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Drop coating deposition Raman spectroscopy of liposomes with different compositions
Antalík, Andrej ; Kočišová, Eva (advisor) ; Holoubek, Aleš (referee)
Method of drop-coating deposition Raman spectroscopy is a special tech- nique that employs deposition of a small droplet of the sample on a hydro- phobic surface with subsequent solvent removal by evaporation. As a result the concentrated material forms a ring at the edge of the droplet. This techni- que has proven to be efficient for liposome studies at low concentrations. We focused on a study of spectral changes and ring formation in phosphatidylcho- line liposomes due to increasing amount of cholesterol. Higher concentration of cholesterol didn't cause significant changes in membrane structure, such as phase transition, however the ring wasn't formed. Second part of this bache- lor thesis concerns with the search for some alternative surface to previously used SpectRIM TM DCDR plates made by Tienta Sciences, Inc. which are no longer available. The polished calcium fluoride glass (CaF2) seems to be the most suitable for the further use from all tested surfaces. Raman signal at the other surfaces was either weak or spectral bands that don't belong to lipid were observed.
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Nanofibrous scaffolds in controlled delivery of autologous growth factors
Buzgo, Matej ; Amler, Evžen (advisor) ; Gášková, Dana (referee)
Platelet preparations are a source of various autologous growth factors and have numerous applications in tissues engineering. The aim of this work was to development electrospun nanofiber scaffolds with platelet preparations. Scaffolds based on the adhesion of platelets on nanofiber meshes were developed. The scaffolds were able to enhance chondrocyte proliferation in vitro. The main disadvantage of this system is the burst release of growth factors immediately after adhesion. To overcome this, we developed coaxially electrospun scaffolds with incorporated alpha granules. Alpha granules are novel platelet preparations with high amounts of growth factors. This system was able to stimulate chondrocyte proliferation and maintain TGF- 1 concentrations for 7 days. Additionally, a novel drug delivery system with coaxially incorporated liposomes was developed. Liposomes incorporated into nanofibers remain intact and can be used for the delivery of various molecules. The ability to maintain HRP activity was compared to systems based on coaxial electrospinning with liposomes, coaxial electrospinning without liposomes and blend electrospinning. When compared to other systems, coaxial electrospinning with liposomes preserves enzyme activity twice as long. These results clearly indicate the potential of...
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Effect of membrane lipid composition on resistance against surfactin
Pinkas, Dominik ; Seydlová, Gabriela (advisor) ; Sýkora, Jan (referee)
Surfactin is an antibiotic produced by several strains of B. subtilis. Its broad range of biological activities is interesting from perspective of medicine, food industry and bioremediation and is based on its surface-active properties and interaction with biological membranes. The latter means mainly forming ion channels, conductive pores and with increasing concentration eventually disrupting membrane structure in detergent-like manner. Mechanism of resistance of producing strain against its own toxic product is not yet fully understood. This work shows that it could be based on surfactin target modification - which means altering membrane lipid composition. We were able to recognize surfactin-formed ion channels or pores with a broad range of conductivities spanning from 2 pS to 2 nS using BLM method. Liposome leakage assay with carboxyfluorescein revealed few distinct mechanisms of lysis, differing in amplitude, rate of lysis and cooperativity. Increased content of anionic lipids with conical shape, namely cardiolipin and phosphatidic acid led to substantial increased membrane resistance to surfactin-induced permeabilization. Key words: membrane, surfactin, Bacillus subtilis, cardiolipin, black lipid membranes, liposomes
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Study of properties and effect of some natural antimicrobial substances
Horňáková, Nikola ; Kostovová, Iveta (referee) ; Márová, Ivana (advisor)
This bachelor thesis is focused on the testing of natural antimicrobial substances of plant origin, dealing with their characterisation, their effect against the microorganisms, optimization of antimicrobial testing and increasing of efficiency of their active compounds. Theoretical part of this thesis describes a characterisation of used natural species, their important properties, spectrophotometrical methods applied in determination of active compounds, opportunities of stabilization of natural substances and various techniques of antimicrobial testing. In experimental part 8 kinds of tea and non-tea plants (Pai Mu Tan, Formosa Gunpowder, Gunpowder Black, Honeybush, Lapacho, Aspalathus linearis and Paullinia cupana) were used and combined with 6 herbal species (Syzygium aromaticum, Urtica dioica, Hippophae rhamnoides, Matricaria recutita, Plantago lanceolata and cinnamon), extracted by different solvents (water, ethanol and dimethyl sulfoxide). In these extracts amount of antioxidants and effect to three bacterial cultures (Escherichia coli, Micrococcus luteus and Serratia marcescens) were determined. To improve antimicrobial, antioxidant activity and stability aqueous tea extracts were encapsulated into liposomal particles.
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Characterization of some natural substances with antimycotic effect
Plachá, Monika ; Hlaváček, Viliam (referee) ; Márová, Ivana (advisor)
The introduced bachelor thesis is focused on investigation of the effects of natural substances with potential antimycotic effects and characterization of their constituents. As a part of this work, an overview of mycosis, antimycotics and constituents of natural substances were introduced. Moreover, antifungal susceptibility and genus Candida were described. In the end of the theoretical part the liposomes and their characterization were described. In the experimental part general characteristics of some natural substances and antimycotic test were analyzed. Aqueous, ethanol, methanol and DMSO extracts were tested on yeast strain Candida glabrata. Aqueous extract of the cloves, cloves oil and their combination were encapsulated into liposomes. The encapsulation effectivity, long-term stability and their antifungal activity were determined too. Characterization of liposomes was identified by DLS. Finally, application of liposomes to a preparation of pharmaceutical and cosmetic preparations were presented.
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Use of genotoxicity tests to characterization of some natural substances and particles
Tilšarová, Kamila ; Bokrová, Jitka (referee) ; Márová, Ivana (advisor)
In production of drugs, cosmetics, in food industry and other biotechnologies, testing of genotoxicity (that are changes in genetic information which are not inhereted) is one of the common steps in the process before distribution to the market. This bachelor´s thesis is focused on the characterization of extracts from chosen natural substances with respect to the content of polyfenols, flavonoides and antioxidant activity. These extracts were encapsulated into liposomes and chitosan particles. The aim was testing possible genotoxicity of whole extracts and particles on possible genotoxicity on the basis of SOS Chromotest performed on bacteria E. coli. This genotoxic assay did not show genotoxic effect of neither pure extracts nor tested liposomes.
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Study of natural antimicrobial mixtures and their stabilization
Vejrostová, Petra ; Veselá, Mária (referee) ; Márová, Ivana (advisor)
Present diploma thesis is focused on natural antimicrobial substances. The aim of the experimental part of thesis was testing of antimicrobial activity of selected natural substances in free and encapsulated forms, preparation of particles and monitoring of their long-term stability. Various teas extracts (aqueous, ethanoic and DMSO) and aqueous herbs extracts were used as plant antimicrobial substances. Liposomes and chitosan particles with encapsulated aqueous teas extracts were prepared by ultrasonication. Encapsulation efficiency, size, stability and antioxidant activity for all particles were specified. Consequently, within a two months period, long-term stability for these particles was monitored. Spectrophotometric method for determining total polyphenols was used for evaluating encapsulation efficiency as well as for monitoring of long-term stability. The DLS method was used for determination of size and stability of particles. Antioxidation activity was monitored by using spectrophotometric method using ABTS. For testing antimicrobial activity of various types of teas, herbs extracts and their mixtures and combianitons with lysozyme and antibiotics, broth dilution methods were used while using both Gram positive and Gram negative bacterial test strains. Antimicrobial activity of prepared particles was also monitored, the antimicrobial activity was especially significant with the chitosan particles. Suitable combinations of natural substances whatever with antimicrobial or antioxidation effect either in free or encapsulated form could be used for applications for example in cosmetics industry, food processing or in medicine.
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Phase-separated systems biopolymer-lipid
Kubalová, Barbora ; Pekař, Miloslav (referee) ; Mravec, Filip (advisor)
This diploma thesis deals with experimental study of interaction based on polymer-lipid system. The main goal was to investigate interaction leading to phase separation. Three anionic polyelectrolytes were selected for this purpose: sodium hyaluronate, sodium chondroitin-6-sulfate and sodium poly(4-styrenesulfonate). The liposomes were formulated by sonication of lipid – cationic 1,2-dipalmitoyl-3-trimethylamonium-propane (chloride salt) and zwitterionic 1,2-dipalmitoyl-sn-glycero-phosphocholine. It was found that the phase separation occured at specific ratio of DPTAP and selected polymer. It was also explored that the strong electrostatic interaction which leads to the phase separation can be weaken by increasing the ionic strength in the sample. As we aspected the systems contain the hydrophobic domain and therefore these are able to incorporate hydrophobic substances in their structure.
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Preparation of some application formula of antimicrobial substances
Krempaský, Kamil ; Němcová, Andrea (referee) ; Márová, Ivana (advisor)
The main aim of this study is to determine antimicrobial activity of herbal extracts and also encapsulate these extracts and lysozyme to liposomes. The extracts were prepared from above ground part of wormwood (Artemisia absinthium), aerial part of breckland thyme (Thymus serpyllum), the root of purple coneflower (Echinacea purpurea), liquorice (Glycyrrhiza glabra), corolla of pot marigold (Calendula officinalis) and berries of sea buckthorn (Hippophae rhamnoides). In all extracts antioxidant activity and the amount of total polyphenols and flavonoids was determined. All of these extracts and also lysozyme were used for encapsulation to liposomes. Prepared liposomes were then tested for their stability. Further, encapsulated as well as free extracts were tested for their antimicrobial activity on Escherichia coli, Serratia marcescens, Micrococcus luteus and Bacillus subtilis. Extract from Echinacea purpurea showed the highest antimicrobial activity.
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|Preparation and characterization of micro- and nanoparticles with selected drugs
Sosková, Simona ; Bokrová, Jitka (referee) ; Márová, Ivana (advisor)
The presented bachelor thesis is focused on the way to prepare combined micro- and nanoparticles containing some natural extracts with antioxidant effects, ibuprofen as the anti-inflammatory and analgetic agent and clotrimazole as the agent with an antifungal effect. The theoretical part contains an introduction of the methods of liposome preparation and a review of encapsulation techniques, methods of particles characterization, and analysis of active substances content. Furthermore, the yeasts of the genus Candida and the methods of the antimicrobial activity determination are described. The mentioned components were encapsulated into liposomes, alginate and chitosan particles. The antioxidant activity of the extracts was determined spectrophotometrically. The encapsulation effectivity of active substances into particles and the long-term stability of encapsulated components were determined and compared too. The HPLC method was used for measuring of the concentration of drugs and spectrophotometric method was used to quantify the content of polyphenols. Prepared particles were characterized with DLS method (size) and zeta- potential (stability). Finally, antibacterial properties of prepared particles and extracts using the test system Candida glabrata were studied. The antimycotic effect of particles, disturbed liposomes and extracts was compared. Finally, potential utilization of prepared liposomes in a cosmetic or pharmaceutical product is proposed.
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