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L'objectiu principal a l'inici de la tesi era la creació de mètodes analítics per a l'anàlisi de forma ràpida exacta i amb la major precisió, així com aconseguir límits de detecció suficientment baixos per poder determinar certs compostos i poder extreure conclusions. Per a ell es va seleccionar una llista d'analitos disruptors endocrinos i compostos relacionats (EDCs) de diferents famílies de compostos, però representatius en relació a estrogeinitat, en matrius acútiques. Finalment, s'ha pogut estudiar bastants dels compostos que a priori es pretenien, consiguiendo paràmetres de qualitat acceptables. Aquests mètodes es podrien aplicar en 2 campanyes de mostres de les principals cuencas de la Península Ibèrica, podent per tant extraer bastant informació. L'aigua és un bé cada cop més escaso i imprescindible per a la vida, en les últimes dècades la societat s'ha començat a ser conscient d'ell, sembla que l'activitat humana té un gran impacte sobre el medi ambient i en conseqüència també en el medi aquàtic. . Los estudios medioambientales, por tanto, están cobrados cada vez más relevancia, siendo imprescindibles para la evaluación de la calidad de este bien tan apreciado. Hoy en dia, i degut en part a la creciente regulació de les diferents agències mundials en termes de qualitat de l'aigua, existeixen molts estudis, enfocats a l'avaluació de diferents famílies de contaminants que poden causar cert impacte en el mitjà aquàtic. Un exemple d'ell són els compostos disruptors endocrinos (EDCs), un grup de substàncies exògenes que interfereixen amb el sistema endocrino i interrompen la funció fisiològica de les hormones. Alguns d'aquests contaminants, amb diferents estructures i propietats, es troben en una gran varietat de productes comunament utilitzats a la vida diària i, en conseqüència, es detecten en el medi ambient aquàtic. El estudio de EDCs s'ha enfocat al llarg de les darreres dècades des de diferents angles, aquests contaminants han estat estudiats per exemple com a part d'un grup principal de substàncies que no només tenen rellevància pels efectes endocrins d'alguns dels seus compostos, sinó que presenten altres característiques nocives, graus de toxicitat elevada, persistència i bioacumulació, com és el cas dels pesticides, productes farmacèutics, compostos químics halogenats, entre altres. Aquesta tesi s'ha centrat només en l'avaluació de diferents grups de contaminants des del punt de vista del seu més o menys grau de disrupció endocrina, així com d'altres compostos relacionats, aquests, han estat escollits segons criteris de grau de disrupció i/o per la seva alta producció o ús, essent aquest últim factor important ja que contaminants amb poc potencial disruptor, ja per quantitat o exposició constant poden arribar a ser igualment perjudicials. Las metodologías analíticas disponibles en la literatura para el análisis de EDCs en el medio ambiente acuático son muy variadas, aunque la mayoría de estos métodos cubren solo un o dos grupos de compuestos que contienen polaridades similares, estructuras similares o actividades similares, cada vez que existen más autors que proposen mètodes multicompuestos per a la determinació d'una àmplia gamma d'EDCs, encara que alguns d'aquests, usen diferents mètodes d'extracció i/o maneres d'elució, durant l'anàlisi, el que resulta tediós i lento. Utilitzant la tecnologia d'injecció directa mitjançant una columna de preconcentració o bé una purificació seguida d'una separació cromatogràfica líquida acoplada a l'espectrometría de masses en tàndem (EQuan/TFC-LC-MS/MS), optimitzades en aquesta tesi, es va solventar una part. de les limitacions exposicions anteriorment, creant mètodes en línia automatitzats per a l'anàlisi dels diferents grups d'EDCs en matrius agudes, sediments i lodos. Aquestes metodologies es poden aplicar de forma ràpida i eficaç per a la determinació del nivell de contaminació dels EDC Per a realitzar tota aquesta investigació s'ha comptat amb l'ajuda de grans experts en la matèria, de diferents universitats y centres de recerca El objetivo principal al inicio de la tesis era la creación de métodos analíticos para el análisis de forma rápida exacta y con la mayor precisión, así como logrando límites de detección suficientemente bajos para poder determinar ciertos compuestos y poder extraer conclusiones. Para ello se escogieron una lista de analitos disruptores endocrinos y compuestos relacionados (EDCs) de distintas familias de compuestos, pero representativos en relación a estrogeinicidad, en matrices acuáticas. Finalmente, se pudieron estudiar bastantes de los compuestos que a priori se pretendía, consiguiendo paramametros de calidad aceptables. Estos métodos se pudieron aplicar en 2 campañas de muestro de las principales cuencas de la Península Ibérica, pudiendo por tanto extraer bastante información. El agua es un bien cada vez más escaso e imprescindible para la vida, en las últimas décadas la sociedad parece haber empezado a ser consciente de ello, dado que la actividad humana tiene un gran impacto sobre el medioambiente y en consecuencia también en el medio acuático. Los estudios medioambientales, por tanto, están cobrado cada vez más relevancia, siendo imprescindibles para la evaluación de la calidad de este bien tan preciado. Hoy en día, y debido en parte a la creciente regulación de las distintas agencias mundiales en términos de calidad del agua, existen muchos estudios, enfocados a la evaluación de distintas familias de contaminantes que pueden causar cierto impacto en el medio acuático. Un ejemplo de ello son los compuestos disruptores endocrinos (EDCs), un grupo de sustancias exógenas que interfieren con el sistema endocrino e interrumpen la función fisiológica de las hormonas. Algunos de estos contaminantes, con diferentes estructuras y propiedades, se encuentran en una gran variedad de productos comúnmente utilizados en la vida diaria y, en consecuencia, se detectan en el medio ambiente acuático. El estudio de EDCs se ha enfocado a lo largo de las últimas décadas desde distintos ángulos, estos contaminantes han sido estudiados por ejemplo como parte de un grupo principal de substancias que no solo tienen relevancia por los efectos endocrinos de algunos de sus compuestos, sino que presentan otras características nocivas, grados de toxicidad elevados, persistencia y bioacumulación, como es el caso de los pesticidas, productos farmacéuticos, compuestos químicos halogenados, entre otros. Esta tesis se ha centrado solamente en la evaluación de distintos grupos de contaminantes desde el punto de vista de su más o menos grado de disrupción endocrina, así como de otros compuestos relacionados, estos, han sido escogidos según criterios de grado de disrupción y/o por su alta producción o uso, siendo este último factor importante ya que contaminantes con poco potencial disruptor, ya sea por cantidad o exposición constante pueden llegar a ser igualmente perjudiciales. Las metodologías analíticas disponibles en la literatura para el análisis de EDCs en el medio ambiente acuático son muy variadas, aunque la mayoría de estos métodos cubren solo uno o dos grupos de compuestos que contienen polaridades similares, estructuras similares o actividades similares, cada vez existen más autores que proponen métodos multicompuestos para la determinación de una amplia gama de EDCs, aunque algunos de estos, usan distintos métodos de extracción y/o modos de elución, durante el análisis, lo que resulta tedioso y lento. Utilizando la tecnología de inyección directa mediante una columna de preconcentración o bien una de purificación seguida de una separación cromatográfica líquida acoplada a la espectrometría de masas en tándem (EQuan/TFC-LC-MS/MS), optimizadas en esta tesis, se solventaron parte de las limitaciones expuestas anteriormente, creando métodos online automatizados para el análisis de los distintos grupos de EDCs en matrices acuosas, sedimentos y lodos. Estas metodologías se pudieron aplicar de forma rápida y eficaz para la determinación del nivel de contaminación de EDCs The main objective at the beginning of the thesis was the creation of analytical methods for the analysis of fast and exact form and with the greatest precision, as well as to achieve sufficiently low detection limits to be able to determine certain compounds and to be able to draw conclusions. For him, a list of endocrine disrupting analytes and related compounds (EDCs) from different families of compounds, but representative in relation to estrogenicity, was selected in acute matrices. Finally, it has been possible to study quite a few of the compounds that were intended a priori, achieving acceptable quality parameters. These methods could be applied in 2 sample campaigns of the main basins of the Iberian Peninsula, therefore being able to extract a lot of information. Water is an increasingly scarce and essential commodity for life, in recent decades society has begun to be aware of it, it seems that human activity has a great impact on the environment and in consequence also in the aquatic environment. . Environmental studies, therefore, are gaining more and more importance, being essential for the evaluation of the quality of this much-appreciated asset. Nowadays, and due in part to the increasing regulation of the different world agencies in terms of water quality, there are many studies, focused on the evaluation of different families of pollutants that can cause a certain impact on the aquatic environment. An example of it is the endocrine disrupting compounds (EDCs), a group of exogenous substances that interfere with the endocrine system and interrupt the physiological function of hormones. Some of these pollutants, with different structures and properties, are found in a wide variety of products commonly used in daily life and, consequently, are detected in the aquatic environment. The study of EDCs has been focused over the last decades from different angles, these pollutants have been studied for example as part of a main group of substances that are not only relevant for the endocrine effects of some of their compounds , but have other harmful characteristics, high levels of toxicity, persistence and bioaccumulation, as is the case with pesticides, pharmaceutical products, halogenated chemical compounds, among others. This thesis has focused only on the evaluation of different groups of pollutants from the point of view of their greater or lesser degree of endocrine disruption, as well as other related compounds, these have been chosen according to criteria of degree of disruption and/or because of its high production or use, the latter being an important factor since pollutants with little disruptive potential, due to quantity or constant exposure, can become equally harmful. The analytical methodologies available in the literature for the analysis of EDCs in the aquatic environment are very varied, although most of these methods cover only one or two groups of compounds that contain similar polarities, similar structures or similar activities, whenever they exist more authors who propose multi-compound methods for the determination of a wide range of EDCs, although some of these, use different extraction methods and/or ways of elution, during the analysis, which is tedious and slow. Using direct injection technology through a preconcentration column or a purification followed by a liquid chromatographic separation coupled to tandem mass spectrometry (EKuan/TFC-LC-MS/MS), optimized in this thesis, solved a part. of the limitations previously exposed, creating automated online methods for the analysis of the different groups of EDCs in sharp matrices, sediments and sludges. These methodologies can be applied quickly and effectively for the determination of the level of EDC contamination. To carry out all this research, we have had the help of great experts in the field, from different universities and research centers internationals and nationals.

The presence of xenobiotics - such as pharmaceutically active compounds, endocrine disrupting compounds and pesticides - in the aquatic environment has risen great environmental concern due to their high toxicity even at low concentrations. These so-called micropollutants have been detected in all aquatic compartments and are continuously entering the environment through via point and non-point sources. In particular, the removal of xenobiotics by means of wastewater treatment plants (WWTPs) is known to be incomplete for several compounds, and more efforts are needed in this context. To tackle this problem, the European Union is updating its legislation. The Directive 2013/39/EU set environmental quality standards in freshwater for 76 priority substances, mainly heavy metals, traditional pesticides and industrial chemicals. With regards of the contaminants of emerging concern, 17 candidates for inclusion in the priority substances list have been included in a Watch list (EU Decision 2015/495). These compounds must be monitored by all member states to collect data on their occurrence in freshwater. Risk assessment will be performed as a decision-making tool to determine whether they pose a risk to or via the aquatic environment. This thesis aims at filling knowledge gaps on micropollutants and in particular on the Watch list compounds at three levels simultaneously: analytical, monitoring and removal possibilities. La presència de xenobiòtics, com ara productes farmacèutics, disruptors endocrins i pesticides, en el medi ambient aquàtic ha engendrat una gran preocupació ambiental degut a la seva elevada toxicitat, fins i tot a baixes concentracions. Aquests compostos, anomenats microcontaminants, s'han detectat en tots els compartiments aquàtics i entren al medi de manera continuada a través de fonts puntuals i no puntuals. En particular, se sap que l'eliminació de xenobiòtics en les plantes de tractament d'aigües residuals (EDAR) és incompleta i calen més esforços en aquest context. Per a solucionar aquest problema, la Unió Europea està actualitzant la seva legislació. La Directiva 2013/39 / UE va establir normes de qualitat ambiental en aigua dolça per a 76 substàncies prioritàries, principalment metalls pesants, pesticides tradicionals i productes químics industrials. Pel que fa als contaminants emergents, 17 compostos candidats a ser afegits a la llista de substàncies prioritàries han estat inclosos en una llista de vigilància (Watch list) (Decisió UE 2015/495). Aquests compostos han de ser monitoritzats per tots els estats membres per a recopilar dades sobre la seva presència en aigua dolça. Es durà a terme l’anàlisi de risc com a eina de presa de decisions per determinar si representen un risc per o a través del medi aquàtic. Aquesta tesi té com a objectiu principal omplir els buits de coneixement sobre microcontaminants i, en particular, sobre els compostos de la Watch list a tres nivells: anàlisi, monitorització i eliminació. Programa de Doctorat en Ciència i Tecnologia de l'Aigua

The main objective of this thesis is to evaluate the impact of wastewater discharges on receiving riverine and coastal ecosystems by studying the occurrence and fate of emerging contaminants (ECs). Since there is a myriad of ECs present in the environment, this thesis mostly focused on two main groups of compounds: (i) endocrine disrupting compounds (EDCs), including natural and synthetic hormones, alkylphenols (APs), such as nonylphenol (NP) and octylphenol (OP), and the plasticizer bisphenol A (BPA), and (ii) pharmaceutically active compounds (PhACs). These compounds were selected because they are one of the ECs of major input into the environment, thus being considered as “pseudo-persistent” pollutants, and for their potential deleterious effects to non-target organisms. Besides these target ECs, this thesis also evaluated the occurrence of other groups of contaminants by using high resolution mass spectrometry (HRMS). Finally, an assessment of the environmental risks posed by the identified ECs has been done in order to select the compounds of major ecological concern and those that could be used as relevant markers of wastewater contamination in freshwater and marine ecosystems. To accomplish the thesis objectives, two different analytical approaches have been used: (i) quantitative target analytical methods to determine the two groups of selected ECs (13 target EDCs and 81 PhACs that belong to nineteen different therapeutic groups) using reference analytical standards, and (ii) suspect screening using HRMS and exact mass compound databases (360 ECs) La finalitat principal d'aquesta tesi és l’avaluació de l'impacte de les aigües residuals en els ecosistemes d'aigua dolça i costaners, mitjançant l'estudi de la presència i destí de contaminants emergents (CEs). Atès que hi ha una gran quantitat de CEs presents en el medi ambient, aquesta tesi s'ha centrat principalment en dos grups de contaminants: (i) compostos disruptors endocrins (EDC), que inclouen les hormones naturals i sintètiques, alquilfenols (AP), com el nonilfenol (NP) i el octilfenol (OP), i el plastificant bisfenol A (BPA); ii) fàrmacs (PhACs). Aquests compostos es van seleccionar perquè pertanyen als CEs amb major entrada al medi ambient, de manera que es consideren com a contaminants "pseudo persistents", a banda dels seus possibles efectes adversos per als organismes aquàtics. Apart d'aquests CEs, en aquesta tesi també s’ha estudiat la presència d'altres grups de contaminants mitjançant l'ús d'espectrometria de masses d'alta resolució (HRMS). Finalment, s'ha realitzat una avaluació dels riscos plantejats per la presència dels CEs identificats, amb l'objectiu de seleccionar els compostos de major rellevància ecològica i que es podrien utilitzar com a marcadors de contaminació d'aigües residuals en els ecosistemes d'aigua dolça i marins. Per aconseguir aquests objectius, s'han utilitzat dos enfocaments analítics diferents: (i) l’ús de mètodes analítics target per determinar els dos grups de CEs seleccionats (13 EDCs i 81 PhACs que pertanyen a dinou grups terapèutics diferents), utilitzant estàndards analítics de referència; (ii) suspect screening mitjançant l’ús de HRMS i bases de dades de massa exactes (360 ECs) This thesis was financially supported by the Spanish Ministry of Economy, Industry and Competitiveness (MINECO), the Spanish Agency of Investigation (AEI), the European Regional Development Found (FEDER) and the European Social Fund (FSE) through the projects TRANSFORMCOAST (CGL2014-56530-C4-4-R, AEI/FEDER, UE) and CLEaN-TOUR (CTM2017-85385-C2-1-R, AEI/FEDER, UE). M.Celic acknowledges her doctoral thesis grant FPI, with reference number BES2015-072297 (MINECO/FSE, EU). Partial financial support from the Ministry of Education, Science and Technological Development of the Republic of Serbia, through the project no.172050, is also acknowledged. Catalan Government support is acknowledged via the Economy and Knowledge Department through Consolidated Research Groups (ICRA-ENV 2017 SGR 1124) and Water and Soil Quality Unit (2017 SGR 1404)

Individual hazardous chemicals and substance mixtures with synergistic toxicity ef-fects occur in the dissolved organic matter (DOM) of wastewater and negatively im-pact human health. Yet a large number of chemicals and their treatment by-products in wastewater makes the tracking of individual compounds nearly impossible and de-mands new analytical strategies. The thesis describes the development and evaluation of non-targeted and suspect anal-ysis methods aimed at the transformation of DOM and micro-contaminants of interest during wastewater treatment using liquid chromatography-high-resolution mass spec-trometry (LC-HRMS) data. On one hand, a non-targeted method to track transformations of DOM in a multiphase wastewater treatment using LC-HRMS data was developed. LC-MS signals were ex-tracted, aligned, and had their isotopologues clustered and elemental composition pre-dicted using open license software MZmine 2 in a way that conceptually prioritized the detection of anthropogenic compounds El destino de los microcontaminantes en el tratamiento de aguas residuales ha ido ga-nando atención debido al impacto ecológico y toxicológico de estas sustancias en la salud humana. Adicionalmente a los productos químicos peligrosos individuales, las mezclas de sustancias como las que ocurren en la materia orgánica disuelta (MOD) en las aguas residuales, pueden producir efectos de toxicidad sinérgicos. Sin embargo, el gran número de compuestos naturales y antropogénicos, así como sus productos de transformación de tratamiento (PTs) provocan que el seguimiento de los compuestos individuales sea casi imposible, exigiendo nuevas estrategias para realizar un segui-miento eficaz de transformaciones MOD. Esta tesis describe un método desarrollado y probado para rastrear transformaciones de MOD en aguas residuales, utilizando un análisis no dirigido de cromatografía de líquidos y espectrometría de masas de alta resolución (LC-MS). Potencialmente, el análisis prioriza la detección de compuestos antropogénicos

The main aim of this thesis was to establish a link between the urban origin of chemical contamination (e.g. PhACs) and other stressors, particularly associated to water scarcity (Chapter 1, 2, and 3). Research has been performed in one Alpine (Chapter 1) and two Mediterranean river basins (Chapter 2 and 3). Effects of the river flow variability on the recovery potential of the rivers (natural in-stream attenuation) have been studied in the tributary streams of the lower Ebro River (Chapter 2) and the Evrotas River (Chapter 3). Results have shown that occurrence and spatiotemporal distribution of PhACs in the fragile Alpine and Mediterranean aquatic environments is subjected to a strong intra-annual variability of the stream flow, while effects of multiple stress conditions may be amplified under water scarcity conditions (e.g. drought), thus resulting in the increased concentrations levels of PhACs in river water and sediments L'objectiu principal d'aquesta tesi ha estat establir un vincle entre l'origen urbà de la contaminació química (per exemple, els PhACs) i altres estressors, particularment associats a l'escassetat d'aigua (capítols 1, 2 i 3). Les investigacions s'han realitzat en un riu Alpi (Capítol 1) i dues conques del Mediterrani (Capítol 2 i 3). Els efectes de la variabilitat del flux fluvial sobre el potencial de recuperació dels rius (atenuació natural) han estat estudiats en rius del baix riu Ebre (capítol 2) i del riu Evrotas (capítol 3). Els resultats mostren l'aparició i la distribució espaciotemporal dels PhACs en els fràgils entorns aquàtics alpins i mediterranis, sotmesos a una forta variabilitat intra-anual del flux, mentre que els efectes de múltiples condicions d'estrès poden ser amplificades sota condicions d'escassetat d'aigua, que afavoreixen concentracions més elevades de PhACs en aigües i sediments

[eng] Pharmaceuticals which are used worldwide are designed to facilitate the life for the human society and have an important role in treatment and prevention of disease for both humans and animals. They are ubiquitous in the aquatic environment and are mainly derived from municipal wastewater treatment plants (WWTPs) due to their low removal rate. Therefore, their presence in the environment is directly linked to the human impact. Various biological and abiotic processes in the environment can transform them to transformation products (TPs). In many cases, transformation is already initiated in the human body by a variety of drug-metabolizing enzymes. The metabolites formed through human metabolism present some modifications in their chemical structures that can differ in physicochemical properties to their parent compound. Once they are excreted from the human body, both the unmetabolised parent drug and their metabolites enter WWTPs by means of the sewer system. Since the WWTPs are not designed to remove completely pharmaceutical residues, the fraction not removed after the treatment will eventually end up in the receiving water bodies. Consequently, due to pharmaceutical transformations in the human body, biotransformations in WWTPs and phototransformations in surface water, they can potentially produce a high number of TPs in real world samples which makes their identification a challenge. In this thesis, two different approaches (TPs profiling and suspect screening) based on high resolution mass spectrometry (HRMS) for the detection and identification of TPs of pharmaceuticals were investigated. TPs profiling approach was applied for the identification of phototransformation products of an antiviral zanamivir (ZAN) in batch reactors filled with surface water. On the other hand, suspect screening approach was applied for evaluation of transformation, prioritization and identification of photoTPs of six iodinated contrast media in surface water. Finally, a combination of suspect and TPs profiling approach was applied for the detection of TPs of an anticonvulsant lamotrigine and its main human metabolite lamotrigine N2-glucuronide which were formed as the result of their degradation in both activated sludge and pH dependent hydrolysis. The TPs profiling approach for evaluation of these transformations is illustrated in the example of photodegradation of an antiviral ZAN with identification of its TPs in surface water (Chapter 3.). Here a set of lab- scale experiments was performed in order to determine the susceptibility of ZAN towards photodegradation under simulated and natural sunlight. The identification of the TPs was performed using hydrophilic interaction liquid chromatography coupled to high resolution mass spectrometry (HILIC-HRMS) where four photoTPs were tentatively identified and their proposed structures were rationalized by photolysis mechanisms. Kinetic experiments showed that photodegradation kinetics of ZAN in surface waters would proceed with slow kinetics since upon exposure of aqueous solutions of surface water (20 μg L-1) to simulated sunlight, ZAN was degraded with t1/2 of 3.6 h. Under natural sunlight irradiating surface water, about 30 % of the initial concentration of the antiviral disappeared within 18 days. However, when ZAN and its TPs were retrospectively screened from surface water extracts, neither the parent nor the TPs were detected. The results of this TP profiling used for the identification of TPs of ZAN, although straightforward, suggests that it is not suitable when dealing with a considerably elevated number of TPs formed in batch experiments. However, time and effort needed to be optimised for the structure elucidation of 108 photoTPs of six iodinated contrast media (ICM) (Chapter 3.). Again, the photodegradation study was performed in surface water spiked with the ICMs using a sunlight lab-scale simulator. 108 TPs were generated and each photoTP was characterised by its unique exact mass of the molecular ion and retention time and added to a suspect list. Once the suspect list was generated, the photoTPs were searched in thirteen surface water samples which were extracted using a generic solid- phase extraction method (four cartridges of different chemistries in order to retain ample number of compounds with different chemical properties). Based on their detection frequency (those TPs with the frequency higher than 50 % were deemed important), eleven TPs were prioritized and their structures elucidated by HRMS and NMR (when possible). Out of the eleven prioritised TPs, ten were formed as the result of deiodination (either by deiodination, oxidative deiodination or intramolecular elimination). In the real surface water samples, median concentration of parent compounds was 110 ngL-1 reaching up to 6 µgL-1 for iomeprol while TPs were found at median concentration of 8 ngL-1, reaching up to 0.4 µgL-1 for iomeprol TP651-B. Here detection-based prioritization served as a crucial step to reduce the number of TPs to be identified and thereby reducing costs and time for the subsequent target analysis. This time-effective approach not only guaranties that the degradation products elucidated would be found, but also that they are environmentally relevant. In summary, the proposed screening approach facilitates the evaluation of the degradation of polar compounds at a real scale with a fast detection of TPs without prior availability of the standards. Approach used for detection and identification of TPs of ICM in Chapter 3 was an example of suspect screening where the suspect list of TPs was generated at lab-scale, In Chapter 4, the work started with a suspect screening of lamotrigine (LMG) and related compounds (its human metabolites, synthetic impurities and photoTPs) which were listed from the literature and searched in wastewater and surface water samples. As the result of suspect screening, LMG, three human metabolites and a LMG synthetic impurity (OXO-LMG) were detected in the screened samples. Preliminary results showed significantly higher concentrations of OXO-LMG in wastewater effluent, suggesting its formation in the WWTPs. However, biodegradation reactors amended with mixed liquor at neutral pH showed that LMG is resistant to biodegradation with only about 5 % elimination after 6 days. Since LMG is extensively and predominantly metabolised by phase II metabolism to its N2-glucuronide, this metabolite (LMG-N2-G) was degraded following the same experimental setup. Results showed that this metabolite was the actual source of the TP detected. Additionally, in batch experiments, LMG-N2-G was transformed, following pseudo-first kinetics, to three TPs as a result of i) deconjugation (to LMG), ii) oxidation of the glucuronic acid (to LMG-N2-G-TP430) and iii) amidine hydrolysis in combination with deconjugation (to OXO-LMG). In order to further rationalize the formation of the TP OXO-LMG, the stability of LMG-N2-G and related compounds was studied as a function of pH in the range of 4 – 9. Same as during biodegradation, LMG was stable across the entire pH range tested. However, LMG-N2-G was transformed to three TPs at neutral – basic pH. They were identified as TPs formed after hydrolysis of amidine and guanidine moieties. The third TP detected was an intermediated in the guanidine hydrolysis reaction. Kinetic experiments in wastewater samples at different concentration (20 and 200 nM) and pH (pH 6.5, 7, 8, 8.5 and 9) demonstrated that while the degradation constants were concentration independent, at higher pH, LMG-N2-G degraded at higher rate. The pH-dependent stability experiments of related compounds with different nitrogen N2-substituents on the 1,2,4-triazine ring showed that reaction of amidine and guanidine hydrolysis depends on imine tautomer equilibrium whose formation depends directly on the N2-supsitutent. LMG- N2-G major abiotic TP (amidine hydrolysis TP) was detected in hospital effluent and WWTP influent samples. Having in mind the concentrations of both biotic and abiotic TPs detected, a total mass balance at two- concentration levels batch reactors was closed at 86% and 102%, respectively. In three WWTPs total mass balance of LMG-N2-G ranged from 71-102%. Finally, LMG-N2-G and its TPs were detected in surface water samples with median concentration ranges of 23–186 ngL-1. The work presented in this chapter gives a new insight into the behaviour of glucuronides of pharmaceuticals, suggesting that they might also be sources of yet undiscovered, but environmentally relevant TPs., [cat] Els productes farmacèutics, l'ús dels quals s'estén a nivell mundial, estan dissenyats per millorar la qualitat de vida de la societat i juguen un paper clau en el tractament i la prevenció de malalties, tant en homes com en animals. Aquests compostos químics es troben de forma ubíqua en el medi ambient. Això es deu principalment a les estacions depuradores d'aigua residual (EDARs), les quals no són capaces d'eliminar de manera eficient aquest tipus de compostos, ja que no estan dissenyades amb aquesta finalitat. Per tant, la presència de fàrmacs en el medi ambient està directament relacionada amb l'activitat humana. Un cop al medi ambient, l'estructura d'aquests compostos pot ser modificada per diferents processos biològics i abiòtics, generant-se així els que es coneixen com a productes de transformació (PTs). De fet, la transformació dels fàrmacs pot iniciar-se en alguns casos en el cos humà, després de la seva administració a causa de l'activitat metabòlica dels diferents enzims que posseeix l'home. Els metabòlits formats en aquests processos presenten algunes modificacions en les seves estructures químiques pel que fa al compost original, i, en conseqüència, unes propietats fisicoquímiques diferents. Un cop excretats, tant el fàrmac original no metabolitzat com els seus metabòlits arriben a les EDARs mitjançant la xarxa de sanejament municipal d'aigües residuals. La fracció d'aquests compostos que no s'elimina en els diferents tractaments realitzats en l'EDAR, es descarrega juntament amb l'efluent de la planta als aigües receptores. El gran nombre de transformacions que poden experimentar els fàrmacs en el seu cicle de vida a causa del seu metabolisme en el cos humà, la seva biotransformació per microorganismes i la seva fototransformació per llum solar, pot generar un nombre molt elevat de PTs en el medi ambient, i, per tant, la identificació dels mateixos, necessària per avaluar el destí dels fàrmacs en el medi ambient, és un desafiament. En el desenvolupament d'aquesta tesi es van aplicar dues aproximacions analítiques diferents: a)avaluació de perfils de PTs generats en experiments a escala de laboratori i b) anàlisi qualitativa dirigida suspect screening en mostres reals, tots dues basades en espectrometria de masses d'alta resolució (HRMS) per a la detecció i identificació de PTs de productes farmacèutics. L'aproximació d'avaluació de perfils de PTs en reactors a escala de laboratori es va aplicar per identificar productes de fototransformació (fotoPTs) de l'antiviral zanamivir (ZAN) en aigua superficial. L'aproximació de suspect screening es va aplicar per prioritzar i identificar fotoPTs de sis mitjans de contrast radiològics iodats (ICM) en aigua superficial. Finalment, una combinació de les dues aproximacions es va aplicar per detectar PTs de l’anticonvulsiu lamotrigina (LMG) i del seu principal metabòlit humà, el lamotrigina-N2-glucurònid (LMG- N2-G), resultants de la seva degradació tant en fangs activats com a reaccions d'hidròlisi a diferents valors de pH.

[spa] Los fármacos, compuestos químicos con actividad biológica, y sus productos de transformación, metabolitos y otros derivados en las plantas de tratamiento y en el medio ambiente, con actividad farmacológica remanente, son contaminantes antropogénicos que, en muchos casos, están llegando al medioambiente a través de las aguas residuales, y son las aguas superficiales el primer y principal compartimento ambiental destino. Este es un hecho que, en la actualidad, es ya ampliamente reconocido por la comunidad científica, tras el desarrollo de métodos de análisis capaces de la determinación de estos microcontaminantes en el medio ambiente, y su posterior aplicación en estudios de presencia en diferentes matrices ambientales. Los científicos, por lo tanto, están demandando medidas de control de los mismos por parte de las autoridades. Aunque tímidos, se están dando pasos adelante en materia legislativa en la lucha contra este tipo de contaminación, lo que muestra la toma de conciencia de la importancia en el control de las fuentes de entrada de estos contaminantes emergentes, así como de su eliminación del medioambiente. Esto conlleva un aumento importante en el número de estudios y, por lo tanto, en las muestras a procesar por parte de los laboratorios medioambientales. De ahí nace la necesidad de optimizar la eficiencia y competitividad de los métodos analíticos actuales, de manera que progresen en fiabilidad y sensibilidad, aumentando al mismo tiempo su versatilidad, así como su capacidad y rendimiento. El desarrollo tecnológico en el sector es muy rápido, y en general, es la comunidad científica la que evalúa en primer término la viabilidad y bonanza de las diferentes propuestas tecnológicas. Con la presente tesis se ha pretendido dar respuesta a estas necesidades. Así se ha llevado a cabo: - El desarrollo y validación de métodos analíticos para la determinación de fármacos y sus derivados en aguas medioambientales y residuales, mostrando las ventajas y puntos débiles de cada uno de ellos. En concreto, se han usado dos fuentes de fiabilidad, que no son excluyentes y pueden ser utilizadas simultáneamente. La primera ha consistido en la introducción de la automatización en el tratamiento de muestra, llevando al mínimo la manipulación de las muestras por parte del analista. La segunda ha consistido en la introducción de surrogados y patrones internos adecuados que han puesto de manifiesto las incidencias producidas durante el proceso analítico, para que hayan podido ser corregidas. Por otro lado, se han conseguido métodos que presentan una elevada sensibilidad para la determinación de los microcontaminantes objeto de estudio presentes a bajas concentraciones en matrices complejas. Para ello se ha utilizado espectrómetros de masas de triple cuadrupolo de última generación, que trabajando en modo SRM (Selected Reaction Monitoring), han proporcionado una muy buena sensibilidad, además de una elevada selectividad, que ha allananado el camino de la fiabilidad comentada anteriormente. Además, se apostó también, por el desarrollo de métodos con la suficiente versatilidad como para que hayan podido ser aplicados sobre cualquier tipo de agua ambiental dulce continental, tanto superficial como subterránea, así como a agua residual y potable, con el fin de propiciar la intercalación de muestras de diferente naturaleza analizadas consecutivamente en una misma tanda. Finalmente, se propusieron métodos dotados de una elevada capacidad, de manera que un solo método haya sido capaz de analizar el mayor número de residuos de interés posibles, incluyendo fármacos y sus derivados, metabolitos y otros productos de transformación. Pero a la vez, se ha conseguido un aumento en el rendimiento analítico de estos métodos, es decir, se ha logrado una disminución del tiempo de análisis por muestra y un aumento del número de muestras por día. Para esto se apostó claramente por la automatización, eliminando algunas etapas en el proceso analítico, lo que, además de redundar en fiabilidad como se ha comentado anteriormente, ha proporcionado métodos analíticos más cortos y autónomos y, por lo tanto, capaces de trabajar en ausencia del analista. Otra enfoque en esta dirección fue la utilización de cromatografía rápida. - Otra parte importante de la presente tesis, ha sido la aplicación posterior de los métodos desarrollados, en estudios medioambientales de elevado interés científico por su innovación. En concreto en los siguientes: A) Estudio del impacto medioambiental a escala real, en un escenario de reutilización de agua residual para el mantenimiento del caudal ecológico de un río mediterráneo. Además, se valoró el riesgo ecotoxicológico y de estrogenicidad en las muestras del río. B) Estudio de presencia de fármacos y sus productos de transformación en las aguas superficiales de la cuenca del río Ebro. C) Estudio de la distribución tridimensional y origen de residuos farmacéuticos (fármacos y sus productos de transformación) en las aguas subterráneas bajo la ciudad de Barcelona (España). D) Determinación enantiomérica de 13 residuos farmacéuticos quirales en las aguas superficiales de la cuenca del río Guadalquivir, así como en las aguas de entrada y salida a las principales EDARs asociadas a la cuenca. Además, se discutieron las implicaciones en la toxicidad de las relaciones racémicas y no racémicas., [eng] Pharmaceuticals are compounds with biological activity and their transformation products with remaining activity are arriving the environment through sewage waters, and superficial waters are the main receiving compartment. Some steps towards the fight this kind of contamination. This is creating an increase in the number of studies, and therefore, in the number of samples to be processed by the environmental laboratories. For this reason, the need to optimize the efficiency of current analytical methods, by improving the reliability, sensitivity, versatility, capacity and throughput, is a must. Technologic development is very fast in the sector, and the scientific community is generally the one that first assesses it. This thesis has tried to give answer to these requests. Thus, four multi-residue analytical methods for the analysis of pharmaceuticals and their transformation products in environmental and sewage waters, have been developed making use of cutting edge technology. The advantages and disadvantages of each of them have been discussed. Different online and offline pre-treatment technologies, fast and conventional liquid chromatographies and several kinds of triple quadrupoles for mass spectrometry, have been tested. Other important part of this thesis has been the application of the developed methods with high scientific studies, as follows: - Impact of the discharge of reclaimed sewage water in a Mediterranean river, and assessment of the eco-toxicological consequences. - Presence of pharmaceuticals and their transformation products in Ebro River basin, one of the main ones in the Iberian Peninsula. - Three-dimensional distribution of pharmaceuticals and their transformation products in the ground waters underlying Barcelona city (Spain). - Enantimeric determination of 13 chiral pharmaceutical residues in Guadalquivir River basin, one of the main ones in the Iberian Peninsula, and the assessment of the eco-toxicity issues related.

[eng] Pharmaceuticals have been detected in natural waters for more than forty years, but with improvements in sample preparation procedures and analytical instrumentation, the number of scientific publications on the issue has increased significantly. Even though the concentration of pharmaceutical residues in surface and drinking water is not critical for human health according to the present level of knowledge, the consequences for the environment are not clear. Wastewater treatment plants (WWTPs) have been identified as the primary route of pharmaceuticals to the environment, with households as the major source point for most of the over-the-counter and prescription drugs. In this thesis, the first aim was to study the occurrence, fate and removal of 43 pharmaceuticals during conventional wastewater treatment. The target compounds were selected on the basis of their high consumption in Spain or/and frequently reported detection in wastewaters and the possibility to be analyzed under the same experimental conditions. They belong to different therapeutic classes, i.e. nonsteroidal anti-inflammatory agents and analgesics, lipid modifying agents, psycholeptic and antiepileptic drugs, beta-blocking agents, beta-2-adrenoreceptor agonists, H2-receptor antagonists, antibiotics, angiotensin converting enzyme agents, diuretics and antidiabetic drugs. A wide variation in removal efficiencies was observed even for individual compounds, and across therapeutic classes and treatment processes, without clear conclusion on the removal of any particular compound. According to mass balance calculations and estimated partition coefficients, the loss of the selected pharmaceuticals during biological wastewater treatment can be fully attributed to biodegradation/biotransformation. None of the studied compounds was entirely biodegraded and/or transformed during biological wastewater treatment, but the measured concentrations were below the levels of concern according to available toxicity data. Conventional WWTPs cannot be expected to be the only mechanism for controlling the entry of pharmaceuticals into the environment because they were neither designed nor can provide their complete removal. Therefore, the challenge is to look for solutions that would be the most economical and effective means of preventing further pollution of natural waters by pharmaceuticals. More appropriate management of sewage waters before they enter treatment plants as well as a stricter control of effluent discharges, along with an in-depth investigation on the development of new designs and strategies for the improvement of existing wastewater treatments should be considered. In light of this, as the second objective of this thesis, alternative approaches for the removal of the antiepileptic carbamazepine (CBZ) were studied in aqueous media in two laboratory scale experiments: a) biodegradation using white rot fungus T. versicolor in an air-pulsed fluidized bioreactor operated in batch and continuous modes, and b) advanced oxidation using TiO2-heterogeneous photocatalysis under simulated solar and UV-A irradiation, and under the combined use of ultrasound and UV-A irradiation (sonophotocatalysis). We selected CBZ as a representative example of compounds that are found to be refractory to biological treatment and ubiquitous in various environmental matrices. Both, the fungal and UVA-driven TiO2-photocatalytic treatments, very different in their nature, have been shown to be very effective in degrading carbamazepine in aqueous media. The fungal treatment resulted in average removals of 54 and 96% in batch and continuous reactor, respectively. Acute toxicity test using the bioluminescent marine bacterium Vibrio fischeri showed a decrease in toxicity during the treatment in both types of bioreactor. In the other study, initial CBZ concentrations were reduced for 95% during 120 min of the UV-driven photocatalytic experiment and sonophotocatalysis, while only 10 % of CBZ was photodegraded during under solar irradiation. A slight increase in toxicity in Daphnia magna acute toxicity testing was observed over the time-course of the photocatalytic experiments, which can be associated with the formation of transformation products of CBZ. Ultraperformance liquid chromatography coupled to a quadrupole-time-of-flight mass spectrometry was used for a tentative identification of the transformation products of CBZ formed during the performed experiments. Most of the tentatively identified intermediates exhibited only slight modifications of the CBZ molecular structure. The fungal as well as the photocatalytic processes yielded oxygenated transformation products. In the biological treatment using T. versicolor, transformation products were formed by enzymatic epoxidation and hydroxylation of seven-membered heterocyclic ring of the carbamazepine molecule. During the photocatalytic experiments, CBZ-related transformation products emerged from hydroxylation and further oxidation of different parts of the molecule of carbamazepine. The generated transformation products appeared to be more persistent than their parent compound, as they were present, although at low concentration, until the end of the experiments. The results of the thesis contribute to a better understanding of a) the magnitude of the selected pharmaceuticals that reach the environment through the wastewater and sludge discharge, b) the efficiency of typical conventional wastewater treatment plants regarding the removal of these compounds from raw wastewater, and c) possible developments of alternative technologies for their enhanced elimination., [spa] Desde hace más de cuarenta años se ha detectado la presencia de fármacos en el ciclo de aguas, sobre todo debido a los avances en la química analítica que han permitido el desarrollo de nuevas metodologías analíticas para la determinación de estos compuestos de modo fiable y a bajas concentraciones. Las estaciones depuradoras de aguas residuales (EDARs) han sido identificadas como la ruta principal de entrada de fármacos de origen humano en el medioambiente. Por tanto, el principal objetivo de esta tesis ha sido el estudio de la presencia, destino y eliminación de 43 fármacos seleccionados, durante el tratamiento convencional realizado en las EDARs. Los compuestos estudiados fueron seleccionados en base a los índices de consumo en España, a la frecuencia de detección en aguas residuales y además en base a la posibilidad de ser analizados bajo las mismas condiciones experimentales. Estos compuestos pertenecen a diferentes clases terapéuticas, i.e. antiinflamatorios no esteroideos , los agentes que reducen los lípidos séricos, ansiolíticos y antiepilépticos, los agentes bloqueadores beta-adrenérgicos, agonistas β2 adrenérgico, antagonistas H2 , antibióticos, inhibidores de la enzima convertidora de angiotensina, diuréticos y antidiabéticos. Para el análisis cuantitativo se utilizó la cromatografía de líquidos acoplada a espectrometría de masas en tándem (LC-MS/MS), empleando un sistema híbrido triple quadrupolo/trampa de iones lineal. Como segundo objetivo hemos considerado evaluar tratamientos avanzados alternativos al tratamiento convencional para la eliminación de un fármaco antiepiléptico, la carbamazepina, uno de los compuestos más recalcitrantes al tratamiento biológico convencional. Se procedió a estudiar su degradación en medio acuoso mediante dos procedimientos a escala laboratorio: a) biodegradación utilizando el hongo ligninolitico Trametes Versicolor en un reactor fluidizado por pulsos de aire operando en modo batch y continuo, y b) oxidación avanzada mediante un tratamiento fotocatalitico en presencia de TiO2 bajo irradiación UV-A y solar, y aplicando la radiación UV en combinación con ultrasonidos (sonofotocatálisis). Para la evaluación de los tratamientos alternativos, se identificaron los productos de transformación de la carbamazepina, y se evaluó la toxicidad de las muestras tratadas. Se utilizó la LC-MS/MS con analizador de tipo cuadrupolo-tiempo de vuelo. También se evaluó la toxicidad de las muestras tratadas.