Research team

Toxicological Centre

Expertise

Expertise in development and validation of bioanalytical assays to detect endogenous and exogenous substances in biological matrices. Assays are based on chromatographic separation coupled to mass spectrometric detection

Exposure of Pregnant Women to Halogenated Organic Pollutants (HOPs) and Health Implications in Areas Impacted by the Halogenated Chemical Industry (HOPE). 01/01/2022 - 31/12/2024

Abstract

Halogenated organic pollutants (HOPs) released into the environment may cause adverse effects on the human health. To assess the human exposure risk of HOPs, it is thus necessary to investigate their environmental levels and consider the multiple exposure pathways of people in high-risk areas. Non-target screening (NTS) analytical approaches allow the early detection of potentially harmful novel HOPs. In addition, some novel HOPs may cause remarkable changes in cell metabolites, although they do not exhibit significant dose-effect relationships. Metabolomics may detect low, but critical effects that are not observed by conventional toxicological methods. The application of NTS and metabolomics on sensitive populations (e.g. pregnant women) in the vicinity of halogenated chemical industrial parks would thus provide a screening of high-risk HOPs. Through the cooperation of the Belgian and Chinese teams in this project, we aim at 1) identifying novel HOPs in multiple environmental media relevant for human exposure in the vicinity of chemical industry parks, 2) clarifying the dominant exposure pathways of novel HOPs through the comparison of exposure levels and characteristics of human internal and external exposure, 3) investigating the effects of combined exposure to HOPs on the metabolome of pregnant women, and 4) identifying biomarkers for high-risk HOPs. Altogether, our results will provide the basis for environmental and chemical management of novel HOPs.

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Assessing human exposure to persistent and mobile chemicals. 01/12/2021 - 30/11/2023

Abstract

Persistent and Mobile chemicals (or PMs) are at this moment insufficiently considered in chemical regulation and management. One important group of PM chemicals is the group of per- and polyfluoroalkyl substances (PFAS). In this project, we aim to investigate the identity, occurrence, fate and sources of PMs in order to elucidate the major exposure pathways and to assess the current human exposure to several groups of PM chemicals, including PFAS.

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Pulling the hairs: Investigating the hair metabolome as a promising approach to characterize the progression of chronic alcoholic liver diseases (METABOLHAIR) 01/11/2021 - 31/10/2022

Abstract

Alcoholic liver disease (ALD) is the most prevalent type of chronic liver disease. Despite the high prevalence of ALD, diagnosis and disease staging are still missing sensitive and specific biomarkers. As a result, most patients are diagnosed only at an advanced stage, leading to frequent hospitalization, increased medication use and liver transplantations, a high-burden option for patients and health-care systems. This PhD project (METABOLHAIR) aims to explore the applicability of human hair as a new diagnostic matrix in metabolomics to identify and detect early-stage biomarkers of chronic ALD. This project strives to elucidate the global hair metabolome composition, its inter-individual variations and possible influencing parameters. Identification of ALD biomarkers will be performed using an advanced metabolomics workflow, including a unique analytical platform (two-dimensional liquid chromatography-ion mobility spectrometry-quadrupole time-of-flight mass spectrometry). Furthermore, the applicability of the biomarkers in segmental hair analysis to predict disease progression and diagnose the different stages (alcoholic fatty liver disease and alcoholic steatohepatitis) of ALD will be investigated. The METABOLHAIR project will help improve the current knowledge on the hair metabolome and can be used to screen individuals with alcohol use disorders for ALD to prevent the development of end-stage liver diseases.

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Analytical and environmental novel insights in the human exposure to Synthetic ANTioxidants by Advanced mass spectrometry (SANTA) 01/11/2021 - 31/10/2022

Abstract

Within the indoor environment, humans are exposed to a diverse group of contaminants of emerging concern (CECs), who remain largely understudied. Two groups of ubiquitous CECs, namely the high molecular weight phenolic antioxidants (HMW SPA) and the organic phosphite antioxidants (OPA), have received little attention in literature. These novel antioxidants exposure has only been accessed in Asia and North-America by applying methods that focus on a small subset of novel antioxidants (AOXs). Therefore, the goal of this project is to develop both qualitative and quantitative exposure methods, including a human biomonitoring method for novel AOXs metabolites. These innovative methods will be applied in Flanders, providing the first exposure studies in Europe for the novel AOXs. This goal will be achieved by employing state of the art mass spectrometry (MS) equipment coupled to liquid or gas chromatography. By developing target and non-target MS screening methods and applying novel techniques such as iterative MS within the screening methods. This project will result in the first biomonitoring method of novel AOX metabolites accessing human exposure in Flanders. The developed methods can be used in upcoming European biomonitoring programs, accessing exposure on a larger scale. In conclusion, this project will be of great added value as it will be the first in Europe to access human exposure to novel antioxidants by employing innovative techniques.

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Reviving electrochemical detection for HPLC-analysis of illicit drugs, metabolites and isomers. (REVAMP). 01/10/2021 - 30/09/2024

Abstract

Drug (ab)use continues to have devastating consequences on human health and society. As large changes have occurred recently in the recreational drug market throughout Europe, such as new psychoactive substances, chemical modifications and isomerisations of typical illicit drugs, novel analytical challenges arose. These chemicals contain multiple drugs or even isomers that are specifically designed to evade current on-site test and international drug legislation. The proposed REVAMP project has the ambitious goal to revive electrochemical detection in liquid chromatography (HPLC). The goal is to create and study the coupling of a new electrochemical detector based on a screen printed electrode (SPE) array with HPLC to develop for the first time a mobile benchtop device able to identify drugs on-site with an enhanced selectivity towards isomers and polydrug detection. The main problem of conventional electrochemical detection (reproducibility and polishing) will be tackled by using SPE's. Although electrochemical detection is an inviting approach to detect a wide variety of compounds, given its high sensitivity (low/sub-ng/ml), low cost and miniaturization opportunities, the methodological coupling to LC with SPE's is lacking. The obtained strategies can be transferred to analytes with often similar functionalities such as antibiotics, phenolic compound and explosives.

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Human exposure assessment of persistent and mobile chemicals: an analytical challenge. 01/10/2021 - 30/09/2024

Abstract

Persistent and mobile chemicals (PMs) are man-made and highly polar organic chemicals. Due to potential risks to the environment and humans, policy needs are currently identified. However, there is limited knowledge on PMs, especially on reliable screening and target analytical methods and human exposure pathways. In order to fill knowledge gaps, this research project aims to develop suitable analytical methods using state-of-art mass spectrometric techniques, to screen and prioritize PMs, to explore the occurrence of PMs in environmental matrices relevant for human exposure, and finally to assess the human exposure to PMs. Reliable, sensitive and robust methods using liquid chromatography-ion mobility-high resolution mass spectrometry and liquid chromatography-tandem mass spectrometry will be developed and validated for the screening and quantification of the analytically-challenging PMs. Using these methods, the occurrence and distribution of PMs will then be measured in various environmental samples relevant for human exposure (e.g. dust, fish), consumer products and in human biological samples. Finally, major exposure sources and hazard quotients of PMs will be identified to assess the human exposure to PMs. The outcomes of this research project will further be valuable in establishing policies for the management of human exposure to PMs.

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Advanced research and training network in food quality, safety and security (FoodTraNet). 01/05/2021 - 30/04/2025

Abstract

FoodTraNet is designed to provide high-level training of a new generation of high achieving early stage researchers (ESRs) in a common platform of advanced mass spectrometry tools for food quality, safety and security. Due to the fast growing and changing area in food science researchers need a multidisciplinary, intersectoral grounding in these tools to extract all the potential from traditional and new available methodologies and techniques. Although already available there is still a big gap between research capability and industrial uptake that needs to be bridged to assure their use in more practical and efficient way. The network is designed to bridge this translational gap by creating a multidisciplinary training and research network that will give ESRs the ability to transfer their knowledge from basic sciences to industrial applications. FoodTraNet will cover techniques including stable isotopes, target, suspect, non-target screening and mass spectrometry imaging to obtain the following objectives: 1) identify biomarkers and bioactive compounds to assure food quality and traceability; 2) characterize novel foods to support food safety and security; and 3) developed and characterize new products based on the latest advances in nanotechnology: edible photonic barcodes; nano-sensors; multi-functional polymer nanostructured materials; and new, active and intelligent packaging. These goals will be achieved by a unique combination of "hands-on" research training, non-academic placements/courses and workshops on scientific and complementary transferrable skills facilitated by the academicnon- academic composition of the consortium. Strong industry involvement in the project, with full participation of CONCAST and SMEs from different sectors, will provide ESRs with the transferable skills necessary for thriving careers in a burgeoning area that underpins food sustainability through innovative technological development across a range of diverse disciplines.

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Project website

Validation of autophagy induction as a therapeutic strategy: from drug discovery and preclinical evaluation to safety investigation and biomarker research. 01/01/2021 - 31/12/2024

Abstract

Autophagy is a ubiquitous process that removes unnecessary or dysfunctional cellular components from the cytoplasm. Defective autophagy is currently emerging as a hallmark of many diseases. In this framework, there is strong interest in pharmacological agents that stimulate autophagy (so-called 'autophagy inducers'), as a potential treatment for these diseases. The unequivocal validation of autophagy induction as a therapeutic strategy, however, is far from established. Many obstacles persist, including the lack of druglike, selective autophagy inducers and readily translatable preclinical results that are obtained with such compounds. In addition, the availability of reliable biomarkers for autophagy and additional fundamental safety data for the approach, would strongly contribute to its validation. This proposal addresses existing limitations in the state-of-the art in the domain. We have recently carried out a phenotypic High-Throughput Screen (HTS) on a curated compound library. Members in this library were preselected from different providers based on in silico druglikeness scores. One compound family that was identified in the screen and maximally validated prior to this application, will be further optimized chemically for autophagy induction potency and biopharmaceutical properties. The biopharmaceutical profile of the best new representative will be thoroughly characterized in vivo, both involving PET-based pharmacokinetics and phenotypic pharmacodynamics. The compound will subsequently be investigated in two mouse models of diseases characterized by defective autophagy: atherosclerosis and Charcot-Marie-Tooth periferal neuropathy. In addition, we propose to investigate whether autophagy induction is intrinsically sufficiently safe as a therapeutic strategy. Existing hypotheses that autophagy induction could accelerate tumorigenesis and/or tumor growth will be investigated in vivo. In the same framework, metabolomics will be relied on to monitor eventual cellular stress fingerprints that result from chronic or long-term autophagy stimulation. Finally, metabolomics will also be relied on to identify cellular biomarkers of autophagy induction. The latter will be validated in plasma samples of animals that were systemically treated with autophagy inducers. Combined, we expect the knowledge and tools that are generated by this proposal to have strong impact on the field of autophagy research and ongoing endeavors to validate autophagy induction as a therapeutic strategy.

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COVID-19 National surveillance in wastewater. 09/12/2020 - 14/09/2022

Abstract

Tracking the presence of SARS-CoV-2 in wastewater allows to monitor the spread of the virus in the population at regional and subregional level, several days earlier compared to data obtained from screening of patient samples. This wastewater monitoring is no replacement for current clinical testing, yet it is pivotal in early detection of future changes in the epidemic and the tracking of novel outbreaks in specific regions. As such this testing will contribute to a proactive management and containment of the epidemic.

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Advanced research and training network in food quality, safety and security (FOODTRANET). Emerging chemicals in novel food commodities of animal origin. 01/12/2020 - 30/11/2022

Abstract

The specific project to be developed at UA aims at identifying emerging chemicals in novel food items for human and animal consumption (including edible insects, farmed fish, insect-fed fish, beehive products, and dairy products) using non-target and suspect screening based methods. Year 1. At first, suitable workflows for non-target and suspect screening methodologies based on liquid chromatography and high-resolution mass spectrometry (LC-HRMS) will be developed and harmonized for the identification of emerging chemicals present in the selected food. Such methods are currently in full development for human biological samples (urine and serum), and it is expected that they could serve as basis for the methods to be developed in food. In case of non-target methods, data analysis will be carried out without any a priori defined suspects, while for suspect screening methods a comprehensive list of emerging chemicals that are suspected/expected to be present in the analyzed samples (e.g. emerging pesticides, per- and polyfluoroalkyl substances, chlorinated paraffins, new plasticizers, etc.) will be set up. Then a selection of relevant samples (e.g. n=3 per food category) belonging to each novel food category will be purchased and analyzed. Year 2. Once the results of this first screening will be available, the dataset of samples belonging to the above-mentioned novel food categories will be widened (to n=10 per food category) and analyzed using the optimized methods. The performance of such analytical platforms and the generated results will then be critically assessed by chemometric data processing (combining multivariate analysis – like principal component analysis (PCA), random forest classification and support vector machines – with the application of univariate tests). The results of the statistical analysis will be combined with available information about food production, in order to establish differences and similarities in the patterns of emerging chemicals in novel food items. This information will be also transferred to the European Food Safety Authority (EFSA) to assess the necessity of including these emerging chemicals in regular monitoring schemes. The outcomes of this project will contribute to a preliminary food safety assessment of the potential of such matrices to be used as alternative food sources. Additionally, this specific project will offer insights on the most "promising" novel food categories present on the Belgian market to favor. In parallel, the EU ETN proposal FoodTraNet will be re-submitted and the coordinator of FoodTraNet, Dr. Nives Ogrinc has supplied a letter of support stating this engagement. As such, this proof of concept of contamination with emerging chemicals will be able to increase the chances of success of the resubmitted proposal and will be able to extent the investigation at broader European level.

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Novel mass-spectrometric approaches to investigate human exposure to emerging contaminants. 01/11/2020 - 31/10/2022

Abstract

Owing to the increasing plastic production, humans are exposed to a rising number of plastic additives. Many of these have been associated with neurotoxicity and adverse effects on the endocrine system. In the past, several new compounds linked to plastics have been identified, which are designated as "contaminants of emerging concern" (CECs). This project aims to identify new CECs derived from plastics. A novel, ultra performant analytical approach based on the hyphenation of liquid chromatography, ion mobility spectrometry and high-resolution mass spectrometry (LC-IM-HRMS) will be used for the creation of a database containing collision cross section (CCS) values of known and new CECs. Non-target and suspect screening approaches will be applied for the identification of new CECs in plastic and indoor dust samples. Using in vitro experiments in hepatic systems, Phase I and II metabolites of the new CECs will be identified. Screening of human urine samples will be conducted to confirm the metabolite's occurrence and identify the most relevant biotransformation products in vivo. Furthermore, a quantitative method using LC-MSMS will be developed and validated. Using this method, a proof-of-concept study on a set of 100 human urine samples will be conducted with the purpose to establish time-trends for these new CECs. The results obtained with this workflow will deepen the knowledge about human exposure to CECs and have an influential input on future biomonitoring programs.

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Training and coaching for carrying out wastewater-based epidemiology in selected European Neighbourhood Policy (ENP) countries 24/06/2020 - 31/05/2022

Abstract

The aim of this training and coaching is to engage selected European countries (Neighbourhood Policy countries) in wastewater-based epidemiology studies. We will train these countries in wastewater sample collection, wastewater analysis and data interpretation

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From exposome to effect assessment of contaminants in human and animal models (EXPOSOME). 01/01/2020 - 31/12/2025

Abstract

We will develop a pioneering holistic framework based on innovative approaches to explore the human exposome in terms of exposure leading to adverse effects with a focus on endocrine-modulated neurological and metabolic disorders by: i. Identifying and characterizing the exposure sources of relevant chemicals in the context of the xposome framework; ii. developing and applying in silico, in vitro and in vivo human and animal models to investigate the absorption, distribution, metabolism, and excretion processes after exposure to chemicals; iii. setting up relevant clinical/epidemiological exposure-wide association studies to better understand the associations between exposure and neurological and metabolic disorders in longitudinal and (nested) casecontrol cohorts and including birth cohorts to understand transgenerational mechanisms; iv. using targeted and untargeted omics techniques (e.g. metabolomics and transcriptomics) in human and animal biological systems to aid data-driven discovery of causal factors for adverse health effects; v. linking exposure to mixtures by integrating exposome research with the adverse outcome pathway concept, a novel toxicological framework structuring the cascade of biological events from an initial molecular-level perturbation of a biological system to an adverse health outcome.

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Novel approaches for the estimation of the use of psychoactive pharmaceuticals and illicit drugs by wastewater analysis. 01/01/2020 - 31/12/2023

Abstract

The present project aims at developing innovative analytical and sampling approaches to estimate the (ab)use of illicit drugs and psychoactive pharmaceuticals, e.g. antidepressants, antipsychotics, benzodiazepines, and opioids in the general population. The analysis of human excretion products of substance use in wastewater has the unique properties to deliver such data. The overarching objective is to validate innovative active-passive sampling (APS) and detection strategies for biomarkers of illicit drugs and psychoactive pharmaceuticals to be used for the long-term monitoring of substance use in communities. The hypothesis to be tested is if APS will provide a more convenient sampling strategy compared to daily 24-h composite sampling. We will first develop analytical assays to measure biomarkers of psychoactive pharmaceuticals and drugs in wastewater. Then we will develop APS strategies for the target analytes in influent and effluent wastewater and will investigate the advantages/limitations of the APS strategies in real-life conditions. These new sampling strategies will be compared to the conventional 24-h composite sampling. We will plan sampling campaigns in 5 Slovene and 3 Belgian cities to: estimate for the first time community-wide psychoactive substance use in the Slovene and Belgian cities; evaluate chemical removal efficiencies for the wastewater treatment plants; investigate the utility of APS for improved identification of new psychoactive substances.

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Improving treatment strategies for different depressive disorders by discovery of novel drug target candidates and discriminatory diagnostic biomarkers. 01/01/2020 - 31/12/2022

Abstract

Psychiatric treatment of patients with depressive illness is characterized by very low remission and recovery rates due to inefficient and inaccurate diagnoses along with a high final non-responsiveness to all psychopharmacological medication (30% to close to 64% in psychotic depression ). With this IOF-SBO project, we aim to optimize treatment strategies for uni- and bipolar depression with or without psychotic symptoms by finding alternative entry points for the development of new antidepressant drugs and through the discovery of peripheral biomarkers for accurate and objective discriminatory diagnostics. In order to establish databases of relevant biomarkers as well as putative targets for future development of psychopharmacological drugs, we will conduct a prospective clinical trial in which patients with uni- or bipolar disorder, with or without psychotic symptoms, will receive electroconvulsive therapy (ECT), the treatment of last resort when other antidepressants have failed. In parallel, post-mortem patient brain tissues will be retrospectively investigated. Samples of both research arms will be analysed by proteomic and metabolomic methodologies using state of the art liquid chromatography-mass spectrometry (LCMS). Final comparative analyses of differentially expressed proteins, protein networks and metabolic pathways will result in the establishment of drug target candidate (DTC) databases for each of the aforementioned disorders and one diagnostic database containing biomarkers objectively discriminating between the 4 depressive subtypes. These databases will likely, in follow-up trajectories, lead to the development of novel antidepressant drugs and diagnostic assays to be implemented in a diagnostic device. Preliminary interviews with potential industrial partners revealed a great interest for collaboration with both pharmaceutical and technology investors.

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Electrochemistry and nanostructured electrocatalysts for tackling substance abuse. 01/01/2020 - 31/12/2022

Abstract

Substance abuse remains one of the global health and socioeconomic problems. It is one of the United Nations' Sustainable Development Goals, i.e., SDG 3: "Ensure healthy lives and promote wellbeing for all at all ages". One of the goals within SDG 3.5 is to "Strengthen the prevention and treatment of substance abuse, including narcotic drug abuse and harmful use of alcohol". Specifically, SDG 3d recognizes the need to "strengthen the capacity of all countries, in particular developing countries, for early warning, risk reduction and management of national and global health risks". The key challenge to early warning, risk reduction and management of substance abuse is early and accurate detection. Electrochemical techniques allow for integration in hand-held devices for rapid detection of drugs and alcohol. However, the state-of-the-art fuel cellbased breath alcohol sensor (FCBrAS) still adopts expensive and old technology of the 70's (use of high amount of platinum electrocatalyst) despite the advances in materials electrochemical science and nanotechnology. The objective of this project is to bridge gaps in knowledge in electrochemistry of illicit drugs in oral fluids and alcohols, innovation in the FCBrAS to reduce mass-loading of expensive precious metals, and enhance selectivity and sensitivity in biological fluids.

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Identification of novel biomarkers for the diagnosis of alcohol-associated liver diseases using metabolomics. 01/11/2019 - 31/10/2023

Abstract

Alcohol abuse is a major cause of liver disease worldwide and is the most common cause of acute-on-chronic liver failure. Despite the high prevalence of alcohol-associated liver disease (AALD), there is currently a lack of sensitive and specific early biomarkers, resulting in diagnosis at late (irreversible) stages of disease progression. The objective of this research project is the identification of (a panel of) small-molecule biomarkers to diagnose early stages of AALD through an advanced metabolomics workflow. Identification of mechanistic biomarkers for AALD will be performed in an in vitro experimental setup, using HepaRG® liver cells exposed to ethanol. A unique and ultra-performant analytical system (two dimensional liquid chromatography-ion mobility spectrometry-quadrupole time-of-flight mass spectrometry) will be used to identify possible biomarkers. An in vivo proof-of-concept study will be performed to extrapolate the applicability of the in vitro identified biomarkers to an in vivo situation. For this purpose, plasma and liver tissue samples of patients suffering from alcoholic fatty liver disease and steatohepatitis, will be investigated. Obtained data will help to improve the current knowledge on the mechanisms of action of alcohol-induced hepatoxicity and to provide new input for adverse outcome pathways. The results of this research project can be used as a lead for a clinical cohort-study to diagnose different stages of AALD.

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Europe-wide Sewage analysis to Monitor Emerging drug problems (EuSeME). 15/10/2019 - 14/04/2022

Abstract

Wastewater-based epidemiology (WBE) is now an established approach that is used to monitor the consumption of a selection of illicit drugs. However, WBE can also become a valuable source of information about new drug-related threats such as consumption of new psychoactive substances (NPS), licit and illicit opioids, as well as new trends in drug use (e.g., crack cocaine and cannabis). The goal of EuSeME is to further develop WBE as an early warning system to monitor emerging drug problems across Europe. First, qualitative and quantitative information about the consumption of NPS and opioids across Europe will be obtained through the development of analytical methods and retrospective analysis of wastewater and urine samples. Second, a thorough literature review will be carried out to identify additional relevant biomarkers of crack cocaine use. Analytical methods will be developed and integrated with existing ones to include these new biomarkers and monitor consumption of crack cocaine through WBE. Third, a thorough investigation of the sorption of cannabis biomarkers on suspended matter present in wastewater will be carried out. Outcomes will be used to establish a detailed protocol, providing a robust and harmonised approach to monitor consumption of cannabis through WBE. Finally, two broad sampling campaigns aiming at covering all of Europe will be carried out, focussing on the newly identified biomarkers and on conventional drugs. These campaigns will provide unique insights about emerging threats linked to the use of NPS, opioids, crack cocaine and cannabis, as well as an updated view about the situation of consumption of conventional illicit drugs in Europe. By implementing a practical application of drug-related research, EuSeME seeks to provide support to the activities of the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA), national governments and stakeholders, law enforcement, as well as national focal points.

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Assessing human exposure to chlorinated paraffins in the indoor environment (HECPAR). 01/10/2019 - 30/09/2022

Abstract

Chlorinated paraffins (CPs) are a complex mixture of thousands of individual compounds which have been used as plasticizers, flame retardants and additives to various lubricants, adhesives and metal working fluids. Certain CPs are recognised as potential carcinogens and have exhibited toxic and bioaccumulative properties, culminating in restriction of their use due to their registration as Persistent Organic Pollutants (POPs) under the Stockholm Convention in 2017. As semi-volatile compounds, CPs may be released from consumer products and materials via volatilisation or leaching and tend to bind strongly to particulates, such as indoor dust. Inadvertent ingestion of contaminated dust is considered to be a major pathway of human exposure to CPs. This study aims to a) investigate the current human exposure risks posed by CPs in the indoor environments of Belgium and b) explore the transport and fate of such chemicals. Sensitive, reliable and robust methods for the quantitation of CPs will first be developed and validated using state-of-the-art techniques based on advanced mass spectrometry. The occurrence and distribution of CPs will then be assessed in dust from a range of indoor microenvironments to produce an estimate of human exposure via incidental dust ingestion for the Belgian population. Finally, the distribution of CPs in consumer goods and the migration of CPs to dust will be investigated to aid in mitigating harmful effects to human populations in the future.

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Buiding a robust and high-bioanalytical platform for metabolomics. 01/04/2019 - 31/03/2023

Abstract

This project aims at using the latest technological improvements to develop a robust state-of-the art bioanalytical platform that can be applied in metabolomics research. The project is divided into three work packages (WPs): WP1: Instrumental detection of metabolome based on innovative techniques: An innovative combination of analytical separation techniques – comprehensive two dimensional liquid chromatography (2D-LC) and ion mobility (IM) - coupled with high resolution accurate mass spectrometry will be explored to obtain better qualitative and quantitative data in the context of metabolomics. The implementation of five dimensions of separation (2D-LC-IM-QTOFMS) for metabolomics will allow increased separation together with more confident identification of metabolites. This WP will include optimization of LC columns (different stationary phases), mobile phase gradients, ion mobility parameters, and mass spectrometric parameters (ionization sources, fragmentor and collision energy voltages). WP2: Data analysis workflow This WP is devoted to optimize data processing strategies and explore relevant bioinformatics approaches to maximally extract relevant information from the complex datasets resulting from WP1. This will need subsequent steps such as alignment of chromatograms, subtraction of the noise, and normalization of the signals. Multivariate statistical tests will then be used to detect signals that are relevant for the study set-up (e.g. differences between exposed and non-exposed cells in an exposure experiment). Identification of the signals (i.e. which metabolites) will be performed based on their retention time, CCS value, m/z value, isotope pattern and fragmentation pattern using algorithms (Molecular Formula Generator (Agilent); MetFrag/MetFusion; CFM-ID) in combination with databases (METLIN, LipidMaps, HMDB, IIMDB, MassBank) in order to putatively annotate the correct structure. WP3: Pre-analytical phase Apart from optimizing the instrumental analysis of (parts of) the metabolome (WP1-2), this WP will substantially improve and standardize extraction protocols in order to maximize the number of extracted metabolites in a reproducible way. Sampling and extraction protocols have to be optimized based on the matrix under investigation (e.g. cell lines, tissues, serum), but all should include critical steps such as metabolism quenching and metabolite extraction.

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Research in the domain of toxicology 01/10/2018 - 30/09/2023

Abstract

The planned research activities are based on my experience accumulated in the field of bioanalysis. The research plan consists of further developing of my current research lines, as well as introducing new research directions in order to support a UA frontline (drug discovery and development) and emerging research domain (proteomics, genomics, and metabolomics). I hypothesize that biomarkers present in wastewater produced by communities provide objective information on spatio-temporal patterns on their lifestyle, disease and exposure to toxicants (= wastewater-based epidemiology). This will be tested through innovative research that will (i) select small-molecule biomarkers related to lifestyle, disease and exposure to toxicants; (ii) develop state-of-the-art bioanalytical assays to measure biomarkers in wastewater; and (iii) interpret the data with advanced statistical and modelling techniques. The aim is to implement wastewater-based epidemiology as a flexible and holistic tool to provide objective and timely information on the health status of whole communities. To allow long-term monitoring of alcohol and drug consumption in forensic contexts (e.g. driver's license regranting, organ transplantation procedure), I will continue my research to explore the use of keratinous matrices, i.e. hair and nails. I will investigate differences in hair and nails in accumulating substances. Furthermore, I want to establish reference values for biomarkers of drugs and alcohol in nails to be applied in a forensic context (as they also exist for hair). Therefore, studies with a high number of participants will be performed to correlate alcohol and drug biomarkers. I further plan to establish a robust bioanalytical platform to be used in the preclinical testing and pharmacokinetic profiling of candidate pharmaceuticals. I will focus on optimizing in vitro bioanalytical techniques for the evaluation of aspects of absorption (rapid equilibrium dialysis), distribution (protein binding assays) and metabolism (incubations with liver preparations) of xenobiotics. Furthermore, dedicated bioanalytical assays can/will be developed for the pharmacokinetic modeling of candidate pharmaceuticals (clearance, distribution volume,…). I aim to develop state-of-the-art bioanalytical procedures for metabolomics that combine multi-dimensional chromatographic and ion mobility separation techniques coupled to mass spectrometry (2D-LC-IM-QTOFMS) for the simultaneous detection of endogenous metabolites in a variety of biological matrices (cells, tissues, body fluids). This innovative technological platform will be used to (i) improve currently available bioanalytical metabolomics protocols in the Toxicological Centre for lipids and amino acids and (ii) develop innovative bioanalytical platforms to study subsets of the metabolome that are relevant for cardiovascular and inflammatory diseases: eicosanoids and steroids. Efforts will be made to optimize pre-analytical aspects including metabolism quenching and analyte extraction. These platforms can eventually be used in larger research contexts where metabolic information is crucial and can be integrated with other omics techniques to understand pathophysiological processes.

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Labo protocol. 14/09/2020 - 09/12/2020

Abstract

Tracking the presence of SARS-CoV-2 in wastewater allows to monitor the spread of the virus in the population at regional and subregional level, several days earlier compared to data obtained from screening of patient samples. This wastewater monitoring is no replacement for current clinical testing, yet it is pivotal in early detection of future changes in the epidemic and the tracking of novel outbreaks in specific regions. As such this testing will contribute to a proactive management and containment of the epidemic.

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Wastewater as early warning system for COVID-19 surveillance 13/07/2020 - 31/08/2021

Abstract

This project aims at optimising an analytical protocol for the detection of SARS-CoV-2 RNA in wastewater. This will then applied to wastewater samples collected across Belgium to investigate the early warning character of the methodology. Triangulation of the wastewater data with other datasets (e.g. hospital data, infections) will be performed to investigate the complementarity of the data.

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SARS-CoV-2 RNA quantification in wastewater as complementary epidemiological indicator to monitor the epidemic. 01/06/2020 - 31/05/2021

Abstract

This project aims to detect SARS-CoV-2 RNA in wastewater sampled throughout Belgium as a complementary epidemiological indicator for the rapid and constant follow-up of the current epidemic at high spatial and temporal resolutions. This translational data-driven epidemiological research will provide direct added value to support and evaluate policy initiatives and to detect novel outbreaks. This project fills current knowledge gaps in monitoring the epidemic by providing prevalence estimates in the general population without the need for individual testing. Furthermore, through triangulation of our data with data from diagnostic testing and clinical surveillance, this project will make a more comprehensive prediction and control of the COVID-19 epidemic possible. The idea to measure SARS-CoV-2 RNA in wastewater as a proxy for its infection in the general population is innovative and epidemiologically highly informative as community-wide data can be obtained at high temporal (daily) and spatial (city or town) resolutions. This proposed wastewater surveillance strategy can be regarded as a sensitive early-warning tool and can substantially improve models/predictions of the ongoing epidemic. Specialists in wastewater-based epidemiology and virology are brought together in this consortium to make this project feasible within the proposed timeframe.

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ADME study of lead ferroptosis inhibitor which blocks organ injury and neurodegeneration in mice. 03/02/2020 - 30/06/2021

Abstract

Ferroptosis is an iron-catalyzed form of regulated necrosis, which is shown to be detrimentally involved in several experimental disease models, such as acute kidney/liver injury and neurodegeneration. Glutathione peroxidase 4 (GPX4) is the central enzyme protecting the cell from excessive lipid peroxidation, which is the key execution process in ferroptosis. A high-throughput screening performed by the Stockwell Lab (Columbia University, US) led to the discovery of ferrostatin-1 (Fer1) as a potent in vitro inhibitor of ferroptosis. In vivo however, the molecule suffers from instability. Therefore, we developed ferrostatin-analogues with improved efficacy, solubility and stability. Ongoing research, in the framework of an FWO research project and an FWO-EOS project, illustrates that our patented lead Fer1--analogue UAMC-3203 is superior as compared to the benchmarks in several ferroptosis-driven experimental disease mouse models. The aim of this project is to study aspects of absorption, distribution, metabolism and excretion of the potential lead ferroptosis inhibitor UAMC-3203 in mice and rats. The results of this POC project should deliver an extended ADME-profile, verify its ability to cross the blood-brain-barrier and validate the possibility to administer UAMC-3203 orally. This will increase the valorisation potential of this compound. As the number of potential applications is relatively big, building a spin-off case for evaluation by seasoned investors and business professionals is likely the most suitable valorisation strategy.

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Community-wide assessment of the opioid burden through combined use of wastewater-based analytics and prescription drug monitoring data. 01/01/2019 - 31/10/2019

Abstract

In recent times, the use of opioids has escalated drastically and resulted in a crisis of epidemic proportions in North America. In Europe, the use of opioids has also increased, but in smaller proportions compared to North America. However, the extent to which opioids are (mis)used in Europe needs to be further explored. In this project, we will optimise and develop a highthroughput bioanalytical assay based on μSPE-LC-MS/MS capable of measuring a broad range of biomarkers of opioids use in the ng/L range in wastewater. We aim at obtaining near real-time, objective and complementary information on the use of opioids at high spatio-temporal resolution through a wastewater sampling campaign in different locations in Belgium. By generating highly relevant data on the consumption of opioids together with triangulation of this data with other existing datasets (e.g. Farmanet, Health Interview Survey,...), this project can aid in obtaining a better picture on the opioid use in Belgium which is of importance for policy makers in optimizing and evaluating prevention, treatment and harm-reduction strategies on the subject of pain management. Furthermore, by combining multiple datasets, information on the illegal use of opioids can be obtained. In a last step, the obtained data will be correlated with existing socioeconomicco-variates of the investigated communities to obtain a better insight on the relationship between socio-demographic features, and prevalence of use of opioids.

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Comprehensive Liquid Chromatography - Ion Mobility - Quadrupole-Time-of-Flight Mass Spectrometry for innovative metabolomics. 01/05/2018 - 30/04/2021

Abstract

The requested infrastructure (comprehensive liquid chromatograph-ion mobility-quadrupole time of flight mass spectrometer LCxLC-IM-QTOFMS) combines several state-of-the-art technologies into one platform which aims at bringing metabolomics research to the next level. As such, the infrastructure will deliver a combined five-dimension separation and detection technology, the first of its kind in Belgium. This instrument will be dedicated to metabolomics research, the science of endogenous metabolites in cells, tissues or organisms. The infrastructure will be able to optimally separate, detect and identify the very broad and complex chemical space of metabolites ranging from very polar (e.g. amino acids) to non-polar (e.g. lipids and hormones) at low nanomolar concentration range. Within UA, there is a growing need to combine the currently scattered efforts in metabolomics, an Emerging Frontline Research Domain in the UA research scene. Research ranges from drug discovery (mode of action and pharmacokinetic profiling), biomarker and toxicity studies to advanced data-analysis and systems biology approaches, but a dedicated metabolomics infrastructure to strengthen these studies is currently missing. As such, the investment in a core facility together with the gathering of nine research groups from five departments and two faculties would centralize the metabolomics research. This will position UA as a key player in the academic metabolomics research in the BeNeLux and worldwide.

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Chemical information mining of wastewater – From human exposure to environmental fate of emerging plasticizers and flame retardants 01/10/2017 - 12/10/2018

Abstract

We and our surrounding environment are constantly and increasingly exposed to synthetic chemicals, some of which have been shown to cause adverse biological or toxicological effects. Whilst the toxicity of some chemicals has been recognised, leading to their ban or regulated use, many substitutes, for which toxicity has not yet been fully assessed, are continuously being introduced. Consequently, monitoring human exposure and environmental fate of future/emerging contaminants is crucial. In this regard, wastewater plays a central role. Being, on the one hand, a pooled diluted sample of human urine, its composition can provide valuable information about the types and amounts of contaminants to which humans have been exposed to. On the other hand, it is recognised as one of the main sources of contaminants for the environment. This project aims at mining the chemical information contained in wastewater to obtain valuable geographical and temporal information about the exposure of humans to emerging contaminants, as well as to understand the environmental fate of these compounds. (i) Dedicated analytical methods will be developed to identify human metabolites of targeted emerging contaminants (plasticizers and flame retardants). (ii) Fate of these chemicals during wastewater treatment and their occurrence in wastewater of Flemish cities will be investigated. (iii) The gathered data will then be used to model community-wide exposure to these contaminants.

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Assessing population health from exposure to tobacco-specific carcinogens in Belgium using an innovative wastewater-based epidemiology approach (APOLLO). 01/04/2017 - 31/03/2019

Abstract

Tobacco smoking is the major cause of many chronic diseases, especially cancer. Knowledge on population exposure to tobacco smoking is essential to characterise the population health. National questionnaire surveys are currently the main source of population tobacco smoking. However, surveys have clear limitations due to its dependence on true self-reported usage and representativeness of the whole population. International comparison of surveyed data is also restricted due to different questionnaire formats among nations. An additional approach allowing more objective, efficient and near real-time assessment is necessary to address this important societal issue. Hence, the project APOLLO aims at developing and validating an innovative approach, wastewater-based epidemiology (WBE), for assessing population exposure to tobacco and its related carcinogens in the community. This involves analysis of raw wastewater from targeted communities for tobacco-specific biomarkers of exposure and cancer risk (the tobacco-specific nitrosamines (TSNAs) and their metabolites). The project comprises three main objectives: (1) development and validation of the WBE methodology; (2) evaluation of TSNA and metabolite levels in wastewater from different Belgian communities; (3) assessment of population health from exposure to tobacco-specific carcinogens in Belgium. The completion of the project will advance our knowledge on factual and near-real-time assessment of population exposure to these carcinogens from community to national and international levels using WBE as a standardised approach. This will be also a valuable tool to evaluate the efficacy of intervention actions against tobacco smoking and its related diseases in the population and specific community e.g. prisons, high schools. This project will promote my career prospect with enhanced interdisciplinary network and research mobility in Europe.

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Synergising international research Studies into the environmental fate and behaviour of toxic organic chemicals in the waste stream (INTERWASTE). 01/01/2017 - 31/12/2021

Abstract

The overall vision of INTERWASTE is to develop scientific understanding of issues related to environmental contamination with toxic organic chemicals (specifically flame retardants (FRs) and pharmaceutical and personal care products (PPCPs)) arising from their presence in the waste stream. Specific research objectives include: (a) exchange of knowledge of and best practice in methods for rapid and cost-effective identification of waste items containing restricted FRs; (b) developing scientific understanding of environmental contamination due to processing of waste items containing FRs; and (c) furthering understanding of the sources of PPCPs and FRs in the sewerage system.

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Unraveling the influence of diseases on the metabolism of xenobiotics by the gut microbiome. 01/01/2017 - 31/12/2017

Abstract

The human lives in symbiosis with millions of micro-organisms. A great amount of bacteria is located in the human gut. These micro-organisms, in combination with their unique genes, are called the gastrointestinal microbiome. Only recently the role of the intestinal microbiome in the formation of metabolites of orally ingested compounds has become evident. Fundamental questions concerning the role of the gut microbiome however remain unanswered for the majority of therapeutic, toxic and diet-derived xenobiotics. The composition of the gut microbiome is quite stable in healthy adults and contributes to the maintenance of a healthy state in adulthood. On the other hand, several pathological states are characterized by a modified composition of the intestinal microflora as already demonstrated in obesity and type 2 diabetes. Therefore, the aim of the project is to develop an integrated experimental platform to identify biotransformation due to the microbiome and to evaluate the influence of different disorders (type 2 diabetes, obesity) on this 'microbiotic metabolism'. The fate of the test compounds (simvastatin, tetrabromobisphenol A and epigallocatechin gallate) in a validated in vitro gastrointestinal dialysis system with a colon phase and in well-characterized phenotypes of healthy people and obese and type 2 diabetic patients, will be studied. Detection and identification of the metabolites will be achieved by advanced analytical techniques.

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Waste water analysis report on the stimulant illicit drug markets in the EU (WATCH). 20/12/2016 - 19/06/2018

Abstract

The aims of this project are: 1) Conduct a wastewater monitoring exercise utilising a standardized protocol to describe the European illicit stimulant drug market through city level studies. Explore within countries differences in city level wastewater estimates to provide a better understanding of geographical patterns between and within European countries. 2) In a sample of selected cities conduct exploratory pilot studies with the collection of repeat waste water samples over a sufficiently long time period to assess temporal variations in consumption patterns and explore the implications of this for sampling and analysis for routine monitoring purpose. 3) Develop models of drug consumption that incorporate waste water analysis with other data sources (such as drug purity, epidemiology and demographical information) and assess the utility of these for commenting on the following questions: a) understanding better illicit stimulant drug consumption patterns at the community level, b) improving market size estimates c) coherence between estimates derived from different data sources, d) if changes in the illicit stimulant drug market resulting from changing availability, purity or police activities are possible to detect in wastewater analysis. 4) Conduct a review and pilot study in selected cities to assess the possible utility of waste water analysis to report on synthetic drug production

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The impact of endocrine disruption on vertebrate embryonic and larval development. 01/10/2015 - 30/09/2019

Abstract

Endocrine disruption is a major concern for the health of wildlife populations. Although many studies have shown reproductive impairment as a consequence of endocrine disruption in adult fish, knowledge of the consequences of endocrine disruption for vertebrate embryonic and larval development is scarce. The zebrafish embryo is an ideal model system to investigate the fundamental mechanisms of endocrine disruption. We will first describe the timing of the normal embryonic activation of the hormone synthesis machinery, as well as the hormone profiles, during the early stages of vertebrate development, which has never been done so far. Secondly, using well-described endocrine disrupting compounds, we will specifically disrupt these profiles and propose mechanisms linking the changed profiles to observed developmental effects. We will then validate the proposed mechanisms of toxicity using targeted disruption techniques, including knockout and morpholino knockdown. This project will offer the information that is needed for follow-up projects to develop assays to specifically assess the risk associated with exposure to different classes of EDCs on embryonic and larval development, allowing discrimination among ER and AR agonism and antagonism, as well as aromatase inhibition. Such assays would fit perfectly within both EU and USA regulatory priorities for screening and prioritizing potential EDCs.

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The rise and threat of new psychoactive substances: integrated research to obtain evidence-based information on their properties and actual use. 01/10/2015 - 30/09/2018

Abstract

In recent years increasing amounts of new psychoactive substances (NPS) are detected on the drug market. These substances are chemically based on known illicit drugs (e.g. amphetamine, cannabis), have comparable effects, and are designed specifically to avoid drug regulations. The number of NPS detected keeps rising at a high rate with more than one new NPS reported every week in 2013 in the EU. In most cases, no information whatsoever is available regarding their metabolic aspects, effects, and toxicity, and this has resulted in a significant number of deaths worldwide. Furthermore, solid epidemiological data on the extent of use NPS use is lacking. This project aims at obtaining objective and timely information on the use of NPS in Belgium through a three-tiered approach. In first instance, an evaluation of the Belgian drug market will be made for the presence of NPS through a combination of amnesty bin analysis, purchase analysis and information obtained from the Belgian Early Warning System Drugs. The relevant NPS identified from this research will be subjected to in vitro metabolism experiments in order to gather scientific information on their metabolic fate and to identify biomarkers to target in biomonitoring studies. Biomonitoring will be performed on different population levels through wastewater analysis (communities), pooled urine analysis (specific populations) and individual serum/urine samples in order to obtain data on the extent of NPS use in Belgium.

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Analysis of kynurenines in serum of bipolar patients and their association with affective and cognitive symptoms. 01/02/2015 - 31/12/2015

Abstract

This project will evaluate the association of an upregulated kynurenine pathway with the clinical symptomatology in bipolar disorder (BD). Therefore, an analytical method based on liquid chromatography-mass spectrometry will be developed for the detection of kynurenines in serum. The method will be applied on samples from BD patients in parallel with mood state monitoring. Project results could aid in a better understanding of BD pathophysiology.

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Identification of new psychoactive substances and their metabolites through the analysis of pooled urine and wastewater. 01/10/2014 - 30/09/2015

Abstract

An increasing number of "New Psychoactive Substances" (NPS), claiming to contain only "legal" compounds, has recently appeared on the drug market. One of the major concerns depends on the lack of suitable analytical methods to identify and also to quantify the NPS entering the market. Even using sophisticated analytical techniques, these new drugs may be missed, since reference standards are not always available. In addition, NPS are often extensively metabolized in the body making almost undetectable the parent compounds, which highlights the importance of identifying their metabolites. In the current proposal, we will analyze several suspicious drug preparations by taking advantage of a high-resolution/accurate-mass (HR/AM) spectrometer quadrupole/time-of-flight (QTOF) in order to achieve structural elucidation of their main active components. Metabolites of the identified NPS will be then obtained by in vitro human liver preparations and further characterized by high-performance-liquid chromatography (HPLC) coupled to QTOF mass spectrometry. Next, we will perform an approach based on "sewage epidemiology" to estimate the consumption of NPS via the analysis of NPS and their metabolites in wastewater. For this purpose, we will develop and validate an analytical method based on solid-phase extraction (SPE) and HPLC coupled to tandem mass spectrometry (MS/MS) for accurate analysis of identified NPS and their metabolites in both wastewater and pooled urine samples.

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Expanding the detection window in forensic toxicology: hair and nail analysis. 01/01/2014 - 31/12/2014

Abstract

This project represents a research agreement between the UA and on the onther hand IWT. UA provides IWT research results mentioned in the title of the project under the conditions as stipulated in this contract.

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Sewage analysis as an alternative methodology to estimate drug use in general population. 13/11/2013 - 31/12/2014

Abstract

This project represents a research contract awarded by the University of Antwerp. The supervisor provides the Antwerp University research mentioned in the title of the project under the conditions stipulated by the university.

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Analysis of ethyl glucuronide in hair as biomarker of alcohol intake. 01/02/2013 - 31/12/2013

Abstract

The harmful use of alcohol is a major global contributing factor to death, disease and injury. A key step in the development of effective policies and interventions is an early diagnosis of the hazardous use of alcohol. The analysis of ethyl glucuronide (EtG) in hair has been suggested from the mid 2000s as a candidate marker for the intake of alcohol over several months and can be used in specific cases such as alcohol withdrawal treatment programs, driving license regranting, and excluding alcohol use during and after liver transplantation. This project aims at optimizing and validating an analytical procedure based on chromatography and mass spectrometry for the measurement of ethyl glucuronide in hair. The developed analytical procedure will be applied to hair samples from liver transplantation patients to detect alcohol use before and after the transplantation in order to prevent failure of the transplantation process.

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A new paradigm in drug use and human health risk assessment: sewage profiling at the community level (SEWPROF). 01/10/2012 - 30/09/2016

Abstract

SEWPROF aims to develop interdisciplinary and cross-sectoral research capability for the next generation of scientists working in the newly-emerging fileld of sewage epidemiology. It will provide an integrated approach towards public health monitoring at a community level based on innovative sewage epidemiology techniques.

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Refinement and validation of the sewage epidemiology approach to estimate illicit drug use in the general population. 01/10/2012 - 30/09/2015

Abstract

In this post-doctoral mandate, I will validate and refine an innovative approach for estimating illicit drug use in the general population based on sewage analysis for human excretion products of these substances ("sewage epidemiology"). In the first objective of this proposal, I will validate the approach by simultaneously executing a drug consumption questionnaire (the gold standard at this moment) and sewage epidemiology for a well-defined population and time period. The data resulting from both approaches will be compared and can serve as a validation of sewage epidemiology. The second objective focuses on refining sewage epidemiology back-calculations, by addressing the issue of estimating the population contributing to a specific sewage system in an objective, dynamic and real-time way. Different approaches for estimating the real amount of people contributing to a specific sewage system will be based on the analysis of specific markers in sewage that result from human activity. The last objective deals with applying sewage epidemiology for new and innovative purposes, such as the exploratory investigation of the use of new and emerging substances and through the execution of uniformed Europe-wide monitoring studies that allow reliable international comparisons on the epidemiology of illicit drug use. It is expected that the successful completion of these objectives will have positive implications for international drug policy and public health.

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Estimation of drug abuse in Belgium by analysis of wastewater. 01/10/2010 - 30/09/2012

Abstract

This project aims at the development of models to estimate the use of illicit drugs in Belgium by means of the analysis of illicit drugs and their metabolites in wastewater ("sewage epidemiology"). Sewage epidemiology can deliver important and real-time information about local, regional, national and international illicit drug consumption which can be used in addition to classical socio-epidemiological studies to give policy makers a better picture about trends, patterns and hot-spots of illicit drug use.

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Evaluation of drug abuse in Belgium by analysis of wastewater. 01/10/2008 - 30/09/2010

Abstract

This projects consists of two parts. In the first part, methods of analysis for the determination of several drugs (cocaine, amphetamines, cannabis,...) and their metabolites in waste and surface water are being developped and validated. In the second part, the concentrations of drugs and metabolites found in watersamples will be used to estimate the abuse of drugs in Belgium.

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Estimation of the use of illicit drugs through analysis of waste water. 01/10/2007 - 30/09/2008

Abstract

This projects consists of two parts. In the first part, methods of analysis for the determination of several drugs (cocaine, amphetamines, cannabis,...) and their metabolites in waste and surface water are being developped and validated. In the second part, the concentrations of drugs and metabolites found in watersamples will be used to estimate the abuse of drugs in Belgium

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