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New psychoactive substances (NPS) have become a serious threat for public health due to their ability to be sold in the street or on internet. NPS are either derived from commercial drugs which are misused (recreational rather than medical use) or whose structure is slightly modified. To regulate NPS, it is essential to accurately characterize them, either to recognize molecules that were previously identified or to quickly elucidate the structure of unknown ones. Most approaches rely on the determination of the exact mass obtained by high-resolution mass spectrometry requiring expensive equipment. This motivated us to develop a workflow in which the elucidation is assisted with databases and does not need the exact mass. This workflow combines 1D and 2D NMR measurements performed on a benchtop spectrometer with IR spectroscopy, for creating a multi-technique database to characterize pure and mixed NPS. The experimental database was created with 57 entries mostly coming from seizures, mainly cathinones, cannabinoids, amphetamines, arylcyclohexylamines, and fentanyl. A blind validation of the workflow was carried out on a set of six unknown seizures. In the first three cases, AF, AB-FUBINACA, and a mixture of 2C-I and 2C-E could be straightforwardly identified with the help of their reference spectra in the database. The two next samples were elucidated for the first time with the help of the database to reveal NEK and MPHP substances. Finally, a precise quantification of each characterized NPS was obtained in order to track NPS trafficking networks.

International audience; This work proposes a rapid, simple and accurate analytical method for quantification of the organophosphate flame retardant triphenylphosphate (TPhP) and the biomarker diphenylphosphate (DPhP) in whole blood by a simple and effective sample preparation followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) quantification. A liquid–liquid microextraction (µLLE) from 250 µL of mouse whole blood was developed and validated to quantify TPhP and DPhP. Nature of extraction solvent, grinding for cell lysis and purification step were investigated. The use of distilled solvents proved necessary to reduce and control the contamination of the ubiquitous target compounds throughout the analytical process and to maintain a stable and repeatable signal in the analytical controls.The validation was performed by matrix-matched calibration with a correction by labelled internal standards: the limit of quantification was 1 ng/mL for DPhP and 2 ng/mL for TPhP with good linearity over the range up to 48 ng/mL for each compound. Accuracy was between 92.5% and 106.7% with relative standard deviation < 20%. The µLLE-LC-MS/MS method was successfully tested during short exposures of mice to DPhP or TPhP via drinking water or by intravenous injection.

BackgroundAryl phosphate esters (APEs), a main class of organophosphorus ester molecules, are widely used and commonly present in the environment. Health hazards associated with these compounds remain largely unknown and the effects of diphenyl phosphate (DPhP), one of their most frequent derivatives in human samples, are poorly characterised.ObjectiveOur aim was to investigate whether DPhP per se may represent a more relevant marker of exposure to APEs and determine its potential deleterious biological effects in chronically exposed mice.MethodsConventional animals (FVB mice) were acutely (intravenous or oral gavage) or chronically (0.1 mg.mL-1, 1 mg.mL-1, 10 mg.mL-1 in drink water) exposed to relevant doses of DPhP or triphenyl phosphate (TPhP), one of its main precursors in the environment. Both molecules were measured in blood and other relevant tissues by liquid chromatography-mass spectrometry (LC-MS). Biological effects of chronic DPhP exposure were addressed through liver multi-omics analysis combining mRNA extraction and sequencing to high resolution LC-MS to determine the corresponding metabolic profile. Deep statistical exploration was performed to extract correlated information, guiding further physiological analyses (immunohistochemistry (IHC) and animal growth measurement).ResultsAcute and chronic exposure to DPhP led to significant levels of this molecule in blood and other tissues, an effect missing with TPhP. Multi-omics analysis confirmed the existence of biological effects of DPhP, even at a very low dose of 0.1 mg.mL-1 in drinking water. Chemical structural homology and pathway mapping demonstrated a clear reduction of the fatty-acid catabolic processes centred on acylcarnitine and mitochondrial β-oxidation. Interestingly, mRNA expression confirmed and extended these observations by demonstrating at all tested doses the overall repression of genes involved in lipid catabolic processes and regulated by PPARα, a master regulator of β-oxidation and its associated ketogenesis. IHC analysis confirmed the alteration of these pathways by showing a specific downregulation of Hmgcs2, a kernel target gene of PPARα, at all doses tested, and surprisingly, a strong reduction of the lipid droplet content only at the highest dose. Overall, DPhP absorption led to weight loss, which was significant using the highest dose.ConclusionsOur results suggest that in mice, the effects of chronic exposure to DPhP, even at a low dose, are not negligible. Fatty acid metabolism in the liver in particular is essential for controlling fast and feast periods with adverse consequences on the overall physiology. Therefore, the impact of DPhP on circulating fat, cardiovascular and metabolic disease incidence deserves, in light of our results, further investigations.

The potential ecotoxicological effects of mixtures of contaminants in the aquatic environment are generating a global concern. Benthic invertebrates, such as the crustacean Gammarus fossarum, are key in the functioning of aquatic ecosystems, and are frequently used as sentinel species of water quality status. The aim of this work was to study the effects of a mixture of the most frequently detected surfactants in the bioconcentration kinetics of two pharmaceuticals in G. fossarum, evaluating their potential enhancing or suppressing effects. Laboratory exposure experiments for both pharmaceuticals and surfactants (concentration ratio 1:25) were set up for two individual compounds, the anxiolytic oxazepam and the antibiotic sulfamethoxazole. Gammarid samples were processed using microQuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) extraction. Pharmaceuticals concentration in the organisms was followed-up by means of nanoliquid chromatography coupled to tandem mass spectrometry (nanoLC-MS/MS). Results indicated a similar mode of action of the surfactants in the bioconcentration kinetics of both drugs, decreasing the accumulation rate in the organism. Oxazepam showed a higher accumulation potential than sulfamethoxazole in all cases. Depuration experiments for oxazepam also demonstrated the high depurative capacity of gammarids, eliminating > 50% of the concentration of oxazepam in < 6 h.

In Western Europe, river water quality can be assessed using sentinel species such as the amphipod Gammarus fossarum. In this work of environmental metabolomics, the objective was to develop suitable chemometrics methods, using a limited number of individuals, to assess the modification of the metabolism of G. fossarum exposed to two human pharmaceuticals. Males and females gammarids were exposed to a mixture of the anxiolytic oxazepam and the antiepileptic carbamazepine (1000 ng L−1) for 14 days under laboratory conditions according to a full factorial design 2² (repeated 5 times). They were analyzed at the single individual scale using a method including a μQuEChERS type extraction followed by a nanoliquid chromatography analysis coupled to high-resolution mass spectrometry. The molecular fingerprints obtained were investigated using XCMS. Several corrections of experimental drifts (by using lock mass and Quality Control samples) were tested prior to using APCA + method for the exploitation of the unbalanced designed data. Signal reproducibility was greatly improved by the lock mass normalisation. From the experimental design, a significant effect of both experimental factors “exposure to the mixture” and “gammarid gender” on the signals measured were highlighted by APCA+. Finally, the results obtained made it possible to identify variables responsible for each of the factor effects.

International audience; An innovative analytical method has been developed to quantify the bioaccumulation in an amphipod crustacean (Gammarus fossarum) of three micropollutants regarded as anthropic-pollution markers: carbamazepine, oxazepam, and testosterone. A liquid-liquid extraction assisted by salts, known as QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) was miniaturised and optimised, so it could be adapted to the low mass samples (approximatively 5 mg dry weight). For this same reason and in order to obtain good sensitivity, ultra-trace analyses were carried out by means of nanoliquid chromatography. A pre-concentration system by on-column trapping was optimised to increase the injection volume. In order to improve both sensitivity and selectivity, the multiple reaction monitoring cubed mode analyses (MRM3) were carried out, validated and compared to the classic MRM. To the best of our knowledge, this is the first time that MRM3 is coupled to nanoliquid chromatography for the analysis and detection of organic micropollutants

This work proposes the first method based on hydrophilic interaction liquid chromatography coupled to multiple reaction monitoring with triple stage fragmentation (HILIC-MRM3) to quantify polar organic micropollutants in complex sewage waters. A fast HILIC-MRM3 analytical method, without sample preparation except a dilution step, was developed and validated to quantify seven iodinated contrast media (ICMs) in sewage waters, namely iohexol, iomeprol, ioversol, iopamidol, diatrizoic acid, iopromide and iopentol. Several chromatographic columns and mobile phase conditions were investigated and a good separation of the ICMs was obtained with a mixed-mode column (Acclaim Mixed-mode WAX) used in HILIC conditions. The validation was performed using a synthetic matrix: the limits of quantification (LOQ) were inferior to 1 µg/L and the linearity of each compound was comprised within the [0.5–50] µg/L range. The applicability of the HILIC-MRM3 method was assessed by the analysis of several raw waters. The results highlighted the presence of ICMs in most samples, at concentrations up to several mg/L in hospital sewage waters.

The authors gratefully acknowledge Céline Briand (SIAAP) for her technical assistance during the sampling campaigns and daily operation of the pilot system.; International audience; Among the release solutions for reducing the discharge of organic and persistent contaminants in the aquatic environment, the use of a tertiary treatment in addition to existing conventional wastewater treatment processes is considered. The use of micro-grain activated carbon in a fluidized bed is a promising technique investigated in this study. The effluents from a large-scale pilot system were analyzed by liquid chromatography coupled with high-resolution mass spectrometry (QToF). Several strategies were deployed, namely molecular fingerprint comparison, suspected and non-target analyses, identification of refractory compounds to treatment, and finally, quantification of identified compounds. The evaluation of the molecular fingerprints provided evidence of the overall effect of the tertiary treatment on the treated wastewater quality. The suspected approach highlighted the presence of 83 pharmaceuticals and pesticides as well as transformation products in the effluents. The non-target approaches also highlighted compounds refractory to tertiary treatment, such as illicit drugs or some pharmaceuticals. The identification and quantification of identified compounds underscored the suitability of micro-grain activated carbon in eliminating many classes of pharmaceuticals with various physicochemical properties, such as anti-hypertensive, analgesic, anti-viral, antidepressant and even various pesticides.

International audience; Nanoliquid chromatography (nanoLC) was coupled to high-resolution mass spectrometry (HRMS) to perform a non-targeted investigation on benthic invertebrates, Chironomus riparius exposed to wastewater treatment plant (WWTP) effluents. Insect larvae represent a complex and low-weight matrix that required the use of a miniaturized Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) method of extraction followed by nanoLC-HRMS to perform the analysis. The optimization of this coupling in terms of separation conditions including trapping step, detection conditions and data treatment provided reproducible fingerprints on insect larvae exposed to WWTP effluents with both in situ and ex-situ approaches. Statistical treatments such as principal component analysis highlighted the impact of WWTP effluents on the metabolome of insect larvae and showed the influence of exposure conditions. The identification of discriminating signals (m/z, tR) matched with several potential endogenous biomarkers. These are mainly fatty acids, indicating a change in lipid metabolism that can be correlated with exposure to WWTP effluents. Several xenobiotics have also been detected, including ibuprofen and propranolol, whose identities have been confirmed by analytical standards. This work demonstrates the effectiveness and sensitivity of nanoLC-HRMS based environmental non-targeted approaches in ecotoxicological studies and provides the first profiling data for a very small aquatic invertebrate.

This paper describes the development of an integrative embryo-toxicity assay in Japanese medaka allowing analysis of several toxicological endpoints together in a same individual. In this assay, embryos are topically exposed, and survival, hatching success, malformations, biometry, behaviour, and target gene expression are subsequently analysed in each individual. This assay was applied to oxazepam, an anxiolytic pharmaceutical compound currently found in wastewater treatment plant effluent. Even if oxazepam accumulation in embryos was very low, it caused spinal and cardiac malformations, delayed growth, erratic swimming and deregulation of genes involved in apoptosis, DNA repair and mitochondrial metabolism. Relationship between gene deregulation, abnormal behaviour, and developmental anomalies was demonstrated. This assay is sensitive enough to detect adverse effects at low chemical concentrations and at multiple endpoints in a unique fish embryo. This integrative embryo-toxicity assay is a powerful tool to characterize the spectrum of effects of new chemicals and also to link effects induced at different molecular, tissue and physiological levels.

International audience; The overall objective of this study was to develop an analytical method to contribute to the assessment of the exposure of humans to plastic additives based on the use of a new polydimethylsiloxane-based stir bar extraction method. The method focuses on a list of 21 contaminants, including 18 plastic additives and 3 degradation products. They are analysed by ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) after a stir bar sorptive extraction from urine using a home-made stir bar. Thus, the paper focuses on the optimisation of the extraction method. The limits of quantification are variable from one substance to another, but they fall between 0.1 and 305 ng mL−1. The sample preparations lead to recoveries of up to 90% in water and 80% in urine. The method was successfully applied to 15 human urine samples. Among the targeted plastic additives and their degradation products, 7 compounds were quantified in samples at concentrations ranging from 0.3 ng mL−1 to 42.7 ng mL−1.

National audience

This study explored the sample preparation steps based on QuEChERS (quick, easy, cheap, effective, rugged, and safe) of a complex matrix, such as sewage sludge, regarding their impacts on the chemical signatures of the samples obtained via high-resolution mass spectrometry (HRMS). The effects on the HRMS signal of the intrinsic parameters of the QuEChERS method, such as the extraction solvent, the buffer salt, or the purification procedure, have been evaluated. Different strategies have been used to perform these evaluations, such as a comparison of the total number of HRMS signals, a comparison of the targeted screening based on more than 100 compounds, and a comparison of a nontargeted screening based both on the study of the most discriminating couples [mass-to-charge ratio (m/z); retention time (tR)] and on the most intense peaks. Moreover, two specific methods dedicated to the extraction of surfactants and drugs have been added to this comparison to visualize the impact of specific conditions on the ...

International audience; A large number of polyphenols are consumed with the diet and may contribute to the prevention of chronic diseases such as cardiovascular diseases, diabetes, cancers, and neurodegenerative diseases. More comprehensive methods are needed to measure exposure to this complex family of bioactive plant compounds in epidemiological studies. We report here a novel method enabling the simultaneous measurement in urine of 38 polyphenols representative of the main classes and subclasses found in the diet. This method is based on differential C-12-/C-13-isotope labeling of polyphenols through derivatization with isotopic dansyl chloride reagents and on the analysis of the labeled polyphenols by tandem mass spectrometry. This derivatization approach overcomes the need for costly labeled standards. Different conditions for enzyme hydrolysis of polyphenol glucuronides and sulfate esters, extraction, and dansylation of unconjugated aglycones were tested and optimized. Limits of quantification varied from 0.01 to 1.1 mu M depending on polyphenols. Intrabatch coefficients of variation varied between 3.9% and 9.6%. Interbatch variations were lower than 15% for 31 compounds and lower than 29% for 6 additional polyphenols out of the 38 tested. Thirty seven polyphenols were validated and then analyzed in 475, 24 h urine samples from the European Prospective Investigation on Cancer and Nutrition (EPIC) study. Thirty four polyphenols could be detected and successfully estimated and showed large interindividual variations of concentrations (2-3 orders of magnitude depending on the compound), with median concentrations spanning from 0.01 to over 1000 mu M for all 34 compounds.

Le Syndicat interdepartemental pour l’assainissement de l’agglomeration parisienne (Siaap) et le Laboratoire eau, environnement et systemes urbains (Leesu) etudient, en collaboration avec les equipes de la Saur, le procede CarboPlus, procede de traitement tertiaire des eaux usees base sur les proprietes de sorption du charbon actif. Parallelement au suivi d’un prototype industriel, des experimentations ont ete menees a l’echelle du laboratoire afin d’ameliorer notre comprehension et nos connaissances sur les processus de sorption des micropolluants sur le charbon actif, et en particulier sur le charbon actif en poudre (CAP). Les resultats ont notamment montre que les performances du CAP etaient fortement liees a la surface specifique (BET), qui peut etre aisement estimee par une mesure de la densite apparente. L’etude des processus de sorption a egalement mis en evidence l’influence forte de la dose appliquee en CAP ainsi que la rapidite de la cinetique de sorption. D’un point de vue operationnel, l’ajout de chlorure ferrique semble apporter un leger benefice sur l’adsorption de la plupart des composes mesures, probablement par coagulation de la fraction colloidale de la matiere organique dissoute. Au contraire, la presence et la concentration residuelle en methanol dans l’eau ne semblent pas modifier le devenir des residus medicamenteux. Il a egalement ete montre que la concentration en carbone organique dissous n’est pas toujours suffisante pour expliquer la limitation du processus de sorption, la nature de la matiere organique presente dans l’effluent doit egalement etre consideree. Enfin, ce travail a montre que la prediction du comportement des micropolluants organiques en utilisant l’abattement de l’absorbance UV a 254 nm semblait prometteuse, notamment pour certains composes comme la carbamazepine ou le diclofenac.

International audience; HIGHLIGHTS 18  BET surface and bulk density of activated carbons are correlated to micropollutants adsorption, 19  The activated carbon dose and the contact time have a great influence on adsorption, 20  FeCl3 has a positive influence on micropollutants adsorption by activated carbon (+10-15%), 21  Both the quantity and composition of organic matter impact the adsorption, 22  UV-254 removals are correlated to several micropollutants removals. 23 24 A Present adress: SIAAP, Direction du Développement et de la Prospective

Publication dans le cadre du programme OPUR; International audience; Among the solutions to reduce micropollutant discharges into the aquatic environment, activated carbon adsorption is a promising technique and a large scale pilot has been tested at the Seine Centre (240,000 m3/d — Paris, France) wastewater treatment plant (WWTP). While most of available works studied fixed bed or contact reactors with a separated separation step, this study assesses a new type of tertiary treatment based on a fluidized bed containing a high mass of activated carbon, continuously renewed. For the first time in the literature, micro-grain activated carbon (μGAC) was studied. The aims were (1) to determine the performances of fluidized bed operating with μCAG on both emerging micropollutants and conventional wastewater quality parameters, and (2) to compare its efficiency and applicability to wastewater to former results obtained with PAC. Thus, conventional wastewater quality parameters (n = 11), pharmaceuticals and hormones (PPHs; n = 62) and other emerging pollutants (n = 57) have been monitored in μGAC configuration during 13 campaigns.A significant correlation has been established between dissolved organic carbon (DOC), PPHs and UV absorbance at 254 nm (UV-254) removals. This confirms that UV-254 could be used as a tertiary treatment performance indicator to monitor the process. This parameter allowed identifying that the removals of UV-254 and DOC reach a plateau from a μGAC retention time (SRT) of 90–100 days.The μGAC configuration substantially improves the overall quality of the WWTP discharges by reducing biological (38–45%) and chemical oxygen demands (21–48%), DOC (13–44%) and UV-254 (22–48%). In addition, total suspended solids (TSS) are retained by the μGAC bed and a biological activity (nitratation) leads to a total elimination of NO2−.For micropollutants, PPHs have a good affinity for μGAC and high (> 60%) or very high (> 80%) removals are observed for most of the quantified compounds (n = 22/32), i.e. atenolol (92–97%), carbamazepine (80–94%), ciprofloxacin (75–95%), diclofenac (71–97%), oxazepam (74–91%) or sulfamethoxazole (56–83%). In addition, alkylphenols, artificial sweeteners, benzotriazole, bisphenol A, personal care products (triclocarban and parabens) and pesticides have removals lying in the 50 −> 90% range.Overall, the fluidized bed of μGAC allows obtaining performances comparable to PAC at the same activated carbon dose. Indeed, the average removal of the 13 PPHs found at a high occurrence (> 75%) in WWTP discharges is similar at 20 g/m3 of μGAC (78–89%) and PAC (85–93%). In addition, this recycled μGAC operation leads to several operational advantages (no FeCl3, reactivable, higher SRT, higher treated flow) and has a stronger impact on the overall wastewater quality compared to PAC.

The occurrence of pharmaceuticals in freshwater ecosystems provokes increasing concern due to their potential risk to non-target organisms and to human health. Pharmaceuticals are used in both human and veterinary medicine and are essentially released into the environment via wastewater treatment plants (WWTPs) and from livestock. In this study, 31 pharmaceuticals were analyzed in effluent and surface water upstream and downstream of two WWTPs in the Loue-Doubs rural karstic catchment in Eastern France. Diclofenac (965 and 2476 ng L-1), sulfamethoxazole (655 and 1380 ng L-1) and carbamazepine (566 and 1007 ng L-1) displayed the highest levels in the effluents of both WWTPs. Diclofenac levels were also high in surface water samples 300 and 166 ng L-1 in the River Doubs and the River Loue, respectively, followed by paracetamol (273 and 158 ng L-1) and sulfamethoxazole (126 and 73 ng L-1). In both rivers, the most critical compounds were found to be the antibiotic sulfamethoxazole (risk quotient (RQ) from 23.7 to 51.1) and ofloxacine (RQ from 1.1 to 18.9), which reached levels inducing toxic effects in aquatic organisms. This study showed that WWTP effluents are the major sources of the pharmaceuticals, but raw discharges from human residences, pastures and livestock manure represent significant sources of contamination of surface water and groundwater. The aim of this study was to assist scientists and authorities in understanding occurrence and sources of pharmaceuticals in order to improve water quality management in chalk streams.

The aim of the work was to develop an analytical procedure able to quantify traces of 13 neonicotinoids and pyrethroids as well as carboxamide in beeswax at low levels (ng g−1) to evaluate the contamination. For this purpose, an efficient sample preparation procedure was developed based on solid–liquid extraction using dispersive diatomaceous earth and acetonitrile. This step was followed by a selective and sensitive analysis based on ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (ESI-MS/MS). This analytical procedure was validated based on International Conference on Harmonization guidelines. The limits of quantification ranged from 1 ng g−1 (thiamethoxam, clothianidin, imidacloprid, acetamiprid, thiacloprid and boscalid) to 40 ng g−1 (lambda-cyhalothrin). The method was then successfully applied to 60 samples of beeswax collected in several areas of France. The presence of thiacloprid, boscalid, imidacloprid and deltamethrin in beeswax was confirmed. The most frequently quantified pesticide was boscalid.

International audience; Due to industrialization and the use of chemical products in everyday life, various types of drugs and pesticides are present in our environment, which threaten and cause negative impacts on aquatic ecosystems. The consequences of these pollutants are gradually becoming visible. Recent evidence confirms that long term exposure to environmental pharmaceutical concentrations can induce adverse effects in aquatic vertebrates and invertebrates such as reproductive impairments and collapse wild populations. Consequently, one of the challenges of environmental science is to evaluate the associated risks.In this context, a new methodology has been developed using nano-LC-nano-ESI MS/MS to quantify traces of two pharmaceuticals (a neuropharmaceutical drug, fluoxetine, and an anticonvulsant drug, carbamazepine) in two molluscs. Potamopyrgus antipodarum and Valvata piscinalis, which are both prosobranch gastropods. A simple and quick extraction method was developed based on a modified and miniaturized version of the QuEChERS method. The procedure involves the extraction of approximately 10 mg of wet mollusc tissue by 500 mu L of a mixture of acetonitrile/water/hexane (50/20/30) and 100 mg of buffer salt. Thus, the extraction step was carried out on an individual scale.The sensitivity of this method allowed for the detection of levels as low as 18 ng/g and 128 ng/g for carbamazepine and fluoxetine, respectively, with recoveries of greater than 85% for the two targeted compounds. This method was then applied to both gastropod species exposed to fluoxetine under laboratory conditions. The results provide evidence of bioaccumulation in both P. antipodarum and V. piscinalis and reveal the inter-species differences.

International audience; Functionalization of bio-based oligomers is a relevant approach in order to allow these materials to replace more conventional and nonbiodegradable polymers in a wider range of applications. This article will report on the hydroxylation of isosorbide-based ethersulfone oligomers. The hydroxy end-group was controlled by the variation of polycondensation parameters. Comparative study was conducted to assess the effect of isosorbide excess and the monomer concentration on structural composition and molecular weights of produced oligomers. Depending on reaction conditions, various products were synthesized, mainly alpha,omega-dihydroxyethersulfone oligomers. The characterization of the synthesized polyethersulfone was demonstrated by MALDI-TOF, NMR spectroscopy, differential scanning calorimetry, and size exclusion chromatography.

International audience; Plant protection spray treatments may expose non-target organisms to pesticides. In the pesticide registration procedure, the honey bee represents one of the non-target model species for which the risk posed by pesticides must be assessed on the basis of the hazard quotient (HQ). The HQ is defined as the ratio between environmental exposure and toxicity. For the honey bee, the HQ calculation is not consistent because it corresponds to the ratio between the pesticide field rate (in mass of pesticide/ha) and LD50 (in mass of pesticide/bee). Thus, in contrast to all other species, the HQ can only be interpreted empirically because it corresponds to a number of bees/ha. This type of HQ calculation is due to the difficulty in transforming pesticide field rates into doses to which bees are exposed. In this study, we used a pragmatic approach to determine the apparent exposure surface area of honey bees submitted to pesticide treatments by spraying with a Potter-type tower. The doses received by the bees were quantified by very efficient chemical analyses, which enabled us to determine an apparent surface area of 1.05 cm(2)/bee. The apparent surface area was used to calculate the exposure levels of bees submitted to pesticide sprays and then to revisit the HQ ratios with a calculation mode similar to that used for all other living species. X-tomography was used to assess the physical surface area of a bee, which was 3.27 cm(2)/bee, and showed that the apparent exposure surface was not overestimated. The control experiments showed that the toxicity induced by doses calculated with the exposure surface area was similar to that induced by treatments according to the European testing procedure. This new approach to measure risk is more accurate and could become a tool to aid the decision-making process in the risk assessment of pesticides.

International audience; Aquatic ecosystems are continuously contaminated by agricultural and industrial sources. Although the consequences of this pollution are gradually becoming visible, their potential impacts on aquatic ecosystems are poorly known, particularly regarding the risk of bioaccumulation in different trophic levels. To establish a causality relationship between bioaccumulation and disease, experiments on biotic matrices must be performed. In this context, a multi-residue method for the analysis of 35 emerging pollutants in three benthic invertebrates (Potamopyrgus antipodarum, Gammarus fossarum, and Chironomus riparius) has been developed. Because the variation in response of each individual must be taken into account in ecotoxicological studies, the entire analytical chain was miniaturised, thereby reducing the required sample size to a minimum of one individual and scaling the method accordingly. A new extraction strategy based on a modified, optimised and miniaturised "QuEChERS" approach is reported. The procedure involves salting out liquid-liquid extraction of approximately 10-20 mg of matrix followed by nano-liquid chromatography-nano electospray ionisation coupled with tandem mass spectrometry. The validated analytical procedure exhibited recoveries between 40 and 98% for all the target compounds and enabled the determination of pollutants on an individual scale in the ng g(-1) concentration. The method was subsequently applied to determine the levels of target analytes in several encaged organisms which were exposed upstream and downstream of an effluent discharge. The results highlighted a bioaccumulation of certain targeted emerging pollutants in three freshwater invertebrates, as well as inter-species differences. 18 out of 35 compounds were detected and eight were quantified. The highest concentrations were measured for ibuprofen in G. fossarum, reaching up to 105 ng g(-1).

International audience; New biosourced alternated poly(ether)Ester-Amides PeEA were prepared by polycondensation in solution or in bulk of unprecedented aminoalcohols based on 1,4:3,6-dianhydrohexitols (DAH) with aromatic and aliphatic diacyl chlorides (sebacoyl, terephtaloyl and isophtaloyl). Optimization of the polymerization in solution has been investigated using MALDI ToF MS and NMR spectroscopy to ascertain the structures and end chain nature of the resulting polymers. The influence of the DAH stereochemistry and the polymerization mode on the structures and thermal properties was demonstrated by MALDI ToF and complementary analytical techniques (DSC and SEC). Isomannide based polymers revealed amorphous but showed higher Tg (around 80 °C for sebacoyl and 180 °C for isophtaloyl) whereas the isosorbide based polymers were semi crystalline. Best properties were obtained in bulk using isomannide based aminoalcohol and sebacoyl chloride (Tg of 82 °C and Mw of 32 kDa, PDI = 1.41).

International audience; Beebread is among the matrices suspected of contaminating honeybee. To better understand this contamination, the study aimed to develop an efficient, sensitive and reliable analytical method for the trace analysis of pesticides in beebread. This study focuses specifically on the insecticides pyrethroids and neonicotinoids and some of their metabolites. It describes the development and validation of an original analytical approach that consists of one simple extraction method based on modified QuEChERS followed by a selective and sensitive analysis by UHPLC-MS/MS to determine the target compounds in beebread. The method was validated using a quadratic fit. RSD values below 20% were obtained, except for 5-hydroxy-imidacloprid and imidacloprid at 0.5 ng/g, which exhibited RSDs of 25% and 21%, respectively. The intra-day precision was less than 10% for many of the investigated compounds. The inter-day precision varied between 2% and 36%, depending on the compound and the concentration. The recoveries varied from 53% to 119%, with averages of 83, 81 and 77% for the extraction of beebread samples spiked at 0.5, 5 and 10 ng/g, respectively. The LOD values for all the substances were below ng/g, with the exception of 6-chloronicotinic acid (LOD = 1.7 ng/g). The method was then applied to the analysis of 32 beebread samples and revealed the presence of 7 of the target substances. The most frequently detected pesticides belonged to the neonicotinoid family and were generally present at low concentrations, but in some cases exceeded 170 ng/g (acetamiprid and thiacloprid). Some pyrethroids were also detected (lambda-cyhalothrine and bifenthrine), but at very low levels Highlights * A single method for the analysis of pyrethroids and neonicotinoids in beebread. * An innovative and efficient sample extraction based on modified QuEChERS-UPLC-MS/MS. * Detection of the 13 target pesticides and metabolites at sub-ng/g levels. * 7 Target substances detected in beebread samples collected in hives.

International audience; BACKGROUND: The honey bee, Apis mellifera, is frequently used as a sentinel to monitor environmental pollution. In parallel, general weakening and unprecedented colony losses have been reported in Europe and the USA, and many factors are suspected to play a central role in these problems, including infection by pathogens, nutritional stress and pesticide poisoning. Honey bee, honey and pollen samples collected from eighteen apiaries of western France from four different landscape contexts during four different periods in 2008 and in 2009 were analyzed to evaluate the presence of pesticides and veterinary drug residues. METHODOLOGY/FINDINGS: A multi-residue analysis of 80 compounds was performed using a modified QuEChERS method, followed by GC-ToF and LC-MS/MS. The analysis revealed that 95.7%, 72.3% and 58.6% of the honey, honey bee and pollen samples, respectively, were contaminated by at least one compound. The frequency of detection was higher in the honey samples (n = 28) than in the pollen (n = 23) or honey bee (n = 20) samples, but the highest concentrations were found in pollen. Although most compounds were rarely found, some of the contaminants reached high concentrations that might lead to adverse effects on bee health. The three most frequent residues were the widely used fungicide carbendazim and two acaricides, amitraz and coumaphos, that are used by beekeepers to control Varroa destructor. Apiaries in rural-cultivated landscapes were more contaminated than those in other landscape contexts, but the differences were not significant. The contamination of the different matrices was shown to be higher in early spring than in all other periods. CONCLUSIONS/SIGNIFICANCE: Honey bees, honeys and pollens are appropriate sentinels for monitoring pesticide and veterinary drug environmental pollution. This study revealed the widespread occurrence of multiple residues in beehive matrices and suggests a potential issue with the effects of these residues alone or in combination on honey bee health.

International audience; This study describes the development of acetylcholinesterase (AChE), carboxylesterases (CaE1, CaE2, CaE3), glutathion-S-transferase (GST), alkaline phosphatase (ALP) and catalase (CAT) as enzyme biomarkers of exposure to xenobiotics such as thiamethoxam in the honey bee Apis mellifera. Extraction efficiency, stability under freezing and biological variability were studied. The extraction procedure achieved good recovery rates in one extraction step and ranged from 65 percent (AChE) to 97.3 percent (GST). Most of the enzymes were stable at −20 °C, except ALP that displayed a slight but progressive decrease in its activity. Modifications of enzyme activities were considered after exposure to thiamethoxam at the lethal dose 50 percent (LD50, 51.16 ng bee−1) and two sublethal doses, LD50/10 (5.12 ng bee−1) and LD50/20 (2.56 ng bee−1). The biomarker responses revealed that, even at the lowest dose used, exposure to thiamethoxam elicited sublethal effects and modified the activity of CaEs, GST, CAT and ALP. Different patterns of biomarker responses were observed: no response for AChE, an increase for GST and CAT, and differential effects for CaEs isoforms with a decrease in CaE1 and CaE3 and an increase in CaE2. ALP and CaE3 displayed contrasting variations but only at 2.56 ng bee−1. We consider that this profile of biomarker variation could represent a useful fingerprint to characterise exposure to thiamethoxam in the honey bee A. mellifera. This battery of honey bee biomarkers might be a promising option to biomonitor the health of aerial and terrestrial ecosystems and to generate valuable information on the modes of action of pesticides.

International audience; A novel multiresidue method was developed for the simultaneous analysis of 34 organochlorines, including chlorobenzenes, chlorophenols, chlorinated hydrocarbons and chlorinated olefins, in soil by GC-MS, using a QuEChERS-based extraction. The conventional QuEChERS method was optimised and, for the first time, the use of a non miscible-water solvent was required. The method was compared to ASE extraction, versatile technique widely used for the soils' extraction and QuEChERS-based method was shown to be the most efficient in terms of recoveries, simplicity and rapidity. For ASE, recoveries between 42% and 85% were obtained for the majority of the compounds. However, due to the high pressure, all volatile compounds were lost. In opposite, QuEChERS extraction allowed detection and quantification of all the compounds with recoveries between 60% and 100%. Moreover, no additional clean up by dispersive SPE on PSA was necessary, which allowed reducing the cost of the analysis. Performance of the method was assessed. The method was linear over the range of concentration of 10-5000 μg kg−1. Precision, expressed as intra-day precision and inter-day variation was verified at three concentrations. Limits of detection were from 2 to 50 μg kg−1 and limits of quantification from 7 to 170 μg kg−1 for the majority of the compounds (chlorobenzenes and chlorinated hydrocarbons and olefins), except for chlorophenols. The method was further applied to different soils coming from a contaminated industrial site, where a new environmental remediation process, using phytoremediation, was tested. The results showed that the method could be applied to any kind of soils (mineral or organic) and was appropriate to very volatile compounds which were not available with conventional technique.

International audience; One of the factors that may explain nowadays honeybees' colonies losses is the increasing presence of chemicals in the environment. The aim of this study is to obtain a global view of the presence of environmental contaminants in beehives and, develop a fast, cheap and sensitive tool to analyze environmental contaminants in apiarian matrices. A multi residue analysis was developed to quantify 80 environmental contaminants, pesticides and veterinary drugs, belonging to different chemical classes, in honeys, honeybees and pollens. It consists in a single extraction, based on a modified "QuEChERS method", followed by gas chromatography coupled with Time of Flight mass spectrometry (GC-ToF) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The "QuEChERS method" combines salting-out liquid-liquid extraction with acetonitrile and a dispersive-SPE clean up. It was adjusted to honey and especially to honeybee and pollen, by adding a small fraction of hexane in acetonitrile to eliminate lipids that interfere with mass spectrometry analysis. This method, combined with accurate and sensitive detection, allowed quantification and confirmation at levels as low as 10 ng/g, with recoveries between 60 and 120%. Application to more than 100 samples of each matrix was achieved for a global view of pesticide presence in the honeybee environment. Relatively high percentages of honeys, honeybees and pollens were found to be contaminated by pesticides used to combat varroa but also by fungicides like carbendazim and ubiquitous contaminants.