Amoebicidal and Amoebistatic Effects of Artemisia argyi Methanolic Extracts on Acanthamoeba castellanii Trophozoites and Cysts



The present study aimed to investigate the amoebicidal and amoebistatic efects of Artemisia argyi leaf methanolic extract by testing the effects on trophozoites and on cysts. We also determined cytotoxic effect, enzymatic and non-enzymatic antioxidant activities, total phenolic, lavonoid and antioxidative contents of A. argyi.


A. argyi was harvested from various geographic sites in Ordu province in Turkey. The fresh leaves were subjected to methanolic extraction. In 100 μl culture, different concentrations of A. argyi methanolic extract (in quantities from 1.2, 2.3, 4.7, 9.4, 18.7, 37.4, 74.8 mg/ml) and the same volume of trophozoite/cyst suspension were mixed for the determination of the amoebicidal activity of the plant extract. Human bronchial epithelial cells were treated with the same concentrations of Artemisia extracts to determine cytotoxic potential.


Total phenolic and lavonoid contents of the extract were calculated as 261 mg gallic acid/g dry extract and 29 mg quercetin/g dry extract, respectively. Total antioxidant activity was also calculated as 367 mg ascorbic acid/g dry extract. The growth of trophozoites stopped in A. argyi methanolic extract with 50% inhibitory concentrations (IC50)/8 h for 37.4 mg/ ml and 74.8 mg/ml extract solution and had stronger amoebicidal activity on the cysts with IC50/72 h. Artemisia showed stronger inhibitory effects on bronchial epithelial cells at the concentrations of 9.4, 18.7, 37.4 and 74.8 mg/ml.


The study indicated that A. argyi leaf extract has cytotoxic and anti-amoebic activities.

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  1. 1.

    Akin Polat Z.,Vural A.,Tepe B., Cetin A. 2007. In vitro amoebicidal activity of four Allium species on Acanthamoeba castellani and their cytotoxic potentials on corneal cells. Parasitology Research, 101, 397–402.

    Article  Google Scholar 

  2. 2.

    Arvouet-Grand A., Vennat B., Pourrat A., Legret P. 1994. Standardization of propolis extract and identification of principal constituents. Journal DePharmacie De Belgique, 49, 462–468. PMID:7884635

    CAS  Google Scholar 

  3. 3.

    Badria F.A., Hetta M.H., Sarhan R.M., Ezz El-Din M.H. 2014. Lethal effects of Helianthemum lippii (L.) on Acanthamoeba castellanii cysts in vitro. Korean Journal Parasitology, 52(3), 243–9.

    CAS  Article  Google Scholar 

  4. 4.

    Barney J.N., DiTommase A. 2003.The biology of Canadian weeds. 118. Artemisia vulgaris L. Canadian Journal of Plant Science, 83, 205–215.

    Article  Google Scholar 

  5. 5.

    Beauchamp C., Fridovich I. 1971. Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Analytical Biochemistry, 44, 276–287.

    CAS  Article  Google Scholar 

  6. 6.

    Benzie I.F.F., Strain J.J. 1996. The ferric reducing ability of plasma (FRAP) as a measure of “antioxidant power”: the FRAP assay. Analytical Biochemistry, 239,70–77.

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Carvalho I.S., Cavaco T., Brodelius M. 2011. Phenolic composition and antioxidant capacity of six Artemisia species. Industrial Crops and Products, 33, 382–388.

    CAS  Article  Google Scholar 

  8. 8.

    Chevallier A. (Ed.). 1996. The encyclopedia of medicinal plants. Dorling Kindersley, London.

    Google Scholar 

  9. 9.

    Degerli S., Berk S., Malatyali E., Tepe B. 2012a. Screening of the in vitro amoebicidal activities of Pastinaca armenea (Fisch & C.A. Mey) and Inula oculus-christi (L.) on Acanthamoeba castellanii cysts and trophozoites. Parasitology Research, 110, 565–570.

    Article  PubMed  Google Scholar 

  10. 10.

    Degerli S., Tepe B., Celiksoz A., Berk S., Malatyali E. 2012b. In vitro amoebicidal activity of Origanum syriacum and Origanumla evigatum on Acanthamoeba castellanii cysts and trophozoites. Experimental Parasitology, 131, 20–24.

    Article  PubMed  Google Scholar 

  11. 11.

    Derda M., Hadaś E.,Thiem B. 2009. Plant extracts as natural amoebicidal agents. Parasitology Research, 104(3),705–708.

    Article  PubMed  Google Scholar 

  12. 12.

    Derda M., Hadaś E., Cholewiński M., Skrzypczak Ł., Grzondziel A., Wojtkowiak-Giera A. 2016. Artemisia annua L. as a plant with potential use in the treatment of acanthamoebiasis. Parasitology Research, 115(4), 1635–1639.

    Article  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Dinis T.C.P., Madeira V.M.C., Almeida L.M. 1994. Action of phenolic derivatives (Acetaminophen, Salicylate, and 5-Aminosalicylate) as inhibitors of membrane lipid peroxidation and as peroxyl radical scavengers. Archives of Biochemistry and Biophysics, 315, 161–169.

    CAS  Article  PubMed  Google Scholar 

  14. 14.

    El-Sayed N.M., Ismail K.A., Ahmed S.A., Hetta M.H. 2012. In vitro amoebicidal activity of ethanol extracts of Arachis hypogaea L., Curcuma longa L. and Pancratium maritimum L. on Acanthamoeba castellanii cysts. Parasitology Research, 110(5), 1985–1992

    Article  Google Scholar 

  15. 15.

    Farnsworth N. R., Bunyapraphatsara N. (Ed.). 1992. Thai medicinal plants: recommended for primary health care system. Medicinal Plant Information Center, Bangkok, Thailand, pp.409

    Google Scholar 

  16. 16.

    Firestone G.L., Sundar S.N. 2009. Anticancer activities of artemisinin and its bioactive derivatives. Expert Reviews in Molecular Medicine, 11, e32.

    Article  PubMed  Google Scholar 

  17. 17.

    Fu R., Zhang Y., Guo Y., Liu F., Chen F. 2014. Determination of phenolic contents and antioxidant activities of extracts of Jatropha curcas L. seed shell, a by-product, a new source of natural antioxidant. Industrial Crops and Products, 58, 265–70.

    CAS  Article  Google Scholar 

  18. 18.

    Ge Y. B., Wang Z. G., Xiong Y., Huang X.J., Mei Z.N. 2016. Anti-inflammatory and blood stasis activities of essential oil extracted from Artemisia argyi leaf in animals. Journal of Natural Medicines, 70 (3), 531–538.

    CAS  Article  PubMed  Google Scholar 

  19. 19.

    Gordanian B., Behbahani M., Carapetian J., Fazilati M. 2014. In vitro evaluation of cytotoxic activity of flower, leaf, stem and root extracts of five Artemisia species. Research in Pharmaceutical Sciences, 9 (2),91–96. PMID: 25657777.

    CAS  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Huang H.C., Wang H.F., Yih K.H., Chang L.Z., Chang T.M. 2012. Dual bioactivities of essential oil extracted from the leaves of Artemisia argyi as an antimelanogenic versus antioxidant agent and chemical composition analysis by GC/MS. International Journal of Molecular Sciences, 13, 14679–14697.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  21. 21.

    Kanatt S. R., Chawla S.P., Sharma A. 2014. Antioxidant and radio-protective activities of lemon grass and staranise extracts. Food Bioscience, 6 (24), 24–30.

    CAS  Article  Google Scholar 

  22. 22.

    Kao P.M., Hsu B.M., Chen N.H., Huang K.H., Huang S.W., King K.L., Chiu Y.C. 2012. Isolation and identification of Acanthamoeba species from thermal spring environments in southern Taiwan. Experimental Parasitology, 130(4), 354–358.

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Karimi A., Hadian J., Farzaneh M., Khadivi-Khub A. 2015. Evaluation of genetic variability, rust resistance and marker-detection in cultivated Artemisia dracunculus from Iran. Gene, 554(2), 224–232.

    CAS  Article  PubMed  Google Scholar 

  24. 24.

    Keyhani J., Keyhani E. Anti-oxidative stress enyzmes in golden chanterelle (Cantharellus cibarius), 2012. In: (Eds A. Mendez-Vilas), Microbes in Applied Research. Singapore: World Scientific, pp. 23–27.

    Google Scholar 

  25. 25.

    Khan N.A. (Ed.). 2009. Acanthamoeba biology and pathogenesis, Caister Academic Press, Norfolk, UK, pp. 85.

    Google Scholar 

  26. 26.

    Kim J.K., Shin E.C., Lim H.J., Choi S.J., Kim C.R., Suh S.H., 2015. Characterization of nutritional composition, antioxidative capacity, and sensory attributes of seomaemug wort, a native korean variety of Artemisia argyi H. Lév. & Vaniot. Journal of Analytical Methods in Chemistry, 1–9.

  27. 27.

    Kitagawa K., Nakamura T., Takahashi N., Oikawa Y., Ikeda T. 2003. A novel combination treatment of chlorohexidine gluconate, natamycin (pimaricin) and debridement for a Acanthamoeba keratitis. Japanese Journal of Ophthalmology, 47, 616–617.

    CAS  Article  PubMed  Google Scholar 

  28. 28.

    Koloren O., Koloren Z., Eker S. 2016. Molecular phylogeny of Artemisia species based on internal transcribed spacer (ITS) of 18S-26S rDNA in Ordu Province of Turkey. Biotechnology & Biotechnological Equipment, 30, 929–934.

    CAS  Article  Google Scholar 

  29. 29.

    Lee J. H., Lee J.M., Lee S.H., Kim Y.G., Lee S., Kim S. et al. 2015. Comparison of artemisin in content and antioxidant activity from various organs of Artemisia species. Horticulture Environment and Biotechnology, 56, 697–703.

    CAS  Article  Google Scholar 

  30. 30.

    Magana M.L., Fernandez-Diez J., Magana M. 2008. Cutaneous amebiasis in pediatrics. Archives of Dermatology, 144(10), 1369–72.

    Article  PubMed  Google Scholar 

  31. 31.

    Malatyali E., Tepe B., Değerli S., Berk S., Akpulat H.A. 2012. In vitro amoebicidal activity of four Peucedanum species on Acanthamoeba castellanii cysts and trophozoites. Parasitology Research, 110, 167–174.

    Article  PubMed  Google Scholar 

  32. 32.

    Morrison A.O., Morris R., Shannon A., Lauer S.R., Guarner J., Kraft C.S. 2016. Disseminated Acanthamoeba Infection Presenting With Cutaneous Lesions in an Immunocompromised Patient: A Case Report, Review of Histomorphologic Findings, and Potential Diagnostic Pitfalls. American Journal of Clinical Pathology, 145(2), 266–70.

    Article  PubMed  Google Scholar 

  33. 33.

    Mosmann T. 1983. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods,65 (1–2), 55–63.

    CAS  Article  PubMed  Google Scholar 

  34. 34.

    Nakisah M.A., Ida Muryany M.Y., Fatimah H., Nor Fadilah R., Zalilawati M.R., Khamsah S. 2012. Anti-amoebic properties of a Malaysian marine sponge Aaptos sp. on Acanthamoeba castellanii. World Journal of Microbiology & Biotechnology, 28, 1237–1244.

    CAS  Article  Google Scholar 

  35. 35.

    Nawab A., Yunus M., Mahdi A., Gupta S. 2011. Evaluation of anticancer properties of medicinal plants from the Indian sub-continent. Molecular and Cellular Pharmacology, 3, 21–29.

    Article  Google Scholar 

  36. 36.

    Oliaee D., Boroushaki T.M., Oliaee N., Ghorbani A. 2014. Evaluation of cytotoxicity and antifertility effect of Artemisia kopetdaghensis. Advances in Pharmaceutical Sciences, 5 pages.

    Article  Google Scholar 

  37. 37.

    Oyaizu M. 1986. Studies on products of browning reaction-antioxidative activities of products of browning reaction prepared from glucosamine. Journal of Nutrition, 44,307–315.

    CAS  Article  Google Scholar 

  38. 38.

    Piao G.C., Li Y.X., Yuan H.D., Jin G.Z. 2012. Cytotoxic fraction from Artemisia sacrorum against three human cancer cell lines and separation and identification of its compounds. Natural Product Research, 26 (16), 1483–1491.

    CAS  Article  PubMed  Google Scholar 

  39. 39.

    Saleh A.M., Aljada A., Rizvi S.A., Nasr A., Alaskar A.S., Williams J.D. 2014. In vitro cytotoxicity of Artemisia vulgaris L. essential oil is mediated by a mitochondria-dependent apoptosis in HL-60 leukemic cell line. BMC Complementary and Alternative Medicine, 14, 226.

    Article  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Sanchez-Moreno C., Larrauri J. A., Saura-Calixto F. 1998. A procedure to measure the antiradical efficiency of polyphenols. Journal of the Science of Food and Agriculture, 76, 270–276.

    CAS  Article  Google Scholar 

  41. 41.

    Santos I.G, Scher R., Rott M.B., Menezes L.R., Costa E.V., Cavalcanti S.C. 2016. Amebicidal activity of the essential oils of Lippia spp. (Verbenaceae) against Acanthamoeba polyphaga trophozoites. Parasitology Research, 115(2), 535–40.

    Article  PubMed  Google Scholar 

  42. 42.

    Sarath V.J., So C.S., Young D.W., Gollapudi S. 2007. Artemisia princeps var. orientalis induces apoptosis in human breast cancer MCF-7 cells. Anticancer Research, 27, 3891–3898. PMID:18225547

    PubMed  Google Scholar 

  43. 43.

    Schuster F.L, Visvesvara G.S. 2004. Free-living amoebae as opportunistic and non-opportunistic pathogens of humans and animals. International Journal of Parasitology, 34, 1001–1027.

    Article  PubMed  Google Scholar 

  44. 44.

    Seal D.V. 2003. Acanthamoeba keratitis update-incidence, molecular epidemiology and new drugs for treatment. Eye, 17, 893–905.

    CAS  Article  Google Scholar 

  45. 45.

    Seo H.J., Park K.K., Han S.S., Chung W.Y., Son M.W., Kim W.B., Surh Y.J. 2002. Inhibitory effects of the standardized extract (DA-9601) of Artemisia asiatica Nakaionphorbol ester-induced ornithine decarboxylase activity, papilloma formation, cyclooxygenase-2 expression, inducible nitric oxide synthase expression and nuclear transcription factor kappa B activation in mouse skin. International Journal of Cancer, 100 (4), 456–462.

    CAS  Article  PubMed  Google Scholar 

  46. 46.

    Sharmila K., Padma P.R. 2013. Anticancer activity of Artemisia vulgaris on hepatocellular carcinoma (HepG2) cells. International Journal of Pharmaceutical Sciences, 5 (3), 479–483.

    Google Scholar 

  47. 47.

    Shoaib H.M, Muazzam A.G, Mir A, Jung S.Y, Matin A. 2013. Evaluation of inhibitory potential of some selective methanolic plants extracts on biological characteristics of Acanthamoeba castellanii using human corneal epithelial cells in vitro. Parasitology Research, 112(3),1179–88.

    Article  PubMed  Google Scholar 

  48. 48.

    Sifaoui I., López-Arencibia A., Martín-Navarro C.M., Chammem N., Mejri M., Lorenzo-Morales J., Abderabba M., Piñero J.E. 2013. Activity assessment of Tunisian olive leaf extracts against the trophozoite stage of Acanthamoeba. Parasitology Research, 112, 2825–2829.

    Article  PubMed  Google Scholar 

  49. 49.

    Silva L., Pezzini B.R., Soares L. 2015. Spectrophotometric determination of the total flavonoid content in Ocimum basilicum L. (Lamiaceae) leaves. Pharmacognosy Magazine, 11, 96–101.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Singleton V.L., Rossi J.A. 1965. Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. American Journal of Enology and Viticulture, 16,144–158.

    CAS  Google Scholar 

  51. 51.

    Sun L., Zhang J., Lu X., Zhang L., Zhang Y. 2011. Evaluation to the antioxidant activity of total flavonoids extract from persimmon (Diospyros kaki L.) leaves. Food and Chemical Toxicology, 49 (10), 2689–2696.

    CAS  Article  PubMed  Google Scholar 

  52. 52.

    Taherkhani M. 2015. Chemical constituents and in vitro anticancer, cytotoxic, mutagenic and antimutagenic activities of Artemisia diffusa. Pharmaceutical Chemistry Journal, 48 (11), 727–732.

    CAS  Article  Google Scholar 

  53. 53.

    Taherkhani M. 2017. Chemical constituents, total phenolic content, antimicrobial, antioxidant and radical scavenging properties, chelating ability, tyrosinase inhibition and in vitro cytotoxic effects of Artemisia aucheri Herbs. Pharmaceutical Chemistry Journal, 50 (11), 736–745.

    CAS  Article  Google Scholar 

  54. 54.

    Tepe B., Malatyali E., Degerli S., Berk S. 2012. In vitro amoebicidal activities of Teucrium polium and T. chamaedrys on Acanthamoeba castellanii trophozoites and cysts. Parasitology Research, 110 (5),1773–1778.

    Article  PubMed  Google Scholar 

  55. 55.

    Wanachiwanawin D., Booranapong W., Kosrirukvongs P. 2012. Clinical features of Acanthamoeba keratitis in contact lens wearers and nonwearers. Southeast Asian J Trop Med Public Health, 43(3), 549–556.

    PubMed  Google Scholar 

  56. 56.

    Watson L.E., Bates P.L., Evans T.M., Unwin M.M., Estes J.R. 2002. Molecular phylogeny of subtribe Artemisiinae (Asteraceae), including Artemisia and its allied and segregate genera. BMC Evolutionary Biology, 2,17.

    Article  PubMed  PubMed Central  Google Scholar 

  57. 57.

    Yoder J.S., Verani J., Heidman N., Hoppe-Bauer J., Alfonso E.C., Miller D., Jones D.B., Bruckner D., Langston R., Jeng B.H., Joslin C.E., Tu E., Colby K., Vetter E., Ritterband D., Mathers W., Kowalski R.P., Acharya N.R., Limaye AP., Leiter C., Roy S., Lorick S., Roberts J., Beach M.J. 2012. Acanthamoeba keratitis: the persistence of cases following a multistate outbreak. Ophthalmic Epidemiology, 19(4), 221–225.

    Article  PubMed  Google Scholar 

  58. 58.

    Zhang L.B., Lv J.L., Chen H.L., Yan X.Q., Duan J.A. 2013. Chemical constituents from Artemisia argyi and their chemotaxonomic significance. Biochemical Systematics and Ecology, 50,455–458.

    CAS  Article  Google Scholar 

  59. 59.

    Zhao F., Shi B., Sun D., Chen H., Tong M., Zhang P. 2016. Effects of dietary supplementation of Artemisia argyi aqueous extract on antioxidant indexes of small intestine in broilers. Animal Nutrition, 2 (3),198–203.

    Article  PubMed  PubMed Central  Google Scholar 

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Kolören, O., Kolören, Z., Şekeroğlu, Z.A. et al. Amoebicidal and Amoebistatic Effects of Artemisia argyi Methanolic Extracts on Acanthamoeba castellanii Trophozoites and Cysts. Acta Parasit. 64, 63–70 (2019).

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  • Artemisia argyi
  • Phytotoxic activity
  • Acanthamoeba castellanii
  • Amoebicidal activity
  • Cytotoxic potential
  • Antioxidant activity