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The Study of Biopolymers Stability Using Light Scattering Techniques
Kratochvíl, Zdeněk ; Sedláček, Petr (referee) ; Kalina, Michal (advisor)
The stability of chosen biopolymers (or simple carbohydrates and amino acids) was in-vestigated using light scattering methods. Samples were prepared by dissolving the sub-stance in deionised water or in diluted acetic acid. Firstly, the effect of increasing concen-tration on zeta potential was observed and the optimal concentration for the next measure-ments was determined. Afterwards, the stability and the molecular weight of used sub-stances were studied during long-time storage in different time intervals. It was found out that chitosan, CMC, hyaluronate and sodium alginate were subjected to degradation whereas increase of the molecular weight and the zeta potential was observed in case of BSA. In the end, the samples were exposed to several different external effects, namely the influence of different temperatures, UV radiation and increasing ionic strength. The results showed that rather higher temperature has a significant impact on biopolymers stability. However, used biopolymers were hardly affected by UV radiation except BSA whose molecular weight increased considerably. Decrease of the zeta potential absolute values and the molecular size usually occurred with increasing ionic strength. The change of molecular weight was not detected.
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Experimental study on the release of active ingredients from gel matrices
Volhinava, Anastasiya ; Enev, Vojtěch (referee) ; Sedláček, Petr (advisor)
This thesis deals with the characterization of hydrogels containing humic substances and salicylic acid. Experimental study of the transport of active substances was performed from the gel matrix over the skin and synthetic membranes. During this work was writing literature review focused on the use of gels with humic substances in cosmetic, medical and pharmaceutical industries. The model humic hydrogels was prepared with salicylic acid and their characterization by basic methods of material analysis. These basic methods were and visual assessment of consistency during the preparation and rheology. The study of the release of active and humic substances was performed due to the vertical diffusion cells. The amount of released humic substance was characterized by UV-VIS method and the amount of released salicylic acid from gel matrix was determined by HPLC.
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Utilization of vibrational spectroscopy for in-situ qualitative and quantitative analysis of polyhydroxyalkanoates in microbial biomass
Kevélyová, Barbora ; Enev, Vojtěch (referee) ; Sedláček, Petr (advisor)
The aim of this bachelor thesis was to evaluate the applicability of selected methods of vibrational spectroscopy for qualitative and quantitative analysis of biomass, which contains polyhydroxyalkanoates. Fourier-transform infrared spectroscopy, that uses attenuated total reflection technique (ATR-FTIR), was chosen for this experiment. Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) has been studied in Cupriavidus malaysiensis, Thermomonas hydrothermalis and Aneurinibacillus thermoaerophilus sp. H1. Data were obtained by evaluating the infrared spectra, which were then compared with the results of performed gas chromatography. Visual changes in the spectra were also observed along with crystallinity. No correlation was detected between the data obtained by gas chromatography and by the method ATR-FTIR in quantitative analysis. Thus, this method is not suitable for determining the content of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) in biomass in the set experiment and the bacteria used. On the other hand, the method was successfully used for in-situ qualitative analysis of copolymer in the samples of Aneurinibacillus thermoaerophilus sp. H1. The degree of data correlation was relatively high in the given experiment. With these findings, ATR-FTIR is a suitable analytical method for determining the monomer composition of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) in this strain. Comparison with the PHA producer Cupriavidus malaysiensis showed that this method is not universal for all types of microorganisms observed in this thesis.
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Preparation and characterization of novel application forms of humic substances in combination with water sorbents
Nováčková, Táňa ; Pekař, Miloslav (referee) ; Sedláček, Petr (advisor)
This bachelor’s thesis discusses an applicability of hydrogel forms-based composite materials of humic acids and commercial water sorbent. According to the literature review, methods of incorporation of humic acids into commercial sorbents of water under various conditions and the resulting materials were subsequently characterized. In the characterization, major attention was paid to the determination of the water content and the swelling characteristics. Preparation of the composite materials was directed to use their sorption abilities and ability to control the release of humic substances in agriculture.
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Utilzation of physico-chemical analyses in study on microbial stress-response
Slaninová, Eva ; Pekař, Miloslav (referee) ; Sedláček, Petr (advisor)
This diploma thesis deals with the utilization of physico-chemical analysis in study on microbial stress–response. The main content of this work was to propose and optimize techniques and methods which are generally used in different industries. Two bacterial strains, Cupriavidus necator H16 and its mutant strain Cupriavidus necator PHB-4 producing polyhydroxyalkanoates (PHA) under certain conditions, were used as model microorganisms. Initially, microscopic teques such as transmission electron microscopy (TEM), cryo scanning electron microscopy (cryo SEM) and atomic force microscopy (AFM) were proposed and tested for the characteriozation of morphological differences of bacteria. Furthermore, viscoelastic properties of bacteria were determined and compared by the oscillatory tests of the rheology method considering another type of samples. Thermal analysis methods, differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA) in particular, focused on water transport and behavior of intracellular water influenced by presence of the granules of PHB in bacteria. The last technique, utilized in the study, was dynamic and electrophoretic light dispersion during pH changes for characterization of the surface properties of bacteria such as size, zeta potential and the isoelectric point.
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Preparation and application particles from copolymer of P(3HB-co-4HB)
Ostanina, Kseniia ; Sedláček, Petr (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis deals with preparation of particles from poly(3-hydroxybutyráte-co-4-hydroxybutyráte). Copolymer was dissolved in acetone and then the solution had been dosed into the anti-solvents. Acetone and anti-solvent are perfectly miscible. Theoretical part focuses on the properties of the copolymer, its production and potential use in medical applications. In the experimental part the copolymer was dripped into selected anti-solvents by syringe and pipette above and below the surface of anti-solvent. The following anti-solvent were chosen: water, methanol, dimethylsulfoxid (DMSO), 50 % acetone-water, 75% acetone-water, 50 % methanol-water, 75 % methanol-water mixtures. There were three copolymers used with different compositions of 4HB units (63 %, 87 % a 33 %). Copolymer composition was analyzed by gas chromatography. Copolymers were prepared by bacterial strain Aneurinibacillus sp. H1. Due to the results particles of size approximately 100-500 nm can be prepared by methods described in this work. The produced particles can be successfully used in medical applications.
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Modelling of particle movement in liquid and hydrogel media
Orság, Miroslav ; Sedláček, Petr (referee) ; Pekař, Miloslav (advisor)
Main goal of this thesis was to became familiar with general mathematical description of particle motion in fluid and hydrogel media in COMSOL software. Simulate a basic Brownian motion of particles with various diameter in media with different density. According to software options, simulate motion in polymer structure or in viscoelastic media. Consider an options of the model for use in microrheology and suggest next process in this field.
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Preparation, characterization and evaluation of application potential of semiIPN hydrogels
Papežíková, Hana ; Pekař, Miloslav (referee) ; Sedláček, Petr (advisor)
Aim of this thesis os to optimize set up of hydrogel based on semi-interpenetrated polymer networks. Polyhydroxyethylmethyl acrylate was chosen as an model example of hydrogel due to its common use in biomedicine. At first, thermoiniciation was used for preparation and changed to photoiniciation afterwards. Following the optimisation of the hydrogel set up with proper qualities, the polyelectrolyte sodium polystyrene sulfonate was implanted. Object of research was its impact on structure, flowing and transport features of hydrogels. All qualities were tested by drying, rheology, and fluorescence correlation spectroscopy. Results clearly validate influence of hydrogel features by polyelectrolytes.
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Adaptation to adverse osmotic conditions as a tool for evolutionary enginnering of bacteria
Drotárová, Lenka ; Nováčková, Ivana (referee) ; Sedláček, Petr (advisor)
This bachelor thesis deals with the application of osmotic stress as a tool for evolutionary engineering of PHA producing bacterial strains. The aim of this thesis is to evaluate a bacterial adaptation to hypoosmotic environment, as an engineering tool in order to increase the production of PHA. The theoretical part focuses on the evolutionary engineering principle, methods of the strategy and the effect of physical factors on microorganism. The aim of experimental part was to performed an adaptive evolutionary experiment with the bacterial strain Halomonas halophila CCM 3662. Reduced osmotic pressure was used as a stressing factor during the serial cultivation. In order to generate PHA producing mutant strains, each passage was characterized using spectrophotometric and gravimetric method and by GC-FID. It was found that after the long-term cultivation, the mutant strain HH35, cultivated in 35 g/l NaCl, was associated with the highest biomass and PHB concentration. The 15th and 30th passages, along with the wild type strain H. halophila were subjected to further cell-robustness analysis with the application of hyper- and hypoosmotic shock. The stress response, viability of cells and morphological changes were analyzed using FC and TGA methods. Isolated polymers were characterized using FTIR analysis.
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Structure and properties of bacterial biofilms
Mrázová, Kateřina ; Sedláček, Petr (referee) ; Obruča, Stanislav (advisor)
This thesis deals with the study of basic properties of cells of bacterial biofilm produced by PHA producing bacteria, specifically Burkholderia cepacia and Burkholderia sacchari. Two cultivation approaches were used to compare different types of cells: at first, the cells were cultivated in bioreactor providing both planktonic and biofilm attached cells, secondly, cells grown on Petri dishes were also included in the study. In experimental part the affinity of each type of cells to hydrophobic phase was studied using BATH method. It was found that sessile cells, either biofilm associated cells as well as cells taken from Petri dishes, prefer hydrophobic phases more than planktonic cells. Also size and Zeta potential were measured using Dynamic Light Scattering (DLS). It was found for studied samples that, cells in these samples tend to aggregate and the difference of size between both types of cells occurs only for biofilm of the strain Burkholderia cepacia. Finally the impact of presence various bacterial cells types on surface tension of water as well as amount of PHA cumulated in different types of cells were observed.
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