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Characterization of some natural substances with antimycotic effect
Plachá, Monika ; Hlaváček, Viliam (referee) ; Márová, Ivana (advisor)
The introduced bachelor thesis is focused on investigation of the effects of natural substances with potential antimycotic effects and characterization of their constituents. As a part of this work, an overview of mycosis, antimycotics and constituents of natural substances were introduced. Moreover, antifungal susceptibility and genus Candida were described. In the end of the theoretical part the liposomes and their characterization were described. In the experimental part general characteristics of some natural substances and antimycotic test were analyzed. Aqueous, ethanol, methanol and DMSO extracts were tested on yeast strain Candida glabrata. Aqueous extract of the cloves, cloves oil and their combination were encapsulated into liposomes. The encapsulation effectivity, long-term stability and their antifungal activity were determined too. Characterization of liposomes was identified by DLS. Finally, application of liposomes to a preparation of pharmaceutical and cosmetic preparations were presented.
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Preparation of nanoparticles and nanofibers with antimicrobial components
Sosková, Simona ; Veselá, Mária (referee) ; Skoumalová, Petra (advisor)
The presented diploma thesis is focused on the preparation of new materials with antimicrobial effect. Liposomes and nanofibers from polyhydroxybutyrate containing clotrimazole and natural extracts with good antifungal and antioxidant effects were prepared. The theoretical part contains examples and short description of using nanoparticles and nanofibers in cosmetics and medicine and the description of plants which have positive and potential antimycotic effects. Moreover, methods for particles and fibers characterisation were shortly described. In the experimental part, natural water and lipid extracts were prepared and spectrophotometrically characterised for the content of polyphenols, flavonoids and the antioxidant activity. Liposomes and liposomes containtng PHB were prepared from selected extracts and the encapsulation effectivity, shortterm and longterm stability via determination of polyphenols were determined. Prepared particles were characterized with DLS method (size) and zeta- potential (stability). PHB nanofibers functionalised with selected lipid extracts and clotrimazole were prepared via electrospinning and forcespinning, and examined via FLIM and FTIR-ATR methods and spectrophotometry was used for antioxidant activity and release of active substances determination. Antifungal properties of prepared particles, extracts and fibers using the test system Candida glabrata were studied. Finally, cytotoxicity of selected samples was tested with MTT assay using human keratinocytes.
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|Preparation and characterization of micro- and nanoparticles with selected drugs
Sosková, Simona ; Bokrová, Jitka (referee) ; Márová, Ivana (advisor)
The presented bachelor thesis is focused on the way to prepare combined micro- and nanoparticles containing some natural extracts with antioxidant effects, ibuprofen as the anti-inflammatory and analgetic agent and clotrimazole as the agent with an antifungal effect. The theoretical part contains an introduction of the methods of liposome preparation and a review of encapsulation techniques, methods of particles characterization, and analysis of active substances content. Furthermore, the yeasts of the genus Candida and the methods of the antimicrobial activity determination are described. The mentioned components were encapsulated into liposomes, alginate and chitosan particles. The antioxidant activity of the extracts was determined spectrophotometrically. The encapsulation effectivity of active substances into particles and the long-term stability of encapsulated components were determined and compared too. The HPLC method was used for measuring of the concentration of drugs and spectrophotometric method was used to quantify the content of polyphenols. Prepared particles were characterized with DLS method (size) and zeta- potential (stability). Finally, antibacterial properties of prepared particles and extracts using the test system Candida glabrata were studied. The antimycotic effect of particles, disturbed liposomes and extracts was compared. Finally, potential utilization of prepared liposomes in a cosmetic or pharmaceutical product is proposed.
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Application of natural antibiotics components in hygienic gels
Kubišová, Veronika ; Pavelková, Renata (referee) ; Skoumalová, Petra (advisor)
This bachelor thesis was focused on the preparation and characterization of gels containing a natural component exhibiting an antifungal effect. A part of this work was the research, which was focused on the description of superficial and systemic fungal diseases, antifungal drugs and plant products with a potential antifungal effect. Moreover, biologically active substances found in selected plants was described. Liposome particles that were part of the experimental part were also described and characterized. Considering the subject of this thesis was the preparation of antifungal gel, the basic characteristics of gels such as their drug delivery systems were described. Descriptions of basic methods for the determination of active substances and antifungal activity were not omitted. In the experimental part, three types of extracts (aqueous, ethanolic and lipidic) were prepared from selected plants, which were subsequntly characterized. Also, all kinds of prepared extracts were encapsulated into liposome particles, in which encapsulation efficiency, size and stability by electrokinetic potential were subsequently determined. Prepared extracts in free and encapsulated form were tested for antifungal activity, when testing was performed on Candida glabrata yeast. Moreover, the antimycotic effect of these natural substances was compared with a commercially available antifungal drug, where the drug was also tested in free and encapsulated form. Extracts in which higher active compound content and higher percent inhibitory activity of C. glabrata cells were determined were selected for incorporation into the prepared gels. The result of this work are natural-based gels that could be used to treat infectious fungal diseases. An example of a prepared gel is a gel containing a lipidic extract of marigold or containing liposomes prepared from a lipidic extract of elm pincushion.
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Monogenic susceptibility to infectious pathogens
Bloomfield, Markéta ; Šedivá, Anna (advisor) ; Koziar Vašáková, Martina (referee) ; Litzman, Jiří (referee)
(ENG) The modern approach to studies of monogenic inborn errors of immunity, driven by unprecedented advances of genetic tools, opens vast undiscovered areas of immune system components and functions. In particular, the diseases with striking clinical phenotypes with normal or near normal baseline immunophenotype, such as disorders of innate and intrinsic immunity with susceptibility to single pathogen, provide a unique window into the host-pathogen interactions. This thesis covers various novel aspects of immunopathology, genetics and clinical facets behind some such diseases, namely chronic mucocutaneous candidiasis due to hypermorphic (gain-of-function, GOF) STAT1 mutations, which hamper Th17-associated immune activities, and Mendelian susceptibility to mycobacterial diseases (MSMD) due to impairment of IL-12, IL-23/IFNγ signalling pathway. Moreover, it contributes to the mounting evidence that IL- 6 signalling is non-redundant in anti-staphylococcal immunity. Finally, it explores the novel Paediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS- TS) as a single pathogen-driven life-threatening immunopathology, which most likely develops due to individual, yet unknown, genetic predisposition. The findings presented in this thesis were in several cases translated...
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The effect of microbiota on pathogenesis of gut diseases
Galanová, Natalie ; Kolařík, Miroslav (advisor) ; Hudcovic, Tomáš (referee)
Gut microbiota is considered an important factor in the development of various diseases including inflammatory bowel disease (IBD, n = 127), Ulcerative colitis, Crohn's disease, and colorectal cancer (CRC, n = 64). A part of this thtesis is to prepare clinical material of different sorts (stool, biopsy) for sequencing on Illumina Miseq platform. This is achieved trough DNA isolation, amplification of 16S and internal transcribed spacer (ITS), normalization and ligation of sequencing adaptors. The aim of this project is to describe the differences between microbiota in healthy and diseased subjects in case of IBD or unimpaired and tumorous tissue for CRC patients. This research is also being based on cultivation, where a fresh stool samples (n = 3) are cultivated in a broad range of conditions, which enables us to obtain ecophysiological and species diversity of these samples by traditional and molecular methods. The cultivable fungi are also assigned reliable taxonomy by amplification of relevant genes (ITS1, β tubulin, second largest subunit of RNA polymerase II, RPB2) followed by both-sided Sanger sequencing. Selected species of fungi are processed into lysates, which are used for stimulation of mice macrofage cell line (RAW). Therefore the impact on immunity response is studied in vitro and...
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Alakali-metal-cation homeostasis in pathogenic Candida species
Elicharová, Hana ; Sychrová, Hana (advisor) ; Heidingsfeld, Olga (referee) ; Půta, František (referee)
Several tens of Candida species belong to the opportunistic human pathogens capable of inducing life-threatening infections in immunocompromised patients. Virulence of single Candida species depends among others on their resistance to the variable external conditions. The maintenance of alkali-metal-cation homeostasis, which means the ability to accumulate sufficient amount of potassium cations and on the other hand to survive under high extracellular concentrations of alkali-metal cations, is essential for growth and virulence of Candida cells. We observed the negative effect of fluconazole (FLC) on salt-tolerance of six Candida species and found that it is independent of the species level of FLC- resistance. FLC hyperpolarizes plasma membrane of Candida cells and therefore increases non-specific uptake of alkali-metal cations which results in strongly increased salt-sensitivity of Candida cells. The FLC-induced hyperpolarization also results in an increased sensitivity of Candida cells to the antifungals which are positively charged and are driven into the cells by the membrane potential. The effect of fluconazole on membrane potential and thus on the uptake of alkali- metal cations into the cell turned our attention to the homeostasis of potassium cations whose high intracellular concentration is...
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The effect of antimicrobial peptides on Saccharomyces cerevisiae and other yeast species
Makarova, Anna-Marie ; Hodek, Petr (advisor) ; Malcová, Ivana (referee)
The increased use of antibiotics, antifungal agents and disinfectants in the last decades has resulted in development of microbial resistance to these drugs. Candida species are the fourth most common cause of hospital-acquired bloodstream infection and kill 40% of those patients. Natural antimicrobial peptides are promising candidates for the development of new agents to treat yeast and bacterial infections, as their presumed mechanism of action differs significantly from the mechanism of action of current drugs. This work is focused on several peptides isolated from the venom of wild bees and their synthetic analogues and the identification of the most effective ones against non-pathogenic Saccharomyces cerevisiae and several pathogenic Candida species. Antifungal activity of eight cationic antimicrobial peptides was tested and compared under various conditions. The overall susceptibility of pathogenic yeast species to currently used antifungal drugs and the antimicrobial peptides was screened with the aim to identify potential synergistic and species-specific effects. The effect of antimicrobial peptides on membrane potential was measured by a fluorescent probe (diS-C3(3)), and the relative hyperpolarization of plasma membrane was shown for each peptide. The effect of antimicrobial peptides on...
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Copper metabolism in pathogenic eukaryotic microorganisms
Doležalová, Taťána ; Šuťák, Róbert (advisor) ; Pilátová, Jana (referee)
Copper is an essential trace element for almost every living organism. Its fundamental role in metabolism is caused by the redox properties of this metal. Owing to that it is a vital cofactor of many enzymes participating in cell energy metabolism as well as in the detoxification of reactive oxygen species. Nevertheless, too high copper concentration can damage the cells. The toxic effect of copper is manifested usually as the production of hydroxyl radicals distorting the cell structures. It can also damage Fe-S clusters that are essential for many enzymes. To avoid toxic manifestations the cell must balance on the edge of proper copper import and regulated export. Many specialized transporters and ATPases serve exclusively for copper transport. There are also known metallochaperones binding the potentially toxic metals and transporting them to the target protein. An interesting role in metabolism is played by the metallothioneins that bind cytosolic copper. Understanding copper homeostasis in pathogenic organisms reveals many interesting possibilities for better targeted treatment. For example, diseases caused by opportunistic fungi like Candida albicans, Aspergillus fumigatus or Cryptococcus neoformans contribute significantly to the deaths of immunodeficient patients. Some copper metabolism...
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Application of natural antibiotics components in hygienic gels
Kubišová, Veronika ; Pavelková, Renata (referee) ; Skoumalová, Petra (advisor)
This bachelor thesis was focused on the preparation and characterization of gels containing a natural component exhibiting an antifungal effect. A part of this work was the research, which was focused on the description of superficial and systemic fungal diseases, antifungal drugs and plant products with a potential antifungal effect. Moreover, biologically active substances found in selected plants was described. Liposome particles that were part of the experimental part were also described and characterized. Considering the subject of this thesis was the preparation of antifungal gel, the basic characteristics of gels such as their drug delivery systems were described. Descriptions of basic methods for the determination of active substances and antifungal activity were not omitted. In the experimental part, three types of extracts (aqueous, ethanolic and lipidic) were prepared from selected plants, which were subsequntly characterized. Also, all kinds of prepared extracts were encapsulated into liposome particles, in which encapsulation efficiency, size and stability by electrokinetic potential were subsequently determined. Prepared extracts in free and encapsulated form were tested for antifungal activity, when testing was performed on Candida glabrata yeast. Moreover, the antimycotic effect of these natural substances was compared with a commercially available antifungal drug, where the drug was also tested in free and encapsulated form. Extracts in which higher active compound content and higher percent inhibitory activity of C. glabrata cells were determined were selected for incorporation into the prepared gels. The result of this work are natural-based gels that could be used to treat infectious fungal diseases. An example of a prepared gel is a gel containing a lipidic extract of marigold or containing liposomes prepared from a lipidic extract of elm pincushion.
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