Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans

Detalhes bibliográficos
Autor(a) principal: Treméa, Carolina Martins
Data de Publicação: 2015
Tipo de documento: Dissertação
Idioma: por
Título da fonte: Repositório Institucional da UFG
dARK ID: ark:/38995/001300000c8mz
Texto Completo: http://repositorio.bc.ufg.br/tede/handle/tede/5748
Resumo: Cryptococcosis is a fungal infection caused by species of the complex Cryptococcus neoformans (C. neoformans and C. gattii) with high incidence in patients with acquired immunodeficiency syndrome (AIDS). Antifungal drugs used in therapy of this infection are scarce and, in addition, have evident toxicity and low effectiveness, which motivates the search for new candidates to antifungals. Essential oils (EOs) extracted from the fruit of Litsea cubeba (Litsea cubeba (Lour.) Pers.) and leaves of Palmarosa (Cymbopogon martini (Roxb.) Will. Watson) and its major constituents, citral and geraniol, were investigated as antifungal sources. The aims of this work were to determine the isolated or associated antifungal activity of EOs and major constituents with amphotericin B (AMB) or fluconazole (FLU) on 15 isolates and two standard strains of C. neoformans complex, evaluate the time-kill curve, the action on the biosynthesis of ergosterol and on cell membrane, in addition to verify the cytotoxicity of EOs and major constituents on human erythrocytes. By in vitro susceptibility testing and minimum fungicidal concentration (MFC), the best activities were to L. cubeba and citral. The EOs and major constituents exhibited minimum inhibitory concentration (MIC) between 16 and 512 μg/mL and 64 and 2048 μg/mL, with a MIC90 of 128 μg/mL for L. cubeba and citral, and 512 μg/mL for C. martini and geraniol. To evaluate the association of EOs and major constituents with antifungal agents was performed the Chequerboard method, that showed indifference in the association of AMB and L. cubeba (94.1%) or citral (70.6%). In association with FLU, L. cubeba was antagonistic (76.5%) and indifferent (94%) for citral. C. martini and geraniol showed synergism associated with AMB, respectively, in 52.9 and 70.6% of isolates, and showed indifference when associated with the FLU in 64.7% and 88.2%. C. gattii 24065 time-kill curve under the action of EOs and major constituents showed a rapid fungicidal activity, which occurred after one hour incubation for L. cubeba, C. martini and geraniol, and after two hours for citral. The amount of ergosterol was quantified after contact with the EOs and major constituents and showed that L. cubeba and citral inhibited 100% of sterol biosynthesis in MIC and 64% for C. martini. This is the probably mechanism of action of these agents, except for geraniol, which reduced only 12% in MIC. The major compounds of EOs showed no ability to injure the plasma membrane of C. gattii, since an average of 2.7% propidium iodide labeled cells were detected by flow cytometry. The in vitro cytotoxicity assay by hemolysis showed that EOs and major constituents were not toxic in the MIC and MFCs. Taken together, our results showed that the EOs of L. cubeba and C. martini and their major constituents, citral e geraniol are promising as candidates for new antifungal agents.
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spelling Souza, Lúcia Kioko Hasimoto ehttp://lattes.cnpq.br/4557218510118165Souza, Lúcia Kioko Hasimoto eFernandes, Orionalda de Fátima LisboaFaganello, Josianehttp://lattes.cnpq.br/4130176016821125Treméa, Carolina Martins2016-07-28T11:15:34Z2015-04-13TREMÉA, C. M. Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans. 2015. 97 f. Dissertação (Mestrado em Biologia da Relação Parasito-Hospedeiro) - Universidade Federal de Goiás, Goiânia, 2015.http://repositorio.bc.ufg.br/tede/handle/tede/5748ark:/38995/001300000c8mzCryptococcosis is a fungal infection caused by species of the complex Cryptococcus neoformans (C. neoformans and C. gattii) with high incidence in patients with acquired immunodeficiency syndrome (AIDS). Antifungal drugs used in therapy of this infection are scarce and, in addition, have evident toxicity and low effectiveness, which motivates the search for new candidates to antifungals. Essential oils (EOs) extracted from the fruit of Litsea cubeba (Litsea cubeba (Lour.) Pers.) and leaves of Palmarosa (Cymbopogon martini (Roxb.) Will. Watson) and its major constituents, citral and geraniol, were investigated as antifungal sources. The aims of this work were to determine the isolated or associated antifungal activity of EOs and major constituents with amphotericin B (AMB) or fluconazole (FLU) on 15 isolates and two standard strains of C. neoformans complex, evaluate the time-kill curve, the action on the biosynthesis of ergosterol and on cell membrane, in addition to verify the cytotoxicity of EOs and major constituents on human erythrocytes. By in vitro susceptibility testing and minimum fungicidal concentration (MFC), the best activities were to L. cubeba and citral. The EOs and major constituents exhibited minimum inhibitory concentration (MIC) between 16 and 512 μg/mL and 64 and 2048 μg/mL, with a MIC90 of 128 μg/mL for L. cubeba and citral, and 512 μg/mL for C. martini and geraniol. To evaluate the association of EOs and major constituents with antifungal agents was performed the Chequerboard method, that showed indifference in the association of AMB and L. cubeba (94.1%) or citral (70.6%). In association with FLU, L. cubeba was antagonistic (76.5%) and indifferent (94%) for citral. C. martini and geraniol showed synergism associated with AMB, respectively, in 52.9 and 70.6% of isolates, and showed indifference when associated with the FLU in 64.7% and 88.2%. C. gattii 24065 time-kill curve under the action of EOs and major constituents showed a rapid fungicidal activity, which occurred after one hour incubation for L. cubeba, C. martini and geraniol, and after two hours for citral. The amount of ergosterol was quantified after contact with the EOs and major constituents and showed that L. cubeba and citral inhibited 100% of sterol biosynthesis in MIC and 64% for C. martini. This is the probably mechanism of action of these agents, except for geraniol, which reduced only 12% in MIC. The major compounds of EOs showed no ability to injure the plasma membrane of C. gattii, since an average of 2.7% propidium iodide labeled cells were detected by flow cytometry. The in vitro cytotoxicity assay by hemolysis showed that EOs and major constituents were not toxic in the MIC and MFCs. Taken together, our results showed that the EOs of L. cubeba and C. martini and their major constituents, citral e geraniol are promising as candidates for new antifungal agents.A criptococose é uma infecção fúngica causada por espécies do complexo Cryptococcus neoformans (C. neoformans e C. gattii) com alta incidência em indivíduos com a síndrome da imunodeficiência adquirida (aids). Os antifúngicos utilizados na terapia dessa infecção são escassos e, além disso, possuem toxicidade evidente e baixa eficácia, o que motiva a busca por novos candidatos a antifúngicos. Os óleos essenciais (OEs) extraídos do fruto de Litsea cubeba (Litsea cubeba (Lour.) Pers.) e das folhas de Palmarosa (Cymbopogon martini (Roxb.) Will. Watson) e seus constituintes majoritários, citral e geraniol, foram investigados como fontes de ação antifúngica. Os objetivos foram determinar a atividade antifúngica isolada e associada à anfotericina B (AMB) ou fluconazol (FLU) sobre 15 isolados e duas cepas padrão do complexo C. neoformans, além de avaliar a curva de morte, a ação sobre a biossíntese do ergosterol e sobre a membrana celular do fungo e a citotoxicidade dos OEs e constituintes majoritários sobre hemácias humanas. Por meio de testes de suscetibilidade in vitro e de concentração fungicida mínima (CFM), verificou-se melhor atividade para L. cubeba e citral. Os OEs e constituintes majoritários avaliados exibiram concentração inibitória mínima (CIM) entre 16 e 512 μg/mL e CFM entre 64 e 2048 μg/mL, com CIM90 de 128 μg/mL para L. cubeba e citral e de 512 μg/mL para C. martini e geraniol. Para avaliar a associação dos OEs com antifúngicos foi realizado o método de Chequerboard, onde houve indiferença na associação de AMB a L. cubeba (94,1%) e citral (70,6%), e na associação com FLU, L. cubeba foi antagônico (76,5%) e citral indiferente (94,%). C. martini e geraniol apresentaram sinergismo associados a AMB, respectivamente, em 52,9 e 70,6% dos isolados, e indiferença associado ao FLU em 64,7% e 88,2%. A curva de morte de C. gattii 24065 sob ação dos OEs e constituintes majoritários mostrou uma rápida ação fungicida, que ocorreu após uma hora de incubação para L. cubeba, C. martini e geraniol, e em duas horas para o citral. A quantificação do ergosterol após contato com os OEs e constituintes majoritários mostrou que L. cubeba e citral inibiram 100% da biossíntese desse esterol na CIM e 64% para C. martini, o que provavelmente é um mecanismo de ação destes agentes, exceto para o geraniol, que reduziu apenas 12% na CIM. Os OEs e compostos majoritários não apresentaram capacidade de lesionar a membrana plasmática de C. gattii, visto que em média de 2,7% das células foram marcadas com iodeto de propídio e detectadas por citometria de fluxo. O ensaio de citotoxicidade in vitro por hemólise mostrou que os OEs e constituintes majoritários não foram tóxicos nas CIMs e CFMs. Os resultados observados no presente trabalho mostram que os OEs de L. cubeba e C. martini e seus constituintes majoritários, citral e geraniol, são promissores candidatos a antifúngicos.Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2016-07-28T11:13:37Z No. of bitstreams: 2 Dissertação - Carolina Martins Treméa - 2015.pdf: 3039510 bytes, checksum: 5e06579e2f9fb745276cc6cc6cd1866e (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2016-07-28T11:15:34Z (GMT) No. of bitstreams: 2 Dissertação - Carolina Martins Treméa - 2015.pdf: 3039510 bytes, checksum: 5e06579e2f9fb745276cc6cc6cd1866e (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)Made available in DSpace on 2016-07-28T11:15:34Z (GMT). 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dc.title.por.fl_str_mv Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
dc.title.alternative.eng.fl_str_mv Activity of Litsea cubeba and Cymbopogon martini essential oils against isolates of Cryptococcus neoformans species complex
title Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
spellingShingle Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
Treméa, Carolina Martins
Complexo Cryptococcus neoformans
Óleos essenciais
Litsea cubeba
Cymbopogon martini
Cryptococcus neoformans species complex
Esential oils
Litsea cubeba
Cymbopogon martini
CIENCIAS BIOLOGICAS::PARASITOLOGIA
title_short Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
title_full Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
title_fullStr Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
title_full_unstemmed Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
title_sort Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans
author Treméa, Carolina Martins
author_facet Treméa, Carolina Martins
author_role author
dc.contributor.advisor1.fl_str_mv Souza, Lúcia Kioko Hasimoto e
dc.contributor.advisor1Lattes.fl_str_mv http://lattes.cnpq.br/4557218510118165
dc.contributor.referee1.fl_str_mv Souza, Lúcia Kioko Hasimoto e
dc.contributor.referee2.fl_str_mv Fernandes, Orionalda de Fátima Lisboa
dc.contributor.referee3.fl_str_mv Faganello, Josiane
dc.contributor.authorLattes.fl_str_mv http://lattes.cnpq.br/4130176016821125
dc.contributor.author.fl_str_mv Treméa, Carolina Martins
contributor_str_mv Souza, Lúcia Kioko Hasimoto e
Souza, Lúcia Kioko Hasimoto e
Fernandes, Orionalda de Fátima Lisboa
Faganello, Josiane
dc.subject.por.fl_str_mv Complexo Cryptococcus neoformans
Óleos essenciais
Litsea cubeba
Cymbopogon martini
topic Complexo Cryptococcus neoformans
Óleos essenciais
Litsea cubeba
Cymbopogon martini
Cryptococcus neoformans species complex
Esential oils
Litsea cubeba
Cymbopogon martini
CIENCIAS BIOLOGICAS::PARASITOLOGIA
dc.subject.eng.fl_str_mv Cryptococcus neoformans species complex
Esential oils
Litsea cubeba
Cymbopogon martini
dc.subject.cnpq.fl_str_mv CIENCIAS BIOLOGICAS::PARASITOLOGIA
description Cryptococcosis is a fungal infection caused by species of the complex Cryptococcus neoformans (C. neoformans and C. gattii) with high incidence in patients with acquired immunodeficiency syndrome (AIDS). Antifungal drugs used in therapy of this infection are scarce and, in addition, have evident toxicity and low effectiveness, which motivates the search for new candidates to antifungals. Essential oils (EOs) extracted from the fruit of Litsea cubeba (Litsea cubeba (Lour.) Pers.) and leaves of Palmarosa (Cymbopogon martini (Roxb.) Will. Watson) and its major constituents, citral and geraniol, were investigated as antifungal sources. The aims of this work were to determine the isolated or associated antifungal activity of EOs and major constituents with amphotericin B (AMB) or fluconazole (FLU) on 15 isolates and two standard strains of C. neoformans complex, evaluate the time-kill curve, the action on the biosynthesis of ergosterol and on cell membrane, in addition to verify the cytotoxicity of EOs and major constituents on human erythrocytes. By in vitro susceptibility testing and minimum fungicidal concentration (MFC), the best activities were to L. cubeba and citral. The EOs and major constituents exhibited minimum inhibitory concentration (MIC) between 16 and 512 μg/mL and 64 and 2048 μg/mL, with a MIC90 of 128 μg/mL for L. cubeba and citral, and 512 μg/mL for C. martini and geraniol. To evaluate the association of EOs and major constituents with antifungal agents was performed the Chequerboard method, that showed indifference in the association of AMB and L. cubeba (94.1%) or citral (70.6%). In association with FLU, L. cubeba was antagonistic (76.5%) and indifferent (94%) for citral. C. martini and geraniol showed synergism associated with AMB, respectively, in 52.9 and 70.6% of isolates, and showed indifference when associated with the FLU in 64.7% and 88.2%. C. gattii 24065 time-kill curve under the action of EOs and major constituents showed a rapid fungicidal activity, which occurred after one hour incubation for L. cubeba, C. martini and geraniol, and after two hours for citral. The amount of ergosterol was quantified after contact with the EOs and major constituents and showed that L. cubeba and citral inhibited 100% of sterol biosynthesis in MIC and 64% for C. martini. This is the probably mechanism of action of these agents, except for geraniol, which reduced only 12% in MIC. The major compounds of EOs showed no ability to injure the plasma membrane of C. gattii, since an average of 2.7% propidium iodide labeled cells were detected by flow cytometry. The in vitro cytotoxicity assay by hemolysis showed that EOs and major constituents were not toxic in the MIC and MFCs. Taken together, our results showed that the EOs of L. cubeba and C. martini and their major constituents, citral e geraniol are promising as candidates for new antifungal agents.
publishDate 2015
dc.date.issued.fl_str_mv 2015-04-13
dc.date.accessioned.fl_str_mv 2016-07-28T11:15:34Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/masterThesis
format masterThesis
status_str publishedVersion
dc.identifier.citation.fl_str_mv TREMÉA, C. M. Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans. 2015. 97 f. Dissertação (Mestrado em Biologia da Relação Parasito-Hospedeiro) - Universidade Federal de Goiás, Goiânia, 2015.
dc.identifier.uri.fl_str_mv http://repositorio.bc.ufg.br/tede/handle/tede/5748
dc.identifier.dark.fl_str_mv ark:/38995/001300000c8mz
identifier_str_mv TREMÉA, C. M. Atividade dos óleos essenciais de Litsea cubeba e Cymbopogon martini sobre isolados do complexo Cryptococcus neoformans. 2015. 97 f. Dissertação (Mestrado em Biologia da Relação Parasito-Hospedeiro) - Universidade Federal de Goiás, Goiânia, 2015.
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