In vitro Antibacterial and Antifungal Activity of a Skin Ointment and Its Active Pharmaceutical Ingredients Larch Turpentine, Turpentine Oil, and Eucalyptus Oil

Abstract

Introduction: Turpentine derivatives and eucalyptus oil are herbal substances traditionally used to treat various skin infections. Limited non-clinical data suggest they exert an immunological activity, but only scant information exists on their antibiotic effects. This in vitro study has been carried out to investigate the antibacterial and antifungal activity of a marketed skin ointment; its active pharmaceutical ingredients larch turpentine, eucalyptus oil, and turpentine oil; and their mixture, against bacteria and yeasts commonly present on the skin and causing skin infections.

Methods: The antibiotic activity was tested using the drop dilution assay on the Gram-positive bacteria Staphylococcus aureus (wild type), a methicillin-resistant S. aureus strain, S. epidermidis, S. haemolyticus, Streptococcus pyogenes, the Gram-negative Pseudomonas aeruginosa, and the yeasts Candida albicans and C. tropicalis.

Results: The ointment exerts a strong inhibitory effect on all Gram-positive bacteria at a concentration of 5 g/100 mL in the Müller-Hinton medium. It also has inhibiting effect on both Candida species but does not inhibit P. aeruginosa growth. As for the single active pharmaceutical ingredients, larch turpentine was the most active substance. The mixture of the three ingredients, in the concentrations used in the ointment, had a higher antibiotic effect than any of the individual ingredients studied, suggesting at least an additive activity.

Conclusions: Our study has shown that the herbal ingredients and their combination exert antimicrobial activities, especially against Gram-positive bacteria, that justify their use in the treatment of skin infections.

Keywords: Antibiotic susceptibility testing; Drop test; Essential oils; Methicillin-resistant Staphylococcus aureus; Pull ointment; Turpentine.

Fig. 1.

Antibacterial activity of Müller-Hinton agar (MHA), MHA supplemented with Vaseline, and Fucidin. Graphically, the more a mean of the 8 replicates (represented by a blue or red dot) is farther right, the larger is the inhibitory effect of the compound on the tested strains. Red dots indicate that the strains have been inhibited in their growth and blue dots indicate no inhibition. The scale from 0 to 6 refers to the log of the colony-forming unit (CFU) concentrations, thus indicating indirectly the level of inhibition, i.e., at which concentration the microorganism can grow, with 0: no inhibition (the microorganisms grew in all drops from 10 to 10⁶ CFU/mL) and 6: maximum inhibition (no growth observed at any concentration). Biologically significant inhibition was arbitrarily defined as no or reduced growth at a concentration of 10³ CFU/mL (log 10 value of 3, dashed line).

Fig. 2.

Antibacterial and antifungal activity of the ointment (final concentration in MHA: 2.5% and 5%) and of the mixture of its three active pharmaceutical ingredients (larch turpentine, turpentine oil, and eucalyptus oil). Final concentration of the mixture in MHA: 6.9% and 13.8%). For additional details, see Fig. 1.

Fig. 3.

Individual antibacterial and antifungal activity of larch turpentine, turpentine oil, and eucalyptus oil. The second highest concentrations are those used in the formulation of the commercial ointment. For additional details, see Fig. 1.