Bisphenol A (BPA) is one of the highest volume chemicals produced worldwide and, as a result, is universally found in environmental and human matrixes. Bisphenol A is a known endocrine disruptor that acts as an estrogen agonist and an androgen antagonist. Due to health concerns, BPA is being phased out and replaced by other bisphenol analogues structurally similar to BPA. To date, there have been little to no studies showing the effects of BP analogues on the reproductive organ development of male mice. Thus, this study aimed to compare the effects of BPA and selected analogues (including BPB, BPE, and BPS) on the reproductive organ development in male mice, and determine preliminary toxicity threshold levels, such as the lowest-observed-effect-dose (LOED) and no-observed-effect-dose (NOED). Exposure to BPA, BPB and BPE via subcutaneous injection at a dose of 10 μg/g body weight (bw)/day each significantly caused a decrease in anogenital distance and glans penis length in male mice. Testis weight was also significantly reduced by BPA and BPE. Although BPS did not cause an effect on the glans penis length, anogenital distance or testis weight, histology work indicated that the spines on the glans penis were at a different developmental stage than the control. A similar result was seen with BPA on the glans penis spines. The LOED and NOED of BPA affecting anogenital distance, penis length, or testis weight were determined to be 10 and 5 μg/g bw/day, respectively. These LOED and NOED values are preliminary for BPA, because only five dose levels are used. Further research is needed to estimate more accurate threshold levels for the studied endpoints for BPA as well as other bisphenol analogues. The results indicated that some bisphenol analogues (BPB and BPE) showed comparable effects to BPA on the reproductive organ development of male mice, including anogenital distance and penis length. This could be indicative of more severe reproductive issues later in life and raised a concern on the safety of using these analogues to replace BPA in consumer products. More research is needed to investigate the mechanisms of the observed effects on genetic or molecular levels, determine what the long-term adverse effects of bisphenol analogues are to the reproductive system of male mice, and determine whether similar effects will be seen at dose levels comparable to human exposure rates.

Bisphenol A (BPA) is a compound that is commonly used in the manufacture of epoxy resins and plastics. Because of large scale production and widespread usages, BPA is released into the atmosphere through air, land, and water. BPA is weakly estrogenic in animals and has acute aquatic toxicity even at low concentrations of 1- 10μg/L. Ibuprofen is a widely used analgesic and antipyretic. Ibuprofen and its metabolites are mainly released into the environment by human urinary excretion. Ibuprofen has been detected at low concentrations in surface and waste waters. The environmental and health effects at such concentrations are unclear. The high removal of BPA and ibuprofen in the wastewater treatment plants (WWTPs), suggest that biodegradation might be responsible for the removal of these compounds. Several bacterial strains, isolated from waste water, are known to degrade BPA and ibuprofen. No studies, however, have reported using ammonia oxidizing bacteria for this purpose. Ammonia oxidizing bacteria (AOB) are an important group of microorganisms in nitrifying activated sludge of WWTPs. AOB are known to express ammonia monooxygenase (AMO) to degrade many different aromatic and aliphatic organics via cometobolic degradation (non beneficial mechanism). Nitrosomonas europaea is a widely studied AOB found to degrade synthetic estrogen by a study. This study aims to characterize the biodegradation of BPA and ibuprofen by AOB. The biodegradation ability of N.europaea with respect to BPA and ibuprofen was examined. Experiments were conducted in the presence/absence of the AMO inhibitor (allylthiourea), an external reducing energy source (sodium formate) and different primary substrate (ammonia) concentrations. The second part of the study comprises of biodegradation tests on BPA and ibuprofen using activated sludge from two WWTPs, one with one-sludge activated sludge system and the other one with two-sludge nitrification system. From the experiments conducted BPA at a concentration of 1.6 mg/L was degraded to 0.12 mg/L by N.europaea. BPA at concentrations of 1.0 mg/L and 0.75 mg/L was completely degraded by the cells. Resting cells of N.europaea were, however, unable to degrade BPA. Also ibuprofen of two concentrations, 0.42 mg/L and 0.8 mg/L, were not degraded by the culture. BPA at a concentration of 1 mg/L was degraded to 0.2 mg/L and ibuprofen at 0.5 mg/L was completely degraded by the activated sludge from the combined reactor. The activated sludge from the nitrification tank degraded BPA of concentration 1 mg/L and ibuprofen of concentration 0.5 mg/L completely. Hence, it can be summarized that Bisphenol A was degraded by N.europaea and also by the activated sludge obtained from the WWTPs. Ibuprofen was found incapable of inhibiting ammonia oxidizing bacteria in the case of the pure culture while it was successfully degraded by the mixed culture.

Zugl.: Dresden, Techn. Universiẗat, Diss., 2004.

Bisphenol (BP) A is a common manufacturing chemical in polycarbonate plastics and has been widely used in plastics, epoxy resin liners of canned foods, dental materials, and thermal receipts. Human exposure to BPA is associated with a negative impact on human health including the development and function of the reproductive system due to its action as an endocrine-disrupting chemical (EDC). Numerous experimental studies have demonstrated that BPA impairs both male and female reproductive function, despite the variation in study paradigms such as dose, exposure route, timing, and outcomes measured. Due to the toxicological effects of BPA, BPA analogues such as BPS have been used as alternatives for BPA. However, recent evidence has suggested these BPA analogues can induce similar or even more severe toxic effects as BPA, and health risks of exposure to replacement bisphenols need to be considered. Therefore, my study was designed to examine whether prenatal exposure to BPE and BPS negatively impacts on male and female reproductive function in mice. Pregnant females were orally administrated corn oil (control), BPA, BPE, and BPS (0.5, 20, or 50mg/kg/day) from gestational day 11 (the presence of vaginal plug=1) to birth, and reproductive tissues in F1 mice were collected and analyzed in both neonatal and adult mice. In males, I observed reduced sperm counts and quality, disrupted stages of spermatogenesis in adults and increased germ cell apoptosis in neonatal testis following prenatal BPA, BPE or BPS exposure. Particularly, I found the expression of methyltransferases for DNA methylation and histone modification was also affected by prenatal exposure to BPA, BPE, or BPS in neonatal testis, suggesting a potential of epigenetic alterations in F1 males. In females, prenatal exposure to BPE and BPS accelerated the onset of puberty, disrupted estrous cyclicity, and caused several fertility problems especially in aged mice. In the neonatal ovaries, I also observed that BPE and BPS inhibit germ cell nest breakdown comparable to BPA. These results suggest that prenatal exposure to BPE and BPS with physiologically relevant doses affects male and female reproductive function probably due to germ cell development defects in the developing gonads. Finally, to understand their complete impact on male and female fertility, a study of transgenerational effects of BPE and BPS is performed to examine the transgenerational effects of prenatal exposure to BPA, BPE and BPS on reproductive function in F3 offspring. To be called transgenerational, expression of the specific phenotype will be continued at least across three generations. As described in previous studies, the direct exposure of a pregnant female (F0 generation) results in the exposure of the embryos (F1 generation) and the germline that will generate the next generation (F2 generation). Thus, I orally exposed to control treatment (corn oil), BPA, BPE or BPS (0.5 or 50 μg/kg/day) from gestational day 7 to birth in pregnant females (F0). Mice from F1 and F2 offspring were used to generate the F3 generation. In F3 males, prenatal exposure to BPA, BPE, and BPS induces persistence and even more severe phenotypes in sperm counts and motility in the F3 generation than in the F1 offspring. The expression of DNA and histone methyltransferases were transgenerationally increased by BPA, BPE and BPS exposure in both neonatal and adult testis. In F3 females, prenatal exposure to BPA, BPE, and BPS accelerated the onset of puberty and exhibited abnormal estrous cyclicity, and those females exhibited similar fertility problems as those in the F1 generation. However, BPA, BPE and BPS exposure did not affect neonatal follicular development such as germ cell nest breakdown or follicle numbers in the ovary on postnatal day 4. Taken together, our results suggest that prenatal exposure to BPA analogues, BPE and BPS, have transgenerational effects on male and female reproductive function in mice. Our findings suggest the hypothesis that transgenerational epigenetic alterations in germ cells may lead to reproductive disorders/dysfunction in the F3 generation.

Estrogens maintain the appropriate androgen-estrogen balance for normal regulation of the structure and function of the male reproductive tract, including the prostate gland. This research investigated viability of cells and expression of selected genes in prostate carcinoma cells (PC-3) exposed to bisphenol A (BPA), an estrogen-like substance present in a number of plastic materials. PC-3 cells are able to metabolize BPA at concentrations below 100 µM. BPA exposure at concentrations between 1nM and 100 µM does not increase or significantly reduce cell viability of these cells. Although the genes investigated in this study (GSTP1 and MGMT) did not show a significant change in expression following in vitro exposure to BPA, the positive control ethinyl estradiol (EE2) caused an increase in GSTP1 expression at mRNA level. These results indicate that BPA does not affect the viability of prostate cells, and motivate a need for further research to identify other genes that could be affected by BPA.

Estrogens maintain the appropriate androgen-estrogen balance for normal regulation of the structure and function of the male reproductive tract, including the prostate gland. This research investigated viability of cells and expression of selected genes in prostate carcinoma cells (PC-3) exposed to bisphenol A (BPA), an estrogen-like substance present in a number of plastic materials. PC-3 cells are able to metabolize BPA at concentrations below 100 µM. BPA exposure at concentrations between 1nM and 100 µM does not increase or significantly reduce cell viability of these cells. Although the genes investigated in this study (GSTP1 and MGMT) did not show a significant change in expression following in vitro exposure to BPA, the positive control ethinyl estradiol (EE2) caused an increase in GSTP1 expression at mRNA level. These results indicate that BPA does not affect the viability of prostate cells, and motivate a need for further research to identify other genes that could be affected by BPA.

Würzburg, Univ., Diss., 2008 / Zsfassung in dt. Sprache

Thesis -- Departmental honors in Psychobiology. / Spine title: Effects of bisphenol A on adult female ovariectomized rats. Bibliography: ℓ. 46-55.

Zugl.: Greifswald, Universiẗat, Diss., 2009.

Since recent years, pollution of surface and drinking waters by chemical pollutants fromanthropogenic activities, in particular micropollutants, has become an important environmentalissue in industrialized countries. Indeed the majority of these molecules are toxic toliving organisms, even present at very low concentrations (pg.L−1 to μg.L−1). Some of themare referred as endocrine disrupting compounds. The existing conventional wastewater treatmentsare most of the time not adapted for the removal of these compounds. Among them,advanced oxidative processes give the better removal efficiencies, but with drawbacks such ashigh energetic costs, or the formation of reaction products, sometimes more toxic than theparent compounds. A promising solution for water decontamination could be bioremediation.This work is based on this technology, via the study of the use of lignolytic enzymes, such aslaccases, for the removal of micropollutants. Degradation studies of a lot of micropollutants(bisphenol A, nonylphenol, triclosan, 17-_-estradiol, etc.) have already been investigatedwith different reactor configurations, using free (solubilized) or immobilized enzymes (biocatalysts).However, a decrease in the performance of some continuous long-term degradationprocesses has been highlighted, most of the cases because of biocatalysts deactivation. Oneof the reasons to explain this deactivation was identified as being induced by the deposit ofreaction products onto the surface of the biocatalysts.The general objective of this work was to investigate the degradation of bisphenol A(BPA) by laccases oxidation in different reactor configurations. Especially, the focus wasmade on the study of BPA degradation and reaction products kinetics, and reaction productscharacterization and identification. In the continuity, a second objective was related tothe optimization of a packed-bed reactor developed in our laboratory, through the developmentof a new biocatalyst, and assessment of biocatalyst deactivation.First, a deep literature review has been done in order to summarize the knowledge regardingsoluble and insoluble reaction products formation during laccase oxidation of a listof the most studied recalcitrant phenolic compounds, and their risk assessment. This reviewreported that the majority of reaction products were insoluble, and corresponded to smalloligomers associated by C-C or C-O bonds. In most of the cases, the assessment of theirstructure identification, and evaluation of their toxicity and/or estrogenic activity was missing.Secondly, a research for the selection of a new suitable support for laccase immobilizationhas been investigated. Celite R648 supports were selected for the immobilization of laccasesfrom Trametes versicolor, based on physical, catalytic and economic properties. Characterizationof the packed-bed reactor developed at the TIPs was performed with this support,through evaluation of pressure drop and residence time distributions. This new biocatalystwas selected as a tool for further studies regarding BPA degradation and reaction productsformation kinetics after laccase oxidation.The third part of this work consisted in the kinetic study of BPA degradation, and solubleand insoluble reaction products formation. For BPA degradation kinetic study, experimentaldata were obtained by LC/MS analyses for different initial conditions and reactor configurations.A mathematical model was build, based on a modified Briggs-Haldane model, in orderto identify the limiting step(s) occurring during laccase-catalyzed oxidation of BPA in batchreactor with free enzymes. This model was validated on experimental data, and highlightedthat the limiting step during BPA degradation was the formation of the enzyme-substratecomplex. For the kinetic study of insoluble reaction products, a protocol was developed enablingthe monitoring of these products over time, through their mass, in batch reactor withfree enzymes, and for seven different experimental conditions according to a Doehlert design.A mathematical model was build, expressed by a first-order kinetics, and fitted experimentaldate from 30 minutes of reaction. The modeling of short reaction times was achieved with theuse of the model build to characterize BPA degradation kinetics, with the assumption thatall the BPA degraded was converted into insoluble reaction products. Combination of resultsfrom these two kinetic studies highlighted that insoluble reaction products formation waslimited by BPA conversion, limited itself by the formation of the enzyme-substrate complex.The study of soluble reaction products kinetics was only experimental, and showed majorkinetics differences in batch reactor depending on the use of free enzymes or biocatalysts.Mass transfer limitations could occur in the case of immobilized enzymes.In parallel, experiments have been done in order to characterize and/or identify reactionproducts. DSC, GPC and FTIR analyses were performed for the characterization of insolublereaction products, and LC/MS(-MS) analyses for the characterization of soluble reactionproducts. Results highlighted that our products were in majority polymers, and that theircomposition was varying depending on initial BPA concentration and reaction time.All the information gathered in this work were used and exploited in order to characterizebiocatalysts deactivation, and to propose solutions to optimize the packed-bed reactor andrecover biocatalysts activity. In parallel, we have developed a tool to monitor biocatalystsdeactivation, with the purpose of increasing the degradation performance of the reactor. / Depuis plusieurs années, la pollution des eaux potable et de surface par des polluants chimiquesprovenant d’activités anthropogéniques, en particulier les micropolluants, est devenueune problématique environnementale importante dans les pays industrialisés. En effet, la majoritéde ces substances sont toxiques pour les organismes vivants, même présentes en trèsfaibles concentrations (de l’ordre du pg.L−1 au μg.L−1). Un grand nombre de ces substancessont considérées comme des perturbateurs endocriniens. Les techniques conventionnelles actuellesde traitement des eaux usées ne sont la plupart du temps pas adaptées à l’abattementde ces substances. Parmi ces techniques, les procédés d’oxydation avancée sont ceux quidonnent les meilleurs rendements d’abattement. Cependant, ils présentent certains inconvénients,incluant des coûts énergétiques importants, et la formation de produits de réactionparfois plus toxiques que les molécules de départ. La bioremédiation pourrait constituer unesolution prometteuse. Ce travail est basé sur cette idée, au travers de l’étude de l’utilisationd’enzymes lignolytiques, comme les laccases, pour la dégradation des micropolluants. Desétudes sur la dégradation de nombreux micropolluants (bisphénol A, nonylphénol, triclosan,17-_-estradiol, etc.) ont déjà été menées pour différentes configurations de réacteurs, avecl’usage d’enzymes libres (solubilisées) ou immobilisées (biocatalyseurs). Cependant, à longterme, une diminution des performances de dégradation lors de certains procédés continusa été mise en évidence, la plupart du temps à cause d’une désactivation des biocatalyseurs.Une raison possible pouvant expliquer cette désactivation réside dans le dépôt de produits deréaction sur la surface des biocatalyseurs.L’objectif général de ce travail a été d’étudier la dégradation d’un micropolluant, le bisphénolA (BPA), via oxydation par les laccases, dans différentes configurations de réacteurs.Plus précisément, nous nous sommes intéressés à l’étude des cinétiques de dégradation dubisphénol A et de formation de ses produits de réaction, ainsi que leur caractérisation etindentification. Dans la continuité, un second objectif a consisté en l’étude de l’optimisationd’un réacteur à lit fixe développé dans notre laboratoire, au travers du développement d’unnouveau biocatalyseur, et l’évaluation de la désactivation enzymatique de ces biocatalyseurs.Premièrement, une revue détaillée de la littérature a été effectuée dans le but de résumerles connaissances acquises à propos de la formation des produits de réaction solubles etinsolubles après oxydation par les laccases des composés phénoliques récalcitrants les plusétudiés actuellement, et l’évaluation de leur toxicité. Cette revue a mis en évidence que lamajorité des produits de réaction sont insolubles, et correspondent à de petits oligomères dontles unités constitutives sont associées via des liaisons C-C ou C-O. Dans la plupart des cas,l’identification de leur structure, et l’évaluation de leur toxicité et/ou activité oestrogéniqueest manquante.Dans un second temps, une étude pour la sélection d’un nouveau support pour l’immobilisationde laccases a été menée. Le support Celite R648 a été sélectionné pour l’immobilisationde laccases de Trametes versicolor, sur bases de propriétés physico-chimiques, catalytiques etéconomiques. La caractérisation du réacteur à lit fixe a été réalisée avec ce support, au traversde l’évaluation des pertes de charges et des distributions de temps de séjour. Le nouveaubiocatalyseur développé a finalement été sélectionné comme outil pour l’étude des cinétiquesde dégradation du BPA, et de formation des produits de réaction, après oxydation par leslaccases.La troisième partie de ce travail a consisté en les études cinétiques de la dégradation du BPA, et de la formation des produits solubles et insolubles de réaction. Pour l’étude de la cinétiquede dégradation du BPA, des données expérimentales ont été obtenues via des analysesde LC/MS, pour différentes conditions expérimentales et pour différentes configurations deréacteurs. Un modèle mathématique a été construit, basé sur un modèle de Briggs-Haldanemodifié, dans le but d’identifier la(les) étape(s) limitante(s) se produisant durant l’oxydationdu BPA par les laccases, en réacteur batch et avec enzymes libres. Ce modèle a été validésur les données expérimentales, mettant en évidence que l’étape limitante dans la réactionde dégradation du BPA était la formation du complexe enzyme-substrat. Pour l’étude de lacinétique de formation des produits de réaction insolubles, un protocole a été développé pourle suivi de ces produits au cours du temps, au travers de leur masse, en réacteur batch etavec enzymes libres, dans sept conditions expérimentales différentes, selon le schéma d’undesign expérimental de Doehlert. Un modèle mathématique a été construit, exprimé commeune cinétique de premier ordre, et a été validé sur les données expérimentales à partir de 30minutes de réaction. La modélisation sur des temps de réactions inférieurs à 30 minutes a étéobtenue en utilisant le modèle construit pour la cinétique de dégradation du BPA, en supposantque tout le BPA dégradé était directement converti en produits de réaction insolubles.L’étude combinée des résultats obtenus a mis en évidence que la formation des produits deréaction insolubles était limitée par la dégradation du BPA, elle-même limitée par la formationdu complexe enzyme-substrat. L’étude cinétique de la formation de produits solubles deréaction a uniquement été expérimentale, et a permis de mettre en évidence des différencesmajeures dans les cinétiques de formation en réacteur batch, selon l’usage d’enzymes libresou de biocatalyseurs.En parallèle, des expériences ont été réalisées dans le but de caractériser et/ou d’identifierles produits de réaction. Des expériences de DSC, GPC et FTIR ont été faites pour la caractérisationdes produits insolubles de réaction, et des expériences de LC/MS(-MS) ont étéréalisées pour la caractérisation des produits solubles de réaction. Les résultats ont permisde mettre en évidence que les produits obtenus sont en majorité des petits oligomères, etque leurs compositions varient en fonction de la concentration initiale en BPA et le temps deréaction.Les informations récoltées dans ce travail ont été utilisées et exploitées pour tenter decaractériser la désactivation des biocatalyseurs, et de proposer des solutions pour optimiserle réacteur à lit fixe et récupérer l’activité initiale des biocatalyseurs. En parallèle, nousavons également développé une méthode pour détecter et suivre la désactivation, dans le butd’améliorer les performances de dégradation du réacteur. / Doctorat en Sciences de l'ingénieur et technologie / info:eu-repo/semantics/nonPublished