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Processsing of food supplements based on medical types of mushrooms
Vetchá, Vendula ; Uhlířová, Renata (referee) ; Hoová, Julie (advisor)
This diploma thesis deals with the characterization of nutrition substances in medical mushrooms, specifically Ganoderma lucidum and Hericium erinaceus. It also focuses on the biological impact of these rare mushrooms on human cell lines HaCaT and Caco-2, and investigates the production of pro-inflammatory cytokines IL-6 and IL-8 by immortalized keratinocytes and colon cancer cells after exposure to aqueous extracts prepared from mushroom powder. The theoretical part of the thesis deals with the characterization of mushrooms in terms of morphological characteristics and their taxonomic classification. It further studies the effects of edible and medicinal species and provides a detailed description of medicinal mushrooms such as Ganoderma lucidum, Lentinula edodes, Flammulina velutipes, Grifola frondosa, Hericium erinaceus and Pleurotus ostreatus, thesis informing about their effects, utilization and possibilities for application in the food and pharmaceutical industries. It mentions possible negative effects of mushrooms on consumers' health and presents methods for processing mushrooms and their use in the food industry. Finally, it focuses on the possibilities of in vitro toxicity testing and provides a brief overview of commonly used cytotoxicity tests such as the MTT test and LDH test. The experimental part of the thesis deals with the preparation of water extracts from mushroom powders and their characterization in terms of nutritional and biologically active substances. HPLC analysis is used to monitor the content of lipophilic substances and vitamin C in mushroom samples. To confirm the biological effect on human health, antimicrobial tests were performed using four model microorganisms, and cytotoxicity tests were performed on two human cell lines. Lastly, a sensory analysis of instant vegetable soup enriched with mushroom powder was performed. Biological effects of the analyzed powdered extracts from Ganoderma lucidum mushroom have been confirmed, especially their cytotoxic effect on colon epithelial tumor cells. However, the application of reishi in the food industry is not easy due to the presence of triterpenoids that cause a bitter taste. On the other hand, the safety of using the Hericium erinaceus mushroom in food has been confirmed through cytotoxicity tests. Sensory analysis results have shown a positive effect of Hericium erinaceus on the taste profile of instant vegetable soups.
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Development of novel Constitutive androstane receptor (CAR) ligands
Dušek, Jan ; Pávek, Petr (advisor) ; Hudeček, Jiří (referee) ; Chládek, Jaroslav (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate Mgr. Jan Dušek Supervisor Prof. PharmDr. Petr Pávek, Ph.D. Title of Doctoral Thesis Development of novel Constitutive androstane receptor (CAR) ligands Constitutive androstane receptor is nowadays known as the established nuclear receptor that regulates the expression of several key cytochrome P450 enzymes, predominantly CYP3A4 and CYP2B6. Recently it has been shown that CAR has also essential role in the regulation of endogenous metabolism of glucose, lipids, cholesterole or bile acids. Simultaneously, this receptor is considered to have proliferative effect on human hepatocytes and protective effects against toxic and dietary damage of liver parenchyme. Given the possible therapeutic utilization of CAR, its therapeutic options are being intensively studied. Unfortunately, currently known ligands of human or mouse CAR are either poorly selective or indirect. The aim of my doctoral thesis was to find new ligands of mouse and human CAR, that would enable more detailed study of the receptor.
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In vitro and ex vivo study of drug-drug interactions of antiretrovirals on intestinal ATP-binding drug transporters
Jahodová, Michaela ; Červený, Lukáš (advisor) ; Vokřál, Ivan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Bc. Michaela Jahodová Supervisor: PharmDr. Lukáš Červený, Ph.D. Title of diploma thesis: In vitro and ex vivo study of drug-drug interactions of antiretrovirals on intestinal ATP-binding drug transporters The absorption of orally administered drugs takes place especially in the intestine, where it can affect by the activity of drug's ABC transporters located on the apical membrane of the intestinal epithelium. Study of drug interactions in intestinal ABC transporters is essential to ensure effective and safe pharmacotherapy. Testing of bi- directional transport on Caco-2 cells is generally the preferred method for in vitro evaluation of substrates and inhibitors of ABC transporters. Drawbacks of the Caco-2 model increase the need and necessity to introduce new models. A great potential is the involvement of ex vivo methodologies in the human or rat intestine. The aim of the work was to introduce an in vitro methodology using the Caco-2 cell monolayer and the ex vivo methodology of precision-cut rat intestinal slices. By the bi-directional transport method, we analyzed drug interactions of the model substrate P-gp and BCRP Rhodamine 123 (RHD123) and clinically-used tenofovir...
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Development of novel Constitutive androstane receptor (CAR) ligands
Dušek, Jan ; Pávek, Petr (advisor) ; Hudeček, Jiří (referee) ; Chládek, Jaroslav (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate Mgr. Jan Dušek Supervisor Prof. PharmDr. Petr Pávek, Ph.D. Title of Doctoral Thesis Development of novel Constitutive androstane receptor (CAR) ligands Constitutive androstane receptor is nowadays known as the established nuclear receptor that regulates the expression of several key cytochrome P450 enzymes, predominantly CYP3A4 and CYP2B6. Recently it has been shown that CAR has also essential role in the regulation of endogenous metabolism of glucose, lipids, cholesterole or bile acids. Simultaneously, this receptor is considered to have proliferative effect on human hepatocytes and protective effects against toxic and dietary damage of liver parenchyme. Given the possible therapeutic utilization of CAR, its therapeutic options are being intensively studied. Unfortunately, currently known ligands of human or mouse CAR are either poorly selective or indirect. The aim of my doctoral thesis was to find new ligands of mouse and human CAR, that would enable more detailed study of the receptor.
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In vitro transport of abacavir across the monolayer of Caco-2 cells; interaction with etravirine and rilpivirine
Mlčochová, Alice ; Čečková, Martina (advisor) ; Červený, Lukáš (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Alice Mlčochová Supervisor: PharmDr. Martina Čečková, Ph.D. Title of diploma thesis: In vitro transport of abacavir across the monolayer of Caco-2 cells; interaction with etravirine and rilpivirine. Abacavir belongs among nucleoside reverse transciptase inhibitors (NRTIs) representing a basic component of combined antiretroviral therapy used in treatment of HIV-positive patients [1]. Etravirine and rilpivirine are newer non-nucleoside reverse transcriptase inhibitors (NNRTIs) combined in cART together with NRTI. ATP-dependent transporters, so called ABC transporters, are able to affect pharmacokinetic properties of drugs, thus they are important site of drug-drug interactions affecting absorption, distribution and excretion level. P-glycoprotein (Pgp, ABCB1) and BCRP (ABCG2) belong among the most clinically important ABC transporters able to cause drug-drug interactions. The aim of this thesis was to introduce and optimize the method for evaluation of drug absorption using monolayers of Caco-2human intestine cell lines, whose integrity was verified by evaluating TEER (transepithelial electrical resistance). This model was also used for abacavir transport studies. Significant...
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In vitro and ex vivo study of drug-drug interactions of antiretrovirals on intestinal ATP-binding drug transporters
Jahodová, Michaela ; Červený, Lukáš (advisor) ; Vokřál, Ivan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Bc. Michaela Jahodová Supervisor: PharmDr. Lukáš Červený, Ph.D. Title of diploma thesis: In vitro and ex vivo study of drug-drug interactions of antiretrovirals on intestinal ATP-binding drug transporters The absorption of orally administered drugs takes place especially in the intestine, where it can affect by the activity of drug's ABC transporters located on the apical membrane of the intestinal epithelium. Study of drug interactions in intestinal ABC transporters is essential to ensure effective and safe pharmacotherapy. Testing of bi- directional transport on Caco-2 cells is generally the preferred method for in vitro evaluation of substrates and inhibitors of ABC transporters. Drawbacks of the Caco-2 model increase the need and necessity to introduce new models. A great potential is the involvement of ex vivo methodologies in the human or rat intestine. The aim of the work was to introduce an in vitro methodology using the Caco-2 cell monolayer and the ex vivo methodology of precision-cut rat intestinal slices. By the bi-directional transport method, we analyzed drug interactions of the model substrate P-gp and BCRP Rhodamine 123 (RHD123) and clinically-used tenofovir...
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Cell culture-based model for the evaluation of adhesive properties of probiotic bacteria
Theodorou, Vasiliki ; Havlík, Jaroslav (advisor) ; Tauchen, Jan (referee)
Probiotic microorganisms, defined as living microorganisms which when administered in adequate amounts confer a health benefit to the host, and their adhesion and colonization of intestinal epithelium, are critical factors in maintaining probiotic efficacy. Polyphenols are a large and heterogeneous group of phytochemicals in plant-based foods, such as tea, coffee, wine, cocoa, cereal grains, soy, fruits and berries. In the last decade, there has been much interest in the health benefits of dietary plant polyphenols that arise from their potential ability to promote adhesion of probiotic bacteria to the human intestinal epithelium.
The purpose of this study was to investigate the effect of four polyphenols: isoquercetrin, phloretin, procyanidin B2 and rutin on the adhesion ability of two potentially probiotic strains (Lactobacillus casei, Lactobacillus gasseri) to in vitro human intestinal epithelial model consisting of Caco-2 and mucus-secreting HT29-MTX co-culture.
The adhesion of Lactobacillus casei after treating the co-culture cell lines with isoquercetrin, phloretin, and rutin was increased by 49.76, 72.97, 63.66 % respectively, whereas procyanidin B2 inhibited the adhesion 20.25% compared with the control sample. The adhesion of Lactobacillus gasseri after treatment of the co-culture with isoquercetrin, phloretin, procyanidin B2 and rutin was increased by 35.45, 31.28, 45.69, 25.01 % respectively compared with the control sample.
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In vitro anti-proliferation activity alkaloids the Amaryllidaceae
Panenková, Kristýna ; Havlík, Jaroslav (advisor) ; Jarošová, Veronika (referee)
Summary
Natural phytochemicals are currently used in the treatment of many diseases. Cancers are just ones of them and they are ranked among the most common and the most serious. Phytochemicals in the form of cytostatics are used in chemotherapeutic treatment of cancer. In future there could be included among cytostatics also some alkaloids from the family
of Amaryllidaceae, whose testing for a selective cytostatic effect on tumor cell lines
of colorectal carcinoma Caco-2 and HT-29 and on normal cell lines of human intestinal epithelial FHs 74 Int is a subject of this thesis.
There were tested 17 alkaloids isolated from plants of Chlidanthusfragrans, Zephyranthes robusta and Nerine bowdenii. Particularly alkaloids from plant Zephyranthes robusta namely haemanthamine with this values: IC50 = 0.99 plus/minus 0.14 microM for tumor cells, Caco-2, 0.59 plus/minus 0.01 microM for tumor cells HT-29 and 19.47 plus/minus 8.86 microM for normal cells
FHs 74 Int, Lycorine with values IC50 = 0.99 plus/minus 0.08 microM for tumor cells Caco-2, 1.2 plus/minus 0.01 microM for tumor cells HT-29 and 22.68 plus/minus 0.09 microM for normal cells FHs 74 Int and Haemanthidin with values IC50 = 3.29 plus/minus 0.91 microM to tumor cells Caco-2, 1.72 plus/minus 0.11 microM to tumor cells HT-29, and 11.63 plus/minus 0.86 microM for normal cells FHs 74 Int proved a significant
anti-proliferative activity.
From these results there is evident the selectivity against colorectal cancer cell lines. For this reason, those tested alkaloids are suitable for further testing and for study of their biological activity against tumor cells in the terms of in vitro and in vivo.
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