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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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Utilization of interfacial rheology to study of biological systems
Kachlířová, Helena ; Kalina, Michal (referee) ; Smilek, Jiří (advisor)
The aim of this bachalor thesis is to implement and optimization a relatively new method of interfacial rheology. The optimization of this method was realized on two types of interfaces, specifically liquid-liquid using immiscible systems water-chloroform and water-toluene, and water-air. The applicability of this method was tested by using real samples in form of surfactant septonex and biological system represented by protein (bovine serum albumin). The interfacial layers formed by septonex were purely viscous, where the rigidity of the layer increased slightly with concentration until reaching critical micellar concentration and then the rigidity decreased. In comparison with septonex, the interfacial film made of bovine serum albumin showed predominantly elastic behaviour.
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Electrochemical impedance spectroscopy as a nanostructured bioelectrodes characterization method
Vrbová, Eva ; Urbánková, Kateřina (referee) ; Hrdý, Radim (advisor)
Diploma thesis deals of nanostructured surfaces, nanoparticles and electrochemical characterization methods such as cyclic voltammetry, differential pulse voltammetry and electrochemical impedance spectroscopy. The aim of this thesis is a theoretical research issues of production and characterization nanostructured modified electrodes. The practical part is the production of biomodified nanostructured electrodes by anodi- zation W/Al layers with galvanic deposition of gold or deposition of mercury, a modifi- cation of the electrodes by 11-mercaptoundecanoic acid and by bovine serum albumin (BSA). The thesis includes SEM images of nanostructured electrodes contact angle mea- surements of these electrodes and form an electrical circuit with subsequent simulation waveforms.
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Characterization of hyaluronan interactions with albumin
Valentová, Kristýna ; Kratochvílová, Romana (referee) ; Chytil, Martin (advisor)
This bachelor thesis focuses on the research of physical interactions between hyaluronic acid and a protein albumin. In order to investigate these interactions following methods were used: pH and conductivity measurement, viscosimetry, turbidimetry and rheometry. The interactions were observed in solutions of fixed albumin concentration 1.0 gdm-3 and varying concentrations of hyaluronic acid from 0.1 to 2.0 gdm-3. Results were compared with a solution of hyaluronic acid without the addition of albumin. The results show that there are interactions between hyaluronic acid and albumin solutions, especially for those containing high molecular weight hyaluronic acid. The interactions exhibited a change in viscosity of solutions. We can see a reduction in viscosity after adding albumin to the reference solutions. The reduction is noticeable even in the low molecular weight HA solutions, but is not so significant.
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Synthesis of quantum dots for proteins detection
Šibíková, Anna ; Fohlerová, Zdenka (referee) ; Pekárková, Jana (advisor)
This thesis is focused on synthesis of quantum dots (QDs) for protein detection. It comprises three parts. The first part summaries the theory of QDs, their synthesis, functionalization, interactions and applications in medicine. In the second part synthesis of CdTe/ZnS core/shell QDs modified by glutathione (GSH) is described, followed by the conjugation with biomolecules BSA and IgG. Several coupling agents such as EDC with NHS and CDI were used. In the last part, the final products were characterized by fluorescence spectroscopy and capillary electrophoresis. The results show the dependence of the fluorescence intensity of the QDs on pH range, concentration of BSA and IgG concentrations using different crosslinkers.
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Separation and Detection of Bioconjugated Quantum Dots Using on a Chip Electrophoresis
Pejović, Jelena
Semiconductor nanocrystals, quantum dots (QDs), are nanoscale particles that have been attract a lot of attention due to their unique optic and electronic properties. Due to very diverse and numerous applications, it is really important to make a tool for precise and controlled detection of QDs. In this study, we investigate on a chip detection and separation of QDs bioconjugated with BSA (bovine serum albumin) using homemade equipment based on principle of capillary electrophoresis (CE) and optical detection. Quenching effect of different concentration of BSA on fluorescence intensity of QDs was monitored. It was found that with increasing concentration of BSA fluorescence intensity of QDs is decreasing. This research can lead to a better understanding of interaction between different size QDs and biomolecules.
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Utilization of interfacial rheology to study of biological systems
Kachlířová, Helena ; Kalina, Michal (referee) ; Smilek, Jiří (advisor)
The aim of this bachalor thesis is to implement and optimization a relatively new method of interfacial rheology. The optimization of this method was realized on two types of interfaces, specifically liquid-liquid using immiscible systems water-chloroform and water-toluene, and water-air. The applicability of this method was tested by using real samples in form of surfactant septonex and biological system represented by protein (bovine serum albumin). The interfacial layers formed by septonex were purely viscous, where the rigidity of the layer increased slightly with concentration until reaching critical micellar concentration and then the rigidity decreased. In comparison with septonex, the interfacial film made of bovine serum albumin showed predominantly elastic behaviour.
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Synthesis of quantum dots for proteins detection
Šibíková, Anna ; Fohlerová, Zdenka (referee) ; Pekárková, Jana (advisor)
This thesis is focused on synthesis of quantum dots (QDs) for protein detection. It comprises three parts. The first part summaries the theory of QDs, their synthesis, functionalization, interactions and applications in medicine. In the second part synthesis of CdTe/ZnS core/shell QDs modified by glutathione (GSH) is described, followed by the conjugation with biomolecules BSA and IgG. Several coupling agents such as EDC with NHS and CDI were used. In the last part, the final products were characterized by fluorescence spectroscopy and capillary electrophoresis. The results show the dependence of the fluorescence intensity of the QDs on pH range, concentration of BSA and IgG concentrations using different crosslinkers.
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Electrochemical impedance spectroscopy as a nanostructured bioelectrodes characterization method
Vrbová, Eva ; Urbánková, Kateřina (referee) ; Hrdý, Radim (advisor)
Diploma thesis deals of nanostructured surfaces, nanoparticles and electrochemical characterization methods such as cyclic voltammetry, differential pulse voltammetry and electrochemical impedance spectroscopy. The aim of this thesis is a theoretical research issues of production and characterization nanostructured modified electrodes. The practical part is the production of biomodified nanostructured electrodes by anodi- zation W/Al layers with galvanic deposition of gold or deposition of mercury, a modifi- cation of the electrodes by 11-mercaptoundecanoic acid and by bovine serum albumin (BSA). The thesis includes SEM images of nanostructured electrodes contact angle mea- surements of these electrodes and form an electrical circuit with subsequent simulation waveforms.
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Characterization of hyaluronan interactions with albumin
Valentová, Kristýna ; Kratochvílová, Romana (referee) ; Chytil, Martin (advisor)
This bachelor thesis focuses on the research of physical interactions between hyaluronic acid and a protein albumin. In order to investigate these interactions following methods were used: pH and conductivity measurement, viscosimetry, turbidimetry and rheometry. The interactions were observed in solutions of fixed albumin concentration 1.0 gdm-3 and varying concentrations of hyaluronic acid from 0.1 to 2.0 gdm-3. Results were compared with a solution of hyaluronic acid without the addition of albumin. The results show that there are interactions between hyaluronic acid and albumin solutions, especially for those containing high molecular weight hyaluronic acid. The interactions exhibited a change in viscosity of solutions. We can see a reduction in viscosity after adding albumin to the reference solutions. The reduction is noticeable even in the low molecular weight HA solutions, but is not so significant.
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