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Institute
Investigating the environmental fate of iodinated X-ray contrast media in the urban water cycle
(2010)
Iodinated X-ray contrast media (ICM) are a group of emerging contaminants which have been detected at elevated concentrations in the aquatic environment. These compounds are excreted unmetabolized into hospital wastewater, and eventually treated at wastewater treatment plants (WWTPs). The removal of ICM in WWTPs has not been very effective and therefore the ICM enter the aquatic environment via WWTP effluent discharges. Research has investigated the removal of selected ICM via abiotic and biotic processes, however limited work has attempted to determine the fate of these compounds once released into the environment. This thesis investigates the biotransformation of four selected ICM (diatrizoate, iohexol, iomeprol, and iopamidol) in aerobic soil-water and sediment-water systems as well as in different environmental matrices. Iohexol, iomeprol and iopamidol were biotransformed to several TPs in the aerobic batch systems, while no biotransformation was observed for the ionic ICM, diatrizoate. In total 34 biotransformation products (TPs) of the three non-ionic ICM were identified. The combination of semi-preparative HPLC-UV, hybrid triple quadrupole-linear ion trap mass spectrometry (Qq-LIT-MS) was found to be an accurate approach for the structural elucidation of ICM TPs. The ICM TPs resulted in microbial transformation occurring at the side chains of the parent ICM, with the iodinated aromatic ring unmodified.
Key mechanisms for the release of metal(loid)s from a construction material in hydraulic engineering
(2017)
Hydraulic engineering and thus construction materials are necessary to enable the navigability of water ways. Since, a variety of natural as well as artificial materials are used, this materials are world wide tested on a potential release of dangerous substances to prevent adverse effects on the environment. To determine the potential release, it is important to identify and to understand key mechanisms which are decisive for the release of hazardous substances. A limited correlation between the conditions used in regulatory tests and those found in environmental systems is given and hence, often the significance of results from standardised tests on construction materials is questioned, since they are not designed to mimic environmental conditions.
In Germany industrial by-products are used as armour stones in hydraulic engineering. Especially the by-product copper slag is used during the last 40 years for the construction of embankments, groynes and coastal protection. On the one hand, this material has a high density and natural resources (landscape) are protected. One the other hand, the material contains high quantities of metal(loid)s. Therefore the copper slag (product name: iron silicate stones) is very suitable as test material. Metal(loid)s examined were As, Sb and Mo as representatives for (hydr)oxide forming elements and Cd, Co, Cu, Fe, Ni, Pb and Zn were studied as representatives for elements forming cations during the release.
Questions addressed in this Thesis were: (i) can we transfer the results from batch experiments to construction scenarios under the prevalent environmental conditions, (ii) which long-term trends exist for the release of metal(loid)s from copper slags and (iii) how environmental conditions influence the leaching of metal(loid)s from water construction materials?
To answer the first question the surface depending release of the metal(loid)s from the construction materials was examined. Therefore, batch leaching experiments with different particle sizes and a constant liquid/solid ratio were performed. In a second step a comparison between different methods for the determination of the specific surface area of armour stones with a 3D laser scanning method as a reference were performed. In a last step it was possible to show that via a roughness factor the results of the specific surface area from small stones, measured with gas adsorption, can be connected with the results from armour stones, determined with an aluminium foil method. Based on calculations of the specific surface area, it was possible to significantly improve catchment scale calculation about the release of metal(loid)s and to evaluate a potential impact of construction materials in hydraulic engineering on the water chemistry of rivers and streams.
To answer the second question long-term leaching diffuse gradient in thin films supported experiments were performed for half a year. Diffuse gradients in thin films (DGT) is an in situ method to passive sample metal(loid)s in water, sediments and soils. They were used as a sink for metal(loid)s in the eluate to provide solution equilibriums. Thus the exchange of the eluent, which is performed normally in long-term experiments, was superfluous and long-term effects under undisturbed conditions were studied. The long-term leaching experiments with DGT have proven to be capable (i) to differentiate between the depletion of the material surface and the solution equilibriums and (ii) to study sorption processes with or without a further release of the analytes. This means for the practically relevant test material copper slag that: (i) the cations Cd, Co, Cu, Ni and Pb are confirmed to be released from the slag over the whole time period of six months, (ii) a surface depletion of Zn was detected, and (iii) that the (hydr)oxide forming elements As, Mo and Sb were released from the slag over the hole periods of six months but the release was masked by adsorption to Fe-oxide colloids, which were formed during the leaching experiments. It was confirmed, that sulphide minerals are the main source for long-term release of Cd, Cu, Ni, Pb and Mo.
To answer the third question short-term leaching experiments simulating environmental conditions in hydraulic engineering were performed. One factor is the salinity. The influence of this parameter was tested in batch experiments with sea salt solution (30 g/l), river Rhine water, ultra pure water and in addition with different NaCl concentration (5, 10, 20 and 30 g/l). In general, the ionic strength is an important factor for the metal(loid) release but the composition of the water (e.g. the HCO3- content) may superimpose this effect. Therefore, the concentrations of the metal(loid)s in the experiments with ultra-pure water spiked with sea salt or native river water and the ultra-pure water spiked with NaCl were significantly different. In a second experiment the influence of the environmental parameters and the interactions between the environmental parameters pH (4–10), sediment content (0 g–3.75 g), temperature (4 °C–36 °C) and ionic strength (0 g/l–30 g/l NaCl) on the release of metal(loid)s from the test material was examined. The statistical Design of Experiments (DoE) was used to study the influence of these factors as well as their interactions. All studied factors may impact the release of metal(loid)s from the test material to the eluent, whereas the release and the partitioning between sediment and eluate of metal(loid)s was impacted by interactions between the studied factors. The main processes were sorption, complexation, solubility, buffering and ion exchange. In addition, by separating the sediment from the slag after the experiments by magnetic separation, the enrichment of metal(loid)s in the sediment was visible. Thus, the sediment was the most important factor for the release of the metal(loid)s, via pH, temperature and ionic strength, because the sediment acted as a sink.
Water scarcity is already an omnipresent problem in many parts of the world, especially in sub-Saharan Africa. The dry years 2018 and 2019 showed that also in Germany water resources are finite. Projections and predictions for the next decades indicate that renewal rates of existing water resources will decline due the growing influence of climate change, but that water extraction rates will increase due to population growth. It is therefore important to find alternative and sustainable methods to make optimal use of the water resources currently available. For this reason, the reuse of treated wastewater for irrigation and recharge purposes has become one focus of scientific research in this field. However, it must be taken into account that wastewater contains so-called micropollutants, i.e., substances of anthropogenic origin. These are, e.g., pharmaceuticals, pesticides and industrial chemicals which enter the wastewater, but also metabolites that are formed in the human body from pharmaceuticals or personal care products. Through the treatment in wastewater treatment plants (WWTPs) as well as through chemical, biological and physical processes in the soil passage during the reuse of water, these micropollutants are transformed to new substances, known as transformation products (TPs), which further broaden the number of contaminants that can be detected within the whole water cycle.
Despite the fact that the presence of human metabolites and environmental TPs in untreated and treated wastewater has been known for a many years, they are rarely included in common routine analysis methods. Therefore, a first goal of this thesis was the development of an analysis method based on liquid chromatography - tandem mass spectrometry (LC-MS/MS) that contains a broad spectrum of frequently detected micropollutants including their known metabolites and TPs. The developed multi-residue analysis method contained a total of 80 precursor micropollutants and 74 metabolites and TPs of different substance classes. The method was validated for the analysis of different water matrices (WWTP influent and effluent, surface water and groundwater from a bank filtration site). The influence of the MS parameters on the quality of the analysis data was studied. Despite the high number of analytes, a sufficient number of datapoints per peak was maintained, ensuring a high sensitivity and precision as well as a good recovery for all matrices. The selection of the analytes proved to be relevant as 95% of the selected micropollutants were detected in at least one sample. Several micropollutants were quantified that were not in the focus of other current multi-residue analysis methods (e.g. oxypurinol). The relevance of including metabolites and TPs was demonstrated by the frequent detection of, e.g., clopidogrel acid and valsartan acid at higher concentrations than their precursors, the latter even being detected in samples of bank filtrate water.
By the integration of metabolites, which are produced in the body by biological processes, and biological and chemical TPs, the multi-residue analysis method is also suitable for elucidating degradation mechanisms in treatment systems for water reuse that, e.g., use a soil passage for further treatment. In the second part of the thesis, samples from two treatment systems based on natural processes were analysed: a pilot-scale above-ground sequential biofiltration system (SBF) and a full-scale soil aquifer treatment (SAT) site. In the SBF system mainly biological degradation was observed, which was clearly demonstrated by the detection of biological TPs after the treatment. The efficiency of the degradation was improved by an intermediate aeration, which created oxic conditions in the upper layer of the following soil passage. In the SAT system a combination of biodegradation and sorption processes occurred. By the different behaviour of some biodegradable micropollutants compared to the SBF system, the influence of redox conditions and microbial community was observed. An advantage of the SAT system over the SBF system was found in the sorption capacity of the natural soil. Especially positively charged micropollutants showed attenuation due to ionic interactions with negatively charged soil particles. Based on the physicochemical properties at ambient pH, the degree of removal in the investigated systems and the occurrence in the source water, a selection of process-based indicator substances was proposed.
Within the first two parts of this thesis a micropollutant was frequently detected at elevated concentrations in WWTPs effluents, which was not previously in the focus of environmental research: the antidiabetic drug sitagliptin (STG). STG showed low degradability in biological systems and thus it was investigated to what extend chemical treatment by ozonation can ensure attenuation of it. STG contains an aliphatic primary amine as the principal point of attack for the ozone molecule. There is only limited information about the behaviour of this functional group during ozonation and thus, STG served as an example for other micropollutants containing aliphatic primary amines. A pH-dependent degradation kinetic was observed due to the protonation of the primary amine at lower pH values. At pH values in the range 6 - 8, which is typical for the environment and in WWTPs, STG showed degradation kinetics in the range of 103 M-1s-1 and thus belongs to the group of readily degradable substances. However, complete degradation can only be expected at significantly higher pH values (> 9). The transformation of the primary amine moiety into a nitro group was observed as the major degradation mechanism for STG during ozonation. Other mechanisms involved the formation of a diketone, bond breakages and the formation of trifluoroacetic acid (TFA). Investigations at a pilot-scale ozonation plant using the effluent of a biological degradation of a municipal WWTP as source water confirmed the results of the laboratory studies: STG could not be removed completely even at high ozone doses and the nitro compound was formed as the main TP and remained stable during further ozonation and subsequent biological treatment. It can therefore be assumed that under realistic conditions both a residual concentration of STG and the formed main TP as well as other stable TPs such as TFA can be detected in the effluents of a WWTP consisting of conventional biological treatment followed by ozonation and subsequent biological polishing steps.
Method development for the quantification of pharmaceuticals in aqueous environmental matrices
(2021)
As a consequence of the world population increase and the resulting water scarcity, water quality is the object of growing attention. In that context, organic anthropogenic molecules — often defined as micropollutants— represent a threat for water resources. Among them, pharmaceuticals are the object of particular concerns due to their permanent discharge, their increasing consumption and their effect-based structures. Pharmaceuticals are mainly introduced in the environment via wastewater treatment plants (WWTPs), along with their metabolites and the on-site formed transformation products (TPs). Once in the aquatic environment, they partition between the different environmental compartments in particular the aqueous phase, suspended particulate matter(SPM) and biota. In the last decades, pharmaceuticals have been widely investigated in the water phase. However, extreme polar pharmaceuticals have rarely been monitored due to the lack of robust analytical methods. Moreover, metabolites and TPs have seldom been included in routine analysis methods although their environmental relevance is proven. Furthermore, pharmaceuticals have been only sporadically investigated in SPM and biota and adequate multi-residue methods are lacking to obtain comprehensive results about their occurrence in these matrices. This thesis endeavors to cover these gaps of knowledge by the development of generic multi-residue methods for pharmaceuticals determination in the water phase, SPM and biota and to evaluate the occurrence and partition of pharmaceuticals into these compartments. For a complete overview, a particular focus was laid on extreme polar pharmaceuticals, pharmaceutical metabolites and TPs. In total, three innovative multi-residue methods were developed, they include analytes covering a broad range of physico-chemical properties. First, a reliable multi-residue method was developed for the analysis of extreme polar pharmaceuticals, metabolites and TPs dissolved in water. The selected analytes covered a significant range of elevated polarity and the method would be easily expendable to further analytes. This versatility could be achieved by the utilization of freeze-drying as sample preparation and zwitterionic hydrophilic interaction liquid chromatography (HILIC) in gradient elution mode. The suitability of HILIC chromatography to simultaneously quantify a large range of micropollutants in aqueous environmental samples was thoroughly studied. Several limitations were pointed out: a very complex and time-consuming method development, a very high sensitivity with regards to modification of the acetonitrile to water ratio in the eluent or the diluent and high positive matrix effects for certain analytes. However, these limitations can be overcome by the utilization of a precise protocol and appropriate labeled internal standards. They are overmatched by the benefits of HILIC which permits the chromatographic separation of extreme polar micropollutants. Investigation of environmental samples showed elevated concentrations of the analytes in the water phase. In particular, gabapentin, metformin, guanylurea and oxypurinol were measured at concentrations in the µg/L range in surface water. Subsequently, a reliable multi-residue method was established for the determination of 57 pharmaceuticals, 47 metabolites and TPs sorbed to SPM down to the low ng/g range. This method was conceived to cover a large range of polarity in particular with the inclusion of extreme polar pharmaceuticals. The extraction procedure was based on pressurized liquid extraction (PLE) followed by a clean-up via solvent exchange and detection via direct injection-reversed-phase LC-MS/MS and freeze-drying HILIC-MS/MS. Pharmaceutical sorption was examined using laboratory experiments. Derived distribution coefficients Kd varied by five orders of magnitude among the analytes and confirmed a high sorption potential for positively charged and nonpolar pharmaceuticals. The occurrence of pharmaceuticals in German rivers SPM was evaluated by the investigation of annual composite SPM samples taken at four sites at the river Rhine and one site at the river Saar between the years 2005 and 2015. It revealed the ubiquitous presence of pharmaceuticals sorbed to SPM in these rivers. In particular, positively charged analytes, even very polar and nonpolar pharmaceuticals showed appreciable concentrations. For many pharmaceuticals, a distinct correlation was observed between the annual quantities consumed in Germany and the concentrations measured in SPM. Studies of composite SPM spatial distribution permitted to get hints about specific industrial discharge by comparing the pollution pattern along the river. For the first time, these results showed the potential of SPM for the monitoring of positively charged and nonpolar pharmaceuticals in surface water. Finally, a reliable and generic multi residue method was developed to investigate 35 pharmaceuticals and 28 metabolites and TPs in fish plasma, fish liver and fish fillet. For this matrix, it was very challenging to develop an adequate clean-up allowing for the sufficient separation of the matrix disturbances from the analytes. In the final method, fish tissue extraction was performed by cell disruption followed by a non-discriminating clean-up based on silica gel solid-phase extraction(SPE) and restrictive access media (RAM) chromatography. Application of the developed method to the measurement of bream and carp tissues from German rivers revealed that even polar micropollutants such as pharmaceuticals are ubiquitously present in fish tissues. In total, 17 analytes were detected for the first time in fish tissues, including 10 metabolites/TPs. The importance of monitoring metabolites and TPs in fish tissues was confirmed with their detection at similar concentrations as their parents. Liver and fillet were shown to be appropriate for the monitoring of pharmaceuticals in fish, whereas plasma is more inconvenient due to very low concentrations and collection difficulties. Elevated concentrations of certain metabolites suggest possible formation of human metabolites in fish. Measured concentrations indicate a low bioaccumulation potential for pharmaceuticals in fish tissues.
The sediments of surface waters are temporary or final depository of many chemical compounds, including trace metals and metalloids (metal(loid)s) from natural and anthropogenic sources. Whether they act as a source or sink of metal(loid)s depends strongly on the dynamics of the biogeochemical processes that take place at the sediment-water interface (SWI). Important information on biogeochemical processes as well as on the exposure, the fate and the transport of pollutants at the SWI can be obtained by determining chemical concentration profiles in the sediment pore water. A major challenge is to conduct experiments with a spatial resolution, which allows to adequately record existing gradients and to log all the parameters needed, to describe and better understand the complex processes at the SWIs. At the same time, it is from major importance to prevent the formation of any artifacts during sampling, which may occur due to the labile nature of the SWIs and the very steep biogeochemical gradients.
In this context, in the first part of this work, a system was developed and tested that enables the automated, minimal invasive sampling of sediment pore water of undisturbed or manipulated sediments while simultaneously recording parameters such as redox potential, oxygen content and pH value. In an incubation experiment the impact of acidification and mechanical disturbance (re-suspension) on the mobility of 13 metal(loid)s was investigated using a triple quadrupole inductively coupled plasma-mass spectrometry (ICP-QQQ-MS) multi-element approach. Most metals were released as consequence of sulfide weathering whereas mechanical disturbance had a major impact on the mobility of the oxide forming elements As, Mo, Sb, U and V. Additionally, options were demonstrated to address with the system the size fractionation of metal(loid)s in pore water samples and the speciation of As(III/V) and Sb(III/V).
In the second part, the focus, with a similar experimental design, was placed on the processes leading to the release of metal(loid)s. For this purpose, two incubation experiments with different oxygen supply were conducted in parallel. For the first time the nonmetals carbon, phosphorus and sulfur were analyzed simultaneous to 13 metal(loid)s in sediment pore water by ICP-QQQ-MS. Throughout the experiment metal(loid) size fractionation was monitored. It was confirmed that resuspension promotes the mobility of metalloids such as As, Sb and V, while the release of most metals was largely attributed to pyrite weathering. The colloidal (0.45-16 μm) contribution in terms of mobilization was only relevant for a few elements.
Finally, the sampling system was used as part of a new approach to sediment risk assessment. Undisturbed sediment cores from differently contaminated positions in the Trave estuary were examined, considering 16 metal(loid)s, the non-metals C, P and S and the ions NH4+, PO43- and SO42-. By the first in-depth comparison with in-situ dialysis-based pore water sampling the ability of the suction-based approach to represent field conditions was proven. The pore water studies together with supplementing resuspension experiments in bio-geochemical microcosms and sequential extraction identified the most “pristine” sediment of the study area as posing the greatest risk of metal(loid) release. However, the potentially released amounts per kg of sediment are only a few parts per thousand of the average daily loads of the Trave river.
Thousands of chemicals from daily use are being discharged from civilization into the water cycle via different pathways. Ingredients of personal care products, detergents, pharmaceuticals, pesticides, and industrial chemicals thus find their way into the aquatic ecosystems and may cause adverse impacts on the ecology. Pharmaceuticals for instance, represent a central group of anthropogenic chemicals, because of their designed potency to interfere with physiological functions in organisms. Ecotoxicological effects from pharmaceutical burden have been verified in the past. Therapeutic groups with pronounced endocrine disrupting potentials such as steroid hormones gain increasing focus in environmental research as it was reported that they cause endocrine disruption in aquatic organisms even when exposed to environmentally relevant concentrations. This thesis considers the comprehensive investigation of the occurrence of corticosteroids and progestogens in wastewater treatment plant (WWTP) effluents and surface waters as well as the elucidation of the fate and biodegradability of these steroid families during activated sludge treatment. For the first goal of the thesis, a robust and highly sensitive analytical method based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed in order to simultaneously determine the occurrence of around 60 mineralocorticoids, glucocorticoids and progestogens in the aquatic environment. A special focus was set to the compound selection due to the diversity of marketed synthetic steroids. Some analytical challenges have been approved by individual approaches regarding sensitivity enhancement and compound stabilities. These results may be important for further research in environmental analysis of steroid hormones. Reliable and low quantification limits are the perquisite for the determination of corticosteroids and progestogens at relevant concentrations due to low consumption volumes and simultaneously low effect-based trigger values. Achieved quantification limits for all target analytes ranged between 0.02 ng/L and 0.5 ng/L in surface water and 0.05 ng/L to 5 ng/L in WWTP effluents. This sensitivity enabled the detection of three mineralocorticoids, 23 glucocorticoids and 10 progestogens within the sampling campaign around Germany. Many of them were detected for the first time in the environment, particularly in Germany and the EU. To the best of our knowledge, this in-depth steroid screening provided a good overview of single steroid burden and allowed for the identification of predominantly steroids of each steroid
type analyzed for the first time. The frequent detection of highly potent synthetic steroids (e.g. triamcinolone acetonide, clobetasol propionate, betamethasone valerate, dienogest, cyproterone acetate) highlighted insufficient removal during conventional Summary wastewater treatment and indicated the need for regulation to control their emission since the steroid concentrations were found to be above the reported effect-based trigger values for biota. Overall, the study revealed reliable environmental data of poorly or even not analyzed steroids. The results complement the existing knowledge in this field but also providednew information which can beused particularly for compound prioritization in ecotoxicological research and environmental analysis. Based on the data obtained from the monitoring campaign, incubation experiments were conducted to enable the comparison of the biodegradability and transformation processes in activated sludge treatment for structure-related steroids under aerobic and standardized experimental conditions. The compounds were accurately selected to cover manifold structural moieties of commonly used glucocorticoids, including non-halogenated and halogenated steroids, their mono- and diesters, and several acetonide-type steroids. This approach allowed for a structure-based interpretation of the results. The obtained biodegradation rate constants suggested large variations in the biodegradability (half-lifes ranged from < 0.5 h to > 14 d). An increasing stability was identified in the order from non-halogenated steroids (e.g. hydrocortisone), over 9α-halogenated steroids (e.g. betamethasone), to C17-monoesters (e.g. betamethasone 17-valerate, clobetasol propionate), and finally to acetonides (e.g. triamcinolone acetonide), thus suggesting a strong relationship of the biodegradability with the glucocorticoid structure. Some explanations for this behavior have been received by identifying the transformation products (TPs) and elucidating individual transformation pathways. The results revealed the identification of the likelihood of transformation reactions depending on the chemical steroid structure for the first time. Among the identified TPs, the carboxylates (e.g. TPs of fluticasone propionate, triamcinolone acetonide) have been shown persistency in the subsequent incubation experiments. The newly identified TPs furthermore were frequently detected in the effluents of full-scale wastewater treatment plants. These findings emphasized i) the transferability of the lab-scale degradation experiments to real world and that ii) insufficient removals may cause adverse effects in the aquatic environment due to the ability of the precursor steroids and TPs to interact with the endocrine system in biota. For the last goal, the conceptual study for glucocorticoids was applied to progestogens.
Here, two sub-types of the steroid family frequently used for hormonal contraception were selected (17α-hydroxyprogesterone and 19-norstestosterone type). The progestogens showed a fast and complete degradation within six hours, and thus empathizes pronounced biodegradability. However, cyproterone acetate and dienogest Summary have been found to be more recalcitrant in activated sludge treatment. This was consistent with their ubiquitously occurrence during the previous monitoring campaign. The elucidation of TPs again revealed some crucial information regarding the observed behavior and highlighted furthermore the formation of hazardous TPs. It was shown that 19-nortestosterone type steroids are able to undergo aromatization at ring A in contact with activated sludge, leading to the formation of estrogen-like TPs with a phenolic moiety at ring A. In the case of norethisterone the formation of 17α-ethinylestradiol was confirmed, which is a well-known potent synthetic estrogen with elevated ecotoxicological potency. Thus, the results indicated for the very first time an unknown source of estrogenic compounds, particularly for 17α-ethinylestradiol. In conclusion, some steroids were found to be very stable in activated sludge treatment, others degrade well, and others which do degrade but predominantly to active TPs depending on their chemical structure. Fluorinated acetal steroids such as triamcinolone acetonide and fluocinolone acetonide are poorly biodegradable, which is reflected in high concentrations detected ubiquitously in WWTP effluents. Endogenous steroids and their most related synthetic once such as hydrocortisone, prednisolone or 17α-hydroxyprogesterone are readily biodegradable. Regardless their high influent concentrations, they are almost completely removed in conventional WWTPs. Steroids between this range have been found to form elevated quantities of TPs which are partially still active, which particularly the case for betamethasone, fluticasone propionate, cyproterone acetate or dienogest. The thesis illustrates the need for an extensive evaluation of the environmental risks and carried out that corticosteroids and progestogens merit more attention in environmental regulatory and research than it is currently the case
Within aquatic environments sediment water interfaces (SWIs) are the most important areas concerning exchange processes between the water body and the sediment. These spatially restricted regions are characterized by steep biogeochemical gradients that determine the speciation and fate of natural or artificial substances. Apart from biological mediated processes (e.g., burrowing organisms, photosynthesis) the determining exchange processes are diffusion or a colloid-mediated transport. Hence, methods are required enabling to capture the fine scale structures at the boundary layer and to distinguish between the different transport pathways. Regarding emerging substances that will probably reach the aquatic environment engineered nanomaterials (ENMs) are of great concern due to their increased use in many products and applications. Since they are determined based on their size (<100 nm) they include a variety of different materials behaving differently in the environment. Once released, they will inevitable mix with naturally present colloids (< 1 μm) including natural nanomaterials.
With regard to existing methodological gaps concerning the characterization of ENMs (as emerging substances) and the investigation of SWIs (as receiving environmental compartments), the aim of this thesis was to develop, validate and apply suitable analytical tools. The challenges were to i) develop methods that enable a high resolution and low-invasive sampling of sediment pore water. To ii) develop routine-suitable methods for the characterization of metal-based engineered nanoparticles and iii) to adopt and optimize size-fractionation approaches for pore water samples of sediment depth profiles to obtain size-related information on element distributions at SWIs.
Within the first part, an available microprofiling system was combined with a novel micro sampling system equipped with newly developed sample filtration-probes. The system was thoroughly validated and applied to a freshwater sediment proving the applicability for an automatic sampling of sediment pore waters in parallel to microsensor measurements. Thereby, for the first time multi-element information for sediment depth profiles were obtained at a millimeter scale that could directly be related to simultaneously measured sediment parameters.
Due to the expected release of ENMs to the environment the aim was to develop methods that enable the investigation of fate and transport of ENMs at sediment water interfaces. Since standardized approaches are still lacking, methods were developed for the determination of the total mass concentration and the determination of the dissolved fraction of (nano)particle suspensions. Thereby, validated, routine suitable methods were provided enabling for the first time a routine-suitable determination of these two, among the most important properties regarding the analyses of colloidal systems, also urgently needed as a basis for the development of appropriate (future) risk assessments and regulatory frameworks. Based on this methodological basis, approaches were developed enabling to distinguish between dissolved and colloidal fractions of sediment pore waters. This made it possible for the first time to obtain fraction related element information for sediment depth profiles at a millimeter scale, capturing the fine scale structures and distinguishing between diffusion and colloid-mediated transport. In addition to the research oriented parts of this thesis, questions concerning the regulation of ENPs in the case of a release into aquatic systems were addressed in a separate publication (included in the Appendix) discussing the topic against the background of the currently valid German water legislation and the actual state of the research.
Water is the basis of all life, a biotope for a variety of organisms and an important component of the
natural balance. For this reason, it is essential to protect water from contamination by anthropogenic
organic micropollutants (MPs) and to develop various innovative strategies for the treatment and reuse
of wastewater. Conventionally biologically treated municipal wastewater contains many organic MPs
that pose a potential threat to aquatic ecosystems and drinking water resources. To date, the focus has
been on physicochemical processes such as activated carbon treatment or ozonation to improve the
removal of MPs in wastewater treatment plants (WWTPs), while knowledge of biological removal
processes and ways to optimize the biological removal of organic MPs is comparatively limited.
Especially in arid and semi-arid regions, treated wastewater is also an important resource for wastewater
reuse for irrigation or drinking water treatment. In consequence of increasing droughts due to climate
change, the reuse of treated wastewater is also increasingly being discussed in Germany. Often the water
is reused after targeted groundwater recharge, while concepts for optimizing the removal of MPs during
soil passages are still scare. In order to assess the potential and limitations of biological wastewater
treatment as well as targeted groundwater recharge for the removal of organic MPs, a better knowledge
of the biological biotransformation processes and the environmental conditions influencing them is
required. Microbial communities and their enzyme pools play a key role in these processes. However,
the influence of environmental conditions on the composition and functional characteristics of microbial
communities, and how this in turn affects the biotransformation potential of MPs with different
structural characteristics, has been little studied. In addition, knowledge about MP-biotransforming
bacteria is often derived from enrichment or even pure culture studies, which are not directly transferable
to the environment.
Against this background, this dissertation focuses on the systematic investigation of the
biotransformation of MPs in contact with activated sludge (simulation of biotransformation processes
in biological treatment stages of WWTPs) and filter materials (simulation of processes in groundwater
recharge systems) under defined laboratory conditions. The aim is to elucidate relationships between
the composition and functional characteristics of microbial communities, microbiologically determined
biotransformation processes and their optimal process conditions, as well as the chemical structure and
primary enzymatically catalyzed biotransformation reactions of MPs.
The first part of the work focuses on the composition of microbial communities and the
biotransformation of MPs depending on defined process parameters in five differently operated
wastewater reactors, divided into two reactor cascades at pilot scale and one full-scale WWTP. The first
cascade consisted of three reactors, the first of which served as a reference reactor with conventional
activated sludge (CAS) treatment. The following reactors operate under anoxic to anaerobic and strictly
anaerobic conditions, respectively. The other cascade consists of two reactors, the first anaerobic and the second aerobic, followed by a simplified vessel to prevent nitrate output, as half of the effluent was
recirculated to the first anaerobic reactor. All five reactors and the WWTP were analyzed for the
biotransformation of 33 MPs and the composition of the microbial community by LC-MS/MS and 16S
rRNA gene sequencing, respectively. The results showed a slight but significant improvement in the
overall biotransformation of MPs in the reactor cascades (about 20%) compared to the WWTP. In
particular, the biotransformation of compounds that are not or only slightly degradable (< 30%) in
conventional wastewater treatment, such as diatrizoate, venlafaxine and diclofenac, was significantly
improved (about 70%). Twelve of the 33 MPs showed an increase in biotransformation of at least 30%
compared to the reference reactor and the WWTP. In detail, the reactor cascade consisting of the
anaerobic pre-treatment showed three times more MPs than the reactor cascade with the anaerobic post-
treatment. Although the environmental conditions (nutrient availability and redox conditions) differed
considerably between the reactors, molecular analysis of the microbial community revealed a core
community of 143 genera, with 54% of the taxa belonging to the phylum Proteobacteria, with the beta
subdivision as the most dominant class. On the other hand, a specialized community consisting of 90
genera was identified that contributed most to the differences between the reactor communities. These
genera were shown to reflect the prevailing nutrient, redox and operating conditions of each treatment.
It was also found that the relative abundances of several genera of the specialized community correlated
with the biotransformation of certain MPs as well as with process parameters (especially redox
conditions). These genera may not necessarily be directly involved in the biotransformation of MPs, but
could be promising biological indicators for the establishment and control of operating conditions
favorable to the biotransformation of certain MPs. For the two reactor cascades analyzed, it was
concluded that the redox conditions as well as the carbon supply were essential factors for the
composition of the specialized community and the biotransformation of the MPs. In addition, the 16S
rRNA gene amplicon sequencing proved to be a complementary tool to confirm process conditions by
correlation. In conclusion, the efficiency of MP biotransformation in conventional WWTPs depends on
key parameters such as redox conditions, biodiversity or the presence of several genera of specialized
microbial communities identified as indicator organisms.
Against the background of previous knowledge that biofilms from moving bed biofilm reactors
(MBBRs), which have been used so far in a few WWTPs, are more efficient in biotransforming certain
MPs, the second study used laboratory batch experiments to investigate the extent to which the
biotransformation potential of carrier-attached biofilms and suspended sludge from hybrid MBBRs
differ between three WWTPs, and whether these differences were also reflected in the composition of
the bacterial communities. The 31 MPs investigated were grouped according to their biotransformation
rates and examined for relationships between the biotransformation behavior and the known primary
biotransformation reactions are discernible. In general, the study confirmed the potential of hybrid
MBBRs for improved biotransformation of a variety of MPs. This could be attributed to an increased
biotransformation potential of the carrier-attached biofilms, especially for oxidatively degradable substances such as trimethoprim, diclofenac and mecoprop. Correlation analyses showed statistically
significant relationships between the occurrence of certain bacterial genera (e.g. Acidibacter, Nitrospira
or Rhizomicrobium) and the biotransformation rates of certain MPs. Thus, some of the identified genera
were also discussed as suitable indicators for the biotransformation potential of suspended sludge or
carrier-attached biofilm.
In the third part of the work, the biotransformation of up to 78 MPs was investigated under different
redox and substrate conditions in a defined column system at laboratory scale as well as ex situ under
uniform incubation conditions by incubating the column filter material in batch experiments. MPs were
categorized based on their biotransformation behavior and how well these categories matched primary
transformation reactions known from the literature or predicted using the Eawag pathway prediction
system for modelling microbial biotransformation pathways. The column system investigated consisted
of two large columns filled with technical sand, an intermediate aeration and four smaller columns, all
connected in series. In the first column, characterized by a carbon-rich environment with high biomass
and strong oxygen depletion, 23 MPs were efficiently removed (>80% removal), while 19 substances
were better or exclusively removed under carbon-limited oligotrophic conditions in the columns after
re-aeration. While the MPs removed predominantly in the first column were mostly attributed to
biotransformation by comparatively ubiquitous and fast transformation reactions such as the oxidation
of alcohols or amide hydrolysis, the biotransformation of MPs removed more efficiently under the oxic
and oligotrophic conditions in the rear columns was characterized by slower transformation reactions
such as N-dealkylation of primary and secondary amines or the hydroxylation of aromatic rings. In
addition, several specific reactions described in the literature, such as the cleavage of C-Cl and C-O
bonds, could only be identified under oligotrophic conditions. The results suggest that conditions of
limited carbon availability may favor the development of alternative metabolic biotransformation
pathways. Furthermore, the observed association between system-specific MP removal and the expected
primary biotransformation reactions from the literature and prediction systems could serve as a basis for
accurately identifying the relevant enzymes in future studies, using metagenomic or metatranscriptomic
data.
Based on these considerations, molecular biological investigations were carried out in the fourth part of
this work by generating sequence-based data complementary to the biotransformation rate constants
during the batch experiments carried out in the third study. A first aim was to identify a statistical relation
between the microbial composition, the process conditions and the nutrient availability as well as the
biotransformation potential of 42 MPs depending on the biomass in the columns of the system.
Furthermore, the metabolic activities of the microbial communities of the first two columns of the
system were analyzed and the identified active enzymes were assigned to known metabolic pathways
and the activities were compared. In addition, the extent to which the biotransformation potential of the
columns was also reflected in the activity of enzymes that have been proven to belong to biotransformation pathways of MPs was investigated. To this end, modern sequencing methods were
used to generate data on the composition of the microbial community (16S rRNA gene sequencing) and
its actual activity in the form of expressed genes (metatranscriptome sequencing). The biotransformation
rates from the batch experiments of the third study were used by normalizing them with the gene copy
numbers (kbio), which were representative of the biomass. For half of the MPs, the highest normalized
kbio values were identified in the rear columns with increased carbon-limiting conditions. Furthermore,
most of the MPs investigated, such as sulfathiazole, sulfamethoxazole or rufinamide, showed an
improved biotransformation potential in these columns. The higher biotransformation potential under
carbon-limiting conditions was also confirmed by metatranscriptomic analysis, where an increased
metabolic capability and a higher functional activity to degrade MPs were observed in the carbon-
limiting column compared to the first nutrient-rich column. Taxonomic analysis revealed a decreasing
trend in biodiversity with a simultaneous increase in carbon-limiting conditions. Furthermore, the
microbial community differed depending on the nutrient and process conditions of the column materials
analyzed. However, within the carbon-limiting columns, the microbial community was similar and were
dominated by the genus Pseudomonas. By relating the biotransformation of MPs to the relative
abundance of taxa in the carbon-limiting columns, the genera of the phylum Acidobacteria and the
classes Alpha- and Gammaproteobacteria showed particularly high associations. In addition, four genera
showed a statistically positive correlation with the two structurally similar MPs sulfamethoxazole and
sulfathiazole.
Overall, no general improvement in biotransformation could be found for all MPs under the conditions
considered, with system- and condition-specific changes observed for individual compounds. However,
the studies indicated favorable environmental conditions for groups of MPs that showed increased
biotransformation linked with high abundant taxa, especially under carbon-limiting conditions and in
carrier-attached biofilms.
Microplastics (MP), i.e., plastic particles < 5 mm, are perceived as a threatening envi-
ronmental and human health issue. Growing public interest in this class of contaminants
requires standardized and harmonized methods for their quantification. While an abun-
dance of analytical methods (both particle-based and mass-based) for the detection of
microplastics is available, existing studies on the quantity of MP in the environment lack
comparability. Therefore, the aim of this work was to establish a fast, reliable screen-
ing method for the quantification of the most common synthetic polymers in complex
environmental samples.
This was accomplished by a two-step pressurized liquid extraction (PLE) followed by
analysis via pyrolysis coupled to gas chromatography and mass spectrometry (Py–GC–
MS). In the first extraction step, a large part of the organic matrix was removed with
methanol at 100 ∘C and 100 bar, followed by a second step with tetrahydrofuran at
185 ∘C and 100 bar to extract the polymers that were subsequently adsorbed to silica
gel and measured with Py–GC–MS. With the developed method, limits of quantification
in an environmentally relevant concentration range of 7–8 μg g−1 for the most common
thermoplastic polymers polyethylene (PE), polypropylene (PP), and polystyrene (PS)
were achieved.
In order to improve the robustness of the method, poly(styrene-d5) (PSd5) was initially
applied as internal standard. However, further analyses revealed a deuterium–hydrogen
exchange during Py–GC–MS measurement, which was catalyzed by the inorganic ma-
trix. This effect was thereupon systematically investigated and poly(4-fluorostyrene) was
established as a new, stable internal standard.
While the developed method enabled the quantification of PE, PP, and PS, several other
polymers had to be excluded. In particular, the quantification of poly(ethylene tereph-
thalate) (PET) proved challenging via Py–GC–MS. A variety of catalytic effects by the
inorganic matrix was revealed and systematically investigated, e.g., changes in pyrolysis
product distribution. Several different sample preparation approaches failed to resolve
these issues. PLE led to a depolymerization of PET which was also catalyzed by the
inorganic sample matrix.
After further optimization and reduction of false positives, the developed method has
the potential to be included in future standardized procedures for MP quantification. It
provides a fast, robust analysis of MP in complex samples, while also considering widely overlooked matrix effects. Potential quantification approaches for other polymers that are
not included in the developed method (e.g., tire wear particles, paint particles) are also
discussed in this thesis.
The presence of synthetic chemicals in the environment can affect both ecosystems and
human health. In particular, the increasing contamination of the aquatic environment by
complex mixtures of anthropogenic trace substances has become a major global concern.
Once released into the environment, these compounds can undergo diverse
transformation processes to form a wide range of transformation products (TPs), which
are commonly unknown. Transformation inevitably alters the pattern of contamination and
exposure, as new substances are formed with frequently different physicochemical
properties, environmental behavior and toxicity in comparison to their precursor
compounds. For instance, TPs can exhibit significantly greater persistence and mobility in
the aquatic environment, posing a threat to both aquatic ecosystems and drinking water
resources. Therefore, TPs need to be considered in the risk assessment and authorization
process of chemicals. However, due to a combination of predictive, analytical, and
regulatory challenges, TPs currently remain largely unrecognized and unregulated. By
addressing these challenges, this thesis comprehensively characterizes the entry paths,
occurrence, fate, and (eco)toxicological relevance of selected TPs in the aquatic system in
Germany. These TPs have been largely overlooked in environmental studies and aquatic
monitoring programs for decades, despite their precursors being produced and used in
large quantities on a global scale.
The highly persistent and mobile substance trifluoroacetate (TFA) has garnered
significant attention in recent years due to its diverse sources, widespread occurrence in
the aquatic environment, and the lack of economically viable options to remove TFA from
contaminated waters. One of the most frequently discussed diffuse sources is the
formation of TFA in the atmosphere through the oxidation of volatile precursors and its
subsequent scavenging from the atmosphere by wet deposition. Despite the previously
reported occurrence of TFA in precipitation, the lack of recent and comprehensive data
has severely limited the understanding of the significance of wet deposition as a source of
TFA to the (aquatic) environment. Thus, in the present work, a nationwide field monitoring
campaign covering all precipitation events over a one-year sampling period was
conducted at eight sites across Germany. Samples were analyzed for TFA using ion
exchange chromatography (IC) coupled to negative-ion electrospray tandem mass
spectrometry (ESI-MS/MS). Of the analyzed samples, 16% exhibited TFA concentrations
≥ 1 μg/L. The precipitation-weighted average TFA concentration of 0.34 μg/L highlighted
that wet deposition alone is responsible for approximately 0.3 to 0.4 μg/L of TFA in
surface waters in Germany. The annual wet deposition fluxes ranged from 91 to
400 μg/m², with the highest fluxes observed in densely populated regions. The annual wet
deposition of TFA for Germany during the observation period was estimated to be 68 t.
The sampling revealed a pronounced seasonality, with the highest concentrations and wet
deposition fluxes of TFA observed in summer. Pearson correlation analyses indicated that
the transformation of TFA precursors in the troposphere is enhanced in summer due to
elevated concentrations of photochemically generated oxidants, primarily •OH, which
ultimately results in increased atmospheric TFA deposition. Overall, the study provided the
first published data on TFA in precipitation in Germany since 1995/96. The derived data
serves as a benchmark for future studies. In addition, it allows for the establishment of
mass balances and can be used to develop models to predict the loads of TFA entering
the aquatic environment from multiple sources.
The lack of robust historical data on the wet deposition fluxes of TFA also impeded long-
term trend analyses. Specifically, a postulated increase in atmospheric formation and
deposition of TFA due to substantial emission increases of numerous volatile TFA
precursors in recent decades remained unquantified. To address this knowledge gap,
archived plant samples were analyzed to evaluate the long-term temporal trends in the
atmospheric deposition of TFA in Germany. A robust and highly sensitive analytical
method for TFA in plant matrices was developed and validated. The method
encompassed a three-step sequential extraction procedure followed by the analysis of the
diluted sample extracts using IC-ESI-MS/MS. Subsequently, archived leaf samples of
various tree species and sampling sites from the German Environmental Specimen Bank
(observation period: 1989−2020) were analyzed for TFA. Statistical analysis revealed
significant (p < 0.05) positive trends in TFA concentrations in plant leaves, which is likely
the result of both phytoaccumulation and increasing emissions of gaseous TFA precursors
over the observation period. The concentrations increased by factors of up to 12 from
1989 to 2020. The highest concentrations (up to ∼1,000 μg/kg dry weight) were found in
Lombardy poplar leaves. Overall, the study presents the first trend analysis of TFA in biota
and raises awareness of the escalating atmospheric deposition of TFA over the past three
decades.
Sulfamate has previously been identified as a TP of the artificial sweeteners cyclamate
and acesulfame in wastewater and drinking water treatment. The preliminary results
indicated that sulfamate concentrations in wastewater treatment plant (WWTP) effluent
are substantially higher than those of other wastewater-borne contaminants. However,
despite its high global production and usage, no information was available on the sources,
occurrence, and environmental significance of sulfamate in the aquatic system in
Germany. To close this knowledge gap, a quantitative monitoring approach of different
urban water cycle compartments was conducted. Target analysis based on IC-ESI-MS/MS
revealed exceptionally high concentrations of sulfamate in wastewater (up to 1,900 μg/L),
surface water (up to 580 μg/L), and finished drinking water (up to 140 μg/L) in Germany.
Considering the limited data on short-term ecotoxicity, approximately 30% of the
sulfamate concentrations detected in groundwater and surface water samples exceeded
the derived predicted no-effect concentration (PNEC) of sulfamate. Therefore, the
potential impact of sulfamate on the aquatic ecosystem in Germany cannot be excluded.
Municipal WWTP effluent was identified as the primary source of sulfamate for the aquatic
system, as its concentrations correlated positively (r > 0.77) with the municipal wastewater
tracer carbamazepine in samples from different waterbodies. Ozonation and activated
sludge experiments demonstrated that sulfamate can be formed through chemical and
biological degradation of various precursors containing a sulfonamide group.
Nevertheless, the transformation of precursors to sulfamate in WWTPs and receiving
waters was found to be quantitatively insignificant, due to the substantial direct use of
sulfamic acid as a descaling agent in domestic and industrial applications. Laboratory
batch experiments, in conjunction with the findings from the sampling conducted at full-
scale waterworks, demonstrated that the commonly applied drinking water treatment
techniques, including ozonation and activated carbon filtration, are largely ineffective in
removing sulfamate. Bank filtration was identified as the only option to efficiently eliminate
sulfamate from contaminated raw water resources (removal: 62% to 99%). Overall, the
study presents the first comprehensive analysis of sulfamate in the urban water cycle and
suggests that there may be other high production volume inorganic chemicals that are
currently overlooked in environmental studies and monitoring programs.
Despite pantoprazole (PPZ) being one of the most widely prescribed human
pharmaceuticals globally, consistently low concentrations of this proton-pump inhibitor in
environmental water samples have been documented. This can be attributed to the
extensive metabolism of PPZ within the human body, with only minor amounts of the
parent compound being excreted. Since environmental monitoring and risk assessment
for regulatory purposes focus on the parent substances of pharmaceuticals, it was
assumed that the current environmental exposure associated with the use of PPZ is
considerably underestimated. In the presented thesis, 4′-O-demethyl-PPZ sulfide (M1)
was identified as the most relevant PPZ metabolite for environmental analysis. This was
achieved by applying reversed-phase high-performance liquid chromatography (RP-
HPLC) coupled to high-resolution mass spectrometry (HRMS) to urine samples of a PPZ
user, as well as to municipal wastewater. M1, which had not been investigated in previous
monitoring studies, was found to be ubiquitous in WWTP influent and effluent (max.:
3 μg/L, detention frequency: 100%) as well as in surface water (max.: 1.2 μg/L; detection
frequency: 97%) in Germany. Its average surface water concentration was approximately
30 times higher than that of the parent compound PPZ. Moreover, quantitative structure-
toxicity relationship (QSTR) modeling indicated a lower preliminary freshwater PNEC for
M1 (4.8 μg/L) compared to PPZ (28 μg/L). The analysis of archived suspended particulate
matter (SPM) samples from the Rhine at Koblenz revealed that the concentrations of M1
increased significantly from 2005 to 2015 and were positively correlated with the
prescription volume of PPZ. Conventional biological wastewater treatment was found to
be insufficient to remove M1 (average removal: 22%). Laboratory-scale experiments and
the analysis of samples taken after different treatment steps of an advanced full-scale
WWTP demonstrated that post-treatment with activated carbon as well as ozonation can
significantly improve the removal of M1 and PPZ during wastewater treatment, thereby
reducing their release to the aquatic environment. During ozonation, a rapid oxidation of
M1 was observed, accompanied by the formation of several ozonation products, which
were proposed for the first time. The identity of the main ozonation TPs of M1 was
confirmed through the synthesis of reference compounds. Their detection in samples
collected after the ozonation step of a full-scale WWTP demonstrated the transferability of
the laboratory-scale ozonation experiments. M1 was found to be sufficiently removed from
contaminated source waters (max. raw water concentration: 0.25 μg/L) by bank filtration
under different redox conditions (removal ≥ 80 %) and by other commonly applied
purification processes in drinking water production. In summary, this study revealed that
the environmental exposure and risk associated with the use of PPZ have been previously
underestimated, which likely extends to other human pharmaceuticals. Therefore, these
findings call for more sophisticated approaches to environmental monitoring and risk
assessment of pharmaceuticals that take TPs into account.
This thesis provides an in-depth understanding of the entry paths, occurrence, fate, and
environmental significance of selected TPs in the aquatic system in Germany. It
significantly advances our understanding of the introduction of TFA into the water cycle,
by characterizing the source of wet deposition and elucidating long-term temporal trends
of atmospherically deposited TFA. Additionally, the thesis gives comprehensive insights
into the formation, behavior, removability, and potential (eco)toxicological risks of
sulfamate, PPZ and its TPs. The thesis addresses key challenges in assessing and
integrating TPs into chemical management and presents solutions to overcome these
challenges. Finally, it highlights the urgent need for increased focus on TPs in research,
aquatic monitoring, and regulation to safeguard the environment and human health.