterpenoid components and different approaches for determining cell membrane damage are discussed.

Aims: This study compared the antimicrobial activity of Melaleuca alternifolia (tea tree) oil with that of some of its components, both individually and in two‐component combinations. Methods and Results: Minimum inhibitory concentration and time‐kill assays revealed that terpinen‐4‐ol, the principal active component of tea tree oil, was more active on its own than when present in tea tree oil. Combinations of terpinen‐4‐ol and either γ‐terpinene or p‐cymene produced similar activities to tea tree oil. Concentration‐dependent reductions in terpinen‐4‐ol activity and solubility also occurred in the presence of γ‐terpinene. Conclusions: Non‐oxygenated terpenes in tea tree oil appear to reduce terpinen‐4‐ol efficacy by lowering its aqueous solubility. Significance and Impact of the Study: These findings explain why tea tree oil can be less active in vitro than terpinen‐4‐ol alone and further suggest that the presence of a non‐aqueous phase in tea tree oil formulations may limit the microbial availability of its active components.

Pseudomonas aeruginosa is less susceptible to the antimicrobial properties of tea tree oil than many bacteria and its tolerance is considered to be due to its outer membrane. Polymyxin B nonapeptide (PMBN), which has no antibacterial action, was used to permeabilize the outer membrane. The addition of PMBN to Ps. aeruginosa NCTC 6749 markedly increased this organism’s susceptibility to tea tree oil and to its normally inert hydrocarbons, p‐cymene and γ‐terpinene.