Three formulations of lipophilic ointments containing St.John's Wort oil, suitable for dermal or vaginal application, were prepared. Their antibacterial effect has been evaluated by using the method of diffusion on blood-agar plate. Polysorbate 80 was used as a diffusion-enhancing agent. Antibacterial effect was tested on six bacterial strains: Streptococcus pyogenes (two strains), Streptococcus viridans, Micrococcus luteus ATCC9341, Moraxella catarrhalis and on Lactobacillus acidophillus. Inhibition of bacterial growth was registered for all tested strains, with exception of Lactobacillus acidophilus, which is a part of normal vaginal bacterial flora.

St John's Wort oil is widely used herbal medicine in treatment of bruises, injuries and burns, which is based on its antibacterial, anti-inflammatory and epitheliant effect (1,2). Certain vaginal disorders could be also treated by using St.John`s Wort preparations (3,4).

The most often used procedure for obtaining St. John's Wort oil is maceration of fresh flowers of Hypericum perforatum L. in oil, exposed under sunlight for several weeks. The oil extract is brilliant red with orange-red fluorescence. The first official monograph of St.John's Wort oil could be found in the first London pharmacopoeia, under the title Oleum Hyperici.

Regarding the topical application of the St.John's Wort oil, the main components of the oil extract, responsible for its therapeutic effect are hyperforins and flavonoids (5,6).

The aim of this study was to evaluate the antibacterial effect of three ointment formulations containing St. John's Wort oil.

Preparation of the ointments

Three ointment formulations containing oil extracts of Hypericum perforatum (Hyperici oleum), potentially applicable on skin or vaginal mucosa, have been prepared. Quantity of Hyperici oleum in the formulation has been varied within the range of 30-50%. The oil extract was incorporated in an ointment basis of lipophilic nature, composed of solid and liquid components.

Evaluation of the antibacterial effect

Antibacterial effect of prepared ointments was tested using the method of diffusion on a blood-agar plate. Mixtures of the ointment sample and Polysorbate 80 have been prepared prior to test application. The presence of surface active agent enables the diffusion into a hydrophilic medium. Antibacterial effect was tested on clinical and referent bacterial strains with previously established sensitivity to antibiotics. Four pathogenic and one referent bacterial strains were selected: Streptococcus pyogenes (two strains), Streptococcus viridans, Moraxella catarrhalis and Micrococcus luteus ATCC9341. Also, the effect of the ointments on Lactobacillus acidophillus, which is regularly present in the vaginal flora was evaluated. Growth inhibition zones were measured after 24h of incubation on 37°C.

All three ointment formulations containing 30%, 40% and 50% of Hyperici oleum, met the criteria for topical semisolid preparations (previously evaluated via melting point, leaking point, rheological measurements etc.) (7).

The results obtained from the microbiological tests on evaluating of the antibacterial effect were presented in table 1.

As it can be seen, an inhibition of bacterial growth was registered on all tested strains, with exception of Lactobacillus acidophillus. Having in mind that Lactobacillus acidophillus is normally present in vaginal secretion, the vaginal application of the prepared ointments will not distort the normal vaginal flora. As expected, by increasing the quantity of Hyperici oleum in ointment formulation, antibacterial effect increased as well.

It should be mentioned that the results from our previous experiments concerning the concentration of total flavonoids and antibacterial effect of different samples of Hyperici oleum (8,9), indicated that certain correlation between those two parameters exists.

The prepared ointments containing Hyperici oleum expressed antibacterial effect on Streptococcus pyogenes, Streptococcus viridans, Micrococcus luteus ATCC 9341 and Moraxella catarrhalis, while the inhibition of bacterial growth of Lactobacillus acidophillus was not observed.

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2. Šaljić J. (1975): Ointment for tretment of burns, Ger. Offen. 2,406,452 (C1.A 61K), (CA 83:197797).

3. Minkov E., Lambov N., Ovčarov R. Bantunova I. (1994): Osnovi na Biofamacijata, Venel Medik.

4. Stanković R. et al. (1998): Arh. Farm 6, 964-965.

5. Berghoefer R., Hoelzel J. (1987): Biflavonoids in Hypericum perforatum, Part 1. Isolation of I3, II8-biapigenin, Planta Med. 53(2), 216-17.

6. Maizenbacher P., Kovar K.A. (1991): Analysis and Stability of Hyperici Oleum, Planta Med.,58(4), 351-354.

7. Peeva-Naumovska V., Fredro-Kumbaradži E., (1999): Formulacija i ispituvanje na fizički parametri na masti so kantarionovo maslo, Book of papers of the 16^th Congress of Chemists and Technologist of Macedonia, 299-303.

8. Peeva-Naumovska V.,Grdanovska T., Panovski N., Fredro-Kumbaradži E. (1999): Antimikrobno deluvanje na kantarionovo maslo, Bulletin of the Macedonian Pharmaceutical Association, 1-2, 21.

9. Peeva Naumovska Vida (1999): Specijalistički trud, Farmacevtski fakultet, Skopje.