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| Author(s): |
Camila Cristina de Foggi
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Araraquara. 2018-04-17. |
| Institution: | Universidade Estadual Paulista (Unesp). Faculdade de Odontologia. Araraquara |
| Defense date: | 2017-03-10 |
| Advisor: | Ana Lucia Machado |
| Abstract | |
Due to the increased tolerance of the microrganisms to the available antimicrobial drugs, there is a need of development of alternative methods for infection control. In the first part of this study, bimetallic oxides were synthesized by hydrothermal microwave-assisted and coprecipitation methods, varying synthesis conditions, as presence or absence of surfactants and types of solvents used. The different microcrystals of α-Ag2WO4 and β-Ag2MoO4 obtained were characterized by X-ray diffraction (XRD), field-emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), energy diffraction spectroscopy (EDS), Raman spectroscopy and photoluminescence measurements (PL) and UV-Vis. Their antimicrobial activity was evaluated against planktonic cells of methicillin - resistant Staphylococcus aureus - MRSA, Escherichia coli (E. coli) and Candida albicans (C. albicans) by the determination of inhibitory concentrations (MIC) and bactericidal/fungicidal concentrations (MBC/MFC). Based on the MIC, MBC and MFC values, the microcrystals that provided the best results (α-Ag2WO4 irradiated and α-Ag2WO4 and β-Ag2MoO4, both synthesized in alcohol) were also evaluated for their ability to inhibit biofilm of MRSA, E. coli and C. albicans, by counting the CFU/mL, dry weight determination, and analysis using confocal laser scanning microscopy (CLSM) and FE-SEM. Other method for infection control evaluated in the second part of this research, was the deposition of SiO2/Ag thin films on the surface of an denture base acrylic resin by co-sputtering plasma, using two deposition times (7 and 15 minutes). Pure Ag and SiO2 films were also deposited for comparison and all thin films were characterized by XRD, thickness measurement, roughness determination, hydrophobicity and surface free energy (SFE), Fourier transformed infrared spectroscopy (FTIR), SEM and atomic force microscopy (AFM). C. albicans adhesion was evaluated by CFU/mL counting, CLSM and MEV. Results from TEM and EDS confirmed the growth of nanofilaments of metallic Ag on the surface of the irradiated α-Ag2WO4. All microcrystals of α-Ag2WO4 and β-Ag2MoO4 analysed showed crystallinity, without the presence of secondary phases, with structural order and symmetry at short (Raman) and long range (XRD). The band gap values derived from UV–Vis spectra ranged from 2.4 eV to 3.12 eV for the irradiated α- Ag2WO4 and those synthesized in alcohol and from 3.29 eV to 3.33 eV for the microcrystals of β-Ag2MoO4 synthesized in ammonia and ethanol, respectively. PL measurements indicated 12 that the maximum absorption was observed at the orange region (613 nm) and red region (750 nm) for the α-Ag2WO4 synthesized in water, while for those obtained in alcohol and ammonia, the highest peaks were in the blue region (447 nm). For the β-Ag2MoO4, the maximum absorption was seen at the blue region, mainly when the synthesis was performed in alcohol. FE-SEM images revealed that the microcrystals of α-Ag2WO4 showed an agglomerated nature, in the shape of elongated hexagonal nanorods, being organized in flower-like structures when synthesized in ammonia and alcohol. For β-Ag2MoO4, the microcrystals showed were round-shaped, typically in the micrometer size. The microscopic images also revealed that, depending on the solvent used, the microcrystals presented different crystallographic faces: [001], [101] and [010] for the α-Ag2WO4 and [011], [001] and [111] for the β-Ag2MoO4. Microbiological tests performed with planktonic cells showed that all synthesized microcrystals exhibit antimicrobial activity. However, the MIC/MBC value for MRSA for the irradiated α-Ag2WO4 (31.25 μg/mL) was lower that that obtained for the non-irradiated (125 μg/mL). Intermediate value (62.50 μg/mL) was required to inactivate C. albicans, for the α-Ag2WO4 synthesized with PVP and β-Ag2MoO4 without PVP, both obtained by co-precipitation method. The values found for the α-Ag2WO4 microcrystals synthesized in alcohol, ammonia and water were, respectively, 3.90 g/mL, 7.81 g/mL and 7.81 g/mL for C. albicans, 15.62 g/mL, 62.50 g/mL and 125 g/mL for MRSA and 0.48 μg/mL, 0.48 μg/mL and 0.97 μg/mL for E. coli. For the β-Ag2MoO4 microcrystals synthesized in alcohol, ammonia and water the values were, respectively, 7.81 g/mL, 7.81 g/mL e 15.62 g/mL for C. albicans, 31.25 μg/mL, 62.50 μg/mL and 500 μg/mL for MRSA and 0.49 μg/mL, 1.95 μg/mL and 3.91 μg/mL for E. coli. In the tests performed with biofilms, (irradiated α-Ag2WO4, α-Ag2WO4 and β-Ag2MoO4, both synthesized in alcohol) were able to inhibit C. albicans biofilm formation at the following concentrations: 15.62 μg/mL, 15.62 μg/mL and 125 μg/mL, respectively. The concentrations required for MRSA were, respectively, 31.25 μg/mL, 62.50 μg/mL and 250 μg/mL, while for E. coli were, respectively, 1.95 μg/mL, 3.90 μg/mL and 7.81 μg/mL. The selected microcrystals, at the same concentrations, also reduced the dry weight of the biofilms of the three microorganisms. CLSM and SEM images showed results that were consistent with each other and with the data obtained in the CFU/mL assay. In addition, SEM analyses indicated morphological alterations, which were more evident for the C. albicans cells and with the α-Ag2WO4 synthesized in alcohol. The results from the second part of this study showed that it was not possible to detect the thickness of the deposited films. The Ag/SiO27, Ag and SiO2 thin films 13 increased the surface roughness (Ra) (3.4 μm, 4.2 μm and 4.1 μm, respectively), whereas the Ag/SiO2 decreased (2.8 μm), in comparison to control, (PMMA) without coating (3.2 μm). The water contact angle of the PMMA was 100.83 (hydrophobic) while those of the deposited films were lower (Ag/SiO27 = 61.57, Ag/SiO2 = 93.97, Ag = 83.66e SiO2 = 47.01). The SFE of the PMMA and the films Ag/SiO215 and Ag were similar and lower than those of the films Ag/SiO27 and SiO2. FE-SEM and MFA revealed the presence of grains with different sizes in the films Ag/SiO27 and Ag/SiO215 , showing a base layer of SiO2 (small granules), covered by Ag nanoparticles (bigger grains) in the surface. The structures of the Ag/SiO27 and Ag/SiO2 films were in the amorphous state (XRD) and FTIR analysis demonstrated the presence of peaks related to Si-O and Si-H bonds. The mean values of Log10 CFU/mL of C. albicans observed for the Ag/SiO27, Ag and SiO2 were similar and not different from the control, but the Ag/SiO2 film promoted a reduction in the number of viable cells more than 2 logs compared to control. Based on the obtained results, the more effective antimicrobial activity was observed for the α-Ag2WO4 microcrystals synthesized in alcohol and for the Ag/SiO215 thin film. (AU) | |
| FAPESP's process: | 15/03654-7 - Antimicrobial activity of bimetallic oxides and of plasma-deposited film on denture base acrylic resin |
| Grantee: | Camila Cristina de Foggi |
| Support Opportunities: | Scholarships in Brazil - Doctorate (Direct) |