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Synthesis of fluorinated serine derivatives and study of their effect on skin barrier function
Župina, Richard ; Vávrová, Kateřina (advisor) ; Opálka, Lukáš (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Organic and Bioorganic Chemistry Supervisor: Prof. PharmDr. Kateřina Vávrová, Ph.D. Consultant: Dr. Georgios Paraskevopoulos, Ph.D. Author: Richard Župina Title of the thesis: Synthesis of fluorinated serine derivatives and study of their effect on skin barrier function The human skin serves as a barrier against physical, chemical and microbial injury. When the skin barrier is perturbed, there is a possibility of microbial infiltration and biochemical damage. Ceramides, as essential components of the skin lipid layer, help to maintain the barrier integrity. Decreased content of skin ceramides in epidermis is connected with skin diseases such as atopic dermatitis and psoriasis. Previous findings of our research group proved that a ceramide analogue with similar steric and hydrophobic parameters to skin ceramides could selectively supplement the recovery of perturbed human skin barrier. The goal of this study was to design a convenient synthesis of ceramide analogues derived from L-serine with two long aliphatic chains. The first chain is amidically bound lignoceric acid and the second chain is either a 10 or a 14 carbon polyfluorinated chain conjugated with the rest of the molecule by an ester bond. Furthermore, the aim of this...
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Study of Topical Ceramide Application
Kulhánková, Roksolana ; Kováčik, Andrej (advisor) ; Vraníková, Barbora (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Technology Author: Roksolana Kulhánková Supervisor: doc. PharmDr. Andrej Kováčik, Ph.D. Title of diploma thesis: Study of topically administred ceramides The human stratum corneum (SC) is the uppermost layer of the epidermis, which is made up of several layers of corneocytes. The space between the corneocytes is filled with barrier lipids, i.e., ceramides, cholesterol, and fatty acids, which are represented in an equimolar ratio. The arrangement of intercellular lipids has a significant effect on the barrier properties of the skin. Damaged skin barrier (e.g., in atopic dermatitis) is associated with a lack of lipids in the SC lipid matrix, which is reflected in increased trans-epidermal water loss and deterioration of skin quality (e.g., severe dryness, flaking, and cracks). One of the potential ways to restore the skin barrier is the topical administration of barrier lipids, especially ceramides (Cer). The goal of this work was to study the effect of topically administered Cer (very long-chain Cer AP alone or in combination with ultra-long-chain Cer EOS) on model-damaged SC and their effect in the process of restoring barrier function. Several topically administered formulations (creams) were prepared, which...
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Preparation of the human skin barrier model using thin lipid film deposition
Suciu-Šubert, Karina ; Pullmannová, Petra (advisor) ; Opálka, Lukáš (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biophysics and Physical Chemistry Student: Karina Suciu-Šubert Supervisor: Mgr. Petra Pullmannová, Ph.D. Title of diploma thesis: Preparation of the human skin barrier model by using thin lipid film deposition The uppermost part of the human skin - the stratum corneum, which protects the body from the external environment, is made up of cornified cells - corneocytes, which are surrounded by the extracellular matrix of highly ordered lipids: ceramides (Cer), fatty acids and cholesterol. The protective barrier is constantly renewed with so-called keratinocytes, which differentiate to corneocytes and finally desquamate from skin to surface. The corneocyte envelope replaces plasma membrane of corneocytes. It consists of proteins (eg. involucrin) and a lipid monolayer consisting mainly of ω-hydroxyceramides which are attached to the protein segments by covalent bonds. The task of this diploma thesis was the preparation of the human skin barrier model by using thin lipid film deposition. Two different lipid mixtures were based on Cer NS or Cer EOS and NS. The mixtures were applied on the hydrophilic or hydrophobic substrate by spraying with Linomat V. The amount of lipids and consequently the number of theoretical lipid layers on the...
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The human skin barrier model containing stripped stratum corneum
Soukupová, Romana ; Pullmannová, Petra (advisor) ; Kováčik, Andrej (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biophysics and Physical Chemistry Author: Romana Soukupová Supervisor: Mgr. Petra Pullmanová, Ph.D. Title of diploma thesis: The human skin barrier model containing stripped stratum corneum The uppermost layer of the epidermis, the stratum corneum, acts as a protective skin barrier. It contains a lipid matrix, which is made up of ceramides, cholesterol, and free fatty acids. The composition and arrangement of these lipids are necessary for the proper function of the skin barrier. Corneocytes, which do not have nuclei, are distributed in the lipid matrix. They are surrounded by insoluble proteins with covalently bound lipids on their surface, creating the corneocyte lipid envelope. The aim of this diploma thesis was to prepare models of the human skin barrier using an isolated stratum corneum with a preserved corneocyte lipid envelope and without a preserved corneocyte lipid envelope, and then monitor and compare the lipid arrangements in these models. Obtaining the stratum corneum from the skin samples was necessary to prepare the models. Human skin lipids were extracted from isolated stratum corneum and part of this delipidated stratum corneum was saponified. Skin lipids were purificated by column chromatography, and their...
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Synthesis and evaluation of human 6-hydroxyceramides
Majcher, Adam ; Opálka, Lukáš (advisor) ; Krátký, Martin (referee)
Title of the Master thesis: Synthesis and evaluation of human 6-hydroxyceramides Candidate: Adam Majcher Supervisor: PharmDr. Lukáš Opálka, Ph.D. Consultant: PharmDr. Andrej Kováčik, Ph.D. Charles University, Faculty of Pharmacy in Hradec Králové Skin Barrier Research Group, Department of Organic and Bioorganic Chemistry Ceramides (Cer), the members of sphingolipid family, occur in all human cells and play an important role in cell signaling. In high concentrations, Cer can also be found in the uppermost layer of epidermis called stratum corneum, along with free fatty acids and cholesterol (in equimolar ratio), where they form the intercellular multi-lamellar lipid matrix. The key function of stratum corneum is to ensure a permeability barrier, thus, to provide water and electrolyte homeostasis, and to prevent entry of harmful substances into the organism. Cer are composed of a sphingoid base and an acyl part derived from a long-chain fatty acid. Cer based on 6-hydroxysphingosine (H) are amongst the most unusual sphingolipids. In contrast to sphingosine-based Cer, 6-hydroxysphingosine-based Cer (H-Cer) are unique for the epidermis and, in addition, H-Cer are not typical for all mammals. Moreover, the function and biosynthesis of H-Cer in the skin is still not completely understood. Several...
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Evaluation of preparation of monolayer lipid skin barrier models
Růžičková, Karolína ; Švecová, Barbora (advisor) ; Vraníková, Barbora (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Technology Author: Karolína Růžičková Supervisor: PharmDr. Barbora Švecová, Ph.D. Consultant: Mgr. Anna Nováčková Title of thesis: Evaluation of preparation of monolayer lipid skin barrier models Skin, the protective barrier of human body, consists of several layers. The uppermost one is the stratum corneum, part of epidermis, whose extracellular matrix is composed mainly of ceramides, cholesterol and free fatty acids. The composition and arrangement of skin lipids are essential for the proper skin barrier function. Various multilayer and monolayer models are used to study skin lipids at the molecular level. Some of the evaluation methods are Langmuir monolayers at the air interface. In this work I dealt with the behavior of monolayer lipid models at four different pH values of the liquid subphase. Lipids isolated from human skin, lipid mixture prepared from the individual components, and a mixture of fatty acids were compared as well. Langmuir isotherms and the Brewster angle microscopy at different compression rates were used for this purpose. The results showed that pH of the subphase has no major effect on lipids arrangement. Lipids were most likely to form a tight monolayer at neutral pH 7,0, at a...
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Optimization of the skin barrier model with isolated ceramides of human stratum corneum
Dulanská, Lucia ; Pullmannová, Petra (advisor) ; Zbytovská, Jarmila (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biophysics and Physical Chemistry Author: Lucia Dulanská Supervisor: Mgr. Petra Pullmannová, Ph.D Title of thesis: Optimization of the skin barrier model with isolated ceramides of human Stratum corneum Stratum corneum (SC), the uppermost layer of the skin, regulates transcutaneous water loss and protects against outer conditions and harmful substances. It consists of cornified cells - corneocytes and extracellular lipid matrix, which is responsible for the barrier functions. Corneocytes are covered with covalently bound lipids creating the corneocyte lipid envelope (CLE). CLE is considered to interconnect the extracellular lipids with corneocytes and to have a templating effect. We aimed to optimize a skin lipid model simulating also the presence of CLE. The lipidic part of the model was prepared from an equimolar mixture of isolated human skin ceramides (hCer), cholesterol and free fatty acids (FFA, either protonated or deuterated) with 5 weight % of cholesteryl sulfate. hCer were extracted from the isolated human SC and purified by the column chromatography. The composition of hCer was determined by the high- performance thin-layer chromatography. The reverse-phase and normal phase silica gel particles served as the CLE...
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Study of free sphingoid bases in skin barrier
Jarešová, Zuzana ; Kováčik, Andrej (advisor) ; Vraníková, Barbora (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Technology Author: Zuzana Jarešová Supervisor: PharmDr. Andrej Kováčik, Ph.D. Consultant: PharmDr. Lukáš Opálka, Ph.D. Title of diploma thesis: STUDY OF FREE SPHINGOID BASES IN SKIN BARRIER The skin barrier, localized in the stratum corneum (SC), consists of corneocytes and an intercellular matrix formed from three types of lipids - ceramides, free fatty acids, and cholesterol, represented in an equimolar ratio. The overall arrangement of lipids is organized and highly specialized. Ceramides are structurally formed from the fatty acid acyl attached to a sphingoid base. In minor but not insignificant amounts, free sphingoid bases can also be found in the skin barrier. Several studies show that there is an increased concentration of free sphingoid bases in skin barrier disorders, such as atopic dermatitis. Although it is assumed that the presence of free sphingoid bases affects the skin barrier, it is not elucidated the way of their participation till today. The lack of studies or their diverse results leads us to the main goal of this thesis - to clarify how free sphingoid bases influence the skin barrier. In this work, the model membranes were prepared by the isolation of human SC ex vivo. Sphingosine (S),...
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Synthesis and evaluation of human 6-hydroxyceramides
Majcher, Adam ; Opálka, Lukáš (advisor) ; Krátký, Martin (referee)
Title of the Master thesis: Synthesis and evaluation of human 6-hydroxyceramides Candidate: Adam Majcher Supervisor: PharmDr. Lukáš Opálka, Ph.D. Consultant: PharmDr. Andrej Kováčik, Ph.D. Charles University, Faculty of Pharmacy in Hradec Králové Skin Barrier Research Group, Department of Organic and Bioorganic Chemistry Ceramides (Cer), the members of sphingolipid family, occur in all human cells and play an important role in cell signaling. In high concentrations, Cer can also be found in the uppermost layer of epidermis called stratum corneum, along with free fatty acids and cholesterol (in equimolar ratio), where they form the intercellular multi-lamellar lipid matrix. The key function of stratum corneum is to ensure a permeability barrier, thus, to provide water and electrolyte homeostasis, and to prevent entry of harmful substances into the organism. Cer are composed of a sphingoid base and an acyl part derived from a long-chain fatty acid. Cer based on 6-hydroxysphingosine (H) are amongst the most unusual sphingolipids. In contrast to sphingosine-based Cer, 6-hydroxysphingosine-based Cer (H-Cer) are unique for the epidermis and, in addition, H-Cer are not typical for all mammals. Moreover, the function and biosynthesis of H-Cer in the skin is still not completely understood. Several...
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Study of sphingosine, dihydrosphingosine and phytosphingosine in skin barrier models
Kubátová, Denisa ; Kováčik, Andrej (advisor) ; Svačinová, Petra (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Technology Author: Denisa Kubátová Supervisor: PharmDr. Andrej Kováčik, Ph.D. Consultant: PharmDr. Lukáš Opálka, Ph.D. Title of diploma thesis: Study of sphingosine, dihydrosphingosine and phytosphingosine in skin barrier models The stratum corneum (SC), the uppermost layer of the skin, localized in the uppermost part of the epidermis, represents the skin barrier of the organism. SC is composed of corneocytes and an intercellular lipid matrix, which is formed by ceramides (Cer), free fatty acids (FFA), and cholesterol (Chol) in an equimolar ratio. Substances from the group of sphingolipids - Cer, are sphingoid bases (for example, sphingosine (S), dihydrosphingosine (dS), phytosphingosine (P)) acylated with a fatty acid (for example, lignoceric acid (LIG)). In the lipid matrix, the metabolic products of Cer (free sphingoid bases) are also present, but their role in SC barrier functions is not clear. Some studies show that Cer with different sphingoid bases, and increased presence of free sphingoid bases, can lead to a change in the permeability of the skin barrier. This work aimed to study the effect of permeability of sphingoid bases on the model membrane permeability. Nine types of membranes were prepared; they...
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