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The permeability and microstructure of model stratum corneum lipid membranes: comparison of non-hydroxylated and (R)- and (S)-α-hydroxylated ceramides
Šilarová, Michaela ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Ceramides are lipid components in the uppermost layer of the skin, stratum corneum and they are essential for correct function of a skin barrier. In the stratum corneum, ceramides with cholesterol and free fatty acids are in equimolar ratio. Ceramides contain sphingoid bases, which are amino alcohols sphingosine, phytosphingosine, dihydrosphingosine or 6- hydroxysphingosine. These sphingoid bases are N-acylated by non-hydroxylated, α- hydroxylated or ω-hydroxylated fatty acid, mostly by lignoceric (C24) acid. The aim of this work was to study the permeability and microstructure of the model membranes containing non-hydroxylated ceramides. Moreover, we aimed to study the effect of additional α-hydroxyl group in ceramides including the effect of stereochemistry in position 2. We prepared model membranes based on Cer, free fatty acids (C16-24), cholesterol and a small amount of cholesteryl sulfate (5 wt%). We investigated four permeability markers: electrical impedance, water loss through the membrane, flux of theophylline and flux of indomethacin. The microstructure and miscibility of ceramides with other lipids were studied by infrared spectroscopy and X-ray powder diffraction. The results from experiments showed some differences between ceramides. For example the effect of the type of sphingoid...
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Characterization of liposomal formulations for brain-targeting.
Labajová, Andrea ; Šklubalová, Zdeňka (advisor) ; Zbytovská, Jarmila (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of: Department of Pharmaceutical Technology Consultant: Assoc. Prof. Zdeňka Šklubalová, Ph.D. Robin Tremmel, Ph.D. student Student: Andrea Labajová Title of Thesis: Characterization of liposomal formulations for brain- targeting This paper deals with characterization of liposomal formulations with encapsulated triethylenetetramine (TETA), which is selective CuII - chelator used in the treatment of Wilson's disease for decades. Liposomal formulations were prepared by a film hydration method with subsequent dual asymmetric centrifugation with the addition of 2.5 mol/l TETA solution dissolved at pH3 or/and pH7, respectively. Size exclusion chromatography (SEC) was performed to separate free-TETA from the encapsulated-one at day 1, 2, 3, 4, 5 and 8. Two methods of liposome purification, the one column method and the two columns method, were used. The size and the size distribution of prepared liposomes were measured by photon correlation spectroscopy (PCS) at each day of storage. The concentration of encapsulated TETA, as well as the concentrations of cholesterol before the SEC and after the SEC were determined by HPLC in order to express the encapsulation efficiency. No influence of pH or the method of purification on the...
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The permeability and microstructure of model stratum corneum lipid membranes: comparison of non-hydroxylated and (R)- and (S)-α-hydroxylated ceramides
Šilarová, Michaela ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Ceramides are lipid components in the uppermost layer of the skin, stratum corneum and they are essential for correct function of a skin barrier. In the stratum corneum, ceramides with cholesterol and free fatty acids are in equimolar ratio. Ceramides contain sphingoid bases, which are amino alcohols sphingosine, phytosphingosine, dihydrosphingosine or 6- hydroxysphingosine. These sphingoid bases are N-acylated by non-hydroxylated, α- hydroxylated or ω-hydroxylated fatty acid, mostly by lignoceric (C24) acid. The aim of this work was to study the permeability and microstructure of the model membranes containing non-hydroxylated ceramides. Moreover, we aimed to study the effect of additional α-hydroxyl group in ceramides including the effect of stereochemistry in position 2. We prepared model membranes based on Cer, free fatty acids (C16-24), cholesterol and a small amount of cholesteryl sulfate (5 wt%). We investigated four permeability markers: electrical impedance, water loss through the membrane, flux of theophylline and flux of indomethacin. The microstructure and miscibility of ceramides with other lipids were studied by infrared spectroscopy and X-ray powder diffraction. The results from experiments showed some differences between ceramides. For example the effect of the type of sphingoid...
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Study on phase behaviour of selected lipid mixtures
Černochová, Klára ; Zbytovská, Jarmila (advisor) ; Matějka, Pavel (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Technology Consultant: Mgr. Jarmila Zbytovská, Dr. rer. nat. Student: Klára Černochová Title of Thesis: Study on phase behaviour of selected lipid mixtures Ceramides, cholesterol and fatty acids are the main components of the stratum corneum (SC) intercellular lipid matrix. These lipids play the major role in maintaining the skin barrier function. The goal of this work is to study the SC lipid mixtures on the molecular level, characterization of their miscibility and their temperature phase behavior. For this purpose, lipid model systems imitating natural structure of the SC intercellular lipid matrix were prepared. The membranes contained one or more ceramides (CerAP, CerNP, CerNS), cholesterol, stearic acid and cholesterol sulfate. The mixtures were studied by infrared spectroscopy using a heated ATR-crystal. From the measured data, IR spectra and temperature dependency of the position of the particular IR-modes were evaluated. The results showed the presence of hydrogen bindings within the ceramides. These bonds were also observable in the equimolar lipid mixture based on CerNP. Otherwise, lipid mixtures containing CerAP showed very good miscibility of the individual components. The two- and three-...
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Study of lipid membranes with ceramide precursors as models of atopic dermatitis
Staňková, Klára ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Charles University in Prague, Faculty of pharmacy in Hradec Králové Department of inorganic and organic chemistry Candidate: Klára Staňková Supervisor: doc. PharmDr. Kateřina Vávrová, Ph.D. Title of diploma thesis: Study of lipids membranes containing ceramide precursors as models of atopic dermatitis Ceramides (Cer) are important determinants for both water-retention function and permeability-barrier function in the stratum corneum (SC). They are synthesized from their precursors, i.e. sphingomyelin (SM) and glycosylceramide (GC) by the enzymes sphingomyelinase (SMase) and glucocerebrosidase (GCase). Their reduced levels have been found in the skin diseases e.g. in atopic dermatitis, and are also a causative factor for the dry and barrier-disrupted skin. The aim of this study was to prepare model skin lipid membranes simulating the defect of SMase and GCase and study their permeability. Models of SC lipid membranes were prepared as an equimolar mixture of Cer or their precursors (either SM or GC) in different ratios, cholesterol, fatty acid mixture and 5 % of cholesterol sulfate. The permeability of these model membranes was evaluated using permeation of theophylline and indometacin, transepidermal water loss and electrical impedance in Franz diffusion cells. The amount of model drug permeated...
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Study of lipid membranes with lysolipids as models of atopic dermatitis
Pospíšilová, Markéta ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of inorganic and organic chemistry Candidate: Pospíšilová, Markéta Supervisor: Assoc. Prof. PharmDr. Kateřina Vávrová, Ph.D. Title of Diploma Thesis: Study of lipid membranes with lysolipids as models of atopic dermatitis We are protected against the harmful influence of the external environment by a skin barrier, which is located in the stratum corneum. The skin barrier also prevents excessive water loss through the skin. Ceramides have the most important role in the barrier function of the skin. The healthy skin's ceramides are synthesized of glucosylceramide and sphingomyelin via enzymes sphingomyelinase and glucocerebrosidase. If there is a lack of ceramide in the stratum corneum, which means, if these mechanisms work insufficiently, it gives rise to atopic dermatitis. In the stratum corneum of a patient who suffers from atopic dermatitis are glucosylceramide and sphingomyelin hydrolyzed by glucosylceramide/sphingomyeline deacylase which leads to free fatty acids and lysolipids (glucosylsphingosine and sphingosinephosphorylcholine) generation. In this work, we prepared model membranes which simulate a composition of lipids of the stratum corneum of a patient with atopic dermatitis, in which ceramides are replaced by...
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Evaluation of the in vitro skin permeation of selected compounds using static Franz diffusion cells
Dvořáková, Tereza ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Inorganic & Organic Chemistry Student: Tereza Dvořáková Title of diploma thesis: EVALUATION OF THE IN VITRO SKIN PERMEATION OF SELECTED COMPOUNDS USING STATIC FRANZ DIFFUSION CELLS The work focuses testing the transdermal permeation of selected substances - tacrine and 7-methoxytacrine (7-Meota). Tacrine is an acetylcholinesterase inhibitor approved for the treatment of Alzheimer's disease. For its hepatotoxicity and side- effects is no longer used today. Transdermal administration could be a good alternative to reduce those side effects. 7-Meota is tacrine derivative, which in previous in vitro testing demonstrated lower toxicity in comparison with tacrine. The aim of this thesis was to clarify if and to what extent has tacrine and 7-Meota ability for transdermal permeation. Experiments were performed in vitro by using Franz diffusions cells. As receptor fluid was used for tacrine a phosphate buffer solution and for 7-Meota was a phosphate buffer solution for one part of the trial and for a second part was water. Substances were applied in an infinite dose in aqueous solution onto the surface of pig skin, which was cut to a thickness of 500 µm and 1000 µm. For the comparison was part of the experiments conducted in a...
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Study of transdermal and dermal absorption of 2,6-diaminopurine acyclic nucleoside phosphonates
Diblíková, Denisa ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Acyclic nucleoside phosphonates (ANP) are broad-spectrum antivirals highly effective against herpes-, retro- and hepadnaviruses. They also exhibit cytostatic, antiparasitic, immunomodulatory activities. Their transdermal delivery offers an attractive and advantageous route of administration, but is limited due to the polar character of their phosphonate moiety. The aim of this work was to study the possibility of both transdermal and dermal application of a series of 2,6-diaminopurine derivatives including (R)-PMPDAP and (S)-PMPDAP, (S)-HPMPDAP, (S)-8-azaHPMPDAP, cyclic (S)-HPMPDAP and lysolipid prodrugs, i.e., hexadecyloxypropyl (HDP) esters of (R)-HDP-PMPDAP and (S)-HDP- HPMPDAP. Ability of ANP to penetrate trough the skin by themselves is generally very low. For this reason the influence of permeation enhancer dodecylester of 6- (dimethylamino)hexanoic acid (DDAK) through and into human skin was investigated. The evaluation was performed in vitro by using Franz diffusion cells and human skin. The results of this work confirm that ANP (60 mM in 60 % propylene glycol) delivery through the skin is very low (flux 0.53-1.40 nmol/cm2 /h), except for the lysolipid prodrugs (R)-HDP-PMPDAP and (S)-HDP-HPMPDAP), which were not detected in the acceptor phase at all. 1 % DDAK enhanced transdermal flux of...
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Study of transdermal and dermal absorption of 2,6-diaminopurine acyclic nucleoside phosphonates
Diblíková, Denisa ; Vávrová, Kateřina (advisor) ; Zbytovská, Jarmila (referee)
Acyclic nucleoside phosphonates (ANP) are broad-spectrum antivirals highly effective against herpes-, retro- and hepadnaviruses. They also exhibit cytostatic, antiparasitic, immunomodulatory activities. Their transdermal delivery offers an attractive and advantageous route of administration, but is limited due to the polar character of their phosphonate moiety. The aim of this work was to study the possibility of both transdermal and dermal application of a series of 2,6-diaminopurine derivatives including (R)-PMPDAP and (S)-PMPDAP, (S)-HPMPDAP, (S)-8-azaHPMPDAP, cyclic (S)-HPMPDAP and lysolipid prodrugs, i.e., hexadecyloxypropyl (HDP) esters of (R)-HDP-PMPDAP and (S)-HDP- HPMPDAP. Ability of ANP to penetrate trough the skin by themselves is generally very low. For this reason the influence of permeation enhancer dodecylester of 6- (dimethylamino)hexanoic acid (DDAK) through and into human skin was investigated. The evaluation was performed in vitro by using Franz diffusion cells and human skin. The results of this work confirm that ANP (60 mM in 60 % propylene glycol) delivery through the skin is very low (flux 0.53-1.40 nmol/cm2 /h), except for the lysolipid prodrugs (R)-HDP-PMPDAP and (S)-HDP-HPMPDAP), which were not detected in the acceptor phase at all. 1 % DDAK enhanced transdermal flux of...
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Study of the Termotropic Behaviour of Short-chain Ceramid Analogues
Vavrysová, Helena ; Zbytovská, Jarmila (advisor) ; Doležal, Pavel (referee)
Ceramides of stratum corneum (SC) play a crucial role in maintaining the barrier function of the skin. This work deals with series short-chain analogues of ceramides with structure identical to naturally occuring ceramide NS, acyl chain length from 2 to 12. Acyl chain length is very important for barrier properties. Ceramides with chain length 4-8C lose barrier function, conversely were able to increase skin peremability. Ceramides with 2 and 12C showed no increase in permeability. Calorimetric and infrared spectroscopic measuring data were used for studying thermotropic phase behaviour analogues of ceramides and compared to the physiological cermide NS. At physiological temperatures, all crystalline phases of ceramides exhibit lamellar structures with highly ordered hydrocarbon chains. Structure-activity relationships and potential permeable aktivity of these ceramides were observed. Short-chain ceramides do not act as natural ceramides and their biological activity has been widely investigated.
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