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The aim of this study is to utilize novel and powerful workflows with publicly available tools to efficiently process data and facilitate rapid acquisition of knowledge on environmental fate studies. Taking imatinib (IMA) as an example, we developed an efficient workflow to describe IMA biodegradation with activated sludge (AS) from wastewater treatment plants (WWTP). IMA is a cytostatic pharmaceutical; a selective tyrosine kinase inhibitor used to treat chronic myeloid leukemia. Its reported ecotoxic, endocrine and genotoxic effects imply high risk for aquatic wildlife and human health, however its fate in the environment is not yet well known. The study was conducted in a batch biotransformation setup, at two AS concentration levels and in presence and absence of carbon source. Degradation profiles and formation of IMA transformation products (TPs) were investigated using UHPLC-QqOrbitrap-MS/MS which showed that IMA is readily biodegradable. TPs were determined using multivariate statistical analysis. Eight TPs were determined and tentatively identified, six of them for first time. Hydrolysis of amide bond, oxidation, demethylation, deamination, acetylation and succinylation are proposed as major biodegradation pathways. TP235, the product of amide bond hydrolysis, was detected and quantified in actual wastewaters, at levels around 1 ng/L. This calls for more studies on the environmental fate of IMA in order to properly asses the environmental risk and hazard associated to IMA and its TPs., This work has received funding from the European Commission through the projects CytoThreat (265264) and SOLUTIONS (603437), from Slovenian Research Agency - ARRS program P1-0143: Cycling of substances in the environment, mass balances, modelling of environmental processes and risk assessment, ARRS projects J1-8140 (Endocrine disrupting and genotoxic potential of protein kinase inhibitors: relevance for environmental hazard and human health) and J1-6744 (Development of Molecularly Imprinted Polymers and their application in environmental and bio-analysis), the Spanish Ministry of Science and Innovation (Project CEX2018-000794-S), and the Generalitat de Catalunya (Consolidated Research Group 2017 SGR 01404-Water and Soil Quality Unit). Žiga Tkalec acknowledges the Slovenian Research Agency – ARRS for the Young researcher grant. The work reflects only the authors' views. The Community in not liable for any use that may be made of the information contained therein. The authors thank Vesna Mislej from Central Wastewater Treatment Plant in Ljubljana and Marjetka Levstek from Wastewater treatment plant Domžale-Kamnik for providing wastewater samples. Special thanks to Metka Filipič from the National institute of biology for providing the imatinib consumption data.

This study analyses the presence of 17 cytostatic agents from seven different groups, based on their different mechanisms of action, in the effluent from a medium-sized hospital located in eastern Spain. Analysis of the compounds found in the effluents studied involved solidphase extraction (SPE) coupled on-line to a high performance liquid chromatograph tandem mass spectrometer (HPLC-MS/MS). The environmental risk of the compounds studied was then assessed by calculating the hazard quotient (HQ), combining the measured environmental concentrations (MECs) with dose-response data based on the predicted no effect concentrations (PNECs). In addition, the environmental hazard associated was evaluated in accordance with their intrinsic characteristics by calculating the PBT (Persistence Bioaccumulation Toxicity) index. The results of this study showed the presence of seven of the 17 compounds analysed in a range of between 25 and 4761 ng/L. The highest concentrations corresponded to ifosfamide (58-4761 ng/L), methotrexate (394-4756 ng/L) and cyclophosphamide (46-3000 ng/L). Assessment of the environmental hazard showed that the three hormonal agents (tamoxifen and its metabolites endoxifen and hydroxytamoxifen) exhibited a maximum PBT value of 9 due to their inherent harm to the environment resulting from their characteristics of persistence, bioaccumulation and toxicity. A combined evaluation of the risk and environmental hazard showed that three of the 17 compounds studied, namely, ifosfamide, imatinib and irinotecan, all of which exhibited HQ values higher than 10 and PBT indices of 6, indicative of a particularly high potential to harm the environment, deserve special attention.

The growing evidence supporting a link between exposure to the naturally occurring toxin β-N-methylamino-l-alanine (BMAA) and progressive neurodegenerative diseases, has recently arisen the interest of the scientific community. Latest investigations suggest that dietary exposure to this algal toxin may have been largely underestimated. This paper reviews the state of the art regarding BMAA, with special attention paid to its neurotoxicity, its concentration levels in food, and human exposure. As for other environmental toxins, dietary intake is most likely the main route of exposure to BMAA for the general population. However, data concerning BMAA levels in foodstuffs are still scarce. It is concluded that further investigations on dietary intake and potential human health effects are clearly necessary to assess the risks to public health associated with BMAA exposure. Some critical remarks and recommendations on future research in this area are provided, which may help to identify approaches to reduce dietary BMAA exposure.

Liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QToF-MS) was used for the accurate identification (

The current work reports the development and validation of a qualitative screening method for 156 pesticides in Atlantic salmon feeds based on the use of ultrahigh-performance liquid chromatography (UHPLC) coupled to traveling-wave ion mobility spectrometry (TWIMS)/quadrupole time-of-flight mass spectrometry (QTOFMS). This novel hyphenation allows a straightforward estimation of the collision cross section (CCS), a physicochemical property that recently has been proposed as an additional identification point for pesticides. In order to increase the overall efficiency of the screening workflow, data processing parameters and tolerances involved in the automated analyte detection were first optimized. The use of the CCS as an additional identification point greatly improved the method selectivity, decreasing the rate of false-positive detections in blank feed samples. Qualitative validation was then performed in accordance with the EU document SANTE/11945/2015 on analytical quality control and validation procedures for pesticide residues analysis in food and feed. Feed samples were spiked with 156 pesticides at three levels (0.01, 0.05 and 0.20 mg/kg) and analyzed by the proposed method. Under the optimized conditions, screening detection limits (SDLs) ≤0.05 mg/kg were achieved for 82% of the investigated pesticides. The applicability of this new approach for the screening of pesticides was finally proven by analyzing several commercial Atlantic salmon feed samples as well as various plant-based feed ingredients. Pirimiphos-methyl and ethoxyquin were among the most frequently detected analytes in the investigated samples.

The purpose of this work was to investigate the in vitro metabolism of nitracaine, a new psychoactive substance, using human liver microsome incubations, to evaluate the cytochrome P450 (CYP) enzyme isoforms responsible for the phase-I metabolism and to compare the information from the in vitro experiments with data resulting from an authentic user's urine sample. Accurate mass spectra of metabolites were obtained using liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) and were used in the structural identification of metabolites. Two major and three minor phase-I metabolites were identified from the in vitro experiments. The observed phase-I metabolites were formed through N-deethylation, N,N-deethylation, N-hydroxylation, and de-esterification, with CYP2B6 and CYP2C19 being the main enzymes catalyzing their formation. One glucuronidated product was identified in the phase-II metabolism experiments. All of these metabolites are reported for the first time in this study except the N-deethylation product. All the in vitro metabolites except the minor N,N-deethylation product were also present in the human urine sample, thus demonstrating the reliability of the in vitro experiments in the prediction of the in vivo metabolism of nitracaine. In addition to the metabolites, three transformation products (p-nitrobenzoic acid, p-aminobenzoic acid, and 3-(diethylamino)-2,2-dimethylpropan-1-ol) were identified, as well as several glucuronides and glutamine derived of them.

Ethoxyquin (EQ) is an additive present in fish feed and its fate in fish should be carefully characterized due to food safety concerns regarding this compound. Therefore, the objective of this work was to identify the transformation products (TPs) of EQ in Atlantic salmon. Salmon in independent tanks were given feed containing ethoxyquin concentrations of 0.5 mg/kg, 119 mg/kg or 1173 mg/kg for 90 days. After the feeding trial, salmon fillets were extracted in acetonitrile and analyzed by liquid chromatography with traveling-wave ion mobility spectrometry coupled to high resolution mass spectrometry (UHPLC-TWIMS-QTOFMS). EQ was transferred from the feed to salmon fillets and 23 TPs were characterized, resulting from dimerization, oxygenation, cleavage, cleavage combined with oxygenation, cleavage combined with conjugation, and other uncategorized alterations. Moreover, EQ and some TPs were also detected in commercial salmon randomly sampled from different Norwegian fish farms. This study confirmed that the dimer 1,8′-EQDM was the main TP of EQ and, together with previous research, brought the overall number of characterized TPs to a total of 47.

Concerns about the presence of anticancer drugs in the environment are rapidly increasing mainly due to their growing use in the developed countries and their known cytotoxic effects. Vinca alkaloids are widely used in cancer therapy; however, very scarce information is available on their occurrence, environmental fate and toxicological effects on aquatic organisms. Even less attention has been paid to their potential transformation products, which can exert higher toxicity than the parent compounds. Thus, in the present work, the reactivity of vincristine, vinblastine, vinorelbine and its metabolite 4- O -deacetyl vinorelbine during water chlorination processes has been investigated for the first time. Under the studied chlorination conditions, vincristine was fairly stable whereas vinblastine, vinorelbine and 4- O -deacetyl vinorelbine were quickly degraded. A total of sixty-five disinfection by-products were tentatively identified by ultra-high performance liquid chromatography coupled to high-resolution hybrid quadrupole-Orbitrap tandem mass spectrometry. Among them, twenty by-products corresponded to mono-chlorinated compounds, eight to di-chlorinated compounds and two to tri-chlorinated compounds, which may be of major environmental concern. Other disinfection by-products involved hydroxylation and oxidation reactions. Although the structures of these by-products could not be positively confirmed due to lack of commercial standards, their chemical formulas and product ions can be added to databases, which will allow their screening in future monitoring studies.

The results of an inter-laboratory comparison exercise to determine cytostatic anticancer drug residues in surface water, hospital wastewater and wastewater treatment plant effluent are reported. To obtain a critical number of participants, an invitation was sent out to potential laboratories identified to have the necessary knowledge and instrumentation. Nine laboratories worldwide confirmed their participation in the exercise. The compounds selected (based on the extent of use and laboratories capabilities) included cyclophosphamide, ifosfamide, 5-fluorouracil, gemcitabine, etoposide, methotrexate and cisplatinum. Samples of spiked waste (hospital and wastewater treatment plant effluent) and surface water, and additional non-spiked hospital wastewater, were prepared by the organising laboratory (Jožef Stefan Institute) and sent out to each participant partner for analysis. All analytical methods included solid phase extraction (SPE) and the use of surrogate/internal standards for quantification. Chemical analysis was performed using either liquid or gas chromatography mass (MS) or tandem mass (MS/MS) spectrometry. Cisplatinum was determined using inductively coupled plasma mass spectrometry (ICP-MS). A required minimum contribution of five laboratories meant that only cyclophosphamide, ifosfamide, methotrexate and etoposide could be included in the statistical evaluation. z-score and Q test revealed 3 and 4 outliers using classical and robust approach, respectively. The smallest absolute differences between the spiked values and the measured values were observed in the surface water matrix. The highest within-laboratory repeatability was observed for methotrexate in all three matrices (CV ≤ 12 %). Overall, inter-laboratory reproducibility was poor for all compounds and matrices (CV 27-143 %) with the only exception being methotrexate measured in the spiked hospital wastewater (CV = 8 %). Random and total errors were identified by means of Youden plots.

Residues of anti-neoplastic drugs represent new and emerging pollutants in aquatic environments. Many of these drugs are genotoxic, and it has been postulated that they can cause adverse effects in aquatic ecosystems. 5- Fluorouracil (5-FU) is one of the most extensively used anti-neoplastic drugs in cancer therapy, and this article describes the results of the first investigation using a two-generation toxicity study design with zebrafish (Danio rerio). Exposure of zebrafish to 5-FU (0.01, 1.0 and 100 mg/L) was initiated with adult zebrafish (F0 generation) and continued through the hatchings and adults of the F1 generation, and the hatchings of the F2 generation, to day 33 post-fertilisation. The exposure did not affect survival, growth and reproduction of the zebrafish ; however, histopathological changes were observed in the liver and kidney, along with genotoxic effects, at all 5- FU concentrations. Increases in DNA damage determined using the comet assay were significant in the liver and blood cells, but not in the gills and gonads. In erythrocytes, a significant, dose-dependent increase in frequency of micronuclei was observed at all 5- FU concentrations. Whole genome transcriptomic analysis of liver samples of F1 generation zebrafish exposed to 0.01 mg/L and 1 mg/L 5-FU revealed dose-dependent increases in the number of differentially expressed genes, including up-regulation of several DNA-damage- responsive genes and oncogenes (i.e., jun, myca). Although this chronic exposure to environmentally relevant concentrations of 5- FU did not affect the reproduction of the exposed zebrafish, it cannot be excluded that 5-FU can lead to degenerative changes, including cancers, which over long- term exposure of several generations might affect fish populations. The data from this study contribute to a better understanding of the potential consequences of chronic exposure of fish to low concentrations of anti- neoplastic drugs, and they demonstrate that further studies into multi-generation toxicity are needed.

Recent studies have demonstrated the role of biofilms on the stability of drug residues inwastewater. These factors are pertinent in wastewater-based epidemiology (WBE) when estimating community-level drug use. However, there is scarce information on the biotransformation of drug residues in the presence of biofilms and the potential use of transformation products (TPs) as biomarkers in WBE. The purpose of this work was to investigate the formation of TPs in sewage reactors in the presence of biofilm mimicking conditions during in-sewer transport. Synthetic cathinones (methylenedioxypyrovalerone, methylone, mephedrone) and phenethylamines (4-methoxy-methamphetamine and 4-methoxyamphetamine) were incubated in individual reactors over a 24 h period. Analysis of parent species and TPs was carried out using liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QToFMS). Identification of TPs was done using suspect and non-target workflows. In total, 18 TPs were detected and identified with reduction of beta-keto group, demethylenation, demethylation, and hydroxylation reactions observed for the synthetic cathinones. For the phenethylamines, N- and O-demethylation reactions were identified. Overall, the experiments showed varying stability for the parent species in wastewater in the presence of biofilms. The newly identified isomeric forms of TPs particularly for methylone and mephedrone can be used as potential target biomarkers for WBE studies due to their specificity and detectability within a 24 h residence time. (C) 2018 Elsevier B.V. All rights reserved.

Once discharged into the sewage system, many pharmaceuticals may undergo degradation reactions in the presence of chemical disinfectants, generating by-products that may possess enhanced toxicity relative to the parent compounds. For this reason, the stability of the widely used cytostatic etoposide in chlorinated water has been investigated for the first time in the present work. Taking advantage of the high-resolution/accurate-mass capabilities of the hybrid quadrupole-Orbitrap mass spectrometer Q Exactive, two new oxidation by-products of etoposide were reliably identified. The time course of etoposide and its by-products was followed at different pH values, free chlorine concentrations and water matrices. Finally, the occurrence of etoposide and its major identified by-product (3'-O-desmethyl etoposide) was investigated in real water samples by on-line solid-phase extraction-liquid chromatography-tandem mass spectrometry using a 4000QTRAP hybrid quadrupole-linear ion trap mass spectrometer. The etoposide by-product was found in various river and wastewater samples at levels between 14 and 33ngL-1, whereas etoposide was not detected in any sample This study has been financially supported by the EU through the FP7 projects CytoThreat (265264) and SOLUTIONS (603437), by the Spanish Ministry of Economy and Competitiveness through the project SCARCE (Consolider-Ingenio 2010 CSD2009-00065), and by the Generalitat de Catalunya (Consolidated Research Groups “2014 SGR 418—Water and Soil Quality Unit” and 2014 SGR 291—ICRA)

A comprehensive and efficient method is presented for the determination of 9 polybrominated diphenyl ether congeners (PBDEs), 2,4,4′-tribromodiphenyl ether (PBDE-28), 2,3′,4,4′-tetrabromodiphenyl ether (PBDE-47), 2,2′,4,4′,6-pentabromodiphenyl ether (PBDE-66), 2,2′,4,4′,5-pentabromodiphenyl ether (PBDE-100), 2,2′,3,4,4′-pentabromodiphenyl ether (PBDE-99), 2,2′,4,4′,5,6′-hexabromodiphenyl ether (PBDE-85), 2,2′,4,4′,5,5′-hexabromodiphenyl ether (PBDE-154), 2,2′,4,4′,5,5′-hexabromodiphenyl ether (PBDE-153) and 2,2′,3,4,4′,5,6′-heptabromodiphenyl ether (PBDE-183), in mollusc bivalve samples. An experimental design was used to evaluate the influence of several operational parameters on the extraction performance and to obtain the best experimental conditions. Under the final working conditions, freeze-dried mollusc was placed in the Soxtec thimble in order to extract PBDEs with 100 mL of n-hexane-acetone (4: 1) after 1.5 h and 1 h of boiling and rinsing times, respectively. Soxtec extracts were additionally purified on a multilayered column and after injected in the gas chromatography combined with mass spectrometry (GC-MS) system. Quantitative recoveries of the analytes at three concentration levels ranged between 80 and 108%, and limits of quantification of the method (LOQs) between 0.5 and 20 ng/g. Analysis of mussel and clam samples showed that PBDE-47 and PBDE-99 were presented in almost all samples with concentrations between 0.6 and 2.6 ng/g dry weight. Moreover, a certified reference material (NIST 2974a) was used. The obtained concentrations were in agreement with the certified values.

Polyfluoroalkyl phosphate esters (PAPs) are high production volume surfactants used in the food contact paper and packaging industry. PAPs may transform to persistent perfluoroalkyl carboxylic acids (PFCAs) under biotic conditions, but little is known about their fate and behavior in aquatic organisms. Here we report for the first time on the uptake, tissue distribution, and biotransformation of 8:2 polyfluoroalkyl phosphate diester (8:2 diPAP) in fish. Gilt-head bream (Sparus aurata) were dosed via the diet (8:2 diPAP at 29 mu g/g) for 7 days, during which time 8:2 diPAP and its transformation products were monitored in plasma, liver, muscle, gills, bile and brain. 8:2 diPAP tended to accumulate in liver, plasma and gills, and to a lesser extent in muscle, bile and brain. Several transformation products (observed previously in other organisms) were also observed inmost tissues and biofluids, including both saturated and unsaturated fluorotelomer acids (8:2 FTCA, 8:2 FTUCA, 7:3 FTCA), and perfluorooctanoic acid (PFOA). 8:2 FTCA was the major metabolite in all tissues/biofluids, except for bile, where PFOA occurred at the highest concentrations. Unexpectedly high PFOA levels (up to 3.7 ng/g) were also detected in brain. Phase 2 metabolites, which have been reported in fish following exposure to fluorotelomer alcohols, were not observed in these experiments, probably due to their low abundance. Nevertheless, the detection of PFOA indicates that exposure to PAPs may be an indirect route of exposure to PFCAs in fish. (C) 2017 Elsevier B.V. All rights reserved.

Concerns about cytostatic anticancer drugs in the environment are increasing, mainly due to the lack of knowledge about the fate and impact of these cytotoxic compounds in the water cycle. In this context, the present work investigated the occurrence of 13 cytostatics and 4 metabolites in wastewater samples from various wastewater treatment plants (WWTPs) and from a large hospital from Spain. The target compounds belong to five different classes according to the Anatomical Therapeutic Classification (ATC), namely, alkylating agents, antimetabolites, plant alkaloids and other natural products, cytotoxic antibiotics and related substances, and other antineoplastic agents. Some of them have been classified as carcinogens in humans by the International Agency for Research on Cancer (IARC). These compounds were determined by an automated on line solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) method. Results showed the presence of methotrexate (MET), ifosfamide (IF), cyclophosphamide (CP), irinotecan (IRI), doxorubicin (DOX), capecitabine (CAP), tamoxifen (TAM) and the metabolites endoxifen (OH-D-TAM), hydroxytamoxifen (OH-TAM) and hydroxypaclitaxel (OH-PAC) at levels ranging from 2 ng L(-1) (for MET) to 180 ng L(-1) (for TAM). Some of these compounds were found to be efficiently removed after wastewater treatment, e.g. MET, DOX and IRI, whereas other compounds, such as TAM, CP and IF remained largely unaltered. The behaviour of the target compounds during the common filtration step of the water samples was also investigated with the finding that some compounds are strongly adsorbed to nylon filters, while cellulose acetate appears as the best choice for the filter material. The aquatic environmental risk associated to the detected compounds was also assessed. To the best of the authors' knowledge, this is the first report of the presence of the metabolites OH-D-TAM and OH-TAM in the water cycle.

Grapefruit is a rich source of flavonoids but also contains furanocoumarins, which are known to strongly interact with a variety of medications. Thus, characterization of grapefruit furanocoumarin metabolites may help in a better understanding of grapefruit-drug interactions. In the present work, identification of the main metabolites of grapefruit juice furanocoumarins in urine was performed by ultraperformance liquid chromatography (UPLC) coupled to linear ion trap-Orbitrap mass spectrometry (LTQ-Orbitrap). Glucuronides of 6',7'-dihydroxybergamottin and a hydroxybergamottin-like metabolite were identified for the first time as grapefruit juice metabolites. Afterward, a fast and sensitive method based on solid-phase extraction (SPE) and UPLC coupled to triple quadrupole-tandem mass spectrometry (QqQ-MS/MS) was developed for determination of the identified metabolites in urine. The proposed method was applied to urine samples of five volunteers after intakes of moderate doses of grapefruit, lemon, and orange juices. Furanocoumarin metabolites were only detected in urines after consumption of grapefruit juice.

A low solvent consumption method for the determination of eight ultraviolet (UV) filters, displaying low to medium polarities, in freeze-dried samples of marine bivalves and fish is proposed. Matrix solid-phase dispersion (MSPD) and gas chromatography with mass spectrometry (GC-MS) were used as sample preparation and determination techniques, respectively. This work describes the influence of several parameters (type and amount of dispersant and clean-up sorbents, as well as elution solvent) on the yield and the selectivity of the MSPD extraction. Under optimized conditions, samples (0.5 g) were ground with 2 g of Florisil in a mortar with a pestle and transferred into a polypropylene syringe, which contained 1 g of C18 as clean-up sorbent. Analytes were eluted with 5 mL of acetonitrile. This extract was concentrated to dryness, re-constituted with 1 mL of ethyl acetate and injected in the GC-MS system without any further clean-up. The global average recoveries, measured for three different biota samples,...

A multianalyte liquid chromatography-electrospray-tandem mass spectrometry (LC-ESI-MS/MS) method for determination of 19 cytostatics and 5 metabolites, from 6 different therapeutic families, has been developed, and the structures of the main characteristic fragment ions have been proposed. Instrumental limits of detection and quantification are in the range 0.1-10.3 and 1.0-34.3 ng mL−1, respectively. Moreover, the stability of the compounds in aqueous solution was investigated in order to establish the best conditions for preparation and storage of both calibration standards and water samples. Dimethylsulfoxide (DMSO) was selected as solvent for preparation of the stock solutions. At room temperature (25 ºC), 11 of the 24 target compounds were shown to be unstable in water (percentage of organic solvent 4%), with concentration losses greater than 20% in less than 24 h. At 4 ºC (typical storage temperature for water samples) all compounds, except MTIC and chlorambucil, were stable for 24 h, but the number of stable compounds decreased to 10 after 9 days. Freezing of the aqueous solutions improved considerably the stability of various compounds: after 3 months of storage at -20 ºC, 10 compounds, namely, 5-fluorouracil, carboplatin, gemcitabine, temozolomide, vincristine, vinorelbine, ifosfamide, cyclophosphamide, etoposide, and capecitabine, remained stable (in contrast to only carboplatin and capecitabine at 4 ºC). The addition of acid improved the stability of methotrexate and its metabolite hydroxymethotrexate but not that of the rest of compounds. The addition of organic solvent (50% methanol or DMSO) prevented the degradation at 4 ºC of the otherwise unstable compounds oxaliplatin, methotrexate, erlotinib, doxorubicin, tamoxifen, and paclitaxel. To the authors' knowledge, five of the analytes investigated have never been searched for in the aquatic environment (imatinib, 6α-hydroxypaclitaxel, endoxifen, (Z)4- Hydroxytamoxifen, and temozolomide), and for many of them the stability data provided, and even the analytical LC-MS/MS conditions, are the first ever published., This study was financially supported by the EU through the EU FP7 project CytoThreat(Fate and effects of cytostatic pharmaceuticals in the environment and the identification of biomarkers for and improved risk assessment on environmental exposure. Grant agreement No.: 265264), by the Spanish Ministry of Economy and Competitiveness through the projects SCARCE (Consolider-Ingenio 2010 CSD2009-00065) and CEMAGUA (CGL2007-64551/HID), and by the Generalitat de Catalunya (Consolidated Research Group: Water and Soil Quality Unit 2009-SGR-965). Nicola Mastroianni acknowledges his CSIC-JAE predoctoral fellowship. It reflects only the author’s views. The Community is not liable for any use that may be made of the information contained therein. Merck is acknowledged for the gift of LC columns, respectively. Damià Barceló acknowledges financial support from the Visiting Professor Program of the King Saud University, Riyadh, Saudi Arabia.

Feed additives are typically used in intensive farming production over long periods, and hence, they can accumulate in farmed animal tissues. Concerns regarding the use of ethoxyquin as an antioxidant feed additive, have recently arisen due to its potential conversion into a series of transformation products (TPs). The aim of this work was to characterize the TPs of ethoxyquin in fish feed by a novel approach based on the use of traveling-wave ion mobility spectrometry (TWIMS) coupled to high-resolution quadrupole time-of-flight mass spectrometry (QTOFMS). First, ethoxyquin was oxidized under controlled conditions and the generated TPs were added to a comprehensive database. Atlantic salmon feeds were then screened for ethoxyquin TPs using both targeted and untargeted approaches. Twenty-seven TPs were tentatively identified during the oxidation experiments, fifteen of them also being present in the feed samples. In addition, ten other potential TPs were detected in fish feed following the untargeted approach. Thirty-one of these TPs have been reported for the first time in this work through the oxidation experiments and the feed samples. Therefore, this study provides valuable information on the oxidative fate of ethoxyquin in feed, which can be used for future evaluations of potential risk related to this additive.

Ion mobility spectrometry allows for the measurement of the collision cross section (CCS), which provides information about the shape of an ionic molecule in the gas phase. Although the hyphenation of traveling-wave ion mobility spectrometry (TWIMS) with high-resolution quadrupole time-of-flight mass spectrometry (QTOFMS) has been mainly used for structural elucidation purposes, its potential for fast screening of small molecules in complex samples has not yet been thoroughly evaluated. The current work explores the capabilities of ultrahigh-performance liquid chromatography (UHPLC) coupled to a new design TWIMS-QTOFMS for the screening and identification of a large set of pesticides in complex salmon feed matrices. A database containing TWIMS-derived CCS values for more than 200 pesticides is hereby presented. CCS measurements showed high intra- and interday repeatability (RSD1%), and they were not affected by the complexity of the investigated matrices (ΔCCS ≤ 1.8%). The use of TWIMS in combination with QTOFMS was demonstrated to provide an extra-dimension, which resulted in increased peak capacity and selectivity in real samples. Thus, many false-positive detections could be straightforwardly discarded just by applying a maximum ΔCCS tolerance of ±2%. CCS was proposed as a valuable additional identification point in the pesticides screening workflow. Several commercial fish feed samples were finally analyzed to demonstrate the applicability of the proposed approach. Ethoxyquin and pirimiphos-methyl were identified in most of the analyzed samples, whereas tebuconazole and piperonil butoxide were identified for the first time in fish feed samples.

[EN] The kinetics of the ozone consumption for the pretreatment of hospital wastewater has been analysed in order to determine the reaction rate coefficients between the ozone and the readily oxidisabled organic matter and cytostatic compounds. The wastewater from a medium size hospital was treated with ozone and peroxone methodologies, varying the ozone concentration, the reaction time and the hydrogen peroxide doses. The analysis shows that there are four cytostatic compounds, i.e. irinotecan, ifosfamide, cyclophosphamide and capecitabine, detected in the wastewaters and they are completely removed with reasonably short times after the ozone treatment. Considering the reactor geometry, the gas hydrodynamics, the mass transfer of ozone from gas to liquid and the reaction of all oxidisable compounds of the wastewater it is possible to determine the chemical ozone demand, COzD, of the sample as 256 mg O-3 L-1 and the kinetic rate coefficient with the dissolved organic matter as 8.4 M (1) s(-1). The kinetic rate coefficient between the ozone and the cyclophosphamide is in the order of 34.7M(-1) s(-1) and higher for the other cytostatics. The direct economic cost of the treatment was evaluated considering this reaction kinetics and it is below 0.3 (sic)/m(3) under given circumstances. (C) 2016 Elsevier B.V. All rights reserved., J. Ferre-Aracil acknowledges the support of the doctoral fellowship from the Universitat Politecnica de Valencia (UPV-PAID-FPI-2010-04). Y. Valcarcel acknowledges the financial support provided by the Spanish Ministry for Economy and Competitiveness, through the Carlos III Health Institute with the program "Projects on Health Research 2011-2012 FIS (PI11/00180)". N. Negreira, M. Lopez de Alda and D. Barcelo acknowledge the financial support by the EU through the EU FP7 projects CytoThreat (265264) and SOLUTIONS (603437), by the Spanish Ministry of Economy and Competitiveness through the project SCARCE (Consolider-Ingenio 2010 CSD2009-00065), and by the Generalitat de Catalunya (Consolidated Research Groups "2014 SGR 418 - Water and Soil Quality Unit" and 2014 SGR 291 - ICRA). Merck is acknowledged for the gift of LC columns.

This work describes an effective, low solvent consumption and affordable sample preparation approach for the determination of eight UV filters in surface and wastewater samples. It involves sorptive extraction of target analytes in a disposable, technical grade silicone disc (5 mm diameter × 0.6 mm thickness) followed by organic solvent desorption, large volume injection (LVI), and gas chromatography-mass spectrometry determination. Final working conditions involved overnight extraction of 100-mL samples, containing 10% of methanol, followed by analytes desorption with 0.2 mL of ethyl acetate. The method provides linear responses between the limits of quantification (from 0.003 to 0.040 ng mL(-1)) and 10 ng mL(-1), an intra-day precision below 13%, and low matrix effects for surface, swimming pool, and treated sewage water samples. Moreover, the extraction yields provided by silicone discs were in excellent agreement with those achieved using polydimethylsiloxane-covered stir bars. Several UV filters were found in surface and sewage water samples, with the maximum concentrations corresponding to octocrylene.

This work presents an effective sample preparation method for the determination of eight UV filter compounds, belonging to different chemical classes, in freeze-dried sludge samples. Pressurized liquid extraction (PLE) and gas chromatography–mass spectrometry (GC–MS) were selected as extraction and determination techniques, respectively. Normal-phase, reversed-phase and anionic exchange materials were tested as clean-up sorbents to reduce the complexity of raw PLE extracts. Under final working conditions, graphitized carbon (0.5 g) was used as in-cell purification sorbent for the retention of co-extracted pigments. Thereafter, a solid-phase extraction cartridge, containing 0.5 g of primary secondary amine (PSA) bonded silica, was employed for off-line removal of other interferences, mainly fatty acids, overlapping the chromatographic peaks of some UV filters. Extractions were performed with a n-hexane:dichloromethane (80:20, v:v) solution at 75 °C, using a single extraction cycle of 5 min at 1500 psi. Flush volume and purge time were set at 100% and 2 min, respectively. Considering 0.5 g of sample and 1 mL as the final volume of the purified extract, the developed method provided recoveries between 73% and 112%, with limits of quantification (LOQs) from 17 to 61 ng g −1 and a linear response range up to 10 μg g −1 . Total solvent consumption remained around 30 mL per sample. The analysis of non-spiked samples confirmed the sorption of significant amounts of several UV filters in sludge with average concentrations above 0.6 μg g −1 for 3-(4-methylbenzylidene) camphor (4-MBC), 2-ethylhexyl-p-methoxycinnamate (EHMC) and octocrylene (OC).

The performance of the dispersive liquid-liquid microextraction (DLLME) technique for the determination of eight UV filters and a structurally related personal care species, benzyl salicylate (BzS), in environmental water samples is evaluated. After extraction, analytes were determined by gas chromatography combined with mass spectrometry detection (GC-MS). Parameters potentially affecting the performance of the sample preparation method (sample pH, ionic strength, type and volume of dispersant and extractant solvents) were systematically investigated using both multi- and univariant optimization strategies. Under final working conditions, analytes were extracted from 10 mL water samples by addition of 1 mL of acetone (dispersant) containing 60 μL of chlorobenzene (extractant), without modifying either the pH or the ionic strength of the sample. Limits of quantification (LOQs) between 2 and 14 ng∈L-1, inter-day variability (evaluated with relative standard deviations, RSDs) from 9% to 14% and good linearity up to concentrations of 10,000 ng∈L-1 were obtained. Moreover, the efficiency of the extraction was scarcely affected by the type of water sample. With the only exception of 2-ethylhexyl-p-dimethylaminobenzoate (EHPABA), compounds were found in environmental water samples at concentrations between 6∈±∈1 ng∈L-1 and 26∈±∈2 ng∈mL-1. [Figure not available: see fulltext.] © 2010 Springer-Verlag.

A simple, inexpensive sample preparation procedure, based on the matrix solid-phase dispersion (MSPD) technique, for the determination of six UV filters: 2-ethylhexyl salicylate (EHS), 3,3,5-trimethylcyclohexyl salicylate (Homosalate, HMS), 3-(4-methylbenzylidene) camphor (4-MBC), isoamyl-p-methoxycinnamate (IAMC), 2-ethylhexyl-p-methoxycinnamate (EHMC) and octocrylene (OCR), in dust from indoor environments is presented and the influence of several operational parameters on the extraction performance discussed. Under the final working conditions, sieved samples (0.5 g) were mixed with the same amount of anhydrous sodium sulphate and dispersed with 2 g of octadecyl bonded silica (C18) in a mortar with a pestle. This blend was transferred to a polypropylene solid-phase extraction cartridge containing 2 g of activated silica, as the clean-up co-sorbent. The cartridge was first rinsed with 5 mL of n-hexane and the analytes were then recovered with 4 mL of acetonitrile. This extract was adjusted to 1 mL, filtered and the compounds were determined by gas chromatography combined with tandem mass spectrometry (GC-MS/MS). Recoveries for samples spiked at two different concentrations ranged between 77% and 99%, and the limits of quantification (LOQs) of the method between 10 and 40 ng g(-1). Analysis of settled dust from different indoor areas, including private flats, public buildings and vehicle cabins, showed that EHMC and OCR were ubiquitous in this matrix, with maximum concentrations of 15 and 41 microg g(-1), respectively. Both UV filters were also quantified in dust reference material SRM 2585 for first time. EHS, 4-MBC and IAMC were detected in some of the analyzed samples, although at lower concentrations than EHMC and OCR.

A sensitive procedure for the determination of three UV filters: ethylhexyl salicylate (EHS), 3,3,5-trimethylcyclohexyl salicylate (Homosalate, HMS), 2-hydroxy-4-methoxybenzophenone (BP-3) and two related hydroxylated benzophenones (2,4-dihydroxybenzophenone, BP-1 and 2,2'-dihydroxy-4-methoxybenzophenone, BP-8) in water samples is presented. Analytes were first concentrated on the coating of a solid-phase microextraction (SPME) fibre, on-fibre silylated and then determined using gas chromatography combined with tandem mass spectrometry (GC-MS/MS). Factors affecting the performance of extraction and derivatization steps are thoroughly evaluated and their effects on the yield of the sample preparation discussed. Under final working conditions, a PDMS-DVB coated SPME fibre was exposed directly to 10 mL of water, adjusted at pH 3, for 30 min. After that, the fibre was placed in the headspace (HS) of a 1.5 mL vial containing 20 microL of N-methyl-N-(trimethylsilyl)-trifluoroacetamide (MSTFA). On-fibre silylation of hydroxyl groups contained in the structure of target compounds was performed at 45 degrees C for 10 min. The whole sample preparation process was completed in 40 min, providing limits of quantification from 0.5 to 10 ng L(-1) and acceptable precision (RSDs under 13%) for samples spiked at different concentrations. All compounds could be accurately determined in river and treated wastewater (relative recoveries from 89 to 115%) using standards in ultrapure water, whereas standard addition is recommended to quantify their levels in untreated wastewater. Analysis of wastewater revealed the systematic presence of BP-3 and BP-1 in raw samples with maximum concentrations close to 500 and 250 ng L(-1), respectively.

This work studies the stability of three UV filters: 2-ethylhexyl salicylate (ES), 2-ethylhexyl 4-(dimethylamino) benzoate (EHPABA) and 2-hydroxy-4-methoxybenzophenone (BP-3), in water samples containing low concentrations of free chlorine. Moreover, 2,4-dihydroxybenzophenone (2,4-DHBP), a metabolite of BP-3, was also included in some of the performed assays. Experiments were carried out considering free chlorine and analytes concentrations at the μg mL −1 and ng mL −1 level, respectively. Gas chromatography with mass spectrometry was used to follow the time course of target compounds and to identify their halogenated by-products. Concentration of water samples with solid-phase extraction cartridges and derivatization (silylation) of some species were also employed to improve their detectability. Under the experimental conditions explored in this work, ES showed an acceptable stability whereas the rest of species reacted with free chlorine at significant rates following pseudo-first-order kinetics. Their half-lives ranged from 0.4 to 25 min depending on the UV filter, chlorine concentration, water pH and presence of bromide traces. For EHPABA a relatively simple degradation pathway was established. It consisted of aromatic substitution of one atom of hydrogen per chlorine or bromide. The same reaction pattern was observed for BP-3 leading, in this case, to mono- and di-halogenated by-products. In addition, several halogenated forms of 3-methoxyphenol were identified as BP-3 cleavage by-products.

Concerns about new psychoactive substances (NPS) are increasing due to the rising frequency of serious intoxications. Analysis of biological fluids (urine) is necessary to get reliable information about the use of these substances. However, it is a challenging task due to the lack of analytical standards and the dynamic character of the NPS market In the present work, a qualitative screening of NPS was carried out in 23 pooled urine samples collected from a city center in the UK and festivals in the UK and Belgium. The analytical method was based on data-independent acquisition mode using liquid chromatography coupled to quadrupole time-of-flight mass spectrometry. An in-house library was used with >1500 entries corresponding to NPS, classical drugs and metabolites. All samples contained 53 and 28 compounds of interest from the UK and Belgium respectively. Of the different compounds detected, about 70% were confirmed using retention time and product ions while the remaining compounds were identified using elucidated fragmentation pathways. The highest numbers of NPS identified in both countries were from the cathinone and phenylethylamine families, with a higher number being detected in samples from the festival in the UK Moreover, several cathinone metabolites in human urine were detected and identified. The screening method proved useful to detect a large number of compounds and determine the use of NPS. (C) 2016 Elsevier B.V. All. rights reserved.

The residues of antineoplastic drugs are considered as new and emerging pollutants in aquatic environments. Recent experiments showed relatively high toxicity of 5-fluorouracil (5-FU), imatinib mesylate (IM), etoposide (ET) and cisplatin (CP) that are currently among most widely used antineoplastic drugs, against phytoplankton species. In this study, we investigated the toxic potential of the mixture of 5-FU + IM + ET against green alga Pseudokirchneriella subcapitata and cyanobacterium Synechococcus leopoliensis, and the stability and sorption of these drugs to algal cells. Toxic potential of the mixture was predicted by the concepts of 'concentration addition' and 'independent action' and compared to the experimentally determined toxicity. In both test species, the measured toxicity of the mixture was at effects concentrations EC10-EC50 higher than the predicted, whereas at higher effect concentration (EC90), it was lower. In general, P. subcapitata was more sensitive than S. leopoliensis. The stability studies of the tested drugs during the experiment showed that 5-FU, IM and CP are relatively stable, whereas in the cultures exposed to ET, two transformation products with the same mass as ET but different retention time were detected. The measurements of the cell-linked concentrations of the tested compounds after 72 h exposure indicated that except for CP (1.9 % of the initial concentration), these drugs are not adsorbed or absorbed by algal cells. The results of this study showed that in alga and cyanobacteria exposure to the mixture of 5-FU + ET + IM, in particular at low effect concentration range, caused additive or synergistic effect on growth inhibition, and they suggest that single compound toxicity data are not sufficient for the proper toxicity prediction for aquatic primary producers.

Following the recent prohibition on the use of BPA for the production of polycarbonate (PC) baby bottles in the European Union, baby bottles made of alternative materials, such as polypropylene, polyethersulphone, polyamide, Tritan (TM) or silicone, have appeared in the market. An initial study done in previous work applying a liquid-liquid extraction (LLE) followed by GC-MS analysis identified a wide variety of migrating compounds. Based on these screening results, the monitoring and quantification of the most important migrating compounds was further pursued. We describe the optimisation, validation and application of an LLE method for the extraction of these migrating compounds adapted to low concentrations. Generally, absolute recoveries between 70 and 110 % with RSDs < 20 % were obtained. Further, specific and quantitative GC-QqQ-MS and LC-QqQ-MS methods were developed and validated by evaluating the following parameters: sensitivity, selectivity, linearity, accuracy, precision, recoveries and matrix effects. Limits of quantification (LOQs) ranged between 0.6 and 8 mu g kg(-1) for the majority of the selected compounds. Linearity ranged using Mandel's fitting test between coefficients of determination (R (2)) of 0.995 and 0.999. Both repeatability and reproducibility were demonstrated to be satisfactory considering the Horwitz equation. Finally, the methods were applied on a selection of baby bottles representative for the Belgian market. Several compounds previously identified by the screening approach were confirmed and adequately quantified. For instance, azacyclotridecan-2-one had a maximum migration value of 1091 mu g kg(-1) whereas 4-propylbenzaldehyde migrated at 27 mu g kg(-1) and 2,2,4-trimethyl-1,3-pentanediol diisobutyrate (TXIB) at 348 mu g kg(-1). However, most of the targeted compounds were not detected or below the LOQ.

Ethylphenidate is a new potent synthetic psychoactive drug, structurally related to methylphenidate. Using human liver microsomes and cytosol, we have investigated for the first time the Phase-I and Phase-II in vitro metabolism of ethylphenidate. The structure of the metabolites was elucidated by hybrid quadrupole time-of-flight mass spectrometry. Overall, seven Phase-I, but no Phase-II metabolites were detected. Ethylphenidate underwent hydroxylation forming two primary mono-hydroxylated metabolites and, subsequently, dehydration and ring opening with an additional hydroxylation, forming secondary metabolites. The involvement of different human cytochrome P450 (CYP) enzymes in the formation of ethylphenidate metabolites was investigated using a panel of human recombinant CYPs (rCYPs). rCYP2C19 was the most active recombinant enzyme involved in the formation of all seven ethylphenidate metabolites detected, although other rCYPs (rCYP1A2, rCYP2B6, rCYPC9, rCYP2D6, and rCYP3A4, but not rCYP2E1) played a role in the metabolism of ethylphenidate. All metabolites identified in the present study can be considered as potential specific biomarkers of ethylphenidate in toxicological studies. Additionally, ritalinic acid and methylphenidate were formed by non-enzymatic hydrolysis and trans-esterification, and, therefore, they cannot be considered as (oxidative) metabolites of ethylphenidate. The presence of methylphenidate and ritalinic acid cannot be exclusively associated to the use of ethylphenidate, since methylphenidate is a drug itself and ritanilic acid can be formed from both ethylphenidate and methylphenidate. (C) 2015 Elsevier B.V. All rights reserved.

The in vitro oxidative metabolism of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), 2,2',4,4',5-pentabromodiphenyl ether (BDE-99), and the individual α-, β- and γ-hexabromocyclododecane (HBCD) isomers catalyzed by cytochrome P450 (CYP) enzymes was studied using chicken liver microsomes (CLMs). Metabolites were identified using a liquid chromatography-tandem mass spectrometry method and authentic standards for the oxidative metabolites of BDE-47 and BDE-99. Six hydroxylated tetra-BDEs, namely 4-hydroxy-2,2',3,4'-tetrabromodiphenyl ether (4-OH-BDE-42), 3-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (3-OH-BDE-47), 5-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (5-OH-BDE-47), 6-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (6-OH-BDE-47), 4'-hydroxy-2,2',4,5'- tetrabromodiphenyl ether (4'-OH-BDE-49), and 2'-hydroxy-2,3',4,4'-tetrabromodiphenyl ether (2'-OH-BDE-66), were identified and quantified after incubation of BDE-47 with CLMs. 4'-OH-BDE-49 was the major metabolite formed. Three hydroxylated penta-BDEs (5'-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (5'-OH-BDE-99), 6'-hydroxy-2,2',4,4',5- pentabromodiphenyl ether (6'-OH-BDE-99), and 4'-hydroxy-2,2',4,5,5'-pentabromodiphenyl ether, 4'-OH-BDE-101, were formed incubating BDE-99 with CLMs. Concentrations of BDE-99 metabolites were lower than those of BDE-47. More than four mono-hydroxylated HBCD (OH-HBCD), more than four di-hydroxylated HBCD (di-OH-HBCD), more than five mono-hydroxylated pentabromocyclododecenes (OH-PBCD), and more than five di-hydroxylated pentabromocyclododecenes (di-OH-PBCD) were detected when α-, β-, or γ-HBCD were individually incubated with CLMs. Response values (the ratio between the peak areas of the target compound and its internal standard) for OH-HBCD were 1-3 orders of magnitude higher than those for OH-PBCD, di-OH-HBCD, and di-OH-PBCD, suggesting that OH-HBCD might be the major metabolites of α-, β- and γ-HBCD produced by CLMs. No diastereoisomeric or enantiomeric bioisomerisation was observed incubating α-, β- or γ-HBCD with CLMs. Collectively, our data suggest that (i) BDE-47 is metabolized at a faster rate than BDE-99 by CLMs, (ii) OH-HBCD are the major hydroxylated metabolites of α-, β- and γ-HBCD produced by CLMs and (iii) the diastereoisomeric or enantiomeric bioisomerisation of α-, β- and γ-HBCD is not mediated by chicken CYP enzymes.

Flavor sensations in food are highly influenced by the aroma and taste compounds. Reviewing the extensive literature of recent years in this field has shown that the reconstitution of flavor based on aroma and taste compounds poses numerous problems. These are of different nature and include among others (a) chemical transformations among these compounds, (b) changes in the concentrations of the compounds responsible for the perceived flavor, (c) interactions among the chemical compounds that enhance or reduce a specific flavor sensation, and finally, (d) the complexity of the different food matrices and its influence in the flavor perception. Another difficulty that flavor scientists must face is how to properly model and visualize the complex relationships existing between the chemical composition of foods and the flavor perception. These problems have repercussions on the reconstitution of the flavor signature of food based on the natural concentrations of its key aroma and taste compounds. Therefore, the main aim of this review is to deal with all these issues to propose potential solutions for a robust transformation in a science-based quality approach.

This work analyses the presence of forty-eight emerging pollutants, including twenty-five drugs of abuse and metabolites, seventeen cytostatic drugs and six iodinated contrast media, in tap water from the Madrid Region. Analysis of the target compounds in the tap water was performed by means of (on-line or off-line) solid-phase extraction followed by analysis by liquid chromatography–tandem mass spectrometry. A preliminary human health risk characterization was undertaken for each individual compound and for different groups of compounds with a common mechanism of action found in tap water.The results of the study showed the presence of eight out of the twenty-five drugs of abuse and metabolites analysed, namely, the cocainics cocaine and benzoylecgonine, the amphetamine-type stimulants ephedrine, 3,4-methylenedioxymethamphetamine and methamphetamine, the opioid methadone and its metabolite 2-ethylene-1,5-dimethyl-3,3-diphenylpyrrolidine and, finally caffeine at concentrations ranging from 0.11 to 502 ng L−1. Four out of the six analysed iodinated contrast media, namely, diatrizoate, iohexol, iomeprol and iopromide, were detected in at least one sample, with concentration values varying between 0.4 and 5 ng L−1. Cytostatic compounds were not detected in any sample. Caffeine was the substance showing the highest concentrations, up to 502 ng L−1, mainly in the drinking water sampling point located in Madrid city. Among the other drugs of abuse, the most abundant compounds were cocaine and benzoylecgonine, detected at concentrations ranging from 0.11 to 86 ng L−1 and from 0.11 to 53 ng L−1, respectively. Regarding iodinated contrast media, iohexol was the most ubiquitous and abundant compound, with a frequency of detection of 100% and concentrations from 0.5 to 5.0 ng L−1 in basically the same range in all sampling points.Taking into account the results and types of treatment applied, ozonisation plus granular activated carbon filtration appears to be efficient in the removal of cocaine and benzoylecgonine. For the amphetamine-type stimulants, opioids and caffeine, ozonisation plus granular activated carbon filtration and ultrafiltration plus reverse osmosis showed higher removal efficiency than sand filtration.The human health risk characterization performed indicates that the lifetime consumption of the tap waters analysed has associated a negligible human health concern. Keywords: Pharmaceuticals, Illicit drugs, Anti-cancer drugs, Drinking water, Human health risk characterization

The selective estrogen receptor modulator tamoxifen is the most commonly used drug for the treatment and prevention of breast cancer. Tamoxifen is considered as a pro-drug since it is known to exert its pharmacological effect through its major active metabolites, 4-hydroxy-tamoxifen and 4-hydroxy-N-desmethyl-tamoxifen, which are mainly excreted in the urine in the days following administration. In the present work, the reactivity of tamoxifen and its major active metabolites in free chlorine-containing water was investigated for the first time. Under the studied chlorination conditions, tamoxifen was fairly stable whereas its metabolites were quickly degraded. A total of thirteen chlorinated byproducts were tentatively identified by ultra-high performance liquid chromatography coupled to high-resolution hybrid quadrupole-Orbitrap tandem mass spectrometry. Time-course profiles of the identified byproducts were followed in real wastewater samples under conditions that simulate wastewater disinfection. A preliminary assessment of their acute aquatic toxicity at two trophic levels by means of quantitative structure-activity relationship models showed that the identified byproducts were up to 110-fold more toxic than the parent compounds.

Chemical transformations of four alkyl esters of p-hydroxybenzoic acid, parabens, in chlorinated water samples are investigated. Quantification of the parent species and identification of their reaction by-products were performed using gas chromatography in combination with mass spectrometry. Experiments were accomplished considering free chlorine and paraben concentrations at the mg L(-1) and microg L(-1) level, respectively. Concentration of water samples, using solid-phase extraction, and silylation of the target species were carried out in order to improve the detectability of parent species and their possible transformation products, achieving quantification limits at the low ng L(-1) level. Under employed experimental conditions, the decrease in the concentrations of parabens followed pseudo-first-order kinetics. Half-lives values obtained for model ultrapure water solutions were in good agreement with those observed in tap water samples. For the first type of sample, only two by-products were detected for each paraben. They corresponded to chlorination of the aromatic ring in one or two carbons situated in ortho-positions to the hydroxyl group. Both species were also generated after the addition of parabens to chlorinated tap water. Moreover, three new transformation products were noticed for each parent compound. They were identified as bromo- and bromochloro-parabens, formed due to the existence of traces of bromide in tap water sources. Experiments carried out by mixing paraben-containing personal care products with tap water, containing free chlorine, confirmed the formation of all above described halogenated by-products. In addition, the presence of the di-chlorinated forms of methyl and propyl paraben has been detected for first time in raw sewage water samples.

The aim of the present study was to identify the in vitro Phase I and Phase II metabolites of three new psychoactive substances: alpha-pyrrolidinovalerophenone (alpha-PVP), methylenedioxypyrovalerone (MDPV), and methedrone, using human liver microsomes and human liver cytosol. Accurate-mass spectra of metabolites were obtained using liquid chromatography-quadrupole time-of-flight mass spectrometry. Six Phase I metabolites of alpha-PVP were identified, which were formed involving reduction, hydroxylation, and pyrrolidine ring opening reactions. The lactam compound was the major metabolite observed for alpha-PVP. Two glucuronidated metabolites of alpha-PVP, not reported in previous in vitro studies, were further identified. MDPV was transformed into 10 Phase I metabolites involving reduction, hydroxylation, and loss of the pyrrolidine ring. Also, six glucuronidated and two sulphated metabolites were detected. The major metabolite of MDPV was the catechol metabolite. Methedrone was transformed into five Phase I metabolites, involving N- and O-demethylation, hydroxylation, and reduction of the ketone group. Three metabolites of methedrone are reported for the first time. In addition, the contribution of individual human CYP enzymes in the formation of the detected metabolites was investigated.

Erlotinib is a highly potent tyrosine kinase inhibitor used in the treatment of the most common type of lung cancer. Due to its recent introduction, very scarce information is available on its occurrence, environmental fate and toxicological effects on aquatic organisms. During chlorination processes normally carried out in wastewater treatment plants and in the pretreatment of hospital effluents, chlorinated transformation products can be formed with an enhanced toxicity relative to the parent compound.Thus, the reactivity of the cytostatic drug erlotinib in free chlorine-containing water was investigated for the first time in the present work. A non-targeted screening approach based on the use of differential profiling tools was applied in order to reveal its potential transformation products. Structural elucidation of the detected transformation products was performed by ultra-performance liquid chromatography coupled to high-resolution hybrid quadrupole-Orbitrap tandem mass spectrometry. The proposed approach allowed detecting a total of nineteen transformation products, being eighteen of them described for the first time in this work, which demonstrates its potential in environmental analysis. Among them, six compounds presented chlorine atoms in their structures, which may be of major concern. Other transformation products involved hydroxylation and oxidation reactions. Time-course profiles of erlotinib and its transformation products were followed in real wastewater samples under conditions that simulate wastewater disinfection. Although the structures of these transformation products could not be positively confirmed due to lack of standards, their chemical formulas and product ions can be added to databases, which will allow their screening in future monitoring studies This study has been financially supported by the EU through the FP7 projects CYTOTHREAT (265264) and SOLUTIONS (603437), by the Spanish Ministry of Economy and Competitiveness through the project SCARCE (Consolider-Ingenio 2010 CSD2009-00065), and by the Generalitat de Catalunya (Consolidated Research Groups “2014 SGR 418dWater and Soil Quality Unit” and 2014 SGR 291dICRA

N-[(1S)-1-(aminocarbonyl)-2-methylpropyl]-1-(cyclohexylmethyl)-1H-indazole-3-carboxamide (AB-CHMINACA) is a recently introduced synthetic cannabinoid. At present, no information is available about in vitro or in vivo human metabolism of AB-CHMINACA. Therefore, biomonitoring studies to screen AB-CHMINACA consumption lack any information about the potential biomarkers (e.g. metabolites) to target. To bridge this gap, we investigated the in vitro metabolism of AB-CHMINACA using human liver microsomes (HLMs). Formation of AB-CHMINACA metabolites was monitored using liquid chromatography coupled to time-of-flight mass spectrometry. Twenty-six metabolites of AB-CHMINACA were detected including seven mono-hydroxylated and six di-hydroxylated metabolites and a metabolite resulting from N-dealkylation of AB-CHMINACA, all produced by cytochrome P450 (CYP) enzymes. Two carboxylated metabolites, likely produced by amidase enzymes, and five glucuronidated metabolites were also formed. Five mono-hydroxylated and one carboxylated metabolite were likely the major metabolites detected. The involvement of individual CYPs in the formation of AB-CHMINACA metabolites was tested using a panel of seven human recombinant CYPs (rCYPs). All the hydroxylated AB-CHMINACA metabolites produced by HLMs were also produced by the rCYPs tested, among which rCYP3A4 was the most active enzyme. Most of the in vitro metabolites of AB-CHMINACA were also present in urine obtained from an AB-CHMINACA user, therefore showing the reliability of the results obtained using the in vitro metabolism experiments conducted to predict AB-CHMINACA in vivo metabolism. The AB-CHMINACA metabolites to target in biomonitoring studies using urine samples are now reliably identified and can be used for routine analysis.

Identification of new psychoactive substances (NPS) is challenging. Developing targeted methods for their analysis can be difficult and costly due to their impermanence on the drug scene. Accurate-mass mass spectrometry (AMMS) using a quadrupole time-of-flight (QTOF) analyzer can be useful for wide-scope screening since it provides sensitive, full-spectrum MS data. Our article presents a qualitative screening workflow based on data-independent acquisition mode (all-ions MS/MS) on liquid chromatography (LC) coupled to QTOFMS for the detection and identification of NPS in biological matrices. The workflow combines and structures fundamentals of target and suspect screening data processing techniques in a structured algorithm. This allows the detection and tentative identification of NPS and their metabolites. We have applied the workflow to two actual case studies involving drug intoxications where we detected and confirmed the parent compounds ketamine, 25B-NBOMe, 25C-NBOMe, and several predicted phase I and II metabolites not previously reported in urine and serum samples. The screening workflow demonstrates the added value for the detection and identification of NPS in biological matrices.

This study describes the biotransformation of cytostatic and immunosuppressive pharmaceutical methotrexate. Its susceptibility to microbiological breakdown was studied in a batch biotransformation system, in presence or absence of carbon source and at two activated sludge concentrations. The primary focus of the present study are methotrexate biotransformation products, which were tentatively identified by the ultra-high performance liquid chromatography-quadrupole - Orbitrap-MS. Data-dependent experiments, combining full-scan MS data with product ion spectra were acquired, in order to identify the molecular ions of methotrexate transformation products, to propose the molecular formulae and to elucidate their chemical structures. Among the identified transformation products 2,4-diamino-N10-methyl-pteroic acid is most abundant and persistent. Other biotransformation reactions involve demethylation, oxidative cleavage of amine, cleavage of C-N bond, aldehyde to carboxylate transformation and hydroxylation. Finally, a breakdown pathway is proposed, which shows that most of methotrexate breakdown products retain the diaminopteridine structural segment. In total we propose nine transformation products, among them eight are described as methotrexate transformation products for the first time., a Jožef Stefan Institute, Department of Environmental Sciences, Jamova 39, Ljubljana, Slovenia b Water and Soil Quality Research Group, Department of Environmental Chemistry, Institute of Environmental Assessment and Water Research (IDAEA-CSIC), Jordi Girona 18-26, Barcelona, Spain c Catalan Institute for Water Research (ICRA), Scientific and Technological Park of the University of Girona, H2O Building Emili Grahit 101, Girona, Spain

A fully automated on-line solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) method has been developed for the determination of 13 cytostatics and 4 metabolites in aqueous matrices, including groundwater, surface water, and raw and treated wastewater. On-line SPE is performed by loading 5mL of water sample at pH 2 through a PLRP-s cartridge. MS/MS is performed with an electrospray (ESI) interface operating in the positive ion mode and registering two selected reaction monitoring (SRM) transitions per compound. Quantification is carried out by the isotope dilution method using 15 different isotope-labelled compounds, specific for the target analytes, as internal standards (IS). The main advantages of the method are high sensitivity, with limits of determination in groundwater, surface water, and raw and treated wastewater below 5ngL-1 for all compounds except for gemcitabine (6.9-9.3ngL-1), temozolomide (26-50ngL-1), imatinib (80-180ngL-1) and etoposide (38-65ngL-1), repeatability, with relative standard deviations (RSDs) in most cases below 15%, and selectivity and reliability of results. The method is also fairly simple and fast, with an analysis time per sample (excluding the manual steps, i.e., sample filtration, pH adjustment, and addition of IS) of 40min. Application of the method to influent wastewater samples collected daily during eight consecutive days from a wastewater treatment plant (WWTP) from Catalonia showed the presence of methotrexate, ifosfamide, capecitabine, tamoxifen and 6(α)-hydroxypaclitaxel but at fairly low concentrations (up to 43ngL-1). © 2013 Elsevier B.V., This study was financially supported by the EU through the EU FP7 project CytoThreat (Fate and effects of cytostatic pharmaceuticals in the environment and the identification of biomarkers for and improved risk assessment on environmental exposure. Grant agreement No.: 265264), by the Spanish Ministry of Economy and Competitiveness through the projects SCARCE (Consolider-Ingenio 2010 CSD2009-00065) and CEMAGUA (CGL2007-64551/HID), and by the Generalitat de Catalunya (Consolidated Research Group: Water and Soil Quality Unit 2009-SGR-965). It reflects only the author's views. The Community is not liable for any use that may be made of the information contained therein. Merck and Biotage are acknowledged for the gift of LC columns and SPE cartridges, respectively. Damià Barceló acknowledges financial support from the Visiting Professor Programme of the King Saud University, Riyadh, Saudi Arabia. Pedro Aguiló, Joaquín Bosque and Begoña Martínez from EMSSA (Barcelona, Spain) are acknowledged for their collaboration in the sampling of wastewaters.

The stability of the UV filter benzophenone-4 (BP-4) in free chlorine-containing water was investigated, for the first time, by liquid chromatography quadrupole time-of-flight mass spectrometry (LC–QqTOF-MS). High mass accuracy and resolution capabilities of this hybrid mass spectrometer were used for the reliable assignation of empirical formulae and chemical structures of BP-4 derivatives. Time-course profiles of the parent compound and its by-products were simultaneously recorded by direct injection of sample aliquots, after quenching the excess of chlorine, in the LC–QqTOF-MS system. At neutral pHs, in excess of chlorine, BP-4 showed a limited stability fitting a pseudo-first-order degradation kinetics. A noticeable reduction in the half-lives of BP-4 was observed when increasing the sample pH between 6 and 8 units and also in presence of bromide traces. The reaction pathway of this UV filter involved a first electrophilic substitution of hydrogen per chlorine (or bromide) in the phenolic ring, followed by oxidation of the carbonyl moiety to an ester group, which induced a further electrophilic substitution in the same aromatic ring. Above reactions were also noticed when mixing a BP-4 containing personal care product with chlorinated tap water and in chlorinated swimming pool and sewage water, previously spiked with a BP-4 standard.

A novel, single step method for the determination of seven ink photo-initiators in carton packed milk samples is described. Solid-phase microextraction (SPME) and gas chromatography (GC), combined with mass spectrometry (MS), were used as sample preparation and determination techniques, respectively. Parameters affecting the performance of the microextraction process were thoroughly evaluated using uni- and multivariate optimization strategies, based on the use of experimental factorial designs. The coating of the SPME fibre, together with the sampling mode and the temperature were the factors playing a major influence on the efficiency of the extraction. Under final conditions, 1.5 mL of milk and 8.5 mL of ultrapure water were poured in a glass vessel, which was closed and immersed in a water boiling bath. A poly(dimethylsiloxane)-divinylbenzene (PDMS-DVB) coated fibre was exposed directly to the diluted sample for 40 min. After that, the fibre was desorbed in the injector of the GC-MS system for 3 min. The optimized method provided limits of quantification (LOQs) between 0.2 and 1 microg L(-1) and a good linearity in the range between 1 and 250 microg L(-1). The inter-day precision remained below 15% for all compounds in spiked whole milk. The efficiency of the extraction changed for whole, semi-skimmed and skimmed milk; however, no differences were noticed among the relative recoveries achieved for milk samples, from different brands, with the same fat content.

A procedure for the determination of six derivatives of 2-hydroxybenzophenone, used as UV absorbers, in water samples is presented. Compounds were first concentrated using a solid-phase extraction (SPE) cartridge and then selectively determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using electrospray ionization (ESI). The effect of different parameters on the performance of concentration and determination steps is discussed. The highly polar and acidic 2-hydroxy-4-methoxybenzophenone-5-sulfonic acid (BP-4) required the use of ammonium acetate as modifier during desorption of SPE cartridges and also to improve the performance of its separation in the LC column. Under optimized conditions, the proposed method provided limits of quantification from less than 1 to 32ngL(-1), depending on the compound and the type of water sample. Recoveries from the SPE step (83-105%) remained unaffected by the nature of the matrix; however, the efficiency of electrospray ionization was compound and sample dependant. Real sample analysis reflected the presence of three of the six investigated species (BP-4, 2-hydroxy-4-methoxybenzophenone, BP-3, and 2,4-dihydroxybenzophenone, BP-1) in the aquatic environment, particularly in raw wastewater samples. In this latter matrix, BP-4 was the compound presenting the highest concentrations; moreover, it was poorly removed in sewage treatment plants and consequently it also appeared in river water.

Etoposide susceptibility to microbiological breakdown was studied in a batch biotransformation system, in the presence or absence of artificial wastewater containing nutrients, salts and activated sludge at two concentration levels. The primary focus of the present study was to study etoposide transformation products by ultra-high performance liquid chromatography coupled to high-resolution hybrid quadrupole-Orbitrap tandem mass spectrometry (MS/MS). Data-dependent experiments combining full-scan MS data with product ion spectra were acquired to identify the molecular ions of etoposide transformation products, to propose the molecular formulae and to elucidate their chemical structures. Due to the complexity of the matrix, visual inspection of the chromatograms showed no clear differences between the controls and the treated samples. Therefore, the software package MZmine was used to facilitate the identification of the transformation products and speed up the data analysis. In total, we propose five transformation products; among them, four are described as etoposide transformation products for the first time. Even though the chemical structures of these new compounds cannot be confirmed due to the lack of standards, their molecular formulae can be used to target them in monitoring studies.

After the prohibition of bisphenol-A-containing polycarbonate baby bottles in the European Union (EU), alternative materials, such as polypropylene, polyethersulphone, Tritan T copolyester, etc., have appeared on the market. Based on an initial screening and in vitro toxicity assessment, the most toxic migrating compounds were selected to be monitored and quantified using validated GC-and LC-QqQ-MS methods. The effect of several 'real-life-use conditions', such as microwave, sterilisation and dishwasher, on the migration of different contaminants was evaluated by means of duration tests. These results were compared with a reference treatment (filling five times with pre-heated simulant at 40 degrees C) and with the legal EU 'repetitive-use conditions' (three migrations, 2 h at 70 degrees C). Analysis of the third migration step of the EU repetitive-use conditions (which has to comply with the EU legislative migration limits) showed that several non-authorised compounds were observed in some baby bottles exceeding 10 mu g kg(-1). However, all authorised compounds were detected well below their respective specific migration limits (SMLs). The reference experiment confirmed the migration of some of the compounds previously detected in the EU repetitive-use experiment, though at lower concentrations. Analysis of extracts from the microwave and dishwasher experiments showed a reduction in the migration during the duration tests. In general, the concentrations found were low and comparable with the reference experiment. Similar observations were made for the two sterilisation types: steam and cooking sterilisation. However, steam sterilisation seems to be more recommended for daily use of baby bottles, since it resulted in a lower release of substances afterwards. Repeated use of baby bottles under 'real-life' conditions showed no increase in the migration of investigated compounds and, after some time, the migration of these compounds even became negligible.