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Phytoremediation potential of higher plants - phytoextraction of paroxetine
Štelbacká, Kristína ; Smrček, Stanislav (advisor) ; Hýsková, Veronika (referee)
Environmental pollution has long been an important issue for humanity. There are many mechanical and chemical pollutants in water and soil that need to be removed. Xenobiotics, which are increasingly found in the environment, are also a problem. This thesis deals with the study of the phytoextraction of the drug paroxetine using the plant maize (Zea mays). The antidepressant paroxetine belongs to the group of selective serotonin reuptake inhibitors and is used for the treatment of depressive disorders. The plants were cultured in sterile medium for the time necessary for sufficient growth. Subsequently, paroxetine was added to the plant medium and samples were collected for 96 hours. For the first experiment the concentration of drug in the medium was 3 mg ∙ l−1 , for the second 2 mg ∙ l−1 and for the third 1 mg ∙ l−1 . To determine the phytoextraction activity, the collected samples were detected by HPLC system with UV detector. The phytoextraction efficiency of paroxetine was determined to be 0.0023 mg/g(fresh weight) for the first experiment and 0.0014 mg/g(fresh weight) for the second experiment. Plants used for phytoremediation have increased potential for oxidative stress and reactive oxygen species formation. After the phytoextraction experiments, the antioxidant capacities of the leaf and...
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Interaction of the anticancer drug lenvatinib with cytochrome P450 subfamily 2C
Srogoň, Jiří ; Dračínská, Helena (advisor) ; Hýsková, Veronika (referee)
Lenvatinib is a multi-targeted kinase inhibitor capable of inhibiting these kinases at nanomolar concentrations. For this reason, it is used as a drug in the treatment of various types of cancer. Like many other xenobiotics, lenvatinib is metabolized by cytochromes P450, which can cause occurrence of drug interactions with other substances. Identification of the most important and clinically relevant drug interactions is essential to ensure the safety of patients already suffering from reduced quality of life due to cancer. The main aim of this bachelor thesis was to investigate the effect of lenvatinib on cytochromes P450, specifically on isoforms of subfamily 2C in humans and rats. The inhibitory effect was measured in vitro using marker rections on rat liver microsome samples for the CYP2C6 isoform and on human recombinant CYP2C9 isoform samples expressed in SuperosomesTM or Bactosomes® . Furthermore, the effect of lenvatinib on CYP2C11 and 2C6 expression in the liver of rats exposed to lenvatinib was investigated. Lenvatinib caused a decrease in the activity of the human recombinant CYP2C9 isoform, whereas no effect was observed on the activity of the CYP2C6 isoform in rat liver microsomes. One of the other objectives was to determine the IC50 for CYP2C9, but the objective was not met. When...
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The role of glutathione in the metabolism of plants exposed to environmental contaminants.
Rychnová, Andrea ; Hýsková, Veronika (advisor) ; Indra, Radek (referee)
Environmental contamination by pharmaceuticals is currently occurring at an unsustainable rate. It is in the public interest that xenobiotics, which enter not only wastewater but also groundwater and drinking water, are removed from the contaminated environment in an efficient and environmentally sound manner. A promising method is phytoremediation, which can meet the requirements. Another contemporary issue is the increasing scarcity of water and the related question of whether it is possible to recycle wastewater and use it for irrigating crops. How plant metabolism responds to pharmaceuticals can be elucidated by biomarkers of oxidative stress caused by xenobiotics. These include the activity of enzymes associated with the detoxification and antioxidant systems of plants. In this work, the effect of currently increasingly used antidepressants (buspirone, sulpiride and paroxetine) was investigated in maize (Zea mays L., DKC 3969) plants grown under sterile conditions in vitro. The work was focused on enzymes related to glutathione metabolism and on ascorbate peroxidase (APX). In plant leaves, the activity of glutathione-S-transferase (GST), a conjugating enzyme of phase II biotransformation of xenobiotics, was increased by all drugs. In addition, a drug-inducible isoform of GST was detected by...
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Effect of fungicides and biological control agens on glutathione metabolism.
Vašková, Marie ; Hýsková, Veronika (advisor) ; Jaklová Dytrtová, Jana (referee)
Triazole fungicides are widely used in agriculture to treat a large number of crops. When they accumulate in soil, plants or water sources, they can also affect non-target organisms, in which they can have a negative effect on the endocrine system or re- production. Much less is known about the effect of triazoles on plants, specifically their antioxidant and detoxification systems. In this work, the effect of penconazole (P), tebuconazole (T) or their combination (PT) on tomato plants Solanum lycopersicum, cv. Cherrola was studied. In contrast to the conventional method of fungicide application by foliar spraying, the effect of soil drench containing P, T or PT was also studied. Soil drenching of fungicides had a worse impact on glutathione metabolism: on the thirty-fifth day after weekly fun- gicide application, the content of reduced thiols and glutathione peroxidase activity were reduced in roots, and the key conjugation enzyme of phase II biotransformation, glutathione-S-transferase (GST), was reduced in leaves by 43 to 20 % (depending on the fungicide) compared with untreated plants. In contrast, the content of reduced thiols and glutathione peroxidase activity were increased in leaves. In the case of spraying with both fungicides (PT), although at the same concentration as P and T alone, a...
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Antioxidant system of tomato plants exposed to triazole fungicides
Žufić, Antoniana ; Hýsková, Veronika (advisor) ; Jaklová Dytrtová, Jana (referee)
Fungal pathogens cause severe crop losses worldwide. Recently, however, chemical fungicides have become dangerous environmental contaminants affecting non-target organisms. The aim of this study was to investigate the effect of triazole fungicides in the form of penconazole (P), tebuconazole (T) or their combination (PT) on tomato plants Solanum lycopersicum L. cv. Cherrola. No significant change in the activity of a key enzyme of the ascorbate-glutathione cycle, ascorbate peroxidase and activity of cytosolic peroxides, was detected in leaves, roots or fruits at the intervals studied, i.e. after two and five weekly treatments of P, T or PT, either as a foliar or a soil application. While slightly increased guaiacol peroxidase activity was found in roots due to T spraying and PT combination, the activity of this enzyme was reduced in leaves similar to catalase. The greatest changes in leaf antioxidant capacity correlated with increased phenolic and flavonoid content were observed after five treatments of penconazole (P) as a soil application both P alone and in combination as PT. The studied triazoles significantly affected the abundance of isoforms of superoxide dismutase and partially total peroxides in leaves, roots and fruits and of acorbate peroxidase in leaves. The highest content of HSP70 was found in...
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Preparation of mitotic inhibitors in the form of nanotransporters
Meskařová, Veronika ; Indra, Radek (advisor) ; Hýsková, Veronika (referee)
Cancer is one of the most widespread diseases that every third person will encounter it. It is a disease arising from the sudden growth and rapid division of own cells. Despite progress in treatment, an effectiveness is not sufficient, and cancer is the second most common cause of death. In addition, the treatment is associated with several side effects. Currently nanomedicine uses nanomaterials for transport drugs in cancer treatment. It is possible to deliver the drug to the target site, but also to reduce the systemic toxicity of the drug by binding the drug to a suitable nanotransporter. This diploma thesis deals with seeking the appropriate mass ratio of antitumor drugs, colchicine and docetaxel, and the nanotransporter apoferritin. The aim was to find suitable in vitro conditions for high drugs encapsulation into the nanotransporter. Drugs encapsulation took place under active and passive conditions. Active encapsulation was under way on opening and closing nanotransporter by changing pH environment in test tube. Drugs were dissolved in DMSO or water with addition of HCl to charge the drug molecules and improve the binding of the drug into nanotransporter. As part of passive encapsulation, drugs were incubated in surroundings of urea or bivalent metal ions. Overall, it can be concluded that...
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Plant alkaloid sanguinarine and its derivatives
Tůmová, Lucie ; Stiborová, Marie (advisor) ; Hýsková, Veronika (referee)
This work is summarizing actual knowledge about sanguinarine and quaternary benzo[c]fenanthridine alkaloids. The quaternary benzo[c]fenanthridine alkaloids were found in roots plants Sanguinaria canadensis and Macleaya cordata. This plants are used in tradicional Chinese medicine for its antimycotic, antibacterial and anti-inflammatory activities since antiquity. Regarding to possibility quaternary benzo[c]fenanthridine alkaloids to induce apoptosis these investigated such as possible agents for cancer treatment. The quaternary benzo[c]fenanthridine alkaloids interact with DNA and proteins. They are able to intercalate to the DNA. The alkaloids can be used like fluorescence DNA probe. Metabolism by sanguinarine and next quaternary benzo[c]fenanthridine alkaloids has not yet completely determinated. The first step sanguinarine detoxication is its conversion to less toxic dihydrosanguinarine. Sanguinarine oxidation is mediated by cytochrome P450 1A1. Key words: Quaternary benzo[c]fenanthridine alkaloids, sanguinarine, apoptosis, intercalate, heterogenous substances, enzymes, cytochrom P450
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Enzyme kinetics and regulation of shikimate dehydrogenase from parsley roots.
Šmeringaiová, Ingrida ; Hýsková, Veronika (advisor) ; Kalendová, Alžběta (referee)
In plants the shikimate pathway followed by the phenylpropanoid pathway leads to the formation of not only aromatic amino acids but also a plenty of secondary metabolites. The crucial enzyme shikimate dehydrogenase is there a part of bi-functional protein 3-dehydroquinate dehy- dratase/shikimate dehydrogenase (DHD/SDH; EC 4.2.1.10 and EC 1.1.1.25). Whereas the regulation of the shikimic pathway is complex, little is known about the feedback regulation of plant SDH. The aim of this project was to find plant source with high SDH activity and to find out, if and in which way this SDH is regulated by compounds of phenylpropanoid metabolism, specifically by simple polyphenols. SDH from parsley root (Petroselinum crispum) has been prepared by 3-step purification to final specific activity 470 mol.min-1 .mg-1 . The enzyme exhibited one band after both isoelectric focusing and red native electrophoresis using detection of the activity. Relative molecular mass of native enzyme was determined by gel chromatography (Mr 60 000) and red native electrophoresis (Mr 63 000). Isoelectric point pI 4.5 was established by isoelectric focusing. Optimal pH for the reaction catalyzed by SDH was determined in rage pH 9.5 - 10.0. The kinetic parameters (Km, Vmax) of SDH for substrate shikimate, and coenzyme NADP+ were...
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Aggregation of bull seminal plasma protein
Boháček, Hanuš ; Liberda, Jiří (advisor) ; Hýsková, Veronika (referee)
Mammalian fertilization is a sequence of unique and fascinating events, during which seminal proteins are of crucial role. In case of bull (Bos taurus), proteins of seminal plasma (BSP), especially its major component PDC-109, are known to be in aggregated forms, but little is known about mechanism of forming aggregates and their biological function. In present thesis we discovered some interesting properties of PDC-109 and BSP proteins. We found that concentration of these proteins influences their aggregation state significantly, which can be of great biological importance. Separation of seminal proteins by size exclusion chromatography revealed three main fractions denoted I, II and III, with apparent molecular weights of Mr > 150 000, Mr = 30 000 and Mr = 13 000, respectively. In case of PDC-109, molecular weights of theese fractions were retained even after purification procedure, which implies very stable interactions in forming of aggregates. In addition, there was a difference in distribution of PDC-109 glycoforms among fractions, which can be related to the fact, that theese fractions have different sperm membrane binding patterns as we determined by fluorescence microscopy. However, further experiments are needed for better understanding this issue.
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