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V doktorski disertaciji smo se osredotočili na identifikacijo potencialnih hormonskih motilcev fenolnih analogov bisfenola A (BPA) iz skupin zdravil, onesnažil in kozmetičnih sestavin, na njihovo pleiotropno delovanje ter preučili hormonske učinke njihovih mešanic. Najprej smo preverili zastavljeno hipotezo doktorske disertacije ali je modulacija hormonskega sistema odvisna od kemijske strukture. Med fenolnimi analogi BPA smo identificirali prepoznavne strukturne elemente odgovorne za interakcijo z jedrnimi receptorji.Z metodo in silico smo s pomočjo prosto dostopnega programskega paketa Endocrine Disruptome smo napovedali afiniteto vezave z 12 različnimi jedrnimi receptorji za izbran niz spojin. Predvsem smo se osredotočili na bromirana zaviralca gorenja TBB in TBPH, njunametabolita (TBBA in TBMEPH) ter aminofenolne analoge BPA (diklofenak (DIC), 4- hidroksidiklofenak (4-HD), paracetamol (PAR) in piceatanol (PIC)). Testirali smo jih in vitro na celičnih linijah s poročevalskim genom za luciferazo: MDA-kb2 celični liniji, ki ima izražen glukokortikoidni (GR) kot tudi androgeni receptor (AR), in GH3.TRE-Luc celični liniji z izraženima tiroidnima receptorjema α in β (ΤRα in TRβ). Za spojine, ki so izkazovale glukokortikoidno in androgeno moduliranje, smo izvedli še potrditveni test vezave in vitro. Med prvimi smo za prej omenjene spojine dokazali njihove različne hormonske učinke na estrogenskem (ER), androgenem, glukokortikoidnem in tiroidnem receptorju pri koncentracijah, katerim smo ljudje realno lahko izpostavljeni. Ugotovili smo, da metabolizem neposredno vpliva na modulacijo androgenega, glukokortikoidnega in tiroidnega receptorja. Za modulacijo glukokortikoidnega receptorja so pomembni: prosta karboksilna skupina in 2-(etilheksil)-oksikarbonilna skupina vezani na aromatski obroč ter prisotnost treh halogenih atomov v orto položaju. Rezultati so pokazali, da je antiandrogena aktivnost odvisna od števila karbonilnih skupin v molekuli in stilbenske strukture brez dodatnih stranskih verig med benzenskima skupinama. Ugotovili smo, da afiniteto vezave na androgeni receptor izkazujejo spojine, ki imajo v svoji strukturi Ph-N-C=O fragment, OH in NH-C=O skupini ter aromatski obroč. Na splošno, spojine, ki so bromirane na položaju 3 in 5 na fenilnem obroču glede na osnovni skelet, imajo antitiroidni učinek. Opazili smo, da je Ph-N-Ph fragment, ki vsebuje še atom Cl na enem od fenilnih obročev, prisoten pri spojinah z mešanim agonističnim/antagonističnim delovanjem na androgeni receptor. Ugotovili smo tudi, da hormonski motilci androgenega, glukokortikoidnega, tiroidnega in estrogenega receptorja v svoji strukturi vsebujejo fenolne skupine kot tudi OH skupino v para položaju glede na –R ( v našem primeru R je –NH-Ph fragment, -NH-C=O fragment, -C=CH-Ph fragment) vezane na fenolni obroč ter da so te prav tako odgovorne za hormonsko aktivnost omenjenih spojin. V drugem delu doktorske disertacije smo potrdili hipotezi, da je hormonski učinek proučevanih spojin na zgoraj omenjene receptorje odvisen od hkratne prisotnosti drugih snovi in da imajo fenolni analogi BPA pleiotropni učinek. Poleg modulacije jedrnih receptorjev smo se osredotočili tudi na njihovo genotoksično in imunomodulatorno delovanje. Izvedli smo teste in vitro na LCL celični liniji za aminofenolne analoge BPA v smislu določanja vpliva izbranih spojin na produkcijo citokinov. Rezultati na LCL celični liniji kažejo, da imajo določeni aminofenolni analogi BPA imunomodulatorno delovanje pri koncentracijah, najdenih v človeški plazmi ter da na njega znatno vpliva metabolizem teh spojin. Dokazali smo, da lahko predvsem fenolni analogi BPA kot hormonski motilci glukokortikoidnega receptorja spremenijo imunski odziv izpostavljenih posameznikov. S pomočjo kvantno-kemijskih metod smo prvi predstavili mehanizem direktne poškodbe DNA toksičnega metabolita paracetamola (NAPQI). Paracetamol lahko uvrstimo med aminofenolne analoge BPA (tako kot diklofenak (DIC), 4-hidroksidiklofenak (4-HD) in piceatanol (PIC)). Ugotovili smo, da izkazuje aminofenolni analog BPA paracetamol preko svojega metabolita NAPQI nizko verjetnost genotoksičnosti, ki je možna le v primeru izčrpanih zalog glutationa v celici ter da so kinoniminske strukture manj reaktivne v primerjavi s kinonskimi. Predstavili smo podroben pregled vseh najpogosteje uporabljenih spojin, ki vsebujejo kinonsko in kinoniminsko strukturo ali vsebujejo to skupino njihovi metaboliti, ter posledice izpostavitve tem spojinam. Učinke mešanic izbranih spojin iz skupin onesnažil (propilparaben (PP), butilparaben (BP), dietilheksil ftalat (DEHP) in tetrametrin (TM)) in kozmetičnih sestavin (2-metilrezorcinol (2MR), avobenzon (AVB) in butil-hidroksianizol (BHA)) smo določili na celičnih linijah na enak način kot hormonske učinke posameznih spojin. Testiranja učinka mešanic smo se lotili na dva načina: proučevali smo mešanice izbranih spojin pri koncentracijah izpostavitve človeka (1 μM in 10 nM) ter pri koncentracijah, v katerih spojine izkazujejo 50 % spodbujevalni ali zaviralni učinek na proučevane receptorje (EC50 ali IC50). Glede na dobljene rezultate smo ugotovili, da testirane komponente mešanic interagirajo ena z drugo. Pokazali smo, da učinki mešanic niso zanemarljivi ter da jih je nujno potrebno vklopiti v oceno tveganja hormonskih motilcev na zdravje ljudi in njihovo okolje. Ti rezultati kažejo, da je potrebno natančno določiti strategijo za testiranje mešanic spojin, ki bo temeljila na ustreznih toksikoloških testih (že obstoječe in silico in in vitro metode, novi biološki testi ter Euromix modelna orodja za testiranje mešanic). In doctoral thesis we focused on the identification of new potential endocrine disrupting compounds, their pleiotropic effects and examined hormonal mixture effects thereof. As target group of tested compounds we chose phenolic analogues of bisphenol A (BPA) from the classes of drugs, pollutants and cosmetic ingredients. At first, we verify the hypothesis of the doctoral thesis whether the modulation of hormonal system depends on the chemical structure of the compounds. Among the phenolic analogues of BPA we identified recognizable structural elements responsible for the interaction with nuclear receptors. Using the freely accessible program package, Endocrine Disruptome, we predicted binding affinity to 12 different nuclear receptors for the selected compounds in silico. In particular, we focused on brominated flame retardants (TBB and TBPH), their metabolites (TBBA and TBMEPH) and aminophenol analogues of BPA (diclofenac (DIC), 4-hydroxydiclofenac (4- HD), paracetamol (PAR) and piceatannol (PIC)). Next, we tested them in vitro on two cell lines with luciferase reporting gene: on the MDA-kb2 cell line, which expresses the glucocorticoid (GR) and the androgen receptor (AR) and the GH3.TRE-Luc cell line expressing thyroid hormone receptors α and β (TRα and TRβ). For compounds which showed modulation of the glucocorticoid and androgen receptor, we additionaly performed an in vitro confirmatory test for binding. Among the first we have proved their different hormonal effects on estrogen (ER) and earlier mentioned receptors at concentrations, which represent real exposure levels on humans. We have found that the metabolism directly affects the modulation of the androgen, glucocorticoid and thyroid hormone receptors. For the modulation of the glucocorticoid receptor, the following functional groups are important: the free -COOH group and 2-(ethylhexyl)-oxycarbonyl group attached to the aromatic ring and the presence of three halogen atoms in the ortho position. The results showed that the antiandrogen activity depends on the number of carbonyl groups in the molecule and stilbenic scaffold without additional side chains between two benzene rings. We have found that the binding affinity for the androgen receptor depends on the presence Ph-N-C=O fragment, OH and NH-C=O groups, and the aromatic ring in their chemical structure. Also, the compounds with the brominated 3,5-substituent in the phenyl group possess antithyroid hormone effect. It has been observed, that the Ph-N-Ph fragment, that contains the Cl atom on one of the phenyl rings, is present in the compounds with mixed agonist/antagonist androgen activity. We have found that the endocrine disruptors of the androgen, glucocorticoid, thyroid hormone and estrogen receptors in their structures contain phenolic groups and also OH groups in para position with respect to the –R (in this case, R is –NH-Ph fragment, -NH-C=O fragment, - C=CH-Ph fragment) attached to a phenolic ring. In the second part of the doctoral thesis, we confirm the hypothesis, that the modulation of the above-mentioned receptors induced by the compounds depends on the presence of other compounds, and that the phenolic analogues of BPA have pleiotropic effect. In addition, we also focus on their genotoxic and immunomodulatory activities. We also performed in vitro assay on the LCL cell line in order to determine the immunomodulatory effect of the aminophenol analogues of BPA. Results obtained using the LCL cell line indicate that certain aminophenol analogues of BPA have immunomodulatory activity at concentrations found in human blood. We proved that the phenolic analogues of BPA as glucocorticoid receptor mimics of the natural ligand, may be associated with altered immune responsiveness. Making use of the available quantum-chemical methods, we first outline the mechanism of direct damage to the DNA by the toxic metabolite of the aminophenol analogue of BPA (NAPQI). Paracetamol can be classified as an aminophenol analogue of BPA (another are diclofenac (DIC), 4-hydroxydiclofenac (4-HD) and piceatannol (PIC)). We have found that the aminophenol analogue of BPA (PAR) possesses low genotoxicity, which is possible only in the case of the depleted glutathione, and that quinone imines are less reactive when compared to quinones. We presented a detailed overview of most commonly used compounds or their metabolites containing quinone and quinone imine structure as well as the consequences of exposure to these compounds. The effects of mixtures of compounds selected from the groups of pollutants (propylparaben (PP), butylparaben (BP), diethylhexyl phthalate (DEHP) and tetramethrin (TM)), and cosmetic ingredients (2-methylresorcinol (2MR), avobenzone (AVB) and butylated hydroxyanisole (BHA)) was determined in vitro in the same way as the hormonal effects of the individual compounds, explained above. We tested the mixture effects in two different ways: (i) selected compounds were mixed at concentrations to which humans are exposed (1 μM and 10 nM), (ii) at their EC50 or IC50 concentrations. Based on the obtained results, we found that the components of tested mixtures interact with each other. We pointed out, that the effects of mixtures are not negligible and that it is necessary to include them into the risk assessment of compounds. Also, it is necessary to specify a strategy for testing mixtures of compounds, which will be based on the relevant toxicological tests (already existing in silico and in vitro methods, new biological tests and Euromix modeling tools for testing mixtures).

Endocrine disrupting chemicals such as natural and synthetic steroid hormones and bisphenols are among the most important pollutants in the aquatic environment. We performed an environmental chemical analysis of five Slovenian water samples, two rivers, one groundwater, and the influent and effluent of wastewater treatment plants, with a highly sensitive analysis of twenty-five endocrine-disrupting compounds belonging to the groups of natural hormones, synthetic hormones, and bisphenols. Since these compounds are simultaneously present in the environment, it is important to study their individual effects as well as the effects of mixtures. We investigated in vitro the estrogenic potency of selected natural and synthetic steroid hormones and bisphenols detected in surface, ground and waste water in Slovenia using the OECD-validated transactivation assay on the cell line Hela9903. We predicted their mixture effects using the concentration addition model and compared them with experimentally determined values. Two mixing designs were used: a balanced design in which chemicals were combined in proportion to their individual EC$_{50}$ values, and an unbalanced design with compounds in proportion to their measured concentrations in the environmental samples. The estrogenic effects of the experimental mixtures followed the concentration addition model. Real water samples exhibited weaker estrogenic effects, showing the great heterogeneity of the real water samples.

Palladium was immobilized on a highly porous copolymer of 4-vinylpyridine and divinylbenzene (polyHIPE-poly(high internal phase emulsion)) using palladium(II) acetate to obtain PolyPy-Pd with 6.1 wt % or 0.57 mmol Pd/g. The immobilized catalyst was able to catalyze the coupling of iodobenzene and phenylboronic acid in ethylene glycol monomethyl ether/water (3:1) within 4 h at rt and complete conversion was observed when 2.5 mol % of Pd per PhI was used. The reaction tolerated a wide range of substituents on the aromatic ring. Iodobenzene derivatives with electron-withdrawing substituents showed higher reactivity, while the opposite was true for the phenylboronic acid series. The polyHIPE-supported Pd catalyst was also used for the direct conversion of phenylboronic acid to biphenyl through an iodination/coupling reaction sequence. The recyclability of the heterogeneous catalyst was also optimized, and by finding a suitable combination of solvents for the loading of Pd, the reaction, and the isolation of the product, the solid-supported catalyst was completely regenerated and used in the next reaction with the same activity.

The volumetric properties of asphalt mixtures are important parameters for evaluating the performance of asphalt. Volumetric properties (grading requirements, air void content, minimum and maximum voids filled with bitumen and voids in mineral aggregate) are usually presented first in specifications. The visualisation of volumetric properties is therefore important for the understanding of the composition of an asphalt mix. A triangular representation of requirements for the volumetric properties of asphalt mixtures is therefore a valuable method to illustrate the volumetric requirements. Such representation gives a better insight into the requirements and helps on decisions when authorities decide about these requirements.

Preko uživanja hrane in vode, vdihovanja plinov in dotikanja stvari v okolici v naše telo nenehno prehajajo številne sintezne in naravne spojine. Nekatere izmed teh lahko vplivajo na delovanje našega endokrinega sistema in s tem povzročajo različne neugodne zdravstvene izide. Ena takih spojin je bisfenol A, ki pa se zaradi številnih odmevnih študij, ki so poročale o njegovih škodljivih učinkih na zdravje, v zadnjem desetletju delno nadomešča z drugimi bisfenolnimi analogi. Posledica tega je, da smo pogosto izpostavljeni mešanicam različnih bisfenolov in še številnim drugim spojinam. V toksikologiji se tako daje vedno večji poudarek na vrednotenje mešanic, saj le-to ponuja natančnejši prikaz toksičnosti, kot tradicionalni način posamičnega testiranja spojin. Cilji našega dela so bili testiranje estrogene aktivnosti posameznih bisfenolov A, AF, C, F, S in Z na estrogenski receptor α (ERα), vrednotenje estrogenih učinkov dvokomponentnih mešanic bisfenola A z drugimi bisfenoli, vrednotenje estrogenega delovanja mešanice vseh šestih testiranih bisfenolov in ocena estrogene aktivnosti produktov bisfenolov A, AF in S po prestali 1. ali 2. fazi metabolizma. Za izvedbo poskusov smo uporabili hERα-HeLa-9903 celično linijo in meritve izvajali s pomočjo luciferaznega testa. Pred pričetkom testiranj smo s kontrolnimi spojinami preverili pravilno delovanje celične linije in opravili test z resazurinom za preverjanje morebitne citotoksičnosti spojin. Za citotoksično se je izkazala le 25 μM mešanica vseh šestih opazovanih bisfenolov, ki smo jo zato iz analize izpustili. Vseh šest testiranih bisfenolov se je po kriterijih OECD 455 protokola izkazalo za agoniste ERα. Jakost njihovega estrogenega delovanja je padala po sledečem vrstnem redu: bisfenol AF, Z, C, A, F, S. Iz meritev jakosti estrogenega delovanja posameznih bisfenolov smo izračunali aditivni model, katerega rezultate smo primerjali z meritvami jakosti estrogenega delovanja mešanic bisfenolov. Vrednosti so se med seboj zelo dobro ujemale, iz česar sklepamo, da imajo mešanice bisfenolov aditiven način součinkovanja na ERα. V testih metabolizma bisfenolov se je izkazalo, da sta najpomembnejša mehanizma metabolne detoksikacije bisfenolov encimski reakciji glukuronidacije in sulfatiranja. V primeru oksidativnega metabolizma bisfenolov, je bilo znižanje estrogene aktivnosti relativno nizko. Vzrok je lahko počasnejša kinetika reakcij ali pa nastanek estrogensko aktivnih produktov. Za določitev tega bi bile potrebne dodatne raziskave. Through the consumption of food and water, the inhalation of air, and the physical contact with objects in the environment, many synthetic and natural compounds are constantly passing into our bodies. Some of these can affect the functioning of our endocrine system and, thus, lead to various adverse health effects. One such compound is bisphenol A, which has been partially replaced by other bisphenol analogs in the past decade due to the numerous attention-attracting studies that reported its adverse health effects. As a result, we are often exposed to mixtures of various bisphenols and many other toxins. Therefore, there is an increasing emphasis in toxicology on the assessment of mixtures, since it offers a more accurate evaluation of toxicity as the individual testing of compounds. The objectives of our work were to test the estrogenic activity of bisphenols A, AF, C, F, S, and Z on the estrogen receptor α (ERα), to evaluate the estrogenic effects of two-component mixtures of bisphenol A with other bisphenols and to determine the estrogenic activity of the mixture of all six bisphenols. Additionally, we evaluated estrogenic activities of the products of bisphenols A, AF, and S after phase 1 or 2 metabolisms. To perform the assays, the hERα-HeLa-9903 cell line was used and measurements were made using luciferase assay. Before testing bisphenols, four control compounds were used to test the proper cell line functioning and a resazurin assay was performed to check for potential cytotoxicity of the tested bisphenols. Only a 25 μM mixture of all six bisphenols proved cytotoxic and was, therefore, omitted from the analysis. All six of the tested bisphenols proved to be ERα agonists. The potency of their estrogenic activity decreased in the following order: bisphenol AF, Z, C, A, F, and S. From the measurements of the estrogenic potency of the individually tested bisphenols, the concentration addition model was calculated. The obtained results were compared to the measured estrogenic potency of the mixtures of bisphenols. The values matched very well, suggesting that the mixtures of bisphenols have an additive mode of action on the ERα. The bisphenol metabolism tests have shown that the most important mechanisms of metabolic detoxification of bisphenols are the enzymatic reactions of glucuronidation and sulfation. In the case of oxidative metabolism of bisphenols, however, the decrease of estrogenic activity was relatively low. This may be due to slower kinetics of the reactions or the formation of estrogen-active products.

Cilj diplomske naloge je bil izdelati in preveriti delovanje sistema senzorjev, s pomočjo katerih bi lahko dolgoročno merili spreminjanje temperature in relativne vlažnosti v kompozitih z mineralnim vezivom. V sodelovanju s Fakulteto za elektrotehniko Univerze v Ljubljani, smo izdelali dokaj preprost merilni sistem, ki je sestavljen iz 8 senzorjev za merjenje temperature in relativne vlažnosti, sistema za sprejem in obdelavo signalov in sistema za napajanje ter prenos podatkov. V prvem delu naloge smo sestavili senzorje in preverili ali delujejo pravilno. Odločili smo se za uporabo senzorskih sond proizvajalca Sensirion, poimenovanih SHT 25. Gre za senzorje z digitalnim I2C vmesnikom, ki ponujajo dolgoročno stabilnost, komercialno dostopnost in sprejemljivo mersko napako. Okolje, v katero bodo senzorji vgrajeni, je zelo agresivno, zato so zaščiteni s posebnim pokrovom filtra in epoksi premazom. Vrednosti se beležijo in shranjujejo na računalniški enoti Raspberry Pi3. Odčitavanje in zapisovanje senzorskih podatkov deluje na podlagi treh napisanih skript, ki se izvršijo vsaki 2 minuti. Vzporedno uporabljamo komercialno poznan merili sistem Ahlborn ALMEMO, s pomočjo katerega na koncu preverjamo verodostojnost podatkov pridobljenih z našim sistemom senzorjev. V drugem delu smo pripravili recepture treh različnih mešanic in sicer : čista apnena malta, podaljšana cementna malta in lahki samozgoščevalni beton. Predhodno smo za namen preiskave pripravili tudi 8 opečnih zidakov. Mešanice smo zamešali po standardnem postopku in vanje vgradili senzorje. Pridobljene vrednosti temperature in relativne vlažnosti obeh merilnih sistemov smo analizirali in med seboj primerjali. Podrobneje smo obravnavali proces hidratacije in z njo povezano teoretično ozadje. S primerjanjem pridobljenih rezultatov meritev je bilo ugotovljeno, da izdelan senzorski sistem kljub zadovoljivemu delovanju potrebuje nekaj izboljšav. Primerjava rezultatov temperature je pokazala dobro delovanje sistema, večja odstopanja in slabše delovanje pa opazimo pri primerjavi rezultatov relativne vlažnosti. S pomočjo preiskave je bilo ugotovljeno, kakšne spremembe in izboljšave potrebujemo oz. bi želeli v procesu razvoja SHT merilnega sistema. The purpose of the diploma thesis was to design and test the functioning of a sensor system, which could be used for long term measurements of the temperature and relative humidity in mineral binder composites. In a cooperation with the Faculty of Electrical Engineering University of Ljubljana, we have developed a fairly simple measuring system consisting of 8 sensors for measuring temperature and relative humidity, a system for receiving and processing signals, a power supply system and data transmission system. In the first part of the thesis we assembled the sensors and checked if they are working properly. We decided to use Sensirion sensor probes, named SHT 25. These sensors are using a digital I2C interface and are offering long-term stability, commercial accessibility and acceptable measurement errors. The environment in which the sensors will be integrated is very aggressive, so sensors have been protected with a special filter cover and epoxy coating all over the sensors. All the values sensors recorded were stored on a Raspberry Pi3 computer unit. All the sensor data readings and recordings works on the basis of three written programs, which are executed every 2 minutes. To compare all the received values with SHT sensor system we used commercially well known sensor system Ahlborn ALMEMO. In the second part of the thesis we prepared three different mixtures: pure lime mortar, cement – lime mortar and lightweight self – compacting concrete. We also prepared 8 brick units. All the mixtures were mixed by standard procedure and the sensors were installed inside them. All the obtained values of temperature and relative humidity of both sensor system were analysed and compared with each other. We discussed the hydration process and its theoretical background in more details. By comparing all the obtained results we found that despite satisfying performance, the sensor system needs some improvements. Worse performance were observed when comparing relative humidity results. Within investigation we identified which changes and improvements are needed or can be done within the measurement system development.

V trajnih nasadih smo pri zatiranju plevelov zelo odvisni od uporabe herbicida glifosat. za boljšo prihodnost pa želimo to spremeniti in poskušamo z alternativno možnostjo zatiranja, kot je setev rastlin in vzdrževanje trajne rastlinske združbe, ki preprečuje razvoj plevelov. Tako smo naredili kratkotrajen poskus, v katerem smo raziskali možnosti preprečevanja rasti plevelov. When combatting weeds in permanent crops we have to rely on the use of glyphosate herbicides. This can be changed by implementing alternative methods of combatting weeds. One alternative is sowing of specific plants and maintaining a permanent community of plants to protect the pears against the yield reduction caused by weeds. For a practical test of this weed control method, an experiment was carried out in a pear plantation.