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Study of interaction of charged catanionic vesicles with oppositely charged polyelectrolytes
Havlíková, Martina ; Lehocký, Marián (referee) ; Táborský, Petr (referee) ; Mravec, Filip (advisor)
The presented dissertation is focused on positively charged catanionic vesicles and their interactions with oppositely charged hyaluronan. The catanionic vesicles were composed of ion pair amphiphile of hexadecyltrimethylammonium-dodecyl sulfate (HTMA-DS), a positively charged double-chain surfactant that electrostatically stabilized the vesicles and cholesterol. Cholesterol may strongly affect the physico-chemical properties of the vesicles, therefore it was necessary to determine its optimal amount in the membrane. Especially, the size and stability of vesicles were studied by dynamic and electrophoretic light scattering, membrane microviscosity by fluorescence anisotropy and hydration of the outer part of the membrane by generalized polarization. It was found that vesicles show ideal properties at a cholesterol content about 40 mol.%. These catanionic vesicles interacted with hyaluronan regardless of its molecular weight. Visible aggregates were observed around their isoelectric point, but they disintegrated with increasing polymer concentration. However, this decay was different for different hyaluronan molecular weights. An increase in ionic strength, or more precisely concentration of sodium chloride above 25 mM had a destabilizing effect on both uncoated and hyaluronan coated vesicles. It was also interesting that increasing the ionic strength affected the microviscosity of the membrane, which became more fluid, but there were no significant changes in the hydration of its outer part. The presented work is also focused on the preparation and characterization of a new type of catanionic vesicles, where the cationic surfactants HTMAB was replaced by Septonex. The same methods were chosen for their characterization as in the case of the HTMA-DS vesicles. This exchange of surfactants had a major effect on the resulting properties of the vesicles, which are stable even at very low cholesterol content. In accordance with long-term stability, a suitable concentration of cholesterol was chosen from 15 mol.% and higher.
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Stability of vesicular complexes against changes in their concentration
Velichová, Veronika ; Marková, Kateřina (referee) ; Mravec, Filip (advisor)
This bachelor thesis is focused on the stability of vesicular systems that consist of ion pair amphiphile. These catanionic vesicles were prepared from single-chained surfactants CTAB cetyltrimethylammonium bromide) and SDS (sodium dodecyl sulphate), which were stabilized by adding double-chained surfactant DODAC (dimethyldioctadecylammonium chloride) with 43 mol.% cholesterol. The aim of the thesis was to verify the stability of vesicular systems against changes in their concentration. For the measurement, a concentration series was prepared by diluting vesicles with deionized water. The stability was evaluated by dynamic (DLS) and electrophoretic (ELS) light scattering measurements at weekly intervals for four weeks. By DLS method were determined the average values of the size of vesicles and their changes during the measurement, the method ELS provided the information about changes of charge with different concentration in time. Samples with a higher concentration, i.e. no dilution till tenfold dilution of the vesicle solution, were marked as stable, higher values of average sizes and lower values of zeta potential were measured at solution with lower concentrations, according to which such diluted vesicles can be described as unstable. There were also visual observations made, but no visible aggregation in the solutions was observed. Furthermore, the aggregation behavior of the vesicles was observed by fluorescence spectroscopy using pyrene as fluorescent probe.
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Solubilization of water-insoluble vitamins into vesicular systems based on ionic amphiphilic pairs
Kolomá, Nikola ; Mravcová, Ludmila (referee) ; Mravec, Filip (advisor)
This thesis deals with the characterization and preparation of catanionic vesicular systems and their interaction with water-insoluble vitamins. Catanionic vesicles systems was prepared from two differently charged sufractants CTAB and SDS in formation ion pair amphiphile. Stability of vesicles was secured by adding cholesterol with positively charged surfactant DODAC. Were selected vitamin A and vitamin E for solubilization into these vesicles. The next part of this work was focused on determining the efficiency of incorporation these vitamins into the HTMA-DS vesicular system. The characterization of these systems was improved by measurement on a UV-VIS spectrometer, DLS and by HPLC with UV-VIS detector. During measurement by UV-VIS and HPLC the efficiency of solubilization of vitamin A and vitamin E into vesicles was determinated. In both cases higher efficiency was determinated for vesicles with incorporated vitamin A. Higher efficiency for both vitamins was determinated by HPLC. The reason is probably the higher sensitivity of the metod, work in labs with dark glass and preparing individuals samples into vials, which were inserted into the device for individual dosing by injector. The work serves primary evaluation results in view of the vesicular system for use in pharmaceutical application like carrier non-polar vitamins.
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Study of microviscosity of membrane systems based on ionic amphiphilic pairs
Moslerová, Lenka ; Venerová, Tereza (referee) ; Mravec, Filip (advisor)
In this master ‘s thesis, catanionic vesicles formed by the pseudo-double-chain complex CTA – DS were investigated from the point of view of microviscosity. Samplesand of cationic vesicles contained 23, 43 and 53 mol. % of cholesterol and the double-chain surfactant DODAC. Cationic vesicles were prepared for visual observation, their stability was determined by DLS and the prepared system was further investigated. Microviscosity was determined from fluorescence anisotropy. To study the outer part of the membrane, laurdan fluorescent probes were used whereas diphenylhexatriene was used for the inner part of the membrane. This method has been proven to be suitable because it reflects the conditions of the membrane. Moreover, a 1,3-bispyrenylpropane probe forming intramolecular excimers was used to study the microviscosity in the vesicle bilayer. The dicyanovinyljulolidine (DCVJ) probe was applied in the case of the molecular rotor technique. It has been shown that in the case of the DCVJ probe, the molecular rotor technique is practically unusable, due to the fact that the probe has a low quantum yield at low temperatures. Also, the excimer formation of P3P probes does not lead to the expected results. The cationic vesicles do not seem to support this formation, as they are too closely related. This type of probe can be used for the selected system with some restrictions.
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Study of the hydration shell of vesicular systems based on ionic amphiphilic pairs
Rašticová, Barbora ; Szabová, Jana (referee) ; Mravec, Filip (advisor)
This bachelor thesis deals with the study of hydration shell of positively charged vesicular systems. The aim of the work was to monitor the hydration shell and to find out the amount of water molecules in the environment at different conditions of the membrane. Furthermore, a comparison of two selected fluorescent probes were studied. Positively charged vesicular systems composed of two types of surfactants were used, namely negatively charged SDS and positively charged CTAB. The two-chain positively charged surfactant was also added, resulting a positively charged vesicular systems. Three different concentrations of cholesterol were selected for the study and added to the system, namely 20, 40 and 60 mol. %. The study was performed by measuring the fluorescent emission as a function of temperature. The temperature range was from 10 to 80 °C. As a fluorescent probe Laurdan and Prodan were selected. For evaluation the method of generalized polarization was used. Due to its structure, Laurdan is mainly found in the membrane environment, so the method of two-wavelength generalized polarization, which includes fluorescence contributions only from the membrane, was sufficient for its evaluation. Compared to Laurdan, Prodan has a shorter hydrocarbon chain, so its distribution is not only in the membrane but also in the aqueous environment. Therefore, the method of three-wavelength generalized polarization, which also includes the contribution of fluorescence from the aqueous environment was used. The results showed that the behaviour of both probes is very similar. In all systems, the values of generalized polarization decreased with increasing temperature. Thus, at low temperatures, the membrane is in a solid ordered phase. With increasing temperature, it transforms into a liquid disordered phase. Thus, with increasing temperature, the number of water molecules increases. As the cholesterol concentration increased, the phase transition interval increased. The results also showed that the values of Prodan’s three wavelength generalized polarization are always higher than the values of Laurdan’s two wavelength generalized polarization.
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Influence of ionic strength on the properties of ionic amphiphilic pairs
Filipová, Lenka ; Smilek, Jiří (referee) ; Mravec, Filip (advisor)
The Bachelor thesis studies catanionic vesicle systems consisting of ion pair amphiphile (IPA) and their properties in the presence of ionic strength. Ion pair amphiphile was prepared from single-chained surfactants: cationic surfactant HTMAB (hexadecyltrimethylammonium bromide) and anionic surfactant SDS (sodium lauryl sulfate). Cationic double-chained surfactant DDAC (dimethyldioctadecylammonium chloride) was added in order to stabilize the vesicle system and make it positively charged. Cholesterol was also added to further stabilize the vesicle system. The system is considered to be relatively stable when consisting of 90 % IPA and 10 % DDAC with 43 mol.% cholesterol in a membrane. The stability of the system can be disrupted by external factors such as ionic strength. The Ionic strength was induced by NaCl solutions of varying concentrations (0.0, 0.5, 1.0, 2.0, 50.0, 100.0, 150.0 and 300.0 mM). The change of properties of a vesicle system, which was induced by ionic strength, was analyzed by electrophoretic and dynamic light scattering, fluorescence anisotropy, and generalized polarization. –potential was obtained by electrophoretic light scattering analysis which determines the stability of the system. Dynamic light scattering measurement resulted in the determination of the size of vesicles. The fluidity of vesicles’ membranes was examined by fluorescence anisotropy with DPH (1,6-diphenyl-1,3,5-hexatriene) as a probe. The hydration shell of vesicles was observed by generalized polarization with Laurdan (2-(dimethylamino)-6-dodecanoylnaphthalene) as a probe. As a result, it was found out that low ionic strength (0.5 to 2.0 mM) causes a decrease in the size of vesicles and high ionic strength (50.0 to 300.0 mM) causes vesicles to grow in size. –potential showed a consistent trend for the whole concentration series – its value rapidly decreasing with increasing ionic strength. The samples with the concentrations of salt 50.0 to 300.0 mM were considered unstable according to the –potential data. Fluorescence anisotropy decreases with increasing temperature and increasing ionic strength. The value of generalized polarization decreases with increasing temperature; hence there is more solvation of a vesicle membrane. The highest value of generalized polarization was measured when the concentration of sodium chloride was relatively high (from 50.0 to 300.0 mM), as a result of which the membrane was less hydrated and therefore, more organized.
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Stability of vesicular complexes against changes in their concentration
Velichová, Veronika ; Marková, Kateřina (referee) ; Mravec, Filip (advisor)
This bachelor thesis is focused on the stability of vesicular systems that consist of ion pair amphiphile. These catanionic vesicles were prepared from single-chained surfactants CTAB cetyltrimethylammonium bromide) and SDS (sodium dodecyl sulphate), which were stabilized by adding double-chained surfactant DODAC (dimethyldioctadecylammonium chloride) with 43 mol.% cholesterol. The aim of the thesis was to verify the stability of vesicular systems against changes in their concentration. For the measurement, a concentration series was prepared by diluting vesicles with deionized water. The stability was evaluated by dynamic (DLS) and electrophoretic (ELS) light scattering measurements at weekly intervals for four weeks. By DLS method were determined the average values of the size of vesicles and their changes during the measurement, the method ELS provided the information about changes of charge with different concentration in time. Samples with a higher concentration, i.e. no dilution till tenfold dilution of the vesicle solution, were marked as stable, higher values of average sizes and lower values of zeta potential were measured at solution with lower concentrations, according to which such diluted vesicles can be described as unstable. There were also visual observations made, but no visible aggregation in the solutions was observed. Furthermore, the aggregation behavior of the vesicles was observed by fluorescence spectroscopy using pyrene as fluorescent probe.
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Solubilization of water-insoluble vitamins into vesicular systems based on ionic amphiphilic pairs
Kolomá, Nikola ; Mravcová, Ludmila (referee) ; Mravec, Filip (advisor)
This thesis deals with the characterization and preparation of catanionic vesicular systems and their interaction with water-insoluble vitamins. Catanionic vesicles systems was prepared from two differently charged sufractants CTAB and SDS in formation ion pair amphiphile. Stability of vesicles was secured by adding cholesterol with positively charged surfactant DODAC. Were selected vitamin A and vitamin E for solubilization into these vesicles. The next part of this work was focused on determining the efficiency of incorporation these vitamins into the HTMA-DS vesicular system. The characterization of these systems was improved by measurement on a UV-VIS spectrometer, DLS and by HPLC with UV-VIS detector. During measurement by UV-VIS and HPLC the efficiency of solubilization of vitamin A and vitamin E into vesicles was determinated. In both cases higher efficiency was determinated for vesicles with incorporated vitamin A. Higher efficiency for both vitamins was determinated by HPLC. The reason is probably the higher sensitivity of the metod, work in labs with dark glass and preparing individuals samples into vials, which were inserted into the device for individual dosing by injector. The work serves primary evaluation results in view of the vesicular system for use in pharmaceutical application like carrier non-polar vitamins.
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Study of microviscosity of membrane systems based on ionic amphiphilic pairs
Moslerová, Lenka ; Venerová, Tereza (referee) ; Mravec, Filip (advisor)
In this master ‘s thesis, catanionic vesicles formed by the pseudo-double-chain complex CTA – DS were investigated from the point of view of microviscosity. Samplesand of cationic vesicles contained 23, 43 and 53 mol. % of cholesterol and the double-chain surfactant DODAC. Cationic vesicles were prepared for visual observation, their stability was determined by DLS and the prepared system was further investigated. Microviscosity was determined from fluorescence anisotropy. To study the outer part of the membrane, laurdan fluorescent probes were used whereas diphenylhexatriene was used for the inner part of the membrane. This method has been proven to be suitable because it reflects the conditions of the membrane. Moreover, a 1,3-bispyrenylpropane probe forming intramolecular excimers was used to study the microviscosity in the vesicle bilayer. The dicyanovinyljulolidine (DCVJ) probe was applied in the case of the molecular rotor technique. It has been shown that in the case of the DCVJ probe, the molecular rotor technique is practically unusable, due to the fact that the probe has a low quantum yield at low temperatures. Also, the excimer formation of P3P probes does not lead to the expected results. The cationic vesicles do not seem to support this formation, as they are too closely related. This type of probe can be used for the selected system with some restrictions.
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Study of the hydration shell of vesicular systems based on ionic amphiphilic pairs
Rašticová, Barbora ; Szabová, Jana (referee) ; Mravec, Filip (advisor)
This bachelor thesis deals with the study of hydration shell of positively charged vesicular systems. The aim of the work was to monitor the hydration shell and to find out the amount of water molecules in the environment at different conditions of the membrane. Furthermore, a comparison of two selected fluorescent probes were studied. Positively charged vesicular systems composed of two types of surfactants were used, namely negatively charged SDS and positively charged CTAB. The two-chain positively charged surfactant was also added, resulting a positively charged vesicular systems. Three different concentrations of cholesterol were selected for the study and added to the system, namely 20, 40 and 60 mol. %. The study was performed by measuring the fluorescent emission as a function of temperature. The temperature range was from 10 to 80 °C. As a fluorescent probe Laurdan and Prodan were selected. For evaluation the method of generalized polarization was used. Due to its structure, Laurdan is mainly found in the membrane environment, so the method of two-wavelength generalized polarization, which includes fluorescence contributions only from the membrane, was sufficient for its evaluation. Compared to Laurdan, Prodan has a shorter hydrocarbon chain, so its distribution is not only in the membrane but also in the aqueous environment. Therefore, the method of three-wavelength generalized polarization, which also includes the contribution of fluorescence from the aqueous environment was used. The results showed that the behaviour of both probes is very similar. In all systems, the values of generalized polarization decreased with increasing temperature. Thus, at low temperatures, the membrane is in a solid ordered phase. With increasing temperature, it transforms into a liquid disordered phase. Thus, with increasing temperature, the number of water molecules increases. As the cholesterol concentration increased, the phase transition interval increased. The results also showed that the values of Prodan’s three wavelength generalized polarization are always higher than the values of Laurdan’s two wavelength generalized polarization.
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