The thesis focuses on the use of passive sampling during an analysis of hormones in drinkable water. In the theoretical part there are described selected passive samplers and the sample of the type of POCIS is described at fuller length. The next chapter engages with steroid hormones and an input of estrogenic hormone into the environment. The thesis contains chapters dealing with possibilities of the determination of hormones in water. It is specialized on the analysis of drinkable and surface water. In the last chapter of the theoretical part there are introduced water works where samples for an experimental assessment were collected from. The experimental part of the thesis presents the specification of the extraction of analytes and an adjustment of used device. In the thesis there are specific calibrations for individual assessment of hormones. The results of the assessment for each collection point are ordered to a well-arranged table. Increased levels of targeted estrogens were not approved in most of sampling points.

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...

<p>Sintetisana su nova jedinjenja, 16- i 17-supstituisani&nbsp;16,17-sekoestratrienski derivati i D-homoestranski&nbsp;derivati, polazeći od 3-benziloksi-17-hidroksi-16,17-sekoestra-1,3,5(10)-trien-16-nitrila. Ispitana je&nbsp;estrogena i antiestrogena aktivnost u eksperimentima <em>in&nbsp;vivo</em>, antiaromatazna aktivnost <em>in vitro</em>, antioksidantna&nbsp;aktivnost DPPH &nbsp;i TBA testom, kao i antiproliferativna&nbsp;aktivnost prema ćelijskim linijama MCF-7 ATCC,&nbsp;MDA-MB-231, HT-29 i MRC-5 novosintetisanih&nbsp;jedinjenja.</p> / <p>Some new compounds, 16- and 17-substituted 16,17-secoestratriene derivatives, as&nbsp; well as D-homoestratriene derivatives, were synthesized, starting from 3-benzyloxy-17-hydroxy-16,17-secoestra-1,3,5 (10)-triene-16-nitrile. The newly synthesized compounds were tested for their <em>in vivo&nbsp;</em>estrogenic and antiestrogenic activity,&nbsp; <em>in vitro&nbsp;</em>antiaromatase activity, antioxidative activity by DPPH and TBA tests, as well as antiproliferative activity against MCF-7 ATCC, MDA-MB-231, HT-29 i MRC-5 cell lines.</p>

(Oberflächen-) Gewässer weltweit sind mit geringen Mengen (ng/L bis wenige µg/L) humaner Pharmazeutika belastet. Diclofenac (DCF; nicht-steroidal, entzündungshemmend) und Metoprolol (MTP; ß-Blocker) gehören entsprechend ihres hohen Verbrauchs zu den am häufigsten gefundenen Substanzen. Deren biologische Aktivität ist nicht auf den Menschen beschränkt. Gut konservierte Enzyme innerhalb der Vertebraten legen Auswirkungen auf Nicht-Zielorganismen wie Fische nahe, die bisher in Langzeituntersuchungen mit umweltrelevanten Konzentrationen unzureichend untersucht wurden. In der vorliegenden Arbeit wurden die physiologischen Effekte von DCF und MTP auf die Nil-Tilapie (Oreochromis niloticus), einem der wichtigsten Aquakulturfische weltweit, untersucht. In vitro konnte anhand primärer Hepatozyten gezeigt werden, dass bereits umweltrelevante Konzentrationen von DCF zu einer erhöhten Genexpression verschiedener Schlüsselenzyme der Detoxifizierung führten. Nach MTP-Exposition waren die Veränderungen weniger eindeutig. Beide Substanzen induzierten die Vitellogenin Genexpression, nur DCF jedoch bereits in umweltrelevanter Konzentration. In vivo wurden in zwei Langzeit-Expositionsversuchen die physiologischen Effekte vom befruchteten Ei bis 80 Tage nach Schlupf in O. niloticus untersucht. Beide Substanzen hatte keinen Einfluss auf Schlupferfolg und Überleben, das Wachstum war nach 80 Tagen nach Schlupf leicht reduziert. Die deutlichsten Auswirkungen waren histopathologische Veränderungen der Kiemen, veränderte Genexpressionen der Gonadotropine und eine erhöhte Expression von Vitellogenin. Die Ergebnisse legen eine stärkere östrogene Aktivität von DCF im Vergleich zu MTP nahe. Zusammenfassend sind die Bedenken gegenüber den Einzelsubstanzen eher gering, negative Auswirkungen auf die Reproduktion und sich verstärkende Effekte bei zeitgleicher Exposition gegenüber DCF und MTP lassen sich jedoch nicht ausschließen und sollten im Weiteren untersucht werden. / Surface waters worldwide are contaminated with low levels (ng/L up to few µg/L) of human pharmaceuticals. Diclofenac (DCF; non-steroidal, anti-inflammatory) and metoprolol (MTP; ß-blocker) are highly consumed and therefore commonly detected. Their biological activity is not restricted to humans. Well conserved enzymes within the vertebrates suggest effects on non-target organisms such as fish, poorly studied in long-term exposure experiments using environmentally relevant concentrations. In the presented work, physiological effects of DCF and MTP on the Nile tilapia (Oreochromis niloticus), an important aquaculture fish species, were studied. Using primary hepatocytes, it was shown in vitro that environmentally relevant concentrations of DCF increased the gene expression of different key enzymes of the detoxification, while MTP exposure had a less clear effect. Both substances induced vitellogenin gene expression, but only after DCF exposure this was significantly elevated already at the environmentally relevant concentration. In vivo, two long-term exposure studies on the physiological effects from the fertilized egg until 80 days post-hatch were evaluated. Both substances did not affect hatching success and survival, while growth was slightly reduced after 80 days post-hatch. Histopathological alterations of the gills, changed gene expression patterns of the gonadotropins and induced vitellogenin gene expression were the most dominant findings. The results indicate a stronger estrogenic mode of action of DCF compared to MTP. Overall, the risk due to a single substance exposure seems to be relatively low but adverse effects on reproduction and additive effects during simultaneous exposure to DCF and MTP cannot be excluded and should be investigated further.

No description available.

<p>Hemijska karakterizacija ekstrakata&nbsp; <em>T. pratense&nbsp;</em>L.&nbsp; određena je LC-MS-MS, a etarskih ulja GC-MS analizom, pri čemu je&nbsp; identifikovano vi&scaron;e&nbsp;bioaktivnih jedinjenja.&nbsp; Spektrofotometrijskim&nbsp;metodama ispitan je antioksidantni potencijal&nbsp;ekstrakata i etarskih ulja. Hepatoprotektivni&nbsp;efekat određen je&nbsp; na&nbsp; homogenatu i hemolizatu&nbsp;jetre laboratorijskih mi&scaron;eva.&nbsp; Budući da&nbsp; ova&nbsp;biljka ima visok sadržaj izoflavona, određena je&nbsp;<br />estrogena, antiestrogena aktivnost, kao i&nbsp;citotoksičnost ekstrakata. Antimikrobna&nbsp;<br />aktivnosta ekstrakata i etarskih ulja ispitana je&nbsp;na&nbsp; &scaron;est bakterijskih sojeva.&nbsp; Ekstrakti i etarska&nbsp;ulja su bili umereno aktivni u pogledu biolo&scaron;kih&nbsp;aktivnosti, osim u slučaju antimikrobne&nbsp;aktivnosti koju nije pokazao nijedan ispitivani&nbsp;uzorak.</p> / <p>The chemical characterization of&nbsp; <em>T.&nbsp; pratense</em> extracts was determined by LC-MS-MS, and the essential oils of the GC-MS analysis, wherein the&nbsp; various&nbsp; bioactive compounds were identified.&nbsp; Antioxidant potential of extracts and essential oils was tested&nbsp; by spectrophotometric methods.&nbsp; Hepatoprotective&nbsp; effect&nbsp; was determined&nbsp; in&nbsp; haemolysate and liver homogenate of&nbsp; laboratory&nbsp; mice.&nbsp; Estrogen, antiestrogen activity, as well&nbsp; as the cytotoxicity of the extracts were&nbsp; determined&nbsp; due to the high&nbsp;content of isoflavones.&nbsp; Antimicrobial activity of extracts and essential oils was&nbsp; investigatied&nbsp; on six bacterial strains. Essential oils and extracts were moderately active in terms of biological activity, except&nbsp; in the case of&nbsp;&nbsp; antimicrobial activity&nbsp; where none of the extracts were active against tested bacterial strains.</p>

MSc (Microbiology) / Department of Microbiology / Background: The occurrence of microorganisms and endocrine disrupting chemicals (EDCs) in water poses a serious concern due to their effects on humans, animals and environment. In recent years, EDCs have been increasingly reported in rivers that receive large amounts of wastewater effluents. Of all the EDCs, natural and synthetic hormones are among those that are recognized for their potential to mimic or interfere with normal hormonal functions of humans and animals. The present study aimed at assessing the occurrence of these hormones in relation to the molecular diversity of Aeromonas and evaluating the resistance of Aeromonas to antibiotics as well as to assess anti-bacterial activity of two selected traditional medicinal plants. Methods: Wastewater, water and fish samples were collected from various sources (rivers, wastewater treatment plants, taps, and dams) for the detection of hormones and isolation of Aeromonas species. The analysis of hormones from various organs of the fish and from water samples was conducted, after extraction using enzymelinked immunosorbent assays (ELISA). Different types of hormones including Estriol, Estradiol, Ethinylesradiol and Testosterone were detected, and their concentrations determined. Aeromonas spp were isolated rom the samples using microbiological methods and Conventional PCR was used for genotyping as well as for detection of the beta-lactamase genes. Kirby-bauer method was used to determine the susceptibility profiles of Aeromonas to different antibiotics. Microdilution assay was used to determine the Anti-bacterial activity of the plant (Annoniceae and Zornia milneana) extracts against Aeromonas species. Results: A total of 144 samples were collected from 23 different locations in two countries: South Africa and Zambia. These included wastewater and treated wastewater, River water, fish and tap water. 17α-ethinylestradiol (EE2) was detected in most of the samples (92.7%) with concentrations varying from 0.59 ng/ml to 65 ng/ml. The hormones were also detected from drinking water, with testosterone detected at high concentrations of up to 140 ng/ml in tap water. Most sewage treatment plants were not able to remove the EE2 from the wastewater as the concentration of this hormone in the final effluent was almost always higher than that in the influent. These homones were also detected in drinking water at high concentrations of up to 53.49 ng/ml in the tap water for EE2 and 1777 ng/ml for E2. The overall detection of Aeromonas species in the samples was 84.5%. A. caviae was the most prevalent species accounting for 73.6%, followed by A. veronii with 64.6%. The bacteria were completely resistant to cefuroxime accounting for 100% resistance. Aeromonas isolates also showed high resistance to trimethroprim (88.7% for A. hydrophila), cefazolin (highest 97.8% for A. cavie), and ceftazidime (83.9% for A. sobria). TEM was the most prevalent beta-lactamase gene with detection rate of 87%. All isolates lacked the presence of the CTX-M3 gene. Also, wastewater had the highest prevalence of A. veronni and A. caviae accounting for 87.5% and 82.5% respectively. Multiple antibiotic resistance was also observed with the Aeromonas isolates being resistant to up to 11 antibiotics. High prevalence of 77.1% of Aeromonas hydrophila was observed in the presence of ethinylestradiol (EE2). Aeromonas veronii and Aeromonas caviae were the most predominant species in the presence of total estriol, A. veronii had a prevalence of 57.1% and A. caviae had a prevalence of 52.8%. Aeromonas hydrophila and Aeromonas caviae had the lower prevalence in the presence of hormones with the percentages of 26.1% and 27.8% respectively. The methanol extracts of both Zornia milneana and Annona species showed good activity against the Aeromonas spp with the lowest MIC of 0.078 mg/ml. Ethyl acetate extracts were the least effective. Conclusion: This study has shown high occurrence of steroid hormones in all types of environmental samples tested. These included tap water, river water, wastewater and fish both in Zambia and South Africa. Therefore, steroid hormones constitute and important health problem in the Southern African Sub-Region. The incapacity of the wastewater treatment plants to remove EE2 is an important problem that needs to be tackled immediately. The prevalence of Aeromonas species is very high in our environmental water as well as in drinking water, with the highest prevalence observed in fish and wastewater. It was also revealed that there is relationship between steroid hormones and Aeromonas species, with the hormones supporting the growth of Aeromonas species. The presence of beta-lactamase genes which causes Aeromonas to be resistant to antibiotics was also noted. Methanol extracts of Zornia milneana and Annona spp were the most effective against Aeromonas spp and could serve as primary sources for the isolation of lead compounds. / NRF