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Effect of pergafast 201 on selected markers of in vitro maturation of porcine oocytes.
Procházková, Bára ; Petr, Jaroslav (advisor) ; Benc, Michal (referee)
Pergafast 201 (N-(p-Toluenesulfonyl)-N'-(3-p-toluenesulfonyloxyphenyl)urea) is a patented color developer that is offered as an alternative to the widely used bisphenols A and S (BPA, BPS). These substances have been shown to have a negative effect on human health and reproduction even in very low doses, and they are endocrine disruptors affecting, among other things, the process of meiotic maturation. The effects of pergafast 201 on the mammalian organism during oogenesis have not yet been monitored. The aim of this work was to evaluate the effect of pergafast 201 on selected specific markers of meiotic maturation after exposure of porcine oocytes to pergafast 201 in low concentrations in vitro. This work is the first to describe the negative effects of pergafast 201 on the maturation of porcine oocytes in vitro, confirming the negative effect of pergafast on the course of meiotic maturation and on the ability of the oocyte to reach the final stage of maturation, as well as its effect on the increased incidence of abnormal dividing spindles. The data show the effect of PF201 on the methylation of epigenetic markers H3K4me2 and H3K9me3. Preliminary results also indicate its effect on the mRNA expression of estrogen receptors α and β. Collectively, these results indicate the possible risks...
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Mixture toxicity of estrogenic endocrine disruptors in samples from the Czech Republic
Černá, Tereza ; Cajthaml, Tomáš (advisor) ; Klusoň, Petr (referee) ; Šváb, Marek (referee)
The environment is contaminated by countless toxic chemicals of mainly anthropogenic origin; therefore, elucidating the impact of chemical mixtures is one of the biggest challenges of current research. Endocrine disruptors (EDs) acting through the oestrogenic nuclear receptor are typical representatives of the mixture toxicity concept of concentration addition, which can be accurately predicted using the latest mathematical models. Oestrogenic activity and oestrogenic EDs have been detected in many matrices across the globe in biologically relevant concentrations; therefore, they are considered a high risk. The presence of synthetic steroids in the aquatic environment in units of ng/l can disrupt the hormonal pathways of individuals and subsequently cause fatal changes in whole populations. However, if the exposure is terminated, the ecosystem will recover in a few years. Wastewater treatment plants (WWTPs) are able to remove a large number of chemicals, including synthetic and natural oestrogenically active compounds; nevertheless, their occurrence in WWTP effluents and in sewage sludges is frequent. Therefore, WWTPs are considered significant sources of agricultural and water ecosystem pollution. This dissertation thesis focuses on the mixture toxicity of oestrogenic EDs and their occurrence in...
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Development of LC-MS method for determination of selected endocrine disruptors
Chvalkovská, Eliška ; Kučera, Radim (advisor) ; Štěrbová, Petra (referee)
Charles University Phaculty od Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Eliška Chvalkovská Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Consultant: Mgr. Martin Novák Title of thesis: Development of LC-MS method for determination of selected endocrine disruptors The thesis is focused on the development and optimization of LC-MS method for the determination of twelve endocrine disruptors in amniotic fluid of pregnant women in the third trimestr of pregnancy. There is a suspision that premature births are connected with an increased incidence of endocrine disruptors in amniotic fluid. The analytes were extracted by liquid-liqud extraction with 4-methylpentan-2- on. The analytical method is based on using reverse phase high-performance liquid chromatography with Hypersil C4 GOLD column (2.1 × 100 mm, particles 1.9 µm). For analysis the gradient elution with the mobile phase A (0.5% (v/v) formic acid aqueous solution) and B (0.5% (v/v) formic acid solution in acetonitrile) at a flow rate of 0.5 ml/min, injection 5 µl and column oven temperature 35 řC was used. The high- resolution mass spectrometer Orbitrap Q Exative Plus with atmospheric pressure chemical ionization in positive mode was used to identify and determine the response of...
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Development of LC-MS method for determination of selected endocrine disruptors
Chvalkovská, Eliška ; Kučera, Radim (advisor) ; Štěrbová, Petra (referee)
Charles University Phaculty od Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Eliška Chvalkovská Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Consultant: Mgr. Martin Novák Title of thesis: Development of LC-MS method for determination of selected endocrine disruptors The thesis is focused on the development and optimization of LC-MS method for the determination of twelve endocrine disruptors in amniotic fluid of pregnant women in the third trimestr of pregnancy. There is a suspision that premature births are connected with an increased incidence of endocrine disruptors in amniotic fluid. The analytes were extracted by liquid-liqud extraction with 4-methylpentan-2- on. The analytical method is based on using reverse phase high-performance liquid chromatography with Hypersil C4 GOLD column (2.1 × 100 mm, particles 1.9 µm). For analysis the gradient elution with the mobile phase A (0.5% (v/v) formic acid aqueous solution) and B (0.5% (v/v) formic acid solution in acetonitrile) at a flow rate of 0.5 ml/min, injection 5 µl and column oven temperature 35 řC was used. The high- resolution mass spectrometer Orbitrap Q Exative Plus with atmospheric pressure chemical ionization in positive mode was used to identify and determine the response of...
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PPCP in water and their influence on water ecosystems
Bláhová, Marie ; Fuksa, Josef (advisor) ; Matoušková, Milada (referee)
The aim of the bachelor thesis is to identify the main representatives of a wide group of Pharmaceuticals and personal care products (PPCP for short) and to explain the reasons why they pose an increasing risk to aquatic ecosystems. In the theoretical part, the work is devoted to defining the concept of PPCP and explaining the main reasons why their concentration in the aquatic environment has been growing steadily in recent decades. It also deals with the division into the main groups of preparations found in the aquatic environment, their characteristics, documented impacts on aquatic organisms and currently detected concentrations in the world's rivers. In the following chapters, the thesis addresses the issue of the effectiveness of the removal of these substances and the mechanisms by which they can be improved. The work briefly summarizes the effects on the organisms caused by both bioaccumulation and endocrine disruption and at the end of the theoretical part deals with the properties of ten selected drugs, which are regularly detected in Czech rivers. The practical part evaluates the volume of sales of ten drugs in the Czech Republic during 2019, their detected concentrations in the rivers Vltava, Elbe, Morava and Dyje and processing these data to assess the risk of concentrations found for...
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Multidisciplinary characterization of interactions between oestrogen hormones and their receptors during sperm capacitation.
Bosáková, Tereza
This doctoral thesis was focused on the multidisciplinary characterizatrion of interactions between hormones and their receptors during sperm capacitation, which is the final step in sperm maturation in mammals and the key event to achieve egg fertilization. Three oestrogenic hormones (17β-estradiol (E2), 17α-estradiol (αE2), 17α-ethynylestradiol (EE2)), which are part of environmental pollution, were selected for the study. These hormones are called endocrine disruptors as they can cause disorders of the hormonal system in animals, thereby adversely affecting physiological processes, including reproduction. A new analytical HPLC-MS/MS method was developed for monitoring of concentration changes of oestrogens (E2, αE2 and EE2) during time-dependent capacitation of mouse and rat sperm in vitro. For individual oestrogens, the concentration dependences of free, unbound hormone on the capacitation time were measured for three concentrations of individual hormones (200, 20 and 2 μg/L). The obtained concentration data were converted into values of relative concentrations, which were subjected to kinetic analysis. Kinetic models were proposed for the interaction of individual hormones with sperm, which were described by a system of differential equations with optimization of rate constants and molar...
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Optimization of endocrine disruptors determination in wastewater treatment plant sludge and application of the method in environmental samples.
Medková, Jaroslava ; Cajthaml, Tomáš (advisor) ; Tesařová, Eva (referee)
Hormonaly active compounds in wastewaters represent nowdays a serious problem. Proceses currently used in watewater treatment plants (WWTP) are unefficient in removing these compounds from contaminated wastewaters. The compounds are supposed to sorb onto solid sludge elements and sediments. In this work seven endocrine disruptors were detected in the sludge samples from WWTPs. A new sensitive method for detection of seven selected endocrine disruptors (4-nonylphenol, bisphenol A, estriol, 17β-estradiol, estrone, 17α- ethynylestradiol, irgasan) was developed. The method is based on accelerated solvent extraction (ASE), gel permeation chromatography (GPC) and solid phased extraction. For final extract analysis, gas chromatography coupled with mass spectrometry (GC/MS) was used. The efficiency of this method was tested using artificially contaminated sludge and the method was used to analyse real samples from several WWTPs in Czech Republic. The effect of sludge age on detection of individual analytes was assessed as well. The concentrations of endocrine disruptors measured in the samples reached up to 1 µg/g. The results are comparable or higher then those reported in other works and they show the necessity of further research on endocrine disruptors in the environment.
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Effect of zearalenone on reproductive parameters and the expression of selected genes in mice.
Dvořáková, Eva ; Pěknicová, Jana (advisor) ; Nedvídek, Josef (referee)
A number of chemicals may have a negative impact on the environment and wildlife. Endocrine disruptors (EDs), which can mimic estrogen, interfere with natural hormones in organism and can have a negative effect on the reproductive system. Such substances include zearalenone (ZEA) - mycotoxin, produced by the fungi Fusarium. Despite the non-steriodal structure of its molecule, ZEA and its derivates possess potent estrogenic activity. The influence of ZEA on reproductive parameters and changes in expression of selected genes were tested in the outbred line of mice. This study showed significant effects of ZEA on number of reproductive performances. This effect was observed at lower examined dose, to which are humans normally exposed, and at higher doses, both showed changes in the tested parameters.
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New flame retardants in the environment
Ezechiáš, Martin ; Cajthaml, Tomáš (advisor) ; Kočí, Vladimír (referee)
In the recent years, many research articles focused on endocrine disrupting compounds in the environment. Some of these compounds are listed in a group named brominated flame retardants. However, only few articles investigated endocrine activity of several "new" brominated flame retardants. These chemicals such as 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE) or bis(2-ethylhexyl) tetrabromophthalate (TBPH) are newly used due to ban of some previously most produced brominated flame retardant mixes. In this study, we used two recombinant yeast screens to measure estrogenic, androgenic, antiestrognic and antiandrogenic activities of some alternative brominated flame retardants. We also used ligninolytic fungi to investigate biodegradation of these compounds. Our results suggest, that 2,4,6-tribromophenol (TBP) may be a new environmental endocrine disruptor. This substance showed antiestrogenic and antiandrogenic activity in our tests. 1,2-bis(2,4,6- tribromophenoxy)ethane (BTBPE) had certain antagonistic activity too. In the biodegradation experiment, only three compounds showed significant degradation during the test period. No biodegradation have been observed for other compounds. In this study, we applied gas chromatography with mass spectrometry to analyze these "new" flame retardants. Method for gas...
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Analysis of biologically active compounds using high performance separation methods
Adamusová, Hana ; Bosáková, Zuzana (advisor) ; Jelínek, Ivan (referee) ; Tůma, Petr (referee)
In the first part of this doctoral thesis, a new analytical HPLC-MS/MS method for monitoring of concentration changes of 17β-estradiol (βE2) during in vitro mouse sperm capacitation was developed. Capacitation was performed for three initial concentrations of βE2 (200, 20 and 2 μg/L). For all the concentrations a similar trend for the total unbound βE2 was observed. In general, the βE2 concentration decreased to reach its minimum and then increased again. The position of the minimum differed for the individual tested βE2 concentrations. Experimentally obtained results were subjected to the kinetic analysis. The curves fitted through the experimentally determined points displayed an autocatalytic character. For the agreement between the curves obtained by fitting through the experimental points and the theoretical calculated curves, it is necessary to assume that the first step is adsorption of βE2 onto the surface of the sperm controlled by Langmuir isotherm. The kinetic study was also used to study the effects of fluorides and aluminium fluoride complexes on the capacitation of mouse sperm. The experimental points were in very good agreement with the shape of the theoretical curves and this fact verifies the mechanism of the mouse sperm capacitation kinetics. In the second part of this work, two...
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