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.

Layer-by-layer (LbL) assembly has proven to be a powerful technique for assembling thin films with a variety of properties including electrochromic, molecular sensing, oxygen barrier, and antimicrobial. LbL involves the deposition of alternating cationic and anionic ingredients from solution, utilizing the electrostatic charges to develop multilayer films. The present work incorporates cationic antimicrobial agents into the positively-charged layers of LbL assemblies. When these thin films are exposed to a humid environment, the antimicrobial molecules readily diffuse out and prevent bacterial growth. The influence of exposure time, testing temperature, secondary ingredients and number of bilayers on antimicrobial efficacy is evaluated here. Additionally, film growth and microstructure are analyzed to better understand the behavior of these films. The antimicrobial used here is a positively-charged quaternary ammonium molecule (e.g. cetyltrimethylammonium bromide [CTAB]) that allow assemblies to be made with or without an additional polycation like polydiallyldimethylamine. While films without this additional polymer are effective, they do not have the longevity or uniformity of films prepared with its addition. All of the recipes studied show linear growth as a function of the number of bilayers deposited and this growth is relatively thick (i.e. > 100 nm per bilayer). In general, 10-bilayer films prepared with CTAB and poly(acrylic acid) are able to achieve a 2.3 mm zone of inhibition against S. aureus bacteria and 1.3 mm against E. coli when test are conducted at body temperature (i.e. 37oC). Fewer bilayers reduces efficacy, but lower test temperatures improve zones of inhibition. As long as they are stored in a dry atmosphere, antimicrobial efficacy was found to persist even when films were used four weeks after being prepared. The best films remain effective (i.e. antimicrobially active) for 4-6 days of constant exposure to bacteria-swabbed plates. This technology holds promise for use in transparent wound bandages and temporary surface sterilization.

This diploma thesis deals with the use of fluorescence correlation spectroscopy technique for microrheological characterization of hydrogel in a system of hyaluronate-cetyltrimethylammonium bromide. Fluorescently labelled particles were used for microrheological FCS analysis. To optimize the method the most appropriate size of particles was chosen on the basis of Newtonian glycerol solutions analysis. Among other things, the discussion was focused on the influence of refractive index change of analysed solutions on analysis results. After hyaluronate solutions analysis it was possible to assess the biopolymer concentration and molecular weight impact on the FCS microrheology results, which could then be compared with analysis results of model hydrogels of hyaluronate and CTAB. Finally, usability and limitations of FCS microrheology have been discussed.

Possibilities for increasing the upper limit of floatable particle sizs in the froth flotation process have been examined since the early beginnings of mineral flotation. The economic implications of such an incresae are far ranging; from decreased grinding costs and increased recoveries to simplified flow-sheet design and increased throughput, all leading to increased revenue. Bubble-particle detachment has been studied to better understand the factors influencing the strength of attachment and the energies involved. Direct measurements of bubble particle detachment were performed using a hanging balance apparatus (KSV Sigma 70 tensiometer) and using a submerged hydrophobic plate in water. Three experiments were used; direct force measurement of bubble-particle detachment, detachment force and energy of a bubble from a submerged hydrophobic plate, and detachment force and energy of a cetyltrimethylammonium bromide coated silica sphere from a flat bubble. Octadecyltrichlorosilane was used as a hydrophobic coating in the first two experimental methods. These experiments were recorded with a CCD camera to identify the detachment processes involved. Energies for both methods were calculated and divided into the two main steps of the detachment process: Three-Phase-Contact pinning and three phase contact line sliding. The first step represents the energy barrier which must be overcome before detachment can begin. It is directly related to contact angle hysteresis. Detachment occurs during the second step, where the solid-vapor interface is replaced by solid-liquid and liquid-vapor. This step corresponds to the work of adhesion. The effects of surface tension, contact angle and hysteresis were well demonstrated with the three experimental methods. Good correlation was found between theoretical work of adhesion and measured energies. / Master of Science

The diploma thesis deals with the preparation and characterization of catanionic complexes and their possible application to pharmaceutics and medicine. The catanionic complexes were prepared by mixing two oppositely charged surfactants with the concentration of 20 mmol·dm­3 in various volume ratios. Two systems were analyzed – CTAB + SDS and Septonex + SDS. Both systems were prepared in an aqueous and physiological environment. The turbidimetric analyses provided results of the intensity of turbidity of the prepared mixtures. The size and stability of the prepared particles were determined by DLS and ELS measurements. CTAB-rich samples showed significant viscosity changes and they were therefore characterized by rheological measurements. The results of this work show that the asymmetry of alkyl chains, surfactant selection, temperature, and ionic strength have a significant influence on the properties and self-assembly of surfactants in catanionic complexes.

This work is focused on interaction between hyaluronan (Hya) and amphiphilic molecules. Using fluorescent probes method were carried out screening of the interaction of various surfactants with Hya. For further examination was chosen cetyltrimethylammonium bromide (CTAB). Interactions in this system have also been studied using fluorescence correlation spectroscopy (FCS), and additionally were performed initial tests of the cytotoxicity of the system. As an alternative CTAB were also used hydrophobically modified amino acids. Results showed their self-aggregation, but their interactions with hylauronanem was not as strong as with CTAB. The interaction of gel systems Hya-CTAB have also been studied, and was found to contain a hydrophobic domain capable solubilize hydrophobic substance. Thermogravimetric have been established that are able to contain up to 98% wt. water and rheologically were characterized their mechanical properties. These are tunable due to molecular weight (MW) of the hyaluronate. With increasing value of MW increases mechanical strength of the gel and decreases its fluidity.

A proteína A de superfície de pneumococo (PspA) é encontrada na superfície de todas as cepas de Streptococcus pneumoniae e candidata promissora para novas vacinas pneumocócicas. Foi desenvolvido um processo de purificação da PspA4Pro cujas etapas iniciais foram: ruptura da biomassa celular, precipitação do homogenato obtido com o detergente brometo de cetiltrimetilamônio (CTAB) e remoção do precipitado por centrifugação. Foram avaliadas cromatografias de troca iônica (aniônica, catiônica), afinidade por metais, interação hidrofóbica e mista de troca catiônica e hidrofóbica. Utilizando precipitação com CTAB, cromatografia de troca aniônica, crioprecipitação em pH4,0 e cromatografia de troca catiônica atingiu-se a pureza requerida de PspA4Pro (>95%) com recuperação entre 14% e 33%. O processo alcançou níveis aceitáveis de endotoxina no produto final e a PspA4Pro purificada foi reconhecida por anticorpos anti-PspA4, manteve sua atividade e sua estrutura secundária. / Pneumococcal surface protein A (PspA) is found in all Streptococcus pneumoniae strains and is a promising candidate to be used in new pneumococcal vaccines. A purification process for PspA4Pro which inicial steps were: cell disruption, precipitation of the homogenate with the cationic detergent cetyltrimethylammonium bromide (CTAB) and pellet removal by centrifugation. The chromatographic techniques tested were ion exchange (anionic and cationic), immobilized metal affinity, hydrophobic interaction and mix mode with hydrophobic and cationic ligands. Using CTAB precipitation, anion exchange chromatography, crioprecipitation in pH4.0 and cation exchange chromatography the PspA reached the required purity (>95%) with recovery between 14% and 33% . The process reached acceptable levels of endotoxin in the final product and the purified PspA4Pro was recognized by anti-PspA4 antibodies and manteined its activity and secondary structure.

A proteína A de superfície de pneumococo (PspA) é encontrada na superfície de todas as cepas de Streptococcus pneumoniae e candidata promissora para novas vacinas pneumocócicas. Foi desenvolvido um processo de purificação da PspA4Pro cujas etapas iniciais foram: ruptura da biomassa celular, precipitação do homogenato obtido com o detergente brometo de cetiltrimetilamônio (CTAB) e remoção do precipitado por centrifugação. Foram avaliadas cromatografias de troca iônica (aniônica, catiônica), afinidade por metais, interação hidrofóbica e mista de troca catiônica e hidrofóbica. Utilizando precipitação com CTAB, cromatografia de troca aniônica, crioprecipitação em pH4,0 e cromatografia de troca catiônica atingiu-se a pureza requerida de PspA4Pro (>95%) com recuperação entre 14% e 33%. O processo alcançou níveis aceitáveis de endotoxina no produto final e a PspA4Pro purificada foi reconhecida por anticorpos anti-PspA4, manteve sua atividade e sua estrutura secundária. / Pneumococcal surface protein A (PspA) is found in all Streptococcus pneumoniae strains and is a promising candidate to be used in new pneumococcal vaccines. A purification process for PspA4Pro which inicial steps were: cell disruption, precipitation of the homogenate with the cationic detergent cetyltrimethylammonium bromide (CTAB) and pellet removal by centrifugation. The chromatographic techniques tested were ion exchange (anionic and cationic), immobilized metal affinity, hydrophobic interaction and mix mode with hydrophobic and cationic ligands. Using CTAB precipitation, anion exchange chromatography, crioprecipitation in pH4.0 and cation exchange chromatography the PspA reached the required purity (>95%) with recovery between 14% and 33% . The process reached acceptable levels of endotoxin in the final product and the purified PspA4Pro was recognized by anti-PspA4 antibodies and manteined its activity and secondary structure.

In this thesis, we have tried to explain the regular and irregular(chaotic) dynamics of worm like micellar solutions on applying shear, through a detailed study of the equation of motion of a nematic order parameter tensor coupled to a hydrodynamic velocity field. We have assumed spatial variations only along one direction i.e. the gradient direction(1D model). The resulting phase diagram shows various interesting steady states or phases such as spatiotemporal chaos, temporal and spatiotemporal periodicities, and alignment of the director axis along the imposed flow field. The coupling of the orientational degrees of freedom of the order parameter with the hydrodynamic flow field holds the key to the appearance of dynamic shear bands in the system. We have solved numerically a set of coupled nonlinear equations to obtain the order parameter stress developed in the system; the magnitude of the order parameter tensor, the biaxiality parameter and the orientation of the director axis of the nemato gens under shear have also been studied in detail. To study the phase diagram obtained by time integration of the equation of motion mathematically, a stability analysis of the fixed point of motion for various parameter values has been performed so that the location of the chaotic-to-aligned phase boundary is verified. Also in the periodic region of the phase diagram, the stability of limit cycles is tested by analysing the fixed point of the corresponding Poincare map. Stability analysis of the periodic orbits leads to the observation that in the parameter space, there are regions of phase coexistence where chaotic or spatiotemporally intermittent behaviour coexists with periodic behaviour. When corrections in the imposed velocity field due to the order parameter stress were taken into account and the order parameter response was looked into at several points in the parameter space, the modified equations of motion were found to reproduce the earlier behaviour in all the different regimes if the value of a dimensionless viscosity parameter is taken to be such that the bare viscous stress overrides the order parameter stress. The phase boundaries are however different from the ones seen in the earlier model. However, for a choice of the viscosity parameter such that the order parameter stress and the bare viscous stress are comparable, we see two distinctly different attractors: a banded, periodic one that is common to both α1equalto 0, and not equal to 0 and a banded chaotic one for α1not equal to 0. Here, α1is a parameter that governs the nonlinearity in the stretching of the order parameter tensor along the direction of the applied shear. Quantitative analysis of the various chaotic attractors throws up not only positive Lyapunov exponents but also that the banded chaos is a “flip-flop” kind of chaos where the switching between two long-lived states of high and lows hear stress is chaotic, where as the behaviour in either of the two states is periodic, with either a single, isolated frequency or a bunch of harmonics. Also, the spatial correlation of the shear stress in the chaotic attractors is of much larger range than the temporal correlation, the latter being almost delta-function-like. On increasing the temperature of the system till it is above the isotropic–nematic transition temperature in the absence of shear, we find that under shear, similar attractors as those in the nematic case are observed, both for passive advection and for the full 1D hydrodynamics. This is an encouraging result since actual experiments are performed at a temperature for which the system is in the isotropic phase in the absence of shear. Thus for the 1D system, the parameter space has been explored quite extensively. Considering spatial variations only along the gradient axis of the system under shear is not enough since experiments have observed interesting behaviour in the vorticity plane in which Taylor velocity rolls were noted. Hence taking the system to 2D was necessary. Our numerical study of the 2D system under shear is incomplete because we came across computational difficulties. However, on shorter time scales we have seen a two-banded state with an oscillating interface and Taylor velocity rolls as well. The methodology used for the 2D study can also be used to reproduce the 1D results by the simple step of taking initial condition with no variation in the vorticity direction. This automatically ensures that no variation in the vorticity direction ever builds up because the equations of motion ensure that these variations in the system do not grow by themselves unless fed in at the start. Using this method, we were able to reproduce all the attractors found in the 1D calculation. Thus the 1D attractors have been observed using two different methods of calculation. Further work on the full 2D numerics needs to be done because we believe that spatiotemporally complex steady-state attractor s exist in the 2D system also for appropriate values of the parameters.

This diploma thesis deals with the preparation and characterization of negatively charged catanionic vesicular systems and their combination with selected polycations. The catanionic vesicular system was prepared by mixing of two oppositely charged surfactants SDS and CTAB. The negative charge as well as the stability of the vesicular system was provided by the incorporation of phosphatidic acid. Polycations, DEAE and TMC, have been selected for use in a pharmaceutical applications. Characterization of the prepared systems was performed by measuring DLS and ELS. The results indicate that we were able to prepare stable negatively charged vesicles that were eligible to non-covalently interact with selected polycations.