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Impact of plasma-treated liquids on selected microorganisms
Klementová, Kamila ; Jirásek, Vít (referee) ; Trebulová, Kristína (advisor)
The subject of this bachelor thesis is the study of the effects of plasma-treated liquids on bacteria Staphylococcus aureus and Pseudomonas aeruginosa. The evolving resistance of bacteria against antibiotics is currently a highly debated topic. Plasma treated liquids have the potential to address this problem due to their antimicrobial effects. However, these effects have not yet been fully explained, therefore they are the focus of this work. The theoretical section of the study is mainly focused on the microbiology part, specifically on the morphology of the bacterial cell and the individual bacteria which was used in the experimental section. Furthermore, plasma, cold atmospheric-pressure plasma (CAP) and plasma-liquid interactions are described. The last part of the theoretical section considers the potential applications of cold plasma and plasma-treated liquids (PTL) in the fields of food and agricultural industry and healthcare. In the experimental section, plasma-treated liquids were prepared from saline solution using two plasma systems. Solutions simulating the composition of the prepared PTL, and standard solutions of nitrites, nitrates and hydrogen peroxide were also prepared. The effects of plasma-treated liquids were compared with those of solutions simulating the composition of plasma-treated liquids and with standard solutions. The effects were tested on standardized bacterial cultures of Staphylococcus aureus and Pseudomonas aeruginosa. Their suspensions were exposed to the plasma-treated liquids, prepared standard solutions and PTL alternatives. At the selected exposure times, the number of viable bacteria was determined by inoculation onto the solid culture medium and after 24 hours of incubation, the antimicrobial effects were evaluated by counting colony forming units using Aurora software. The results of this study proved better antimicrobial efficacy of plasma-treated liquids compared to the solutions simulating their composition. There was a difference in the effects between the two plasma-treated liquids that were tested. Pseudomonas aeruginosa was also shown to be more susceptible to plasma-treated liquids than Staphylococcus aureus.
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Development of cosmetic products with antimicrobial effect
Dribňáková, Barbora ; Wikarská, Monika (referee) ; Márová, Ivana (advisor)
The proposed bachelor's thesis focuses on the preparation and study of nanomaterials with antimicrobial properties. The nanofibers were synthesized from biopolymers using the forcespinning method, and liposomes were prepared using the sonification method. Active ingredients used in this work are tymol, carvacrol, and eugenol. The theoretical section of the thesis discusses the methods for preparing nanofibers and nanoparticles and the materials required for their synthesis. It also covers the encapsulation of active ingredients in nanomaterials. Additionally, this part explores the skin microbiome, antimicrobial effects, and the use of both natural and synthetic antimicrobial agents. The practical part of the thesis focuses on determining the polyphenol content and antioxidant activity of each active ingredient. The gradual release of active ingredients into saline solution and distilled water was determined for nanofibers synthesized from polyhydroxybutyrate with encapsulated active ingredients. Furthermore, it deals with the optimization of nanofiber synthesis from chitosan and polyamide using the forcespinning method. The functionalized liposomes were characterized in terms of their size, stability, encapsulation efficiency, and gradual release of active ingredients. The liposomes showed high stability even after several weeks, and their size ranged from 170.90 ± 1.63 nm to 243.07 ± 7.83 nm. Finally, the antimicrobial activity of the prepared products and active ingredients was tested against Cutibacterium acnes, Staphylococcus epidermidis, and Escherichia coli. The tested active ingredients in pure and encapsulated form, demonstrated antimicrobial effect against both gram-positive and gram-negative bacteria. We assume the possibility of using the prepared nanomaterials in cosmetic products intended for the treatment of acne and other skin diseases, and also its use in the form of preservatives.
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Use of some microbial oils in foods and cosmetics
Požgayová, Viktória ; Szotkowski, Martin (referee) ; Márová, Ivana (advisor)
This bachelor thesis focuses on preparation of microbial oils from oleaginous yeasts and the subsequent characterization of their properties and biological effects on organisms. The thesis is divided into two main parts, theoretical and experimental part. The theoretical part provides an overview of information about microbial oils and the possibilities of their current or future applications in food and cosmetic industry. It also describes other lipophilic metabolites synthesized by carotenogenic yeasts and briefly characterizes the carotenogenic yeast strains as well as analytical methods used in the experimental part. Experimental part was focused on the process of optimization of the extraction of lipophilic active compounds from yeast biomass. The best option proved to be the combination of three solvents such as dimethyl sulfoxide, hexane, and ethanol. Using this approach, the extracts were prepared from Rhodotorula kratochvilovae, Rhodotorula mucilaginosa, Rhodosporidium toruloides, Cystofilobasidium macerans and Sporobolomyces pararoseus biomass. All of the prepared extracts were characterised based on the content of lipophilic metabolites by UV-VIS spectrophotometry and by analysis on HPLC/PDA and GC/FID. Furthermore, the antioxidant activity, SPF, antimicrobial effect, and short-term stability of these extracts were determined. The best temperature to preserve these extracts proved to be 7 °C. Three extracts with the highest SPF values were chosen and added to the prepared cosmetic emulsion which was characterized and the sensory analysis was carried out. The MTT cytotoxicity assay on human HaCaT keratinocytes was realized for the three extracts that were added to the prepared cosmetic product as well as for their combination.
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Development of cosmetic products with antimicrobial effect
Dribňáková, Barbora ; Wikarská, Monika (referee) ; Márová, Ivana (advisor)
The proposed bachelor's thesis focuses on the preparation and study of nanomaterials with antimicrobial properties. The nanofibers were synthesized from biopolymers using the forcespinning method, and liposomes were prepared using the sonification method. Active ingredients used in this work are tymol, carvacrol, and eugenol. The theoretical section of the thesis discusses the methods for preparing nanofibers and nanoparticles and the materials required for their synthesis. It also covers the encapsulation of active ingredients in nanomaterials. Additionally, this part explores the skin microbiome, antimicrobial effects, and the use of both natural and synthetic antimicrobial agents. The practical part of the thesis focuses on determining the polyphenol content and antioxidant activity of each active ingredient. The gradual release of active ingredients into saline solution and distilled water was determined for nanofibers synthesized from polyhydroxybutyrate with encapsulated active ingredients. Furthermore, it deals with the optimization of nanofiber synthesis from chitosan and polyamide using the forcespinning method. The functionalized liposomes were characterized in terms of their size, stability, encapsulation efficiency, and gradual release of active ingredients. The liposomes showed high stability even after several weeks, and their size ranged from 170.90 ± 1.63 nm to 243.07 ± 7.83 nm. Finally, the antimicrobial activity of the prepared products and active ingredients was tested against Cutibacterium acnes, Staphylococcus epidermidis, and Escherichia coli. The tested active ingredients in pure and encapsulated form, demonstrated antimicrobial effect against both gram-positive and gram-negative bacteria. We assume the possibility of using the prepared nanomaterials in cosmetic products intended for the treatment of acne and other skin diseases, and also its use in the form of preservatives.
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Bactericides in the environment: Occurrence of selected bactericides in wastewater and their impact on the aquatic environment
DUŠÁNKOVÁ, Monika
Bactericides are a group of chemically active substances that exhibit antimicrobial effects. Their ability to kill or inhibit the growth of unwanted microorganisms is widely used in industry and healthcare, but also in household. However, the widespread use of bactericides has resulted in the abundant occurrence of these substances in surface water and wastewater, which subsequently supports the development of antibiotic resistance. Specifically wastewater treatment plants (WWTP) significantly influence the entry of antibiotic resistance genes into the environment. This study aimed to determine the occurrence of bactericides in wastewater and surface water samples, sediments and sewage sludge using high-performance liquid chromatography hyphenated with mass spectrometry. The detected concentrations were used to evaluate removal efficiency and impact on the recipient. The positive removal efficiency was confirmed mainly for the group of quaternary ammonium compounds and some antibiotics. Clarithromycin achieved the highest removal efficiency. Although some monitored substances had a high removal efficiency at the WWTP, their final occurrence proved to affect the aquatic environment. Quaternary ammonium compounds were effectively removed from the wastewater. But mainly thanks to their properties, such as strong sorption to solid substances, i.e., sediments and sludge, they were concentrated in sewage sludge. Using sewage sludge on agricultural land can cause problems by retaining some monitored substances in soils or leaching them into the surface and groundwater. This work briefly described the fate of selected bactericides at the sewage treatment plant and the adjacent recipient. The possible risks of using bactericides and their entry into the environment were pointed out. Given the variety and quantity of bactericides and the severe consequences of their excessive use, this topic deserves wider attention.
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Study of plasma and plasma activated water effect for bacterial disease treatment
Korečková, Svatava ; Jirásek,, Vít (referee) ; Kozáková, Zdenka (advisor)
The aim of this master thesis was to prepare plasma-treated liquids (PTL), specifically plasma-treated saline solutions, using different plasma systems. Subsequently, PTL was applies onto selected microorganisms to determine its antimicrobial effect. Systems that use both direct and indirect plasma-liquid interactions were selected, and the plasma was generated at atmospheric pressure. The theoretical part is focused on the plasma treated water, the systems by which PTL can be produced, and a description of the characterization process, from the viewpoint of the amount of active particles or the physical-chemical properties. Here, the microbiological part of this work is also included, namely a description of the microorganisms used for the experimental part. The disease of otitis externa (inflammation of the external auditory canal) and the possibilities of its antibiotic treatment are also described here. In this work, PTL prepared by four different plasma systems were tested: the pinhole discharge with DC or AC high voltage source (PTLAC and PTLDC), the liquid electrode dielectric barrier discharge (PTLDBD), and the microwave surface wave discharge (PTLSW). Prepared PTLs were first characterized in terms of reactive particles (hydrogen peroxide, nitrates and nitrites) and changes in specific conductivity and pH. This was followed by microbiological testing of the antimicrobial efficiency of the prepared PTL. The bacteria Staphylococcus pseudintermedius, Pseudomonas aeruginosa and the yeast Malassezia pachydermatis were selected as test microorganisms. These microorganisms were exposed to PTL and the effects were studied for exposure times from 1 minute to 4 hours. After 24 hours of cultivation, the antimicrobial effects were evaluated by the standard plate count method and also by the image analysis method in the software Aurora. The next step was to compile antibiograms and compare the effect of the prepared PTL in different systems with the effect of antibiotics. In this master thesis, it was proven that the bacterium Pseudomonas aeruginosa is resistant to several antibiotics. Thus, the prepared PTLDC could represent the possibility of its inactivation without the use of antibiotics. Staphylococcus pseudintermedius and Malassezia pachydermatis do not have resistance to any antibiotic (antimycotic), but with incorrect use there is a risk of developing resistance, which is why PTL, with the highest antimicrobial effect, were also found for these microorganisms.
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USE OF COMPLEX EXTRACTS FROM PLANT AND MICROORGANISMS IN FOODS AND COSMETICS
Uhlířová, Renata ; Kovalčík, Adriána (referee) ; Vorlová, Lenka (referee) ; Márová, Ivana (advisor)
The presented doctoral thesis is focused on optimalization of preparation of extracts of various origin (plant and microbial). The influence of extraction agent, disintegration of microbial cells as well as special techniques for enhancement of extraction process effectiveness (ultrasound and microwaves) were monitored. As part of the characterization, considering the source material, the composition of the prepared extracts was analyzed both spectrophotometrically and using liquid and gas chromatography. The most suitable extraction procedure varies depending on the type of material. For herbal samples, the highest content of active substances was achieved when the extraction facilitated by microwaves was used. Contrarily, the optimal method for cyanobacteria is the traditional extraction method, which is carried out for 24 hours at laboratory temperature. The most suitable method for extracting the active substances from yeast was a combination of DMSO, ethanol and hexane. Further, the effect of extracts on microbial cultures of bacteria (E. coli, S. epidermidis, C. acnes) and yeast (C. glabrata) was observed. For cytotoxicity evaluation MTT and LDH assays were performed on HaCaT and Caco2 human cell lines. The antimicrobial tests have demonstrated the antimicrobial, antibacterial and antifungal effect of a numerous extracts, especially the essential oils of Lavandula angustifolia and Mentha piperita. On the other hand, the cytotoxicity tests showed that the most of the extracts were safe for the use in the food industry and cosmetics within a range of up to 10% by volume. To improve stability, the prepared extracts were enclosed into liposomal particles enriched with PHB polymer (0–100%). Using a ZeteSizer NanoZS device, the particles' size, polydispersity index, and colloidal stability were assessed. The cryoTEM was used to examine the particle´s morphology. Liposomes and liposomes with the addition of 20 % PHB (PHB20) were evaluated as the most suitable particles for encapsulation of the prepared extracts. Excellent stability and high encapsulation efficiency were displayed by the produced particles. Genotoxicity testing with SOS chromotest on the model organism E. coli did not reveal any genotoxic effects. As an alternative method to achieve stabilization of active substances, nanofibers from PHB were prepared by three different techniques and gelatin nanofibers by electrospinning. The fibers were monitored for their diameter, uniformity, distribution on the collector using SEM and the gradual release of active substances from the fiber structure. The evaluations revealed that the forcespinning and electrospinning techniques were the most appropriate for further investigation. A continuous release of active ingredients was achieved by incorporating extracts into nanofibers. Several applications of prepared materials in the food industry and cosmetics were designed and developed as part of this work. The base emulsion's stability and general qualities were improved when green coffee and carotenogenic yeast extracts were added. The usage of emulsions containing coffee and yeast extracts would be suitable as sunscreens, and after the addition of Spirulina extract, as day or night anti-wrinkle creams. Energy drinks of the new type based on coffee with the addition of encapsulated plant extracts enable gradual release of caffeine and antioxidants during digestion.
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Use of some microbial oils in foods and cosmetics
Požgayová, Viktória ; Szotkowski, Martin (referee) ; Márová, Ivana (advisor)
This bachelor thesis focuses on preparation of microbial oils from oleaginous yeasts and the subsequent characterization of their properties and biological effects on organisms. The thesis is divided into two main parts, theoretical and experimental part. The theoretical part provides an overview of information about microbial oils and the possibilities of their current or future applications in food and cosmetic industry. It also describes other lipophilic metabolites synthesized by carotenogenic yeasts and briefly characterizes the carotenogenic yeast strains as well as analytical methods used in the experimental part. Experimental part was focused on the process of optimization of the extraction of lipophilic active compounds from yeast biomass. The best option proved to be the combination of three solvents such as dimethyl sulfoxide, hexane, and ethanol. Using this approach, the extracts were prepared from Rhodotorula kratochvilovae, Rhodotorula mucilaginosa, Rhodosporidium toruloides, Cystofilobasidium macerans and Sporobolomyces pararoseus biomass. All of the prepared extracts were characterised based on the content of lipophilic metabolites by UV-VIS spectrophotometry and by analysis on HPLC/PDA and GC/FID. Furthermore, the antioxidant activity, SPF, antimicrobial effect, and short-term stability of these extracts were determined. The best temperature to preserve these extracts proved to be 7 °C. Three extracts with the highest SPF values were chosen and added to the prepared cosmetic emulsion which was characterized and the sensory analysis was carried out. The MTT cytotoxicity assay on human HaCaT keratinocytes was realized for the three extracts that were added to the prepared cosmetic product as well as for their combination.
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Studie antimikrobiálního účinku včelího jedu na druhy borelie z komplexu \kur{B. burgdorferi} sensu lato: izolace genu kódujícího antimikrobiální peptid melitin z jedové žlázy včely medonosné \kur{(Apis mellifera)}
DAVIDOVÁ, Marie
The incidence of tick borne diseases has drastically increased over the past few years. Control of tick borne pathogens, especially spirochetes from \kur{B. bugdorferi} sensu lato complex, is complicated due to a lack of vaccines against them. The emergence of bacterial strains to well established and widely used antibiotics has become a serious public health concern. Analysis of new classes of antimicrobial peptides became the subject of new investigation due to their efficacy against disease causing pathogens. Among possible candidates, melittin from honey bee venom is of particular interest. The aim of this work was to study the effect of whole bee venom on Lyme borreliosis spirochetes. The minimum concentration of the venom that possesed the inhibitory effect on borrelia growth was identified. The gene enconding melittin was isolated from bee venom gland and recombinant melittin was produced in bacterial expression system. The effect of recombinant melittin on live borrelia culture was studied.
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Antimicrobial properties of mesenchymal stem cells
Černá, Kristýna ; Hájková, Michaela (advisor) ; Vobořil, Matouš (referee)
Mesenchymal stem cells (MSCs) were first isolated from bone marrow. Since that they were isolated from almost every tissue in the body. MSCs are multipotent cells with many properties including self-renewal, differentiation into the cells of all three germ lines, trophic and immunomodulatory abilities, specific migration to the site of injury and recently discovered antimicrobial properties. This properties make them a good candidate for cell therapy because they can be used in treatment of many diseases including bacterial infections. This thesis brings a summary of all discovered antimicrobial properties of MSCs. It is primarily focused on their direct and indirect antimicrobial effect. The direct effect is mediated by a secretion of antimicrobial substances and the indirect effect includes an impact on the activity and functions of macrophages and neutrophils. Extracellular vesicles derived from MSCs constitute separate chapter because they can have both direct and indirect antimicrobial effect. Keywords: mesenchymal stem cells, antibacterial effect, bacteria, antimicrobial peptides
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