In this diploma thesis were studied solubilization properties of polysaccharides by using hydrophobic solutes (Sudan Orange G, Sudan Red G, (±)-alpha-Tocopherol, Pyrene, Perylene, Nile red), which were represented by alkyl derivates of hyaluronan. At first, a behaviour of individual hydrophobic solutes was investigated in variously polar solvents (Methanol, 1 Propanol, Chloroforme, Cyklohexane, n Heptane) and in the environment of varying ionic strength (water, 0.1 M and 0.4 M NaCl). Afterwards, solubilization properties of Sodium Dodecyl Sulfate model solubilizated the hydrophobic solutes into a core of micelles was examinate. We were interested in the solubilization capacity as the mol of solubilized molecules per mol micelles of surfactant corresponding with a state of micelles saturation. In the case of the solubilization of (±)-alpha-Tocopherol into the core of micelles, it was not possible to determine the solubilization capacity. So we changed the determination of universally solubilization power. The solubilization power is defined as mol of molecules solubilized per mol surfactant relative to the quantity solubilizate at the micelles saturation. Model system of Sodium Dodecyl Sulfate as a simple surfactant carrying a negative charge as the alkyl derivates of hyaluronan was selected bacause of its characteristics.The surfactant forms unimolar micelles and its critical micelle concentrations and aggregation numbers are tabelated for the investigated microenvironment. The main aim of the study was investigating of hydrophobic domains of alkyl derivates of hyaluronan as free places for incorporation hydropbobic solutes in the microenvironment of varying ionic strength. The critical aggregation concentrations were determined by the Pyrene 1:3 ratio method. For the research of micropolarity of alkyl derivates hyaluronan’s domains were selected two concentrations of derivates for the next research of solubilization experiments - the first concentration near the critical aggregation concentration and the second concentration above it. The effect of concentration of Pyrene on a core polarity of derivates was investigated. We discovered the influence of the concentration and the other we found a stationary area of the concentration. In the end we investigated the influence of preparation of solutions of derivates of hyaluronan on the core polarity by the concentration of pyrene which corresponds to the stationary area. The study of solubilization properties of alkyl derivates of hyaluronan is not a simple case as we assumed. When we measured spectra of the absorbance, higher concentration of derivates of hyaluronan belittle absorbance of solubilizates. At the experiment of solubilization with Sudan Red G we found out that Sudan Red G is not able to solubilizate into the hydrophobic core of micelles of hyaluronan’s derivates because of lipophilic or steric effects. We had to change Perylene as a new solubilizate. From the measured emission spectra we found saturation micelles. We can express the solubilization power of hyaluronan’s derivates for the concentration of Perylene. The main aim of the diploma thesis was to determine optimal way of the preparation of hyaluronan’s derivates solutions with required degree of solubilization.

This diploma thesis investigated thermal stability and the degradation of hyaluronan (HA) in HA with a molecular mass of 90–130 kDa and in HA with a molecular mass of 1 500–1 750 kDa. The following methods were used for the research: rheology, SEC-MALLS, TGA and DSC. Low-molecular HA was subject to time dependency of degradation investigation, where it was dried at a temperature of 90 °C for a period of 30 minutes and 60 minutes prior to the preparation of the solutions itself. High-molecular HA was investigated not only from the point of view of time but from the point of view of temperature dependency of degradation as well. In the case of investigating the time dependency of degradation, high-molecular HA was dried at a temperature of 75 °C at a time range from 15 minutes to 120 minutes prior to the preparation of the solutions. During the preparation of the solutions for discovering the temperature dependency of degradation, the high-molecular HA was then dried for a period of 30 minutes at a temperature range from 60 °C to 90 °C. For low-molecular HA, thermal stability was proven. Therefore, there is no decrease in the molecular mass and the solutions did not demonstrate a significant decrease of viscosity. For high-molecular HA, thermal stability was not proven. Degradation due to the temperature of drying as well as the time of drying occurred, which was demonstrated by a significant decrease in molecular mass and viscosity of the solutions. While in the case of using a drying temperature of 60 °C, a decrease in the molecular mass occurred by approximately 5 %, the molecular mass decreased by approximately 20 % at a drying temperature of 90 °C compared to undried HA. Due to this reason, high-molecular HA was also further investigated by means of the TGA method, where the decrease of humidity of HA samples in relation to the drying temperature was observed. The DSC method was also used. The objective of the DSC method was to find out temperatures, at which evaporation of humidity contained in an HA sample in relation to its form (undried HA, dried HA and lyophilized HA) occurs. This method further finds out the heat necessary to evaporate humidity from an HA sample. To conclude this research, the results obtained for high-molecular HA were compared with the results of other drying processes – lyophilized proved to be a very gentle drying method because a decrease in the molecular mass for lyophilized HA compared with undried HA almost did not occur.

This work is focused on physical interactions of negatively charged polymers with small ionogenic fluorescent molecules. Trying to verify the presence of these interactions using fluorescence correlation spectroscopy (FCS) and provides a comprehensive view of the problem. The aim of this work is to observe the effect of concentration on the diffusion properties. P/D ratio, where P represents number of polymer binding sites and D number of dye binding sites, was chosen for this issue. Hyaluronate, sodium chondroitin sulfate and sodium polystyrene sulfonace were used as polymers and Acridine Orange, and Rhodamine 6G were chosen as fluorescent probes. The reason why this experiment uses these probes, was the assumption, that the positive charge occuring on the fluorescent probe will lead to the electrostatic interaction with the negatively charged polymer. As a result, the bond between acridine orange and polyelectrolyte was not clearly demonstrated, but the interaction with Rhodamine 6G have been proved.

This thesis studies the properties and behaviour of phase-separated hydrogels prepared by interaction of hyaluronan with oppositely charged surfactants. Three representatives of surfactants, namely cetyltrimethylammonium bromide, chloride and p-toluensulfonate (CTAB, CTAC, CTAT), were selected for comparison. Using the method of rheology, the fact that the system of Hya-CTAT forms the most rigid hydrogels has been proved. Higher molecular weight of hyaluronan has also direct influence on the volume and stiffness of the newly formed hydrogels. Preparation methods were compared as well. Mixing the stock solutions of entry components appeard to be the most suitable. Small-angle X-ray scattering was used for determination of shape and size of surfactant's micelles, concluding that it has no effect on the volume of formed gels. The results of this study indicate that given gels are to a certain extent competitive and incorporation of hyaluronan into their structure is desirable with respect to its biological activity. This offers a potential usage of these substances in the field of medical applications.

The master´s thesis deals with the study of the interaction between the polysaccharide hyaluronan of diffrerent molecular weights with the amino acids arginine, lysine, arginine hydrochloride and 6-aminocaproic acid. They are expected interaction between carboxyl groups of hyaluronan and amino groups of amino acids. These interactions were investigated by using ultrasonic spectroscopy, DLS, measuring pH and conductivity. Obtained results were compared with sodium polystyrene sulfonate. With ultrasonic spectroscopy was observed a change of concentration inkrement for titration of amino acid to water or polymers solutions especially for high molecular weight hyaluronan and for NaPSS in combination with 6AKK in concentration range of added amino acid 0–30 mM. The size of this change could mean a degree of interaction between polymers and amino acids. This theory has not been confirmed by other methods. By pH and conductivity measurements interations between arginine and low molecular weight hyaluronan and NaPSS were only confirmed. There was no possibility to make unequivocal conclusions from determination of particle size and zeta potential by DLS. Overall, the issue of the interaction of amino acids with polyanions was proved above expectations complex and will be appropriate to further expand the observations made in this thesis.

This diploma thesis deals with the study of possible interactions between hyaluronan (HA) and plasmid DNA (pDNA). Plasmid DNA was isolated from E. coli JM109 (pUC19) and resuspended in TE buffer as well as high molecular weight hyaluronan. Individual samples of pDNA, HA and pDNA-HA were characterized by gel electrophoresis, CD spectroscopy and high resolution ultrasonic spectroscopy. Agarose gel electrophoresis examined the effect of the addition of hyaluronan to plasmid DNA on the migration of samples to the positive electrode. Structural changes in pDNA-HA samples were examined using CD spectroscopy. Individual CD spectra describes the dependence of the difference in absorption coefficients for left-hand and right-handed elliptic polarized light at wavelength. High resolution ultrasonic spectroscopy has been used to study interactions. It is an analytical method based on ultrasonic velocity and attenuation. We classify this technique as a non-destructive method because the passing waves do not affect the structure of the analyzed sample.

This master thesis is dealing with the use of light scattering techniques in the study on formation of polyelectrolyte complexes in the system biopolymer-surfactant. Sodium hyaluronate was chosen as biopolymer and cetyltrimethylammonium bromide and karbethopendecinium bromide were selected as surfactants. The first precipitation experiments were performed to determine the optimal concentration range of surfactants for subsequent titration measurements performed on Zetasizer Nano ZS and for SEC-MALS analysis. The key parameters obtained from these measurements were the values of critical aggregation concentrations in hyaluronate-surfactant systems. It was found out that the increasing molecular weight of hyaluronan in these systems is causing reduction of obtained values of critical aggregation concentrations. Changes of Z-average particle size, zeta potential, polydispersity index, conformation plot, radius of gyration and molecular weight in the hyaluronan-surfactant system were monitored during experiments.

This diploma thesis deals with study of hydrogels formed by phase separation of hyaluronan with oppositely charged surfactants cetyltrimethylammonium bromide (CTAB) and Septonex. It follows the bachelor thesis and extends the knowledge about the detailed characterisation of the inner environment of the hydrogel by determining the diffusion behaviour of the fluorescent probes Atto 488 and Nile Red using fluorescence correlation spectroscopy (FCS) technique and its modified version dual-focus fluorescence correlation spectroscopy (2f-FCS). Compared results showed that both methods show similar values and probes specifically interact with CTAB but Atto 488 shows only weak interaction with Septonex compared to Nile Red. Additionally, these interactions were not affected by the molecular weight of hyaluronan. In conclusion, it was recommended to measure this type of hydrogel in a small depth of gel using a conventional method.

This diploma thesis was focused on study of barrier and transport properties of selected polyelectrolytes in hydrogel matrices by using diffusion techniques. The study of these properties was performed in horizontal diffusion cells where is observed the change in diffusion probe concentration over time. Diffusion experiments were performed on an agarose hydrogel with the addition of alginate, hyaluronic acid, polystyrene sulfonate, humic acids and as a model probe rhodamine 6G was used. Important parts of this thesis are also the methods which characterize the substances and hydrogel matrices such as rheology and potentiometric titration. The main aim of this diploma thesis was to investigate the effect of interactions between passing model dye (rhodamine 6G) and the appropriate gel (agarose + polyelectrolyte) on the fundamental diffusion parameters (effective diffusion coefficient, lag time, etc.).

This diploma thesis focuses on the study of temperature dependence of -carrageen, hyaluronan of low molecular weight (250–450 kDa) and hyaluronan of high molecular weight (1 500–1 750 kDa) in aqueous solutions. The description of temperature depending changes in properties of these substance is based on rheological method. This work concentrates on the characterization of viscoelasticity through measurement of the storage (elastic) and loss (viscous) modulus using -carrageen and hyaluronan solutions of different molecular weights and different concentrations depending on temperature (20–50 °C, alternatively, in carrageen, 20–80 °C). The temperature dependence was further measured using an uncommon technique called high resolution ultrasonic spectroscopy (HR-US) which measured relative velocity of selected concentrations of -carrageen and hyaluronan of high molecular weight depending on the temperature of heating and cooling (20–80 °C). The measurements of temperature dependence were carried out for varied oscillation frequencies of rheometer sensor/frequency of ultrasonic waves. Knowledge of temperature dependence of the investigated substances may be applied in drug chemistry, primarily with respect to degradation due to high temperatures. The measured data gives information about sol-gel temperature and gel-sol temperature of carrageen.