In vivo Anti-fungal Activity of the Essential Oil Fraction from Thymus Species and in vitro Synergism with Clotrimazole

  • Published : 2007.09.30

Abstract

The antifungal activity of the essential oil fraction from Thymus magus, and its major component thymol, against Candida albicans was investigated in vitro and in vivo. The combined effects of the oils and clotrimazole, a commonly used antifungal drug for treatment of external candidiasis, were evaluated in this study. In experimental vaginal candidiasis the essential oil fraction of T. magnus resulted in relatively milder inhibition of fungal growth following the inoculation of test mice compared to clotrimazole. However, new fungal growth was not detected up to 12 days after cessation of treatment. In contrast, in a similar experiment using clotrimazole, C. albicans was detected in the $12^{th}$ day post-treatment with the sample. This result indicates that T. magnus oil could be a promising drug to control vaginal candidiasis. In checkerboard titer tests, the combination of clotrimazole with the essential oil fraction of T. magus or T. quinquecostatus resulted in significant synergism, with FIC indices between 0.14 and 0.27 against C. albicans, while clotrimazole combined with thymol, the major component of these oils, produced only an additive effect, with FIC indices ranging between 0.50 and 1.00. Thus, the prominent synergistic effects of clotrimazole combined with T. magus essential oil indicate that these compounds may be an effective treatment for C. albicans infections.

Keywords

References

  1. Canuto, M.M. and Rodero, F.G., Antifungal drug resistance to azoles and polyenes. The Lancet Infect. Dis. 2, 550-563 (2002) https://doi.org/10.1016/S1473-3099(02)00371-7
  2. Davidson, P.M. and Parish. M.E., Methods for testing the efficacy of food antimicrobials. Food Technol. 43, 148-155 (1989)
  3. Groll, A. H. and Walsh, T. J., Uncommon opportunistic fungi: new nosocomial threats. Clin. Microbiol. Infect. 2, 8-24 (2001)
  4. Heimark, L., Shipkova, P., Greene, J., Munayyer, H., Yarosh-Tomaine, T., DiDomenico, B., Hare, R., and Pramanik, B.N., Mechanism of azole antifungal activity as determined by liquid chromatographic/mass spectrometric monitoring of ergosterol biosynthesis. J. Mass Spectrom. 37(3): 265-269 (2002) https://doi.org/10.1002/jms.280
  5. Keele, D.J., Crank, C.W., Ernst, E.J., and Klepser, M.E., Evaluation of amphotericin B and voriconazole in combination against fluconazole susceptible and resistant Candida isolates using time-kill methodology. J. Infec.t Dis. Pharmacother. 6, 49-63 (2003) https://doi.org/10.1300/J100v06n01_03
  6. Keele, D.J., DeLallo, V.C., Lewis, R.E., Ernst, E.J., and Klepser, M.E., Evaluation of amphotericin and flucytosine in combination against Candida albicans and Cryoptococcus neoformans using time-kill methodology. Diagn. Microbiol. Infect. Dis. 41, 121-126 (2001) https://doi.org/10.1016/S0732-8893(01)00297-8
  7. Kim, Y.H., Lee, J.C., and Choi, Y.H., Essential oils of Thymus quinquecostatus Celakov. and Thymus magnus Nakai. Korean J. Medicina.l Crop Sci. 2, 234-240 (1994)
  8. Kontoyiannis, D.P. and Lewis, R.E., Antifungal drug resistance of pathogenic fungi. The Lancet 359, 1135-1144 (2002) https://doi.org/10.1016/S0140-6736(02)08162-X
  9. Lee, S.H. and Kim, C.J., Selective combination effect of anethole to antifungal activities of miconazole and amphotericin B. Yakhak Hoeji, 43, 228-232 (1999)
  10. Lim, S. and Shin, S., Synergism in antifungal activity against Candida and richophyton species in combination with the essential oil of Coriandrum sativum L. and antibiotics. Natural Product Sciences 13, 85-89 (2007)
  11. Marchetti, O., Moreillon, P., Glauser, M.P., Bille, J., and Sanglard, D., Potent synergism of the combination of fluconazole and cyclosporine in Candida albicans. Antimicrob. Agents Chemother. 44, 2373-2381 (2000) https://doi.org/10.1128/AAC.44.9.2373-2381.2000
  12. Metzger, S. and Hoffmann, H., Fluconazole-resistant Candida species from HIV-infected patients with recurrent Candida stomatitis: Cross resistance to itraconazole and ketoconazole. Mycoses 40, 56-63 (1997) https://doi.org/10.1111/j.1439-0507.1997.tb00543.x
  13. Shin, S. and Kim, J.H., Antifungal activities of essential oils from Thymus quinquecostatus and T. magnus, Planta Med. 70, 1087-1090 (2004) https://doi.org/10.1055/s-2004-832653
  14. Shin, S. and Pyun, M.-S., Anti-Candida effects of estragole in combination with ketoconazole of amphotericin. Phytotherapy Res. 18, 827-830 (2004) https://doi.org/10.1002/ptr.1573
  15. Singh N., Trends in the epidemiology of opportunistic fungal infections predisposing factors and the impact of antimicrobial use practices. Clin. Infect. Dis. 33, 1692-1696 (2001) https://doi.org/10.1086/323895
  16. Sobel, J.D., Ohmit, S.E, Schuman, P., Klein, R.S., Mayer, K., Duerr, A., Vazquez, J.A., and Rampalo, A., The evolution of Candida species and fluconazole susceptibility among oral and vaginal isolates recovered from human immunodeficiency virus (HIV)-seropositive and at-risk HIV-seronegative women. J. Infect. Dis. 183(2), 286-293 (2001) https://doi.org/10.1086/317936
  17. Suresh, B., Sriram, S., Dhanaraj, S.A., Elango, K., and Chinnaswamy, K., Anticandidal activity of Santolina chamaecyparissus volatile oil. J. Ethnopharmacol. 55, 151-159 (1997) https://doi.org/10.1016/S0378-8741(96)01490-0
  18. White, R.L., Burgess, D.S., Manduru, M. and Bosso, J.A., Comparison of three diffenent in vitro methods of detecting synergy: time-kill, checkerboard, and E test. Antimicrob. Agents Chemother. 40, 1914- 1918 (1996)
  19. Zarzuelo, A. and Crespo, E.T., The medicinal and non-medicinal uses of thyme. In: Thyme-the genus Thymus. Stahl-Biskup E., Saez, F., editors. London, New York: Taylor & Francis, p. 263-292 (2003)
  20. Zhang, J.-D., Xu, Z., Caoa, Y.-B., Chenb, H.-S., Yan, L., Ana, M.-M., Gaoa, P.-H., Wang, Y., Jia, X.M., and Jiang, Y.-Y., Antifungal activities and action mechanisms of compounds from Tribulus terrestris L. J. Ethnopharmaco. 103, 76-84 (2006) https://doi.org/10.1016/j.jep.2005.07.006