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Master's Dissertation
DOI
https://doi.org/10.11606/D.60.2020.tde-28102021-150618
Document
Author
Full name
Glaucia Rigotto Caruso
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2020
Supervisor
Committee
Kress, Marcia Regina von Zeska (President)
Saltoratto, Ana Lucia Fachin
Braga, Gilberto Ubida Leite
Silva, Roberto Santana da
Title in Portuguese
Efeito da inativação fotodinâmica em fungos utilizando fotossensibilizadores fenotiazínicos associados à nanopartículas de prata
Keywords in Portuguese
Candida albicans
Fenotiazínicos
Fusarium keratoplasticum
Fusarium oxysporum
Galleria mellonella
Inativação fotodinâmica antimicrobiana
Nanopartículas de prata
Abstract in Portuguese
A emergência de novas espécies de fungos patogênicos que são resistentes aos antifúngicos comerciais leva à necessidade de desenvolver novas estratégias para o tratamento de infecções fúngicas. A inativação fotodinâmica antimicrobiana (IFA) bem como as nanopartículas de prata (AgNPs) são metodologias que apresentam atividade antimicrobiana e, portanto, são promissoras no controle de microrganismos. Assim, este estudo visa otimizar o efeito antifúngico dessas metodologias pela associação dos fotossensibilizadores (FS) fenotiazínicos azul de metileno (MB), novo azul de metileno N (NMBN) e novo azul de metileno N Zinco (NMBN-Zn) com as AgNPs na IFA. As nanopartículas de prata (AgNPs) foram biossintetizadas por Fusarium oxysporum e associadas aos FS fenotiazínicos (AgNPs-FS). Tanto as AgNPs quanto as associações AgNPs-FS foram caracterizadas físicoquimicamente usando a espectrofotometria Uv-Vis para medir a banda de plasmon de superfície, microscopia eletrônica de transmissão, e o método de espalhamento de luz dinâmico para obter diâmetro e potencial zeta. A toxicidade dos compostos e as associações AgNPs-FS na IFA foi avaliada no modelo invertebrado Galleria mellonella. O método de microdiluição em caldo foi utilizado para determinar a concentração inibitória mínima (CIM) dos FS, AgNPs e AgNPs-FS na IFA contra o fungo leveduriforme Candida albicans e o fungo filamentoso Fusarium keratoplasticum. A caracterização fisicoquímica das AgNPs mostrou banda de plasmon de superfície em torno de 420 nm, diâmetro hidrodinâmico de 86,72 nm e potencial zeta de -28,6 mV. As associações AgNPs-FS mostraram uma banda de plasmon de AgNPs mais larga e deslocada devido aos ligantes dos FSs na superfície. O potencial zeta das associações AgNP-MB, AgNP-NMBN e AgNP-NMBN-Zn diminuiu para -13,9 mV, -17 mV e -9,25 mV, respectivamente. Os FSs, AgNPs e AgNPs-FS na IFA não apresentaram toxicidade às larvas de G. mellonella nas concentrações testadas. No teste de susceptibilidade foi observado que as associações AgNPs-FS apresentam baixas CIM na IFA contra os fungos em comparação com as CIM dos compostos isoladamente. Assim, as baixas concentrações dos compostos e ausência de toxicidade torna a associação AgNPs-FS na IFA uma promissora alternativa terapêutica para tratamento antifúngico.
Title in English
Effect of photodynamic inactivation on fungi using silver nanoparticleconjugated associated photosensitizers
Keywords in English
Antimicrobial photodynamic inactivation
Candida albicans
Fusarium keratoplasticum
Fusarium oxysporum
Galleria mellonella
Phenothiazines
Silver nanoparticles
Abstract in English
The emergence of new species of pathogenic fungi that are resistant to commercial antifungals leads to the need to develop new strategies for the treatment of fungal infections. Antimicrobial photodynamic inactivation (API) as well as silver nanoparticles (AgNPs) are methodologies that have antimicrobial activity and, therefore, are promising in the control of microorganisms. Thus, this study aims to optimize the antifungal effect of these methodologies by the association of phenothiazine photosensitizers (FS) methylene blue (MB), new methylene blue N (NMBN) and new methylene blue N Zinc (NMBN-Zn) with AgNPs in API. The silver nanoparticles (AgNPs) were biosynthesized by Fusarium oxysporum and associated with phenothiazine FS (AgNPs-FS). Both AgNPs and AgNPs- FS were characterized physically and chemically by using Uv-Vis spectrophotometry to measure the surface plasmon band, transmission electron microscopy, and dynamic light scattering method to obtain diameter and zeta potential. The toxicity of the compounds and the AgNPs-FS associations in API was evaluated in the invertebrate model Galleria mellonella. The broth microdilution method was used to determine the minimum inhibitory concentration (MIC) of FS, AgNPs and AgNPs-FS in API against the yeast fungus Candida albicans and the filamentous fungus Fusarium keratoplasticum. The physicochemical characterization of AgNPs showed a surface plasmon band peak at around 420 nm, hydrodynamic diameter of 86.72 nm and zeta potential of -28.6 mV. The AgNPs-FS associations showed a wider and displaced AgNP plasmon band peak due to the FS ligands on the surface. The zeta potential of the associations AgNP-MB, AgNP-NMBN and AgNP-NMBN-Zn decreased to -13.9 mV, -17 mV and -9.25 mV, respectively. The FSs, AgNPs and AgNPs-FS in API did not show toxicity to the G. mellonella larvae at the tested concentrations. In the susceptibility test was observed that the AgNPs-FS associations present low MICs in API against the fungi compared to the MICs of the compounds alone. Thus, the low concentrations of the compounds and the absence of toxicity make the AgNPs-FS association in API a promising therapeutic alternative for antifungal treatment.
 
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Release Date
2022-02-18
Publishing Date
2021-11-10
 
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