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Soil application of hydrogels containing rhizobacteria
Hlaváčková, Barbora ; Černayová, Diana (referee) ; Kalina, Michal (advisor)
Plant growth-promoting rhizobacteria (PGPR) are a group of bacteria that can influence plant growth by their mechanisms. In recent years, there has been an increase in the use of these soil-supporting products in the form of biofertilizers to increase yields and improve crop growth in adverse conditions in an environmentally friendly way. The aim of this bachelor thesis was to assess the effect of the external addition of different application forms of PGPR on soil physico-chemical characteristics and on the growth of model plants of lactuca sativa without irrigation restriction in the first case and in the second case under irrigation restricted conditions. A particular strain of Azotobacter vinelandii CCM 289 was selected for the preparation of four forms of PGPR treatments, namely bacterial biomass suspended in PBS, alginate gel without bacterial culture, gel with bacterial culture and lyophilized gel with bacterial culture. The effect of the different treatments was compared with the negative control without the addition of PGPR, as different forms of treatments may achieve different efficiencies of rhizosphere colonization, which in consequence may have different effect on plant growth. To assess the effect of PGPR addition, soil physico-chemical characteristics (moisture content, pH) and growth characteristics of lactuca sativa such as number of leaves, height and width of aerial parts of these model plants were measured during the cultivation experiments. After the termination of the cultivation experiments, the basic growth parameters of the model plants from each treatment such as total plant length, length of aerial parts and roots of the model plants, as well as total weight of each plant and weight of their aerial parts and roots were determined. After drying, the total dry weight of individual plants and the dry weight of the aerial parts and roots were characterized. After drying, root density was analyzed by performing root scans and compared between treatments. Soil extracts from individual treatments after the cultivation experiments were analyzed to assess the effect of PGPR addition on the microbial activity of the soil used in the experiments by plate cultivation using Petri dishes with agar medium, and the average well colour development (AWCD) value determined using the BIOLOG EcoPlate kit. The obtained results were compared between the individual treatments and also between cultivation experiments conducted with different irrigation conditions. Physico-chemical characterization (pH, conductivity) was performed on soil extracts from the drained soils after the cultivation experiments. The amount of organic and inorganic fractions was analyzed on the dried soils of each treatment by TGA. The results were compared with each other and the effect of different forms of external addition of PGPR on the physico-chemical characteristics of the selected soil under conditions without irrigation limitation and under conditions with irrigation limitation during the experiments was assessed.
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Optimization of the cultivation media for thermophilic bacterium Caldimonas thermodepolymerans
Fajtová, Zuzana ; Černayová, Diana (referee) ; Obruča, Stanislav (advisor)
The subject of the bachelor ‘s thesis is the optimization of the cultivation conditions of the thermophilic bacterium Caldimonas thermodepolymerans. The theoretical part provides a general characterisation of PHA materials, their synthesis, degradation, and applications. In addition, extremophilic microorganisms and the bacterium C. thermodepolymerans are discussed in the theoretical part. The experimental part of the thesis focuses on the production of the PHB by the bacterium C. thermodepolymerans DSM 15344. A series of experiments were carried out to optimise the cultivation conditions. The influence of temperature, inoculum ratio, inoculum age, mineral medium composition, and concentrations of nitrogen sources in the mineral medium on culture growth and PHB production was investigated. It was found that the original conditions set were already close to optimal for the cultivation. A higher yield compared to the control conditions was observed with a combination of 0.5 g/l yeast extract and 1.5 g/l ammonium chloride, resulting in a 7% increase in PHB.
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Analytical methods for qualitative and quantitative determination of PHA in cyanobacteria
Černayová, Diana ; Samek, Ota (referee) ; Sedláček, Petr (advisor)
The diploma thesis is confused to verify the applicability of selected physicochemical and spectroscopic methods for characterization of cyanobacteria, with special emphasis on possibilities of qualitative and quantitative analysis of polyhydroxyalkanoates (specifically polyhydroxybutyrate (PHB)) accumulated in cyanobacterial cells. The sample basis of the work was formed by cultures of cyanobacterial strains of Synechocystis sp. PCC 6803 and Synechocystis salina CCALA 192. The cultures were were cultivated in several ways to cover the widest possible range of physiological conditions and PHB contents, in particular using an autotrophic way of cultivation on shakers and multicultural culture method in a basic culture medium,and in media enriched with 2% salt (NaCl ) as well as mixotrophic culture media with different types of the carbon substrate. After few weeks of cultivation, cyanobacterial cultures were obtained and complexly analyzed by following techniques- cell suspensions were analyzed by flow cytometry and UV-VIS spectrometry (transmission and diffusion transmission mode), dry cell biomass was characterised by gas chromatography to obtain a exact amount of PHB, and then FT-IR spectrometry and thermogravimetric analysis. The work aimed to assess whether any of these methods can be a quick and affordable alternative to the determination of PHB content to the most commonly used method of gas chromatography, but also to assess what additional information about the physiological state of cyanobacterial cells can provide test methods. The highest correlation on PHB content was determined for the parameters determined by infrared spectroscopy, in which specific peaks from the characteristic wavelengths for polyhydroxybutyrate were important. Weak correlations on PHB content were achieved in thermogravimetric analysis and cytometry, using the hydrophobic fluorescent probe BODIPY 439/503, which bound to lipophilic parts of cells. In addition to the determination of PHB, it was possible to determine pigments present in cyanobacteria (such as chlorophyll, phycocyanin and carotenoids) by flow cytometry and UV-VIS diffusion transmission spectrometry. In the end, results from all used techniques were compared by PCA analysis to determine the similarity of all analyzed samples.
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Polyphenols in nutritions and their effect on DNA
Osorio, Juan ; Černayová, Diana (referee) ; Brázda, Václav (advisor)
Epidemiologické studie prokázaly vliv konzumace rostlinných potravin v prevenci široké škály nemocí. Přírodní antioxidanty přítomné v těchto potravinách, mezi nimiž jsou velmi důležité polyfenoly, mohou být zodpovědné za tuto činnost podporující zdraví. Cílem bakalářské práce je ukázat interakci určitých polyfenolů s genetickým materiálem prostřednictvím různých signálních mechanismů, zejména pokud jde o stabilizaci nekanonické struktury DNA G-kvadruplex a poukázat tak na nejselektivnější látku pro inhibici biochemických procesy. Dále práce obsahuje podrobné informace, které mohou pomoci pochopit, jak mohou polyfenolové sloučeniny interagovat s DNA prostřednictvím epigenetických mechanismů a G4 struktur, a které faktory mohou ovlivnit jejich účinnost. Různé experimenty, biologickým a experimentálním opakováním, byly použity k potvrzení interakce mezi sloučeninami a DNA.
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Analytical methods for determination of polyhydroxyalcanoates content in bacterial cells
Černayová, Diana ; Enev, Vojtěch (referee) ; Sedláček, Petr (advisor)
This bachelor thesis deals with methods that are used for determination of polyhydroxyalkanoates (PHA) in cells of bacteria Cupriavidus necator H16. To the applied methods belong Fourier transform infrared spectroscopy, Raman spectoscopy, turbidimetry and thermal analysis (thermogravimetric analysis and differential scanning calorimetry). The results of each method were compared to amount of PHA determinated by gas chromatography. Fourier transform infrared spectroscopy and Raman spectroscopy revealed a linear relation between ratio of characteristic peaks and amount of PHA. FTIR enabled to determine not only the quantitative amount of PHA, but the quality of the sample (cristallinity, amorphous state) as well. Another method was turbidimetry, which has shown correlation between dispersion of UV-VIS light to amount of PHA in bacterial cells. Thermogravimetric analysis was the most effective technique for determination of PHA. The dependence of weight loss of PHA on its content (found by gas chromatography) was determined with the highest accuracy to gas chromatography. Tha last method- DSC was not alternative replacement for quantitative PHA determination. However, DSC indicated crystalline and amorphous state of the polymer.
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Production of polyhydroxyalkanoates by bacterium Schlegelella thermodepolymerans
Černý, Vojtěch ; Černayová, Diana (referee) ; Pernicová, Iva (advisor)
The aim of this work was to study capability of various strains of thermophilic bacteria Schlegelella thermodepolymerans incorporate diverse monomer units to structure of polyhydroxyalkanoates (PHA). Precursors for production mcl-PHA, 4-hydroxybutyrate and 3-hydroxyvalerate were tested. The theoretical part deals with characteristic of studied polymers, properties and applications and usage of thermophilic bacteria in industry. In experimental part there were undergone cultivations with different precursors, which were added in different times and concentrations. Four bacterial strains S. thermodepolymerans from various public collections of microorganisms were used during these cultivations. Levulinic acid, added at start of cultivation, was chosen as the best precursor. Afterwards there was cultivation with concentration line to find out, whether concentration influence composition of copolymer. Increasing the amount of precursor led to decreasing quantity of biomass and PHA, but there were very interesting materials with high amount of 3HV. Addition 8 g/l of precursor led up to 52 %. Subsequently there was found out molecular height of polymers by size-excluion chromatography (SEC) combined with multi-angle light scattering (MALS).
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Polyphenols in nutritions and their effect on DNA
Osorio, Juan ; Černayová, Diana (referee) ; Brázda, Václav (advisor)
Epidemiologické studie prokázaly vliv konzumace rostlinných potravin v prevenci široké škály nemocí. Přírodní antioxidanty přítomné v těchto potravinách, mezi nimiž jsou velmi důležité polyfenoly, mohou být zodpovědné za tuto činnost podporující zdraví. Cílem bakalářské práce je ukázat interakci určitých polyfenolů s genetickým materiálem prostřednictvím různých signálních mechanismů, zejména pokud jde o stabilizaci nekanonické struktury DNA G-kvadruplex a poukázat tak na nejselektivnější látku pro inhibici biochemických procesy. Dále práce obsahuje podrobné informace, které mohou pomoci pochopit, jak mohou polyfenolové sloučeniny interagovat s DNA prostřednictvím epigenetických mechanismů a G4 struktur, a které faktory mohou ovlivnit jejich účinnost. Různé experimenty, biologickým a experimentálním opakováním, byly použity k potvrzení interakce mezi sloučeninami a DNA.
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Analytical methods for qualitative and quantitative determination of PHA in cyanobacteria
Černayová, Diana ; Samek, Ota (referee) ; Sedláček, Petr (advisor)
The diploma thesis is confused to verify the applicability of selected physicochemical and spectroscopic methods for characterization of cyanobacteria, with special emphasis on possibilities of qualitative and quantitative analysis of polyhydroxyalkanoates (specifically polyhydroxybutyrate (PHB)) accumulated in cyanobacterial cells. The sample basis of the work was formed by cultures of cyanobacterial strains of Synechocystis sp. PCC 6803 and Synechocystis salina CCALA 192. The cultures were were cultivated in several ways to cover the widest possible range of physiological conditions and PHB contents, in particular using an autotrophic way of cultivation on shakers and multicultural culture method in a basic culture medium,and in media enriched with 2% salt (NaCl ) as well as mixotrophic culture media with different types of the carbon substrate. After few weeks of cultivation, cyanobacterial cultures were obtained and complexly analyzed by following techniques- cell suspensions were analyzed by flow cytometry and UV-VIS spectrometry (transmission and diffusion transmission mode), dry cell biomass was characterised by gas chromatography to obtain a exact amount of PHB, and then FT-IR spectrometry and thermogravimetric analysis. The work aimed to assess whether any of these methods can be a quick and affordable alternative to the determination of PHB content to the most commonly used method of gas chromatography, but also to assess what additional information about the physiological state of cyanobacterial cells can provide test methods. The highest correlation on PHB content was determined for the parameters determined by infrared spectroscopy, in which specific peaks from the characteristic wavelengths for polyhydroxybutyrate were important. Weak correlations on PHB content were achieved in thermogravimetric analysis and cytometry, using the hydrophobic fluorescent probe BODIPY 439/503, which bound to lipophilic parts of cells. In addition to the determination of PHB, it was possible to determine pigments present in cyanobacteria (such as chlorophyll, phycocyanin and carotenoids) by flow cytometry and UV-VIS diffusion transmission spectrometry. In the end, results from all used techniques were compared by PCA analysis to determine the similarity of all analyzed samples.
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Analytical methods for determination of polyhydroxyalcanoates content in bacterial cells
Černayová, Diana ; Enev, Vojtěch (referee) ; Sedláček, Petr (advisor)
This bachelor thesis deals with methods that are used for determination of polyhydroxyalkanoates (PHA) in cells of bacteria Cupriavidus necator H16. To the applied methods belong Fourier transform infrared spectroscopy, Raman spectoscopy, turbidimetry and thermal analysis (thermogravimetric analysis and differential scanning calorimetry). The results of each method were compared to amount of PHA determinated by gas chromatography. Fourier transform infrared spectroscopy and Raman spectroscopy revealed a linear relation between ratio of characteristic peaks and amount of PHA. FTIR enabled to determine not only the quantitative amount of PHA, but the quality of the sample (cristallinity, amorphous state) as well. Another method was turbidimetry, which has shown correlation between dispersion of UV-VIS light to amount of PHA in bacterial cells. Thermogravimetric analysis was the most effective technique for determination of PHA. The dependence of weight loss of PHA on its content (found by gas chromatography) was determined with the highest accuracy to gas chromatography. Tha last method- DSC was not alternative replacement for quantitative PHA determination. However, DSC indicated crystalline and amorphous state of the polymer.
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