Performance assessment of an equine metabolomics model for screening a range of anabolic agents.

INTRODUCTION: Despite their ban, Anabolic Androgenic Steroids (AAS) are considered as the most important threat for equine doping purposes. In the context of controlling such practices in horse racing, metabolomics has emerged as a promising alternative strategy to study the effect of a substance on metabolism and to discover new relevant biomarkers of effect. Based on the monitoring of 4 metabolomics derived candidate biomarkers in urine, a prediction model to screen for testosterone esters abuse was previously developed. The present work focuses on assessing the robustness of the associated method and define its scope of application. MATERIALS AND METHODS: Several hundred urine samples were selected from 14 different horses of ethically approved administration studies involving various doping agents' (AAS, SARMS, ß-agonists, SAID, NSAID) (328 urine samples). In addition, 553 urine samples from untreated horses of doping control population were included in the study. Samples were characterized with the previously described LC-HRMS/MS method, with the objective of assessing both its biological and analytical robustness. RESULTS: The study concluded that the measurement of the 4 biomarkers involved in the model was fit for purpose. Further, the classification model confirmed its effectiveness in screening for testosterone esters use; and it demonstrated its ability to screen for the misuse of other anabolic agents, allowing the development of a global screening tool dedicated to this class of substances. Finally, the results were compared to a direct screening method targeting anabolic agents demonstrating complementary performances of traditional and omics approaches in the screening of anabolic agents in horses.

Evidence of complementarity between targeted and non-targeted analysis based on liquid and gas-phase chromatography coupled to mass spectrometry for screening halogenated persistent organic pollutants in environmental matrices.

This study explored the complementarity between targeted (TS) and non-targeted screening (NTS) based on liquid and gas-phase chromatography coupled to (high-resolution) mass spectrometry (LC-/GC-(HR)MS) for the comprehensive characterization of organohalogen fingerprints within a set of Lake Ontario lake trout samples. The concentrations of 86 legacy, emerging and novel halogenated compounds (HCs), were determined through 4 TS approaches involving no less than 6 hyphenated systems. In parallel, an innovative NTS strategy, involving both LC and GC-Q-Orbitrap, was implemented to specifically highlight halogenated signals. Non-targeted HRMS data were processed under the HaloSeeker software based on Cl and Br isotopic ratio and mass defect to extend the screening to unsuspected and unknown HCs. A total of 195 halogenated mass spectral features were characterized in the Lake Ontario lake trout, including well known HCs (PCBs, PBDEs, PBBs, DDT and their degradation products), emerging HCs (novel brominated flame retardants, short-, medium- and long-chain chlorinated paraffins) or suggested molecular formula (mainly polychlorinated ones). Among the 122 HCs highlighted by TS, only 21 were identified by NTS. These results fueled a discussion on the potential and limitations of both approaches, and the current position of NTS within environmental and health monitoring programs.

Metabolomics in chemical risk analysis - A review.

Exposure to chemical hazards is a growing concern in today's society, and it is of utmost interest to know the levels of exposure to chemicals and the risks associated with such exposure in order to implement effective health prevention strategies. Chemical risk analysis represents a complex and laborious task due to the large number of known substances, but also unknown compounds and emerging risks that must be addressed. In this challenging scenario, the study of metabolic perturbations induced by exposure to a given chemical hazard has recently emerged as an interesting alternative approach to apply in chemical risk analysis. Specifically, the biomarkers of effect identified by metabolomics are expected to reveal the adverse effects of chemicals and further link exposure to disease development. In this context, analytical chemistry has become an essential part of the strategy to highlight such biomarkers. The corollary is that the relevance of the discovered biomarkers will largely depend on both the quality of the analytical approaches implemented and the part of the metabolome covered by the analytical technique used. This review focuses on describing significant applications of metabolomics in the field of chemical risk analysis. The different risk assessment steps, including hazard identification, dose-response assessment and exposure assessment, and risk management are addressed through various examples to illustrate that such an approach is fit-for-purpose and meets the expectations and requirements of chemical risk analysis. It can be considered as an innovative tool for predicting the probable occurrence and nature of risks, while addressing the current challenges of chemical risk analysis (e.g. replacement, reduction and refinement (3R) of animal testing, effects of exposure to chemical mixtures at low doses, etc.), and with the aim of responding to chemical exposures concerns in a holistic manner and anticipating human health problems.

Do farming conditions influence brominated flame retardant levels in pig and poultry products?

Brominated flame retardants (BFR) are primarily used as flame retardant additives in insulating materials. These lipophilic compounds can bioaccumulate in animal tissues, leading to human exposure via food ingestion. Although their concentration in food is not yet regulated, several of these products are recognised as persistent organic pollutants; they are thought to act as endocrine disruptors. The present study aimed to characterise the occurrence of two families of BFRs (hexabromocyclododecane (HBCDD) and polybrominated diphenyl ethers (PBDE)) in hen eggs and broiler or pig meat in relation to their rearing environments. Epidemiological studies were carried out on 60 hen egg farms (34 without an open-air range, 26 free-range), 57 broiler farms (27 without an open-air range, 30 free-range) and 42 pig farms without an open-air range in France from 2013 to 2015. For each farm, composite samples from either 12 eggs, five broiler pectoral muscles or three pig tenderloins were obtained. Eight PBDE congeners and three HBCDD stereoisomers were quantified in product fat using gas chromatography-high-resolution mass spectrometry, or high-performance liquid chromatography-tandem mass spectrometry, respectively. The frequencies of PBDE detection were 28% for eggs (median concentration 0.278 ng/g fat), 72% for broiler muscle (0.392 ng/g fat) and 49% for pig muscle (0.403 ng/g fat). At least one HBCDD stereoisomer was detected in 17% of eggs (0.526 ng/g fat), 46% of broiler muscle (0.799 ng/g fat) and 36% of pig muscle (0.616 ng/g fat). Results were similar in concentration to those obtained in French surveillance surveys from 2012 to 2016. Nevertheless, the contamination of free-range eggs and broilers was found to be more frequent than that of conventional ones, suggesting that access to an open-air range could be an additional source of exposure to BFRs for animals. However, the concentration of BFRs in all products remained generally very low. No direct relationship could be established between the occurrence of BFRs in eggs and meat and the characteristics of farm buildings (age, building materials). The potential presence of BFRs in insulating materials is not likely to constitute a significant source of animal exposure as long as the animals do not have direct access to these materials.

Steroidogenic potential of human fetal kidney at early gestational age.

Steroidogenic potential of the human fetal kidney (hFK) at the end of first trimester is poorly investigated. Little is known about the ontogeny of steroidogenic enzymes and activities of steroidogenic pathways in the hFK at early pregnancy. Our aim was to explore steroidogenesis and the expression of steroidogenic enzymes in the hFK at gestational weeks (GW) 9-12. Steroids in the hFK were analyzed by gas chromatography/coupled to tandem mass spectrometry. The expression of steroidogenic enzymes in the hFK at GW 9-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We observed that the hFK produced substantial amount of steroids of the Δ5 and Δ4 pathways and several steroid precursors in the biosynthesis of DHT via the backdoor pathway but not DHT itself. The levels of steroids and expression of relevant steroidogenic enzymes (e.g., CYP17A1, HSD3B1, HSD3B2, CYP11B1 and AKR1C4) we significantly higher in the hFK at GW11-12 compared to GW9. We also found the expression of sex steroid receptors (e.g., AR, ERα and ERß) in the hFK at GW9-12. No sex-dependent differences in the levels of all identified steroids and expression of steroidogenic enzymes in the hFK from male and female fetuses were found. Altogether, our data indicate that the hFK at early pregnancy is steroidogenic organ with potential to synthesize multiple steroids that may play an important role in the formation and development of this organ in humans.

Quantitative method for conjugated metabolites of bisphenol A and bisphenol S determination in food of animal origin by Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry.

With the objectives of both generating bisphenols (BPs) conjugates occurrence data in food from animal origin but also investigating the origin of associated contamination, the present study deals with the development of an efficient analytical method aiming at monitoring both BPA and BPS conjugated metabolites in food from animal origin. The objective of such monitoring is to determine the origin of BPs contamination (FCM or animal contamination). The targeted compounds were BPA-monoglucuronide (BPA-1G), BPA-diglucuronide (BPA-2G), BPA-monosulfate (BPA-1S), BPA-disulfate (BPA-2S) and BPS-monoglucuronide (BPS-1G). The developed standard operating procedure includes a preliminary solid-liquid extraction step followed by two successive solid phase extraction (SPE) stages, using successively a non-polar phase and a strong cation exchange polymer. Quantification was achieved according to both the isotopic dilution and surrogated quantification methods, using 13C-BPA-1G and BPA-d6-1S as internal standards. Linearity was validated (R2 > 0.99) for each molecule within the concentration range [0-10] µg kg-1. Detection limits ranged from 0.02 µg kg-1 (BPA-1G in muscle, BPA-1S and BPA-2G in liver) to 0.50 µg kg-1 (BPA-2S in muscle). The strategy was then proven on liver samples collected from pregnant ewes subcutaneously exposed to BPA during 105 days, at 50 µg kg-1 per day. BPA-1G, BPA-2G and BPA-1S were detected and quantified at a concentration of 3.81 µg kg-1, 0.80 µg kg-1 and 0.09 µg kg-1, respectively. The analytical method was finally implemented on fifty unpacked food samples from animal origin in which significant free BPA concentrations were previously measured. Since no metabolites of BPA could be measured (

Ontogenesis of human fetal testicular steroidogenesis at early gestational age.

The onset of steroidogenesis in human fetal testes (HFT) during the first trimester is poorly investigated. One important unresolved question is the ontogeny of steroidogenic enzymes and formation of steroidogenic pathways in the HFT at early pregnancy. Our aim was to explore steroidogenesis, the expression of steroidogenic enzymes and their maturation in the HFT at gestational weeks (GW) 8-12. Steroids in the HFT were analyzed by gas chromatography/coupled to tandem mass spectrometry. The expression of steroidogenic enzymes in the HFT at GW8-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We demonstrated that the HFT at GW8-9 produced low level of testosterone via the Δ4 pathway and progesterone was the major steroid found in the testicular tissue. In contrast, more mature Leydig cells from the HFT at GW11-12 synthesized high levels of androgens via the Δ5 pathway. We also observed a significant upregulation of the expression of StAR, CYP11A1, CYP17A1 and its accessory proteins, P450 oxidoreductase (POR) and cytochrome b5 in the HFT at GW11-12 compared to GW8-9. Altogether, our data suggest that that human fetal Leydig cells differentiate rapidly at the end of the first trimester by acquiring capacity to express high levels of steroidogenic enzymes and switch from the Δ4 to the Δ5 pathways to synthesize high levels of androgens due to maturation of the CYP17-POR-b5 complex.

The human genital tubercle is steroidogenic organ at early pregnancy.

It is generally accepted that androgens produced by fetal Leydig cells (FLC) control proper masculinization of the male external genitalia. Here, we hypothesized that the human genital tubercle (GT) has potential to synthesize androgens independently of FLC at early pregnancy. We observed that human GT of both genders have capacity to synthesize steroids of the Δ4, Δ5 and alternative pathway of DHT synthesis including the androgen itself. The presence of steroids in the GT was associated with the expression of corresponding steroidogenic enzymes. Levels of steroids and the expression of steroidogenic enzymes were similar in the GT from male and female fetuses. In contrast to the GT, the human fetal testis synthesized DHT from testosterone but not via the alternative pathway. Our findings strongly suggest that the human GT at early pregnancy can synthesize DHT via the alternative pathway, which may play an important role in organogenesis of the urethra.

Occurence of legacy and novel brominated flame retardants in food and feed in France for the period 2014 to 2016.

Determination of the occurrence levels of legacy and novel BFRs is today required to better understand the trends of BFRs contamination in food consecutive to the EU PBDEs restrictions and to proceed to a recent human food exposure in parallel. Therefore, concentrations of a large set of brominated flame retardants (BFRs) (n = 27) including PBDEs, HBCDDs, TBBPA and novel flame retardants (nBFRs) have been determined in more than 600 food and feed samples collected between 2014 and 2016 in the context of French monitoring plans. Although legacy BFRs had already been studied in France, such a survey constituted the very first determination of nBFRs occurrence in foodstuffs at the national level. The concentration levels measured in fish and fish products were in general higher than in the other food categories. PBDEs were detected in 70% of the samples and were observed as the most abundant congeners (representing 80% of the sum of the monitored BFRs), while α-HBCDD could also be considered as a predominant congener (up to 26% of the sum of the monitored BFRs in fishes). nBFRs concentration levels were most of the time below the LOQ, except PBT, PBBz and HBBz which were more frequently detected at low levels. Also investigated in the study, BRPs exhibited high concentration levels in crustaceous (maximum value > 2700 pg/g ww).

Development of a Cryptosporidium-arsenic multi-risk assessment model for infant formula prepared with tap water in France.

Tap water is used in France to reconstitute powder infant formula, although it is not sterile and possibly contaminated by microbiological and chemical hazards. The present study aims to quantify risks of using tap water in France for the preparation of infant formula, during the first six months of life. Cryptosporidium and arsenic were selected as hazards of greatest concern in microbiology and chemistry, respectively. A probabilistic model was developed using French (when available) and European (alternatively) data. Second order Monte Carlo simulation was used to separate uncertainty and variability of inputs. Outputs were expressed at the individual level as probability of illness and at the population level, using a common metric, the DALY (Disability Adjusted Life Year). Two scenarios of milk preparation were considered: with un-boiled or boiled tap water. Consuming infant formula rehydrated with un-boiled tap water during the first six months of life led to a total of 2250 DALYs per 100,000 infants (90% uncertainty interval [960; 7650]) for Cryptosporidium due to diarrhea, and 1 DALY [0.4; 2] for arsenic due to expected lifetime risk of lung and bladder cancer as a result of early exposure in life. For the entire population, boiling water would suppress the risk from Cryptosporidium. In contrast, the incremental cancer risk was low at the population level but elevated for 5% of the population exposed to high levels of arsenic. A stringent monitoring of tap water supply points should be continued. This multi-risk assessment model could help public health authorities and managers in evaluating both microbiological and chemical safety issues associated with using infant formula prepared with tap water.

Comparison between liquid chromatography and supercritical fluid chromatography coupled to mass spectrometry for beta-agonists screening in feeding stuff.

ß-agonistic drugs have been forbidden as growth promoters in rearing animals in Europe since the late 1980s (Dir 96/22/EC). Specific and sensitive analytical methods based on UHPLC-MS/MS allow to monitor a large set of these substances. However, optimal performances are not observed for all the target analytes, especially for those exhibiting the highest polarities. We developed an SFC-MS/MS approach to cover the huge elution window of ß-agonists, from the most polar which are usually eluted in the void volume when using reversed phase chromatography in conventional HPLC to the most apolar ones. The objective was to reach performances in accordance with the European Union recommended level in feeding stuff, i.e. 50 µg kg-1. LC/MS and SFC/MS performances were thoroughly compared in terms of analytical validation data (linearity, selectivity, recovery rates, reproducibility, compounds identification, trueness, decision limit (CCα) and detection capability (CCß)) for 6 ß-agonistic drugs, namely bromobuterol, clenbuterol, isoxsuprine, ractopamine, salbutamol and zilpaterol. As a result, the SFC approach appeared complementary to the LC one because the elution order of compounds was totally different from the one obtained with a classical C18 stationary phase. Moreover, the UPLC-MS/MS approach gave a better response linearity and more accurate values, whereas SFC-MS/MS provided greater data for identification purposes, reproducibility and sensitivity. Both analytical approaches enabled the detection of targeted ß-agonists at a lower concentration than the recommended one (50 µg kg-1).

Androgenic potential of human fetal adrenals at the end of the first trimester.

The onset of steroidogenesis in human fetal adrenal glands (HFA) during the first trimester is poorly investigated. An unresolved question is the capacity of the HFA to produce potent androgen DHT via conventional and/or the backdoor pathway(s) at the end of first trimester, when androgen-responsive organs are developed. Our aim was to explore steroidogenesis and the expression of steroidogenic enzymes and transcription factors in HFA at gestational weeks (GW) 9-12 with focus on their androgenic potential. Steroids in the HFA were analyzed by gas chromatography/mass spectrometry. The expression of steroidogenic enzymes and transcription factors in the HFA at GW9-12 was investigated by qPCR, automated Western blotting and immunohistochemistry. We demonstrated that during GW9-12 HFA produced steroids of the ∆5, ∆4 and the backdoor pathways of the biosynthesis of DHT, though the latter was limited to production of 17α-OH-dihydroprogesterone, androsterone and androstanedione without further conversion to DHT. The only androgens identified in the HFA were testosterone and androsterone, a precursor in the biosynthesis of DHT. We also observed higher levels of CYP17A1 but low expression of 3ßHSD2 at GW11-12 in the HFA. Elevated levels of CYP17A1 were associated with an increased expression of SF-1 and GATA-6. Altogether, our data demonstrate that of those steroids analyzed, the only potent androgen directly produced by the HFA at GW9-12 was testosterone. The onset of steroidogenesis in the HFA is a complex process that is regulated by the coordinated action of related transcription factors.

Country-specific chemical signatures of persistent organic pollutants (POPs) in breast milk of French, Danish and Finnish women.

The present study compares concentrations and chemical profiles of an extended range of persistent organic pollutants (dioxins, polychlorobiphenyls, brominated flame retardants and organochlorine pesticides) in breast milk samples from French (n = 96), Danish (n = 438) and Finnish (n = 22) women. Median exposure levels observed in French women (WHO-TEQ2005 PCDD/F = 6.1 pg/g l.w., WHO-TEQ2005 dl-PCB = 4.3 pg/g l.w., sum of 6 ndl-PCB = 85.2 ng/g l.w., sum of 7 i-PBDE = 1.5 ng/g l.w.) appeared overall lower than in Danish and Finnish women for all examined POPs, except for α-HBCD (2-fold higher level at 0.6 ng/g l.w.). Furthermore, the observed exposure levels of dioxins and PCBs were higher in Danish women (WHO-TEQ2005 PCDD/F = 13.2 pg/g l.w., WHO-TEQ2005 dl-PCB = 6.6 pg/g l.w., sum of 6 ndl-PCB = 162.8 ng/g l.w.) compared to Finnish women (WHO-TEQ2005 PCDD/F = 9.0 pg/g l.w., WHO-TEQ2005 dl-PCB = 4.6 pg/g l.w., sum of 6 ndl-PCB = 104.0 ng/g l.w.), whereas the concentrations of PBDEs were similar for Danish and Finnish women (sum of 7 i-PBDE = 4.9 and 5.2 ng/g l.w. respectively). The organochlorine (OC) pesticide contamination profile, determined in a subset of French samples, was dominated by p,p'-DDE (56.6%), followed by ß-HCH (14.2%), HCB (9.7%) and dieldrin (5.2%), while other compounds were only minor contributors (<5%). The three countries appeared to be discriminated by the observed contamination patterns of the PCDD/F versus PCB, and the 1,2,3,6,7,8-HxCDD versus 1,2,3,4,7,8-HxCDD ratios, in addition to the relative contributions of specific congeners to the contamination profile (PCBs #118 and #156, PBDEs #28, #47, #99 and #153). In conclusion, unique chemical signatures were observed for each country on the basis of some POP congeners. Future biomonitoring studies will need to consider the high variability of individual exposure profiles in relation to multiple exposure sources but also physiological and metabolic differences.

Simultaneous determination of 16 brominated flame retardants in food and feed of animal origin by fast gas chromatography coupled to tandem mass spectrometry using atmospheric pressure chemical ionisation.

A gas chromatography tandem mass spectrometry method using atmospheric pressure chemical ionisation was developed for the monitoring of 16 brominated flame retardants (7 usually monitored polybromodiphenylethers (PBDEs) and BDE #209 and 8 additional emerging and novel BFRs) in food and feed of animal origin. The developed analytical method has decreased the run time by three compared to conventional strategies, using a 2.5m column length (5% phenyl stationary phase, 0.1mm i.d., 0.1µmf.t.), a pulsed split injection (1:5) with carrier gas helium flow rate at 0.48mLmin(-1) in one run of 20 min. For most BFRs, analytical data were compared with the current analytical strategy relying on GC/EI/HRMS (double sector, R=10000 at 10% valley). Performances in terms of sensitivity were found to meet the Commission recommendation (118/2014/EC) for nBFRs. GC/APCI/MS/MS represents a promising alternative for multi-BFRs analysis in complex matrices, in that it allows the monitoring of a wider list of contaminants in a single injection and a shorter run time.

Thyroid endocrine status of wild European eels (Anguilla anguilla) in the Loire (France). Relationships with organic contaminant body burdens.

In teleost fish, thyroid function is involved in various critical physiological processes. Given the complexity of the hypothalamo-pituitary-thyroid (HPT) axis, a large number of genes and proteins can be the potential target of endocrine-disrupting compounds (EDCs). The aim of this study was to evaluate, in yellow and silver European eels (Anguilla anguilla), potential effects of EDCs on thyroid status by analyzing the associations between EDC body burdens and thyroid hormones (THs). In yellow individuals, greater free T3/T4 ratios (FT3/FT4) and lower plasma FT4 levels were associated with greater concentrations of polychlorinated biphenyls (PCBs) and polybrominated diphenyl ethers (PBDEs), as highlighted by significant correlations with many congeners. Few positive relationships with alkylphenols were noticed. In contrast, silver eels usually exhibited less significant correlations between THs and contaminant loads. Expression of a series of genes involved in the HPT axis was also investigated in the silver individuals. Concerning mRNA expression in silver females, some main correlations were noticed: thyroid-stimulating hormone (TSHß) gene expression was significantly correlated to numerous PCBs, and hepatic mRNA levels of deiodinase 2 (Deio 2) were negatively correlated to 2-hydroxyfluorene (2-OHF) and 2-naphtol (2-OHNa). Thyroid receptor (TRα and TRß) mRNA levels exhibited weak negative correlations with some PBDEs in silver females and males. Hepatic vitellogenin (Vtg) mRNA levels were detected in all silver males but at lower levels than in silver females. In males, Vtg mRNA levels were positively associated to FT4/TT4. In silver females, strong positive correlations were found between congeners of PCBs, PBDEs and PFAS suggesting potential estrogenic effects. Overall, the observed results indicate that several organic contaminants, mainly dl-, ndl-PCBs and PBDEs, could be associated with changes in thyroid homeostasis in these fish, via direct or indirect interactions with peripheral deiodination, metabolism of T4 and mechanisms involved in TSHß, Deio 2 and Vtg gene transcription.

Occurrence of POPs and other persistent organic contaminants in the European eel (Anguilla anguilla) from the Loire estuary, France.

The chemical contamination of the Loire estuary by three classes of persistent organic pollutants (POPs): the polychlorinated biphenyls (PCBs), the polybrominated diphenyl ethers (PBDEs) and the perfluorinated and polyfluorinated alkyl substances (PFAS), and three families of organic contaminants, the alkylphenols (APs), the polycyclic aromatic hydrocarbon metabolites (OH-PAHs) and the bisphenol A (BPA) were investigated in the muscles and bile of European eel (Anguilla anguilla). Yellow eels (n=30) were caught in three different points along the estuary to highlight variations between sites and sources of contaminations. Silver eels (n=15) were also studied to compare contaminant impregnation between different life stages of the species. Average concentrations in the muscles of the eel ranged between: 857 and 4358 ng/gLW for the PCBs, 26 and 46 ng/gLW for the PBDEs, 130 and 1293 ng/gLW for the PFAS; and in bile: 31 and 286 µg/g protein for the APs, 9 and 26 µg/g protein for the OH-PAHs and ND-1213 µg/g protein for the BPA. Among PCBs, PCB 153 (40% contribution to the sum of PCBs) was predominant in all eel muscles. PBDE 47 (60%) was the most predominant PBDE congeners, while perfluorooctanesulfonic acid (85%) was the most widely detected PFAS. For APs, 4p-nonylphenol (91%) was the most abundant and for the OH-PAHs, it was 1OH-Pyrene (63%). All the eels exceeded the environmental quality standards (EQS) for biota for the PBDEs and about 75% were higher than the EQS specific to PFOS. Finally, 20% of the analyzed eels presented TEQ concentrations above the maximum limits for lipid-rich species. These results supplied new data on the occurrence, levels, and patterns of 53 organic chemicals in the eels from the Loire estuary and they highlighted the need of further investigations focused notably on the potential effects of these chemicals on this species and their analysis in the water and sediments of the estuary.

Development and validation of a specific and sensitive gas chromatography tandem mass spectrometry method for the determination of bisphenol A residues in a large set of food items.

BPA-containing products are widely used in foodstuffs packaging as authorized within the European Union (UE no. 10/2011). Therefore, foods and beverages are in contact with BPA which can migrate from food contact material to foodstuffs. An accurate assessment of the exposure of the consumers to BPA is crucial for a non-ambiguous risk characterization. In this context, an efficient analytical method using gas chromatography coupled to tandem mass spectrometry (GC-MS/MS), in the selected reaction monitoring (SRM) mode, was developed for the quantification of BPA in foodstuffs at very low levels (<0.5µgkg(-1)). A standard operating procedure, based on the combination of two successive solid phase extractions (SPE), was developed for various liquid and solid foodstuffs. The use of (13)C12-BPA as internal standard allowed accurate quantification of BPA by isotopic dilution. Control charts based on both blank and certified materials have been implemented to ensure analytical data quality. The developed analytical method has been validated according to in-house validation requirements. R(2) was better than 0.9990 within the range [0-100µgkg(-1)], the trueness was 4.2%. Repeatability and within-laboratory reproducibility ranged from 7.5% to 19.0% and 2.5% to 12.2%, respectively, at 0.5 and 5.0µgkg(-1) depending on the matrices tested for. The detection and quantification limits were 0.03 and 0.10µgkg(-1), respectively. The reporting limit was 0.35µgkg(-1), taking into account the mean of the laboratory background contamination. The global uncertainty was 22.2% at 95% confidence interval.

Dietary exposure to perfluoroalkyl acids of specific French adult sub-populations: high seafood consumers, high freshwater fish consumers and pregnant women.

Perfluoroalkyl acids (PFAAs) are globally found in various media, including food and especially fishery products. In the present study, the dietary exposure to 15 perfluoroalkyl acids was assessed for 3 French adult populations, namely high seafood consumers, high freshwater fish consumers, and pregnant women. Purified food extracts were analysed by LC-MS/MS and PFBA, PFPA, PFHxA, PFHpA, PFOA, PFNA, PFDA, PFUnA, PFTrDA, PFTeDA, PFBS, PFHxS, PFHpS, PFOS and PFDS were monitored and quantified according to the isotope dilution principle. Under lower bound (LB) hypothesis (i.e. contamination values

Food risk assessment for perfluoroalkyl acids and brominated flame retardants in the French population: results from the second French total diet study.

To determine the exposure of the French population to toxic compounds contaminating the food chain, a total diet study was performed in France between 2007 and 2009. This study was designed to reflect the consumption habits of the French population and covered the most important foods in terms of consumption, selected nutrients and contribution to contamination. Based on French consumption data, the present study reports the dietary exposure to perfluoroalkyl acids (16 congeners) and brominated flame retardants (polybrominated diphenyl ethers, hexabromocyclododecane and polybrominated biphenyls). Comparison of the calculated dietary exposures with the generally accepted health-based guidance values revealed that most compounds do not pose any risk. There are however knowledge gaps for some congeners in these large chemical classes.

Dioxin-like, non-dioxin like PCB and PCDD/F contamination in European eel (Anguilla anguilla) from the Loire estuarine continuum: spatial and biological variabilities.

To characterize the eel contamination by dioxin-like (dl) and non dioxin-like (ndl) polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins and furans (PCDD/Fs), sixty-two eels from the Loire estuary (France) were analyzed. PCB contamination significantly increased from glass eel stage (3.71 ± 1.85 and 15.2 ± 4.2 ng g(-1) dw) to other life stages (for yellow eels: 62.8 ± 34.4 and 382 ± 182 ng g(-1) dw; for silver eels: 93.7 ± 56.3 and 463 ± 245 ng g(-1) dw respectively for dl and ndl-PCBs). An inter-site variability based on PCB levels and profiles was observed among the three studied sites. For glass eels, the profile was mainly characterized by less chlorinated PCBs contrary to the other eels, displaying a different bioaccumulation pathway. Overall, the contamination level in the eels from this estuary was shown to be low for PCDD/Fs and intermediate for dl and ndl-PCBs, compared to other international/national areas. However, more than 60% of the studied silver eels displayed higher values for PCDD/F and dl-PCB WHO2005 TEQ than the EU permissible level of 10 pg g(-1) ww. This statement suggests a potential exposure to PCBs through eel consumption, especially with silver eels, and also points out apparent contamination that could eventually affect the reproductive success of the species.