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Encapsulation of natural antioxidants
Štindlová, Jitka ; Obruča, Stanislav (referee) ; Márová, Ivana (advisor)
In theoretical part of this diploma thesis the basic properties of antioxidants, especially anthocyanins and phenolic compounds are described, as well as the basic features and principles of free radicals formation. The theoretical part also describes some possible ways of encapsulation of antioxidants into polysaccharide and lipid particles. In the experimental part basic characteristics of extracts from selected lyophilized fruits and vegetables (carrots, apples and mixed berries) are described. As group parameters of plant extracts the total antioxidant activity, content of flavonoids and phenolics, carotenoids, anthocyanins and ascorbic acid were determined. In experimental part also various encapsulation techniques were tested, encapsulation effectiveness of each technique was evaluated and the stability and size of the created particles were determined. As the best encapsulation method in terms of encapsulation efficiency in most of samples/parameters ethanol injection was found. On the other hand the particles prepared by ethanol injection are relatively unstable in terms of zeta potential, which is followed by their tendency to aggregate. As the most stable particles prepared by thin layer evaporation (TLE) and reverse phase thin layer evaporation (RP-TLE) were evaluated. Particles prepared by TLE, RP-TLE and chitosan-alginate particles exhibited a negative charge, while particles prepared by ethanol injection stayed uncharged and chitosan particles have a positive charge.
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Bioactive peptides as a component of anti-aging cosmetics
Jatzová, Katarína ; Obruča, Stanislav (referee) ; Muthný,, Tomáš (advisor)
Ageing is a natural part of every human life cycle. During ageing there are lots of changes in the organism. One of the main pillars of the cosmetics industry is the development of active compounds that are fighting signs of skin ageing. The components as bioactive peptides are considered to be promising anti-ageing products, mainly because of the possibility to precisely define their chemical structure and therefore achieve more effective biological targeting. One of the signs of skin ageing is the weakening of the connections between epidermal cells and the extracellular matrix, decreasing expression of adhesion molecules and molecular components of dermo-epidermal connection. The most abundant adhesive receptors in the skin are integrins. Their ligands are extracellular matrix molecules, e. g. laminin or fibronectin. The minimal recognition sequence of integrins receptors is the amino acid motif arginine- glycine- aspartic acid (RGD). This sequence is also very interesting in terms of cosmetic applications because it provides the ability to create new and effective bioactive peptides. The subject of present work is basic safety testing of four peptides with RGD motif. A sequence of three peptides was modified by addition of glycine amino acids, or alanine. The expected effect was to improve interaction with integrin receptors. In each of the sample, levels of endotoxin was determined in order to exclude any possible interfering effects on the viability of cells. Subsequently, in NIH3T3 mouse fibroblasts viability was monitored by MTT assay and morphology. The quantity of obtained protein had been determined to increase data interpretation relevance.
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Controlled biotechnological production of polyhydroxyalkanoates
Šnajdar, Ondřej ; Pekař, Miloslav (referee) ; Obruča, Stanislav (advisor)
Předložená diplomová práce se zabývá produkcí polyhydroxyalkanoátů (PHA) bakterií Cupriavidus necator H16. Cílem práce byla příprava, selekce a charakterizace mutantních kmenů schopných vyšší produkce PHA. V teoretické části byla zpracována literární rešerše zabývající se nejdůležitějšími typy PHA, bakterií Cupriavidus necator a způsoby indukce mutageneze. V experimentální části byly připraveny mutantní kmeny pomocí fyzikální a chemické mutageneze. Mutantní kmeny schopné nadprodukce PHA byly selektovány pomocí kultivace na minerálním médium s olejem. Pro další studium byly vybrány 4 mutantní kmeny schopné nadprodukce PHA. Tyto mutantní kmeny byly dále podrobeny biochemické charakterizaci. Byly naměřeny specifické aktivity vybraných intracelulárních enzymů včetně enzymů podílejících se na biosyntéze PHA. Také byla naměřena resistence mutantů vůči oxidačnímu stresu. Bylo zjištěno, že mutantní kmeny schopné nadprodukce PHA mají vyšší aktivity enzymů produkujících NADPH. NADPH je jeden z klíčových substrátů ovlivňujících směr toku acetyl-CoA metabolizmem. Vyšší intracelulární koncentrace NADPH parciálně inhibuje Krebsův cyklus a aktivuje akumulaci PHA. Aktivity acetoacetyl-CoA reduktázy a PHA syntázy, enzymů zapojených do syntézy PHA, těchto mutantů proto byly také vyšší stejně jako molekulová hmotnost připravených polymerů. Aplikace fyzikálních a chemických mutagenů je způsob, kterým lze připravit biotechnologicky perspektivní mutantní kmeny schopné nadprodukce PHA.
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The chitosan-glucan complex isolated from Schizophyllum commune
Krčmář, Martin ; Obruča, Stanislav (referee) ; Omelková, Jiřina (advisor)
Chitosan-glucan complex is fungal origin copolymer that finds application in medicine and cosmetics. Traditionally mycelium of Aspergillus and Penicillium is considered as industrial chitosan-glucan complex source, though utilization of Micromycetes in biotechnological productions is sometimes undesirable. The aim of the work was to study the possibility of Basidiomycete Schizophyllum commune submerged cultivation for industrial scale chitosan-glucan complex production use. Within the work there was studied effect of cultivation conditions (type and concentration of carbon sources in nutrient medium, ratio of carbon source to nitrogen source, medium initial pH and aeration intensity) on Sch. commune #127 mycelium growth, chitosan-glucan complex formation and exopolysaccharide synthesis. As the result, the method for chitosan-glucan complex production increase and exopolysaccharide synthesis suppression was suggested. Chitosan-glucan complex from Sch. commune #127 submerged mycelium was separated by successive alkali and acid treatments. Effects of alkali concentration and application technique, and type of acid on physical and chemical properties of chitosan-glucan complex were described. Analytical methods for in process control and final product characteristics were suggested.
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Influence of stabilizers and other components on final quality of mixtures for ice-cream production
Sládková, Veronika ; Obruča, Stanislav (referee) ; Márová, Ivana (advisor)
Presented diploma thesis was focused on study of influence of stabilizers and other components on final quality of mixtures for ice-cream production. This thesis was worked out in cooperation with Frujo Co., Ltd., Tvrdonice to find out the most suitable stabilizer with positive influence on the quality of final product. In experimental part six mixtures of stabilizers, three mixtures for ice-cream production and particular ingredients for production of mixtures for ice-cream production were analysed. Saccharides and lipids were analysed only in mixtures of stabilizers. Content of saccharides was analyzed using spectrophotometry and HPLC/RI. Lipid components were analysed by TLC. Viscosity was measured by rotatory viscosimeter and the differences of flow curves of each mixture with different stabilizers were monitored. Based on viscosity measurements some ice-creams with different stabilizers were chosen for sensory analysis. Mixture of stabilizers “Stobicol” was analysed for active compounds and antioxidant stability. Microbiological stability was tested in all of the stabilizers and also in particular ingredients including final mixture for ice-cream production. In three ice-cream mixtures of different flavour protein composition was measured by microfluidic electrophoresis. The highest content of total and reducing saccharides was determined in hydrolyzed samples. The highest values of total saccharides were found in mixture of stabilizers “Meyprogen”, the highest values of reducing saccharides were detected in stabilizer “Swissgum”. Content of triglycerides was found out almost in all of the measured mixtures of stabilizers using TLC method even though according to formulas provided by producers there should not be any. Diglycerides were detected in “Meyprogen” too even though this mixture of stabilizers should contain only saccharides. Based on results of rheology measurements and in connection with sensory analysis “Meyprogen” stabilizer could be recommended as the best material for the industrial production.
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Properties of biocomposites based on flax fibers
Pelánová, Markéta ; Obruča, Stanislav (referee) ; Přikryl, Radek (advisor)
This thesis is focused on composites, which are composed of degradable components. The theoretical part deals with composite materials, treatment of fibers and properties of composites. The purpose of this study is to create a composite consisting of polyhydroxybutyrate, which is reinforced with flax fibers with various modifications. The thesis also evaluates the effect of reinforcing fibers in the composite material on the tensile properties and hydrolytic stability and degradability of the composite in compost in laboratory conditions.
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Hydrolytic cleavage of coffee spent ground
Skřivanová, Veronika ; Hlaváček, Viliam (referee) ; Obruča, Stanislav (advisor)
The bachelor thesis deals with various types of hydrolysis of spent coffee ground for the following biotechnological applications such as the culture medium. The work included mechanical, acid, alkaline and enzymatic hydrolysis. The resulting hydrolysates were analyzed concerning the content of reducing carbohydrates, and potentially inhibitory substances polyphenolic nature. The results show that the best conditions for culturing yeast samples were prepared with a concentration of coffee 150 g/l hydrolysed combination of 2,5% mineral acid by heating at 121 °C for 15 minutes. Using ultrasonic homogenizer significantly increases the effect of hydrolysis. The enzymatic hydrolysis is best placed hydrolyzate prepared cellulase. The hydrolysed spent coffee grounds were used as the production medium for the production of ethanol by Saccharomyces cerevisiae. For samples was determined concentration change of glucose and ethanol by HPLC/RID. The highest concentration of biomass sample contained hydrolyzed cellulose on the other side, the highest yields of ethanol (20,07 g/l) were achieve using hydrolysate prepared by 2,5% hydrochloric acid.
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Study of bioremediation of some organic polutants by Cupravidus necator
Miléřová, Miluše ; Hurtová, Jana (referee) ; Obruča, Stanislav (advisor)
The aim of this work is the study of bioremediation of p-nitrophenol using bacteria Cupriavidus necator. The theoretical part contains a summary of organic pollutants and the bioremediation aproaches used to remove them from the environment. In the experimental part of this work we used bacteria Cupriavidus necator H16 and its mutant strain Cupriavidus necator H16/PHB-4 unable of PHB accumulation. The experiments were performed with different concetrations of p-nitrophenol and also under none-optimal conditions. Apart from concentration of p-nitrophenol, viability of the bacterial cultures was determined by flow cytometry. During cultivation in progress for sub-optimal conditions with a concentration of 50 mg•l-1 p-nitrophenol at 4°C using a flow cytometer seemed more resistant bacteria Cupriavidus necator H16. On the contrary strain Cupriavidus necator H16/PHB-4 was more resistant at 37 °C. The biodegradation assay was also performed in fermentor, during 51 hours lasting cultivation, bacterial culture degraded 65 % of p-nitrophenol and moreover decreased wavelenght of maximum in absorption spectra from 401 m to 384 nm was observed. The quantity of PHB was analyzed by gas chromatography with FID detector along with the p-nitrophenol determined by spectrophotometry, whose maximum content of PHB was achieved in 20 hour of cultivation
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Utilization of the polyelectrolyte complexes in medicine and cosmetics.
Karásková, Iva ; Obruča, Stanislav (referee) ; Sedláček, Petr (advisor)
This thesis is focused on the possibilities of polyelectrolyte complexes of humic acid and chitosan. In the experimental part, based on a literature search, were optimized appropriate procedures for preparing such complexes in hydrogel form. The main aspects of the preparation, the effect was studied, the influence of the direction of the addition, the concentration, pH and ionic strength. Furthermore, the encapsulation method was tested to give chitosan pearls. Work was aimed for cosmetic and medical applications.
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Microbial production of extracellular polymers and their involvement in stress response
Müllerová, Lucie ; Sedláček, Petr (referee) ; Obruča, Stanislav (advisor)
Bachelor’s thesis is focused on the production of extracellular polysaccharide pullulan by microorganism Aureobasidium pullulans, purification of pullulan and its possible use as a cryoprotectant. As a part of this work a description of A. pullulans, pullulan and an overview of involvement in its stress response were provided. In the experimental part growth characteristics of A. pullulans (the strain CCM 8182 was used) and pullulan production during growth in optimal conditions was analyzed. Biomass production was the highest with fructose as a carbon source (3,580 g/l CCM 8182), the highest pullulan production occurred when using sucrose as a carbon source (10,300 g/l F 148). Among three organic solvents used for pullulan precipitation – ethanol, acetone, isopropylalcohol, ethanol was selected as the best for further experiments in ratio 1:2 (fermentation broth:ethanol). Pullulan purity was characterized by HPLC. As a further part of this work, cryoprotectant properties of pullulan at temperatures – 72° C, -18° C, 4° C and 60°C were studied. The presence of pullulan at temperatures – 72° C and 60° C was found to be detrimental to cell viability. At temperatures – 18° C and 4° C the cryoptotectant activity of the polysaccharide was confirmed.
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