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Optimization of 5-fluorouracil determination by high-performance liquid chromatography
Durychová, Eva ; Křížek, Tomáš (advisor) ; Kozlík, Petr (referee)
This thesis is dedicated to the development of a method for the determination of 5-fluorouracil by high-performance liquid chromatography, which could be used for determination of its encapsulation efficiency by liposomes. First, separation of 1 mM standard of 5-fluorouracil was tested on several types of columns. Among tested columns belonged a C18 column, a C18 column with positive surface modification, a phenyl-hexyl column and several fluoride columns together with two HILIC columns. A mixture containing 10 mM CH3COONH4, pH = 4,5 and MeOH in the ratio of 98/2 was initially used as the mobile phase. The results showed, that 5-fluorouracil elutes on all columns too close to the dead time, where different impurities often elute. To avoid possible distortion of 5-fluorouracil signal by impurities potentially occurring in the real sample, several adjustments of chromatographic conditions were tested. The most effective solution was addition of 5 mM ion-pairing agent (namely tetrabutylammonium chloride hydrate) to the mobile phase, together with adjusting pH to 8,0 to support the ionization of the analyte. In the combination with mobile phase modified in this way, a phenyl-hexyl column with retention time 4,36 minutes reached the best result, therefore it was selected for the final determination of...
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Methods of preparation and characterization of liposomal systems
Chýlková, Patricie ; Venerová, Tereza (referee) ; Jugl, Adam (advisor)
In this bachelor's thesis the properties of liposomes prepared by reference thin-film rehydration method and Mozafari method were compared. The aim of the experimental part was to determine whether the Mozafari method is convenient alternative for liposome preparation and whether thin-film rehydration and Mozafari methods produce liposomes with the same properties. The prepared liposomes were characterized in terms of size, polydispersity and zeta potential using dynamic light scattering on certain days for one month since the day of preparation. It was determined that electrostatically or sterically stabilized liposomes of identical composition prepared by either method had mutually corresponding properties.
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Optimization of Combined Chitosan Particles and their Characterization for Cosmetic use
Mrůzková, Barbora ; Dzurická, Lucia (referee) ; Bendová, Agáta (advisor)
The bachelor thesis focused on optimizing preparation and characterization of chitosan coated liposomes with selected active substances and their use for cosmetic applications. The theoretical part of the thesis describes natural substances, their effect on the skin, encapsulation methods, and the preparation and use of liposomes and chitosomes. In the experimental part, the optimization of the preparation of liposomes and chitosomes and their subsequent characterization by measuring particle size, polydispersity index, zeta potential, encapsulation efficiency and gradual release of active substances were performed. The selected active substances in this work were vitamin C, -carotene, caffeine and vitamin E. From the data obtained, the method of adding liposomes dropwise into the 2% chitosan with a short mixing time and ultrasonication for 1 minute was selected as the most suitable method for the preparation of chitosomes usable in cosmetics. Furthermore, the encapsulation efficiency of the particles and the antioxidant capacity of vitamin C, -carotene, caffeine and vitamin E were compared. The best antioxidant properties were determined for vitamins C and E; therefore, the gradual release of chitosomes with these active substances was subsequently measured in an environment simulating cosmetics. At the end of the work, cosmetic emulsions and gels containing selected chitosomes were prepared and tested on the skin by determining changes in desquamation.
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Rheological characterization of polysaccharide gels with incorporated vesicles
Kalendová, Lucie ; Smilek, Jiří (referee) ; Venerová, Tereza (advisor)
This diploma thesis deals with the rheological characterization of polysaccharide gels with incorporated vesicular systems. Positively charged Ion Pair Amphiphile vesicles (IPA) consisting of cetrimethylammonium bromide, sodium dodecyl sulfate, dioctadecyldimethylammonium chloride and cholesterol were chosen for the incorporation. Other vesicles used for incorporation were oppositely charged liposomes made of phosphatidylcholine (DPPC), phosphatidic acid (PA) and cholesterol. The thesis is divided into two sections. The first one deals with preparation, characterization and stability control (temperature and time stability) of both vesicular systems by using dynamic light scattering (DLS). In this part, the vesicles’ stability was confirmed which made them perfect candidates for the following incorporation. The second part of the thesis describes the preparation and rheological characterization of selected gels. Three types of gels were prepared in total, specifically 1% and 2% gellan gum gels and 4% hyaluronic gel. All of them were studied in water, as well as in solutions of IPA vesicles or DPPC liposomes, both of same concentration (1 mmoll-). The subsequent rheological characterization including amplitude, frequency, and stress relaxation sweeps demonstrates there is no significant difference between reference gels and those containing vesicles. This indicates that vesicles used in this thesis were only freely stored in the gel networks, meaning the structure of the gels didn’t affect their transport properties. This is a significant information for using these systems in drug delivery.
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Development of natural cosmetics emulsions with encapsulated active substances
Chrápavá, Edita ; Dzurendová, Simona (referee) ; Márová, Ivana (advisor)
In recent years, there is a increase of consumer interest in products, which in some way help a more favourable ecological sustainability, whether it is a reduction of emissions during mass production, easier breakdown of chemicals in wastewater, or harmlessness of products and ingredients from which products are made. This topic also affected the cosmetic industry in a big way, and they were thus forced to start adapting not only smaller brands of natural cosmetics, but also large companies. As a result, bigger companies started to pay more attention to research of natural substances for cosmetics and the development of natural alternatives to synthetic ingredients. At the same time, there is also increasing interest in the effects of Eastern medicine mushrooms and their beneficial effects on both physical and mental health. This thesis focuses on the connection of these two topics, the analysis of plant extracts and extracts of medicinal mushrooms on one side and their potential cosmetic use on the other. Subsequently, a cosmetic product was designed made from ingredients of natural origin with encapsulated extracts from these natural ingredients as active substances
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Study of stability of antimicrobial nanomaterials in different environment by HPLC
Janderová, Šárka ; Trachtová, Štěpánka (referee) ; Márová, Ivana (advisor)
This thesis deals with the preparation of nanomaterials (nanoparticles, nanofibers) with encapsulated active substances and the stability of these nanomaterials in different environments using liquid chromatography. The development and optimization of methods for monitoring the kinetics of release of encapsulated substances from nanomaterials into various environments play a key role in research aimed at their utilization in the pharmaceutical, food, and cosmetic industries, and also contribute to the development of new antimicrobial nanomaterials limiting antibiotic resistance. The theoretical part focuses on the characterization of nanomaterials (nanoparticles, liposomes, nanofibers, and combined nanomaterials). Another section of the literature review deals with natural (eugenol, carvacrol, curcumin, thymol, vitamin E) and synthetic (ampicillin, streptomycin) antimicrobial substances. The final chapter discusses the instrumental methods used for the characterization of nanomaterials and the evaluation of their stability, released, and degradation products. In the practical part, the antioxidant activity of selected substances was first monitored. Liposomes were prepared, and their stability and encapsulation efficiency were examined. An HPLC method for determining active substances was optimized. Subsequently, the gradual release of the mentioned active substances from prepared liposomes was monitored. Nanofibers were prepared using forcespinning, and their release into selected model environments was also analyzed. Distilled water with minimal ion and impurity content served as a model environment, as well as a physiological solution at 25°C, simulating the temperature of the human body surface, and a physiological solution at 37°C, characteristic of human body temperature. This analysis provides a comprehensive view of the behavior of prepared materials and their potential practical applications. In the final section, combined nanomaterials (based on nanofibers and nanoparticles) with encapsulated active substances were prepared. The gradual release of active substances and any potential synergistic effects were examined for these nanomaterials.
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Optimization of Combined Chitosan Particles and their Characterization for Application Use
Netopilík, Tibor ; Hudečková, Helena (referee) ; Márová, Ivana (advisor)
The thesis deals with the preparation and characterization of nanoparticles for cosmetic and food applications. The thesis explores various methods of preparing nanoparticles, including liposomes and chitosomes, and analyses their properties and stability in different model environments. The experimental part includes the determination of the encapsulation efficiency of different vitamins by HPLC and UV-VIS spectrophotometry. Results show the influence of various factors such as time, type of environment and method of preparation on stability and efficiency of the nanoparticles. Furthermore, the optimization of the preparation is carried out chitosomes using ultrasound and magnetic stirrer. Cosmetic products enriched with these nanoparticles were developed and their sensory analysis was proved. This work contributes to the understanding of nanoparticle preparation processes and provides useful information for the development of new cosmetic and food products with improved properties and stability.
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Preparation of nanoparticles and nanofibers with antimicrobial components
Kubišová, Veronika ; Slaninová, Eva (referee) ; Skoumalová, Petra (advisor)
This thesis addresses the problem of inadequate current wound therapy and presents a solution in the form of nanomaterial-based wound dressings (coverings). Specifically, it focuses on the development and characterization of various nanofibrous materials with integrated liposome particles that could serve as a source of therapeutic agents and be useful specifically in the field of a wound therapy. The review in the introductory part of the thesis first focused on the mentioned problem, which describes the shortcomings of existing conventional dressing materials. A description of human skin from an anatomical and functional point of view was not omitted, nor was the wound healing process itself. Different types of covering materials were also presented. However, a large part was focused on nanomaterials and their use in the field of the skin wound therapy. The nanomaterials mentioned were mainly liposome particles and nanofibres, as the experimental part of the work was focused on these structures. The description dealt with the characterization of these structures as well as the biopolymers used for their preparation. However, the aim was to prepare nanofibers with liposome content, so the method of forming such systems was described. Various therapeutic agents are also an integral part of the covering materials, especially those that suppress the development of infection and reduce wound pain; therefore, the search focused on the antibiotic ampicillin and the analgesic ibuprofen. The experimental part was devoted to the actual production of nanofibers with liposomes and also to the qualitative demonstration of the presence of liposomes in electrostatically prepared nanofibers. The selected polymeric components of these systems were polyhydroxybutyrate (PHB) and gelatine. However, the nanofibers and liposome particles (as well as combined liposomes with PHB) were first prepared in their own form and characterized mainly in terms of the gradual release of the drug substances. The results obtained were then compared with those of therapeutic drug delivery via combined nanofibrous structures with liposomes. For these combined structures, the aim was to achieve synergy in drug delivery between these systems. The aim of this work was to create a biomaterial covering with the controlled drug release. The drugs contained in these materials were the aforementioned ampicillin and ibuprofen. An important part of the work was then to determine the safety of the prepared materials which were tested for cytotoxicity, where the tests performed were MTT test and LDH test. And the actual wound healing ability of the nanofibers was then monitored in a scratch test or a "wound healing" test. At the end of the paper, recommendations for future work on this topic are given.
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Characterization and stabilization of pancreatin
Wurstová, Agáta ; Němcová, Andrea (referee) ; Obruča, Stanislav (advisor)
This work focuses on a study of enzyme mixture pancreatin, its characterization and subsequent encapsulation into liposomes. As a reference proteins bovine serum albumin and trypsin were used. Characterization of pancreatin consisted of two parts. The first part focuses on optimization of methods for the concentration determination by absorption spectrophotometry using basic methods for identifying proteins (Biuret method, Hartree-Lowry method and Bradford method). Moreover, UV spectrums of the protein were measured. As a method for identification of protein´s molecular weight, SDS-PAGE was used. To identify components of pancreatin, LPLC was employed in two modifications, ion-exchange chromatography and size exclusion chromatography. The second part is dedicated to the characterization of pancreatin as enzyme in terms of pH and temperature optimum for the enzyme activities of protease (pH 9, 8 and 50 °C), amylase (pH 7 and 40 °C) and lipase (pH 7 and 50 °C). The last part of this work aimed at an encapsulation of pancreatin into liposomes and DLS analysis of distribution of particles and their zeta potential. Liposomes did not spontaneously release encapsulated enzyme. To confirm that proteins were successfully entrapped into liposomes, their structure was disrupted by application of phospholipase D. In conclusion, liposomes can be utilized as delivery systems for native enzymes.
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Development of a new type of energy drinks containing natural active components
Vysoká, Marie ; Němcová, Andrea (referee) ; Skoumalová, Petra (advisor)
The aim of this thesis is the development and preparation of a new energy drink. The theoretical part of the thesis focuses on the comparison of the drinks available in market, the presentation of the main active ingredients and the proposal of enriching the new energy drink. It also describes the possibility of using an encapsulation and the method used to determine specific compound in energy drink or encapsulated particles. In the experimental part of the work, at the beginning, the main content of energy drinks were characterized, for instance caffeine, sugar, polyphenol and vitamin C. Subsequently the energy drink based on selected types of loose teas that were enriched with the encapsulated extract of acai, acerola, maca, goji and ginseng was designed. All extracts contained large amounts of polyphenols and antioxidants. In addition, acerola extracts contained very high amounts of vitamin C. Two types of particles, alginate particles and liposomes were prepared for encapsulation. For all particles, encapsulation efficiency, size, stability, and antioxidant activity were determined. Long-term stability was then monitored within three months. The particles were also exposed to artificial gastric, pancreatic and bile juice. All tested particles showed very good encapsulation efficiency. In terms of long-term storage stability in an aqueous environment. Particularly liposome particles, which also showed high stability by zeta potential measurement, were suitable. On the contrary, in case of alginate particles, when stored in an aqueous medium, the release of active substances occurred. Finally, an energy drink was created and subjected to sensory analysis. The best energy drink was based on black or green tea with alginate particles acai, goji and ginseng.
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