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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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Microrheology with Fluorescence Correlation Spectroscopy
Kábrtová, Petra ; Sedláček, Petr (referee) ; Mondek, Jakub (advisor)
A comparison of three passive microrheological techniques was made with the emphasis on FCS. Fluorescently labelled and unlabelled polystyrene particles were used to probe a microrheological response of glycerol solutions, Mili-Q water and sodium hyaluronate solutions. In addition, for FCS technique an approximated equation for calculation of MSD values was derived and verified. It was found that FCS outmatches current microrheological techniques of DLS and video-based particle tracking by its ability to gain a broader data range including the area in which, until now, it was impossible to describe a microrheological behaviour of samples reliably.
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Hyaluronan micro- and nanoparticles
Mourycová, Jana ; Marián, Lehocký (referee) ; Pekař, Miloslav (advisor)
The aim of this thesis was to prepare hyaluronic acid micro- and nanoparticles based on electrostatic interactions with oppositely charged molecules. Following parameters were monitored: correlation function behavior, the particle size and zeta potential value. At the beginning, it was necessary to study the behavior of hyaluronan in solution by dynamic light scattering measurement. Micro- and nanoparticles were prepared by mixing different volume ratios of negatively charged hyaluronan and positively charged polyarginine or cetyltrimethylammonium bromide. Micro- and nanoparticles were prepared in aqueous solution as well as in 0,15 M sodium chloride solution (physiological solution). In the case of the hyaluronan solution a polydisperse character of hyaluronan was detected. It was found that the dissolution of hyaluronan in the physiological solution gives us the smaller particle size in opposite to particle size obtained from the same concentrations of hyaluronan dissolved in water. Furthermore, it was found that systems composed of hyaluronan and polyarginine create particle size of about 100 nm. Whereas systems consisting of cetyltrimethylaminoum bromide and hyaluronan form larger particles, in units of hundreds of nanometers, the particle size in physiological solution were smaller than the same systems dissolved in aqueous solution.
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Preparation and characterization of complex liposomal for drug delivery systems
Szabová, Jana ; Pekař, Miloslav (referee) ; Mravec, Filip (advisor)
This diploma thesis deals with the preparation and characterization of stealth liposomes and their combination with trimethylchitosan (TMC). This complex could find application in the field of inhalation administration. Stealth liposomes were prepared from neutral phophatidylcholine, negatively charged fosfatidic acid and polyethyleneglycol bounded to phosphatidylethanolamine. We have managed to prepare stealth liposomes with suitable properties that should guarantee passive targeting without evocation an immune response, despite the content of the negative component. We also found a suitable method of preparation for stealth liposome–TMC complex, where the change of size and zeta potential confirmed the non–covalent bound between two components despite the content of the polyethyleneglycol.
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Use of some encapsulation techniques to controlled release of active substances in food and cosmetics products.
Skoumalová, Petra ; Rittich, Bohuslav (referee) ; Kráčmar, Stanislav (referee) ; Márová, Ivana (advisor)
The presented doctoral thesis is focused on preparation, characterization and application of organic micro- and nanoparticles as transport systems for active components and some their complex natural sources. Active component were packed into liposomes and polysaccharide particles. As active components were used caffeine, some drugs – clotrimazole and ibuprofen, further antioxidants and vitamins. Antimicrobial herbs and spices extract, antimicrobial peptides lysozyme, nisin and other antimicrobial ingredients were encapsulated too. Encapsulation of selected hydrolytic enzymes was tested, too. Particles were also used for encapsulation of probiotic strains Bifidobacterium breve and Lactobacillus acidophilus and prebiotic components. These prebiotics were co-encapsulated into capsules with probiotic cells. Natural extracts were encapsulated e.g. extracts of guarana, ginseng, goji, green barley, propolis, black, green and white tea, coffee, fruit and vegetable extracts. The efficiency of encapsulation was determined by HPLC/PDA and by spectrophotometry. Long-term stability of particles and amount of released component in model/real foods, in model cosmetic conditions and in a model physiological environment were monitored too. Size of prepared liposomes and polysaccharide particles was determined by dynamic light scattering and by light microscopy and electron microscopy, respectively. Stability of the particles was measured using a zeta potential. Also, analytical centrifugation was used to measurement of sedimentation velocity and stability of the prepared particles. The antimicrobial activity were tested using two Gram-positive (Bacillus subtilis, Micrococcus luteus), two Gram-negative (Escherichia coli, Serratia marcescens) bacteria and one fungal strains (Candida glabrata). For determining the antimicrobial properties of active component and prepared particles two the most widely used methods were used - agar diffusion method and broth dilution method. The viability of probiotic strains were performed using flow cytometry and fluorescence microscopy. Encapsulation of active component was successful in all types of particles. Liposome showed a very good long-term stability mainly in water conditions with neutral pH and polysaccharide particles were stable in acidic conditions. Prepared particles showed a very good stability in model stomach environment, while in model intestines environments particles were disintegrated and active component were released. Prepared particles with encapsulated caffeine as well as other tested antioxidants and vitamins could be used to modern types of energy drinks, food supplements and also for some cosmetics applications. Encapsulated antimicrobial components could be used for food application as well as for cosmetics and pharmaceutical application like antimicrobial wound formulation. Encapsulated enzymes can be used for controlled release of proteases in wound healing, as delivery systems in digestive tract and as a part of pharmaceutical preparative and food supplements for enzyme therapy. The study revealed that encapsulation of probiotics and also co-encapsulation of probiotics with prebiotics exhibited longer stability of particles and survival bacterial cells. So, prepared particles are suitable for use to food product with beneficial effects on the human body.
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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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Possibilities of encapsulation of particular types of macromolecules and bacteria
Kapar, Jiří ; Obruča, Stanislav (referee) ; Márová, Ivana (advisor)
Presented diploma thesis is focused on testing encapsulation methods of enzymes and probiotic bacteria. In the theoretical part a summary of different encapsulation techniques used in food industry is given. Further, materials for encapsulation, above all polysaccharides are presented. Next, some procedures of encapsulation of biopolymers and microorganisms – mainly enzymes and probiotic cultures are discussed. In the experimental part methods for preparation of several types of particles based on polysaccharides and liposomes are introduced. Particles were used for encapsulation of selected hydrolytic enzymes and probiotic strains Bifidobacterium breve a Lactobacillus acidophilus. The encapsulation effectiveness was evaluated by analysis of total proteins and enzyme activities. Particles sizes and their stability in water, in selected model foods and model body fluids were observed, too. According to results obtained in this work it was found that encapsulation of enzymes into polysaccharide particles were succesfull in all types of particles (encapsulation effectivness was more than 50 %). Polysaccharide particles showed a very good stability in body fluids as well as in model foods. As the most suitable materials for enzymes encapsulation chitosan and liposomes were found. Polysaccharide particles were used also for the encapsulation of microorganisms. The stability of particles with lactic acid bacteria was similar to particles containig enzymes, very good stability was verified aslo in model foods and model body fluids. Encapsulation enables long-term stabilization of biologically active compounds as well as posibility of their transport and controlled releasing in gastrointestinal tract. Encapsulation of probiotic bacteria could preserve their viability and long-term survival until the product expiration date. Thus, encapsulation is one of the most promissing procedures for production of foods and food suplements of great quality and high additional value.
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Study on Sol-gel Process of Agarose by Classical Rheology and Dynamic Light Scattering
Krňávková, Šárka ; Hnyluchová, Zuzana (referee) ; Smilek, Jiří (advisor)
The main aim of the bachelor thesis was characterization of hydrogels from rheological point of view and by dynamic light scattering. Rheological part of research relates with the determination of the influence of temperature on mechanical properties, gelation temperature and temperature of solidification as well as the effect of aging on mechanical properties of hydrogel. The influence of the particle size on diffusion coefficient was investigated by scattering techniques. The positive correlation between the data obtained by both types of measurement and the potencial usage of dynamic light scattering method for the characterization of hydrogels was discussed as well.
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Influence of particle size of cosmetic pigments on final properties product
Obručová, Kateřina ; Hurčíková, Andrea (referee) ; Zemanová, Jana (advisor)
This diploma thesis is focused on the study of the influence of the of size cosmetics pigments on the properties of the final product. This effect was studied on samples of make-up given by industry partner. Make-up samples were measured by the rheology method and the yield stress of all samples was determined. Scanning electron microscopy was used to determine the size and shape of the pigment particles. Particle size was also measured by dynamic light scattering, but the results were influenced by forming aggregates. To easier determination of the types of pigments in the sample was determined elemental composition of the pigments (Ti, Fe, O, Al) by EDS detector. Due to the unknown composition of make-up, the information was supplemented by X-ray powder diffraction (XRD) analysis, and it was possible to determine the compounds TiO2 (anatase and rutile), FeOOH and Fe2O3. The results obtained by these methods showed what pigments can be found in make-up samples, what their sizes and shapes are. It has been shown that the smaller the pigments are, the higher the viscosity and yield stress of the final product. The higher these quantities, the harder it is to handle the product.
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Study on Interconnection between Structure and Rheological Properties of Hydrogels on Macro and Microscopic Level
Lepíková, Jana ; Klučáková, Martina (referee) ; Kalina, Michal (advisor)
Diploma thesis main goal is to obtain new pieces of knowledge about relationship between hydrogel structures and its flow and transport properties. Thesis is mainly focusing on combining pertinent biopolymers into model hydrogels based on agarose. Then perform correlation of results obtained by diffusion methods, and by rheologic measurements on macroscopic and microscopic level. Properties of hydrogels were measured by selected rheologic measurements, dynamic light scattering method, and correlative fluorescence spectroscopy. From these methods various parameters (MSD modules, values of complex viscosity) were obtained. Afterwards transport properties of prepared hydrogels were studied by observing Rhodamine 6G diffusion. Here two different approaches were used. From macroscopic perspective, simple principles of mass diffusion from dye solution to cuvettes filled with hydrogels containing individual biopolymers were used. From microscopic perspective, dye was added during the sample preparation and then the mass diffusion was investigated using FCS. Based on evaluated results it was discovered that added biopolymers don’t influence properties of carrier medium, in this case agarose hydrogels. During the study of prepared hydrogels’ reactivity and barrier properties some differences were observed. Charge of biopolymer and its charge density were discovered as main factors influencing transport of charged solutes into prepared hydrogels.
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