In Vitro Amoebicidal Activity of Imidazolium Salts Against Trophozoites

  • Laura Führich Fabres
  • Fabiany da Costa Gonçalves
  • Eliane Oliveira Salines Duarte
  • Francisco Kercher Berté
  • Débora Kélen Si lva da Conceição
  • Leonildo Alves Ferreira
  • Henri Stephan SchrekkerEmail author
  • Marilise Brittes RottEmail author
Original Paper



Several strains of the free-living genus Acanthamoeba can cause granulomatous amoebic encephalitis (GAE), a rare chronic and slowly progressive infection of the central nervous system (CNS), and Acanthamoeba keratitis (AK), a sight-threatening eye infectious disease. AK incidence has increased with the popularization of the contact lens wear and its treatment is currently limited and frequently unsuccessful. As imidazolium salts (IS), cationic imidazole derivatives, have promising antimicrobial potential.

Materials and Methods

The present study evaluated the amoebicidal activity of four IS; 1-n-hexadecyl-3-methylimidazolium methanesulfonate (C16MImMeS), chloride (C16MImCl) and bis (triluoromethylsulfonyl) imide (C16MImNTf2 ), and 1-methyl-3-n-octadecylimidazolium chloride (C18MImCl), against the Acanthamoeba castellanii (ATCC30010) environmental strain and a clinical isolate (genotype T4).


Three IS showed being lethal to 100% of the Acanthamoeba trophozoites at the minimum inhibitory concentrations of 125 and 62.5 μg/mL (C16MImMeS), 31.25 and 62.5 μg/mL (C16MImCl), and 125 and 125 μg/mL (C18MImCl) for ATCC30010 and isolate T4, respectively. C16MImNTf2 did not demonstrate amoebicidal activity. All active IS caused the hemolysis of erythrocytes. The cytotoxic effect of the IS was tested in RAW macrophages and human brain microvascular endothelial cells, which demonstrated cytotoxicity in all concentrations tested against both cell lines. As a consequence, these IS with amoebicidal activity presented low selectivity index values (SI) (SI < 1.0), demonstrating lack of parasite selectivity.


Thus, C16MImMeS, C16MImCl, and C18MImCl seem to hold greater promise as components for contact lens cleaning and disinfection solutions, instead of direct human application.


Acanthamoeba Imidazolium ionic liquids Clinical and environmental isolates Keratitis Amoebic encephalitis In vitro cytotoxicity 



This study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES)—Finance Code 001; and the Conselho Nacional de Desenvolvimento Científico e Tecnológico—Brasil (CNPq). Dr. Julio Scharfstein (Instituto de Biofísica Carlos Chagas Filho, UFRJ) is kindly acknowledged for the donation of human brain microvascular endothelial cells.

Supplementary material

11686_2019_161_MOESM1_ESM.docx (723 kb)
Supplementary file1 (DOCX 722 kb)


  1. 1.
    Abjani F, Khan NA, Jung SY, Siddiqui R (2017) Status of the effectiveness of contact lens disinfectants in Malaysia against keratitis-causing pathogens. Exp Parasitol 183:187–193. CrossRefPubMedGoogle Scholar
  2. 2.
    Anderson EB, Long TE (2010) Imidazole- and imidazolium-containing polymers for biology and material science applications. Polymer 51:2447–2454. CrossRefGoogle Scholar
  3. 3.
    Behera HS, Satpathy G, Tripathi M (2016) Isolation and genotyping of Acanthamoeba spp. from Acanthamoeba meningitis/meningoencephalitis (AME) patients in India. Parasit Vectors 9:442. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Biczak R, Pawłowska B, Bałczewski P, Rychter P (2014) The role of the anion in the toxicity of imidazolium ionic liquids. J Hazard Mater 274:181–190. CrossRefPubMedGoogle Scholar
  5. 5.
    Borase HP, Patil CD, Sauter IP, Rott MB, Patil SV (2013) Amoebicidal activity of phytosynthesized silver nanoparticles and their in vitro cytotoxicity to human cells. FEMS Microbiol Lett 345(2):127–131. CrossRefPubMedGoogle Scholar
  6. 6.
    Carrijo-Carvalho LC, Sant'ana VP, Foronda AS, de Freitas D, de Souza Carvalho FR (2017) Therapeutic agents and biocides for ocular infections by free-living amoebae of Acanthamoeba genus. Surv Ophthalmol 62(2):203–218. CrossRefPubMedGoogle Scholar
  7. 7.
    Clarke DW, Niederkorn JY (2006) The pathophysiology of Acanthamoeba keratitis. Trends Parasitol 22:175–180. CrossRefPubMedGoogle Scholar
  8. 8.
    Coleman D, Špulák M, Garcia MT, Gathergood N (2012) Antimicrobial toxicity studies of ionic liquids leading to a ‘hit’ MRSA selective antibacterial imidazolium salt. Green Chem 14:1350–1356. CrossRefGoogle Scholar
  9. 9.
    Coulon C, Collignon A, McDonnell G, Thomas V (2010) Resistance of Acanthamoeba cysts to disinfection treatments used in health care settings. J Clin Microbiol 48:2689–2697. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Fabres LF, Maschio VJ, Santos DL, Kwitko S, Marinho DR, Araújo BS, Locatelli CI, Rott MB (2018) Virulent T4 Acanthamoeba causing keratitis in a patient after swimming while wearing contact lenses in Southern Brazil. Acta Parasitol 63(2):428–432. CrossRefPubMedGoogle Scholar
  11. 11.
    Fang B, Zhou CH, Rao XC (2010) Synthesis and biological activities of novel amine-derived bis-azoles as potential antibacterial and antifungal agents. Eur J Med Chem 45:4388–4398. CrossRefPubMedGoogle Scholar
  12. 12.
    Fuerst PA, Booton GC, Crary M (2015) Phylogenetic analysis and the evolution of the 18S rRNA gene typing system of Acanthamoeba. J Eukaryot Microbiol 62:69–84. CrossRefPubMedGoogle Scholar
  13. 13.
    Gao Y, Vlahakis JZ, Szarek WA, Brockhausen I (2013) Selective inhibition of glycosyltransferases by bivalent imidazolium salts. Bioorg Med Chem 21:1305–1311. CrossRefPubMedGoogle Scholar
  14. 14.
    Gauthier C, Legault J, Girard-Lalancette K, Mshvildadze V, Pichette A (2009) Haemolytic activity, cytotoxicity and membrane cell permeabilization of semisynthetic and natural lupane- and oleanane-type saponins. Bioorg Med Chem 17(5):2002–2008. CrossRefPubMedGoogle Scholar
  15. 15.
    Ghashghaei O, Kielland N, Revés M, Taylor MC, Kelly JM, Di Pietro O, Muñoz-Torrero D, Péres B, Lavilla R (2018) Tetrasubstituted imidazolium salts as potent antiparasitic agents against African and American trypanosomiases. Molecules. CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Gilmore BF (2011) Antimicrobial ionic liquids. In: Kokorin PA (ed) Ionic liquids: applications and perspectives, InTech, Rijeka, pp 587–604. doi: 10.5772/1782.Google Scholar
  17. 17.
    Goellner E, Schmitt AT, Couto JL, Muller ND, Pilz-Junior HJ, Schrekker HS, Silva CE, da Silva OS (2018) Larvicidal and residual activity of imidazolium salts against Aedes aegypti (Diptera: Culicidae). Pest Manag Sci 74(4):1013–1019. CrossRefPubMedGoogle Scholar
  18. 18.
    Hadas E, Ozarowsi M, Derda M, Thiem B, Cholewinski M, Skrzypczak L, Gryszczynska A, Piasecka A (2017) The use of extracts from Passiflora spp. in helping the treatment of acanthamoebiasis. Acta Pol Pharm 74(3):921–928PubMedGoogle Scholar
  19. 19.
    Hassan F, Bhatti A, Desai R, Barua A (2019) Analysis from a year of increased cases of Acanthamoeba Keratitis in a large teaching hospital in the UK. Contact Lens Anterior Eye 42(5):506–511. CrossRefPubMedGoogle Scholar
  20. 20.
    Hernández-Núñez E, Tlahuext H, Moo-Puc R, Torres-Gómez H, Reyes-Martínez R, Cedillo-Rivera R, Nava-Zuazo C, Navarrete-Vazquez G (2009) Synthesis and in vitro trichomonicidal, giardicidal and amebicidal activity of N-acetamide(sulfonamide)-2-methyl-4-nitro-1H-imidazoles. Eur J Med Chem 44(7):2975–2984. CrossRefPubMedGoogle Scholar
  21. 21.
    Hübner DPG, de Brum VP, Frasson AP, Menezes CB, Senger FR, Santos da Silva GN, Baggio Gnoatto SC, Tasca T (2016) Anti-Trichomonas vaginalis activity of betulinic acid derivatives. Biomed Pharmacother 84:476–484. CrossRefPubMedGoogle Scholar
  22. 22.
    Khan NA (2006) Acanthamoeba: biology and increasing importance in human health. FEMS Microbiol Rev 30:564–595. CrossRefPubMedGoogle Scholar
  23. 23.
    Khan NA (2009) Acanthamoeba—biology and pathogenesis. Caister Academic Press, Norfolk, p 290Google Scholar
  24. 24.
    Kolar SS, Manarang JC, Burns AR, Miller WL, McDermott AM, Bergmanson JP (2015) Contact lens care solution killing efficacy against Acanthamoeba castellanii by in vitro testing and live-imaging. Contact Lens Anterior Eye 38(6):442–450. CrossRefPubMedGoogle Scholar
  25. 25.
    Król-Turmińska K, Olender A (2017) Human infections caused by free-living amoebae. Ann Agric Environ Med 24(2):254–260. CrossRefPubMedGoogle Scholar
  26. 26.
    Kullmer CNP, Ta D, Chen CY, Cieker CJ, Annunziata O, Dzyuba SV (2019) Hexadecyl-containing organic salts as novel organogelators for ionic, eutectic, and molecular liquids. ACS Omega 4:9400–9406. CrossRefGoogle Scholar
  27. 27.
    Lis R, Davey DD, Morgan TK Jr, Lumma WC Jr, Wohl RA, Jain VK, Wan CN, Argentieri TM, Sullivan ME, Cantor EH (1987) Synthesis and antiarrhythmic activity of novel 3-alkyl-1-[omega.-[4[(alkylsulfonyl)amino]phenyl]-omega-hydroxyalkyl]-1H-imidazolium salts and related compounds. J Med Chem 30:2303–2309. CrossRefPubMedGoogle Scholar
  28. 28.
    Liu LWH, Riduan SN, Ying JY, Zhang Y (2013) Short imidazolium chains effectively clear fungal biofilm in keratitis treatment. Biomaterials 34:1018–1023. CrossRefPubMedGoogle Scholar
  29. 29.
    Lorenzo-Morales J, Martín-Navarro CM, López-Arencibia A, Arnalich-Montiel F, Piñero JE, Valladares B (2013) Acanthamoeba keratitis: an emerging disease gathering importance worldwide? Trends Parasitol 29(4):181–187. CrossRefPubMedGoogle Scholar
  30. 30.
    Lorenzo-Morales J, Khan NA, Walochnik J (2015) An update on Acanthamoeba keratitis: diagnosis, pathogenesis and treatment. Parasite 22:10. CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Malhotra SV, Kumar V (2010) A profile of the in vitro anti-tumor activity of imidazolium-based ionic liquids. Bioorg Med Chem Lett 20:581–585. CrossRefPubMedGoogle Scholar
  32. 32.
    Marciano-Cabral F, Cabral G (2003) Acanthamoeba spp. as agents of disease in humans. Clin Microbiol Rev 16:273–307. CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Martins RC, Dorneles GP, Teixeira VON, Antonello AM, Couto JL, Rodrigues Júnior LC, Monteiro MC, Peres A, Schrekker HS, Romão PRT (2018) Imidazolium salts as innovative agents against Leishmania amazonensis. Int Immunopharmacol 63:101–109. CrossRefPubMedGoogle Scholar
  34. 34.
    Maycock NJ, Jayaswal R (2016) Update on Acanthamoeba Keratitis: diagnosis, treatment, and outcomes. Cornea 15(5):713–720. CrossRefGoogle Scholar
  35. 35.
    Nava-Zuazo C, Estrada-Soto S, Guerrero-Alvarez J, León-Rivera I, Molina-Salinas GM, Said-Fernández S, Chan-Bacab MJ, Cedillo-Rivera R, Moo-Puc R, Mirón-López G, Navarrete-Vazquez G (2010) Design, synthesis, and in vitro antiprotozoal, antimycobacterial activities of N-{2-[(7-chloroquinolin-4-yl)amino]ethyl}ureas. Bioorg Med Chem 18(17):6398–6403. CrossRefPubMedGoogle Scholar
  36. 36.
    Niederkorn JY, Alizideh H, Leher JP, McCulley JP (1999) The pathogenesis of Acanthamoeba keratitis. Microbes Infect 1:437–443. CrossRefPubMedGoogle Scholar
  37. 37.
    Nikolskaia OV, de Lima AP, Kim YV, Lonsdale-Eccles JD, Fukuma T, Scharfstein J, Grab DJ (2006) Blood-brain barrier traversal by African trypanosomes requires calcium signaling induced by parasite cysteine protease. J Clin Invest 116(10):2739–2747. CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Omaña-Molina M, Vanzzini-Zago V, Hernandez-Martinez D, Gonzales-Robles A, Salazar-Villatoro L, Ramirez-Flores E, Oregon-Miranda E, Lorenzo-Morales J, Martinez-Palomo A (2016) Acanthamoeba genotypes T3 and T4 as causative agents of amoebic keratitis in Mexico. Parasitol Res 15:873–878. CrossRefGoogle Scholar
  39. 39.
    Padzik M, Chomicz L, Szaflik JP, Chruścikowska A, Perkowski K, Szaflik J (2014) In vitro effects of selected contact lens care solutions on Acanthamoeba castellanii strains in Poland. Exp Parasitol 145(Suppl):S98–S101. CrossRefPubMedGoogle Scholar
  40. 40.
    Pendleton JN, Gilmore BF (2015) The antimicrobial potential of ionic liquids: a source of chemical diversity for infection and biofilm control. Int J Antimicrob Agents 46:131–139. CrossRefPubMedGoogle Scholar
  41. 41.
    Polat ZA, Vural A, Cetin A (2007) Efficacy of contact lens storage solutions against trophozoite and cyst of Acanthamoeba castellanii strain 1BU and their cytotoxic potential on corneal cells. Parasitol Res 101(4):997–1001. CrossRefPubMedGoogle Scholar
  42. 42.
    Riduan SN, Zhang Y (2013) Imidazolium salts and their polymeric materials for biological applications. Chem Soc Rev 42:9055–9070. CrossRefPubMedGoogle Scholar
  43. 43.
    Rosales MJ, Ximenis M, Costa A, Rotger C, Romero D, Olmo F, Delgado E, Clares MP, García-España E, Marín C, Sánchez M (2018) In vitro activity of squaramides and acyclic polyamine derivatives against trophozoites and cysts of Acanthamoeba castellanii. J Biosci Med 6(8):1–14. CrossRefGoogle Scholar
  44. 44.
    Schrekker CML, Sokolovicz YCA, Raucci MG, Selukar BS, Klitzke JS, Lopes W, Leal CAM, de Souza IOP, Galland GB, dos Santos JHZ, Mauler RS, Kol M, Dagorne S, Ambrosio L, Teixeira ML, Morais J, Landers R, Fuentefria AM, Schrekker HS (2016) Multitask imidazolium salt additives for innovative poly(L-lactide) biomaterials: morphology control, Candida spp. biofilm inhibition, human mesenchymal stem cell biocompatibility, and skin tolerance. ACS Appl Mater Interfaces 8:21163–21176. CrossRefPubMedGoogle Scholar
  45. 45.
    Schrekker HS, Stracke MP, Schrekker CML, Dupont J (2007) Ether-functionalized imidazolium hexafluorophosphate ionic liquids for improved water miscibilities. Ind Eng Chem Res 46(22):7389–7392. CrossRefGoogle Scholar
  46. 46.
    Schrekker HS, Silva DO, Gelesky MA, Stracke MP, Schrekker CML, Gonçalves RS, Dupont J (2008) Preparation, cation–anion interactions and physicochemical properties of ether-functionalized imidazolium ionic liquids. J Braz Chem Soc 19:426–433. CrossRefGoogle Scholar
  47. 47.
    Schrekker HS, Donato RK, Fuentefria AM, Bergamo VZ, Oliveira LF, Machado MM (2013) Imidazolium salts as antifungal agents: activity against emerging yeast pathogens, without human leukocyte toxicity. Med Chem Commun 4:1457–1460. CrossRefGoogle Scholar
  48. 48.
    Schulz D, Simões MO, Frohner CRA, Gabilan NH, Batista CRV (2005) Citotoxicidade do extrato bruto de Bacillus amyloliquefaciens frente a hemácias de carneiro e células Vero. Alim Nutr 16(2):145–151Google Scholar
  49. 49.
    Siddiqui R, Aqeel Y, Khan NA (2016) The development of drugs against Acanthamoeba infections. Antimicrob Agents Chemother 60(11):6441–6450. CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Siddiqui R, Abjani F, Yeo CI, Tiekink ER, Khan NA (2017) The effects of phosphanegold(I) thiolates on the biological properties of Acanthamoeba castellanii belonging to the T4 genotype. J Negat Results Biomed 16(1):6. CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Sifaoui I, Reyes-Batlle M, López-Arencibia A, Chiboub O, Bethencourt-Estrella CJ, Nicolás-Hernández DS, Expósito RLR, Rizo-Liendo A, Piñero JE, Lorenzo-Morales J (2019) Screening of the pathogen box for the identification of anti-Acanthamoeba agents. Exp Parasitol 201:90–92. CrossRefPubMedGoogle Scholar
  52. 52.
    Sokolovicz YCA, Schrekker CML, Hild F, Bodo LO, Couto JL, Klitzke JS, Maraschin T, Basso NRS, dos Santos JHZ, Dagorne S, Schrekker HS (2019) Organometal-catalyzed synthesis of high molecular weight poly-(L-lactic acid) with a covalently attached imidazolium salt: performance-enhanced reduced graphene oxide-PLLA biomaterials. New J Chem 43:16367–16373. CrossRefGoogle Scholar
  53. 53.
    Toppino A, Bova ME, Crich SG, Alberti D, Diana E, Barge A, Aime S, Venturello P, Deagostino A (2013) A carborane-derivative “Click” reaction under heterogeneous conditions for the synthesis of a promising lipophilic MRI/GdBNCT agent. Chem Eur J 19:721–728. CrossRefPubMedGoogle Scholar
  54. 54.
    Ustüntürk M, Zeybek Z (2014) Amoebicidal efficacy of a novel multi-purpose disinfecting solution: first findings. Exp Parasitol 145(Suppl):S93–S97. CrossRefPubMedGoogle Scholar
  55. 55.
    Visvesvara GS, Moura H, Schuster FL (2007) Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea. FEMS Immunol Med Microbiol 50(1):1–26. CrossRefPubMedGoogle Scholar
  56. 56.
    Visvesvara GS (2010) Free-living amebae as opportunistic agents of human disease. J Neuroparasitol. CrossRefGoogle Scholar
  57. 57.
    Wang D, Galla HJ, Drücker P (2018) Membrane interactions of ionic liquids and imidazolium salts. Biophys Rev 10(3):735–746. CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    Zhang C, Ding Z, Suhaimi NA, Kng YL, Zhang Y, Zhuo L (2009) A class of imidazolium salts is anti-oxidative and anti-fibrotic in hepatic stellate cells. Free Radic Res 43:899–912. CrossRefPubMedGoogle Scholar

Copyright information

© Witold Stefański Institute of Parasitology, Polish Academy of Sciences 2020

Authors and Affiliations

  • Laura Führich Fabres
    • 1
  • Fabiany da Costa Gonçalves
    • 2
  • Eliane Oliveira Salines Duarte
    • 3
  • Francisco Kercher Berté
    • 3
  • Débora Kélen Si lva da Conceição
    • 4
  • Leonildo Alves Ferreira
    • 4
  • Henri Stephan Schrekker
    • 4
    Email author
  • Marilise Brittes Rott
    • 5
    Email author
  1. 1.Programa de Pós-Graduação Em Microbiologia Agrícola E Do Ambiente, Universidade Federal Do Rio Grande Do Sul (UFRGS)Porto AlegreBrazil
  2. 2.Hospital de Clínicas de Porto Alegre (HCPA)Porto AlegreBrazil
  3. 3.Faculdade de FarmáciaUniversidade Federal Do Rio Grande Do Sul (UFRGS)Porto AlegreBrazil
  4. 4.Graduate Program in Chemistry, Laboratory of Technological Processes and Catalysis, Institute of ChemistryUniversidade Federal Do Rio Grande Do SulPorto AlegreBrazil
  5. 5.Departamento de Microbiologia, Imunologia E Parasitologia, Instituto de Ciências Básicas da Saúde, Setor de ParasitologiaUniversidade Federal Do Rio Grande Do SulPorto AlegreBrazil

Personalised recommendations