A new derivatizing reagent for the determination of 5-nitro-2-furaldehyde in trout muscle by liquid chromatography-tandem mass spectrometry.

The 5-nitro-2-furaldehyde (5-NF) is an aldehyde aromatic organic compound that has been envisaged as an alternative marker for detecting nitrofurazone treatment abuse and to avoid the false positive results induced by the semicarbazide. Analyzing 5-NF presents challenges, and its derivatization reaction with hydrazine reagents is required to enhance the capability of its detection and its identification. This study aims at developping an analytical method for 5-NF determination in trout muscle samples based on chemical derivatization prior to analysis by liquid chromatography-tandem mass spectrometry. Four commercially available hydrazine reagents, namely: N,N-Dimethylhydrazine (DMH), 4-Hydrazinobenzoic acid (HBA), 2,4-Dichlorophenylhydrazine (2,4-DCPH) and 2,6-Dichlorophenylhydrazine (2,6-DCPH) were proposed for the first time as derivatizing reagents in the analysis of 5-NF. The derivatization reaction was simultaneously performed along with the extraction method in acidic condition using ultrasonic assistance and followed by liquid extraction using acetonitrile. The efficiency of the chemical reaction with 5-NF was examined and the reaction conditions including the concentration of hydrochloric acid, pH, temperature, reaction time and the concentration of the derivatizing reagents were optimized. Experiments with fortified samples demonstrated that 2,4-DCPH derivatizing reagent at 20 mM for 20 min of ultrasonic treatment under acidic condition (pH 4) gave an effective sample derivatization method for 5-NF analysis. Under the optimized conditions, the calibration curves were linear from 0.25 to 2 µg kg-1 with coefficient of determination >0.99. The recoveries ranged from 89 % to 116 % and precision was less than 13 %. The limit of detection and quantification were 0.1 and 0.2 µg kg-1, respectively.

Occurence of antimicrobial residues in milk and labneh consumed in Lebanon.

Antimicrobials are administered in livestock for different uses leading to milk contamination and several undesirable effects. Because there is a lack of surveillance of antimicrobial residues (AMRs) in milk and dairy products in Lebanon, this study aims to determine the occurrence of AMRs in 90 Lebanese samples of milk and labneh (concentrated yoghurt). Multi-residue screening methods with suitable sample preparations were applied to detect 71 AMRs in milk and labneh, respectively, using LC-MS/MS. Of the total number of samples, 71% was contaminated with AMRs and (fluoro)quinolones and macrolides were the most detected families. Additional confirmation tests proved that 6.7% of the milk samples were non-compliant for the macrolides tilmicosin, tulathromycin and spiramycin. Moreover, some labneh prepared from contaminated milk samples was analysed to determine the fate of AMRs during the manufacturing process. The results showed that some AMRs could be concentrated, eliminated or degraded, based on their physicochemical characteristics.

Stability study of selected veterinary drug residues spiked into extracts from different food commodities.

Analyte stability is more commonly a confounding factor in analytical chemistry than many analysts recognize. In this study, we assessed the stability of 31 common veterinary drugs in water and final extracts of bovine (milk and kidney/liver) and chicken (muscle and egg) matrices. Two different sample preparation methods were evaluated for one-month storage of the final extracts at typical room, refrigerator, and freezer temperatures. Liquid chromatography - mass spectrometry (LC-MS) by triple quadrupole and high-resolution techniques was used for analysis of the extracts spiked at different relevant concentrations for general regulatory purposes (10-1000 ng/g sample equivalent). Comparison of results between two labs demonstrated that stable drugs (≤20% loss) at all tested conditions consisted of danofloxacin, enrofloxacin, florfenicol, flubendazole, hydroxy-flubendazole, flumequine, flunixin, 5-hydroxy-flunixin, lincomycin, and meloxicam. The tested drugs found to be the most unstable (>20% loss at room temperature within a matter of days) consisted of the ß-lactams (ampicillin, cefalexin, cloxacillin, and penicillin G). Curiously, the following antibiotics (mostly macrolides) were apparently more stable in sample extracts than water: emamectin, erythromycin, ivermectin, lasalocid, monensin, tilmicosin, tulathromycin, and tylosin. Those and the other drug analytes (ciprofloxacin, doxycycline, florfenicol amine, 2-amino-flubendazole, oxytetracycline, sulfadiazine, sulfadimethoxine, sulfamethazine, and trimethoprim) were mostly stable for a month in refrigerated extracts, especially at higher concentrations, but not in all cases. In practice, freezer storage of extract solutions was found to be acceptable for at least a month, with a few exceptions.

Fate of Sulfonamides and Tetracyclines in Meat during Pan Cooking: Focus on the Thermodegradation of Sulfamethoxazole.

Although antimicrobials are generally found in trace amounts in meat, the human health risk they bear cannot be ignored. With the ultimate aim of making a better assessment of consumer exposure, this study explored the effects of pan cooking on sulfonamides and tetracyclines in meat. Screening of these antimicrobials in cooked meat was first performed by the European Union Reference Laboratory on the basis of HPLC-MS/MS analyses. A proof of concept approach using radiolabeling was then carried out on the most cooking-sensitive antimicrobial-sulfamethoxazole-to assess if a thermal degradation could explain the observed cooking losses. Degradation products were detected thanks to separation by HPLC and monitoring by online radioactivity detection. HPLC-Orbitrap HRMS analyses completed by 1D and 2D NMR experiments allowed the structural characterization of these degradation compounds. This study revealed that cooking could induce significant antimicrobial losses of up to 45% for sulfamethoxazole. Six potential degradation products of 14C-sulfamethoxazole were detected in cooked meat, and a thermal degradation pattern was proposed. This study highlights the importance of considering the cooking step in chemical risk assessment procedures and its impact on the level of chemical contaminants in meat and on the formation of potentially toxic breakdown compounds.

Development and applicability of a multi-residue method for dyes, including new residue markers, to detect drug misuse in aquaculture.

A qualitative and quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the sensitive and exhaustive analysis of residues from triarylmethane dyes, triarylmethane-derivative dyes, phenothiazines, phenoxazines and xanthenes in aquaculture samples. For a wider and more robust detection of dye misuse on farms, other residue markers were also included the leuco forms of brilliant green, crystal violet and malachite green; one direct metabolite of Victoria pure blue BO and methylene blue and three bile acids, which are endogenous markers of the effects of dye contamination in fish. We optimised the extraction method by comparing several extraction solvents and sample solvents reported in the literature to have the best extraction efficiency. The residues were determined using a positive electrospray ionisation source. We assessed the parameters of this LC-MS/MS method by evaluating the matrix effects, identification and quantitative parameters according to the criteria stipulated in the European Commission Decision No. 2002/657/EC. A study on the applicability of the method was conducted on various aquaculture species and on a positive catfish.

Development of a liquid chromatography-tandem mass spectrometry method to determine colistin, bacitracin and virginiamycin M1 at cross-contamination levels in animal feed.

Cross-contamination of animal feed with antibiotics may occur during manufacturing in feed mills, because shared production lines can be used for medicated and non-medicated feed, but may also occur during transport, storage and at the farm level. This is a major issue in the current context where antimicrobial usage must be controlled in order to maintain their effectiveness. A LC-MS/MS method was developed for the determination of colistin, bacitracin A and virginiamycin M1 in feed for pigs, poultry and rabbits at concentrations similar to those encountered in cross-contamination. After investigating various issues related to colistin behaviour and matrix effects, we successfully validated this method according to the requirements of European regulations in terms of linearity, trueness, precision, limit of quantification and limit of decision. Trueness ranged 88.6-107.8% and precision ranged 12.6-21.2%. We then applied this method to the analysis of medicated pig feed to check the performance of the method on "real" samples of medicated feed. We subsequently analysed non-medicated pig, and rabbit feed samples, collected directly on farms, to check the rate of cross-contamination. No samples were contaminated by colistin, bacitracin, or virginiamycin.

Confirmation of five nitrofuran metabolites including nifursol metabolite in meat and aquaculture products by liquid chromatography-tandem mass spectrometry: Validation according to European Union Decision 2002/657/EC.

LC-MS/MS method for confirmation of nitrofuran metabolites in meat and aquaculture products, including the nifursol metabolite (DNSH), was developed. The nitrofuran metabolites investigated were as follows: 3-amino-2-oxazolidinone (AOZ), 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ), 1-aminohydantoine (AHD), semicarbazide (SEM) and 3,5-dinitrosalicylic acid hydrazide (DNSH). The sample preparation includes a washing step, allowing to analyze only the fraction of protein-bound residues. The final optimized recovery solvent for injection was found to be a mixture of ammonium acetate 2 mM/acetonitrile (60/40 ; v/v). Matrix effects and stability in biological matrix and standard solution for DNSH have been also carried out. Method performances were assessed using criteria of the Decision (EC) No 2002/657 and considering the proposed-for-adoption reference point for action (RPA) of 0.50 µg.kg-1 under Reg 1871/2019. Trueness ranged 86.5%-103.7% and precision ranged 2.0%-6.5% on intra-laboratory reproducibility conditions. Decision limit (CCα) ranged 0.028-0.182 µg.kg-1 and capability of detection limit (CCß) ranged 0.032-0.233 µg.kg-1.

Evaluation of ELISA kits for the screening of four nitrofuran metabolites in aquaculture products according to the European guideline for the validation of screening methods.

The administration of nitrofurans to livestock to treat or prevent animal diseases has been banned in the EU for the production of food of animal origin. The corresponding marker residues are tissue-related metabolites AMOZ, AHD, SEM, and AOZ. The MRPL (minimum required performance limit)/RPA (Reference point for action) was set at 1 µg kg-1 in the EU. Thus, all the laboratories involved in the control of nitrofuran metabolites must detect at least at this analytical limit of performance. The objectives of the work reported here were to evaluate the performance of ELISA kits from two different manufacturers (R-Biopharm, Germany; Europroxima, the Netherlands) for the individual screening of the four nitrofuran metabolites (AOZ, AMOZ; AHD; and SEM) in aquaculture products (fish, shrimps), and then to validate the kits according to the European Decision EC/2002/657 and to the European guideline for the validation of screening methods. The false positive rates were below 9 % for the kits from both manufacturers. The detection capabilities CCß determined were all below the current RPA (1 µg/kg). However, regarding the updated RPA at 0.5 µg/kg that shall apply in 2022, the AMOZ and SEM kits from R-Biopharm and the SEM kit from Europroxima will not be able to reach it.

Tissue distribution, metabolism, and elimination of Victoria Pure Blue BO in rainbow trout: Main metabolite as an appropriate residue marker.

Victoria Pure Blue BO is a dye that bears some therapeutic activity and that can be retrieved in effluent or may be used in aquaculture as a prohibited drug. In this study, the metabolism and tissue distribution during uptake and depuration of VPBO were investigated in order to propose a residue marker of illegal treatment in fish. The dye was administered to rainbow trout (oncorhynchus mykiss) for one day by water bath at a dose of 0.1 mg.L-1. The concentrations of VPBO in all tissues increased rapidly during the treatment period, reaching a Cmax of 567 ± 301 µg.L-1 in plasma and 1846 µg kg-1 ±517 for liver after 2 h. After placing the rainbow trout in a clean water bath for a 64 day-period of depuration, the concentrations in the tissues and plasma decreased to reach comparable levels for muscle and for skin after 33 days. The concentrations measured were still above the LOQ at 2.26 ± 0.48 µg kg-1 for muscle and 2.85 ± 1.99 µg kg-1 for skin at the end of the depuration period. The results indicated the existence of 14 phase I metabolites and one glucuronide conjugated metabolite. Non-compartmental analysis was applied to assess the pharmacokinetic parameters. The half-life in edible muscle of the main metabolite detected, deethyl-leuco-VPBO, was found to be 22.5 days compared to a half-life of 19.7 days for the parent VPBO. This study provides new information to predict a VPBO drug treatment of aquacultured species via a proposed new residue marker.

Evaluation of three ELISA kits for the screening of colistin residue in porcine and poultry muscle according to the European guideline for the validation of screening methods.

Colistin is a polypeptide antibiotic mainly used in porcine and poultry to treat gastrointestinal infections. It has been included by the World Health Organisation (WHO) in the list of critically important human antibiotics of high priority for antimicrobial resistance since 2017. Therefore, it is necessary to develop specific and sensitive screening methods for this molecule. Screening for colistin with immunoassays is an interesting alternative to LC-MS/MS screening methods. The performance of three commercially available ELISA kits was evaluated in poultry and porcine muscles for the detection of colistin in regards to its European maximum residue limit (MRL) (150 µg/kg). The applicability of the three ELISA kits to the detection of colistin at or below the MRL in porcine and poultry muscles was demonstrated. The detection capabilities (CCß) of two kits were or lower than or equal to the MRL (150 µg/kg). The lowest detection capability (30 µg/kg) was achieved with the third ELISA kit. The specificity of the three kits was very satisfactory (false positive rates 0%). The three kits are very specific for the detection of colistin (colistin A and B) and polymyxin B.

Control of Antimicrobials in Feed Using Liquid Chromatography-Tandem Mass Spectrometry: Assessment of Cross-Contamination Rates at the Farm Level.

Antimicrobial cross-contamination of animal feed may occur during feed manufacturing, because shared production lines can be used for the production of medicated and nonmedicated feeds, and also during feed transport, storage at the farm level, and usage. This is a major issue in the current context in which antimicrobial usage must be controlled to maintain their effectiveness. The purpose of this study was to assess the antimicrobial cross-contamination rate of feed at the farm level. Here, we optimized a liquid chromatography-tandem mass spectrometry method for the determination of 11 antimicrobials in feed for pigs, poultry, and rabbits, which were strategically chosen. The method was validated according to European regulations in terms of mass spectrometry identification criteria and quantification criteria (linearity, trueness, precision, limit of quantification, and limit of decision). The results were in compliance with these regulations except for doxycycline, which may be quantified with higher uncertainty. This method was applied to the analysis of 192 nonmedicated pig, poultry, and rabbit feed samples that were collected directly from farms to assess antimicrobials animal exposure. Cross-contamination rates were relatively high with 44% of the samples being contaminated at a concentration above the quantification limit of 0.125 mg/kg and 15% of the samples being contaminated above 1 mg/kg. This result suggests that the current regulations and feed processing recommendations need to be improved, taking into account the risks arising from these contaminations.

In vitro investigations of the metabolism of Victoria pure blue BO dye to identify main metabolites for food control in fish.

Although banned, dyes, such as Victoria pure blue BO (VPBO), are illicitly used in aquaculture to treat or prevent infections due to their therapeutic activities. The present study examined the formation of phase I and phase II metabolites derived from VPBO using trout liver microsomes and S9 proteins. The well-known malachite green (MG) dye was also studied as a positive control and to compare its metabolism with that of VPBO. First, we optimised the incubation conditions for the detection of VPBO and MG metabolites by studying the formation of cytochrome P450 (CYP) substrates. Using the determined conditions (2 h at 20 °C), we incubated VPBO with trout microsomal and S9 fractions induced with ß-naphtoflavone, and analysed the supernatant in a LC-LTQ-Orbitrap-HRMS system. The in vitro assays led to the detection of 16 VPBO metabolites from Phase I reactions, arising in particular from reactions with CYP1A. No metabolites were detected from Phase II reactions. The main metabolite detected, deethyl-VPBO, was CID-fragmented to determine its chemical structure, and thus recommend a potential biomarker for the control of VPBO in farmed fish foodstuffs.

Development of a multi-class method to determine nitroimidazoles, nitrofurans, pharmacologically active dyes and chloramphenicol in aquaculture products by liquid chromatography-tandem mass spectrometry.

LC-MS/MS method was developed for the efficient identification and quantification of 21 banned substances including various nitroimidazoles, nitrofurans, pharmacologically-active dyes and chloramphenicol, respectively in aquaculture products. The sample preparation was started by acid-treatment with 2-nitrobenzaldehyde (NBA) to liberate matrix-bound residues of nitrofurans. A modified QuEChERS method was optimized for the extraction and clean-up of the target analytes. The metabolites of the four conventional nitrofurans (nitrofurantoin, furazolidone, nitrofurazone and furaltadone) and of three other nitrofurans (nifursol, nifuroxazide, and nitrovin), and an underivatizable nitrofuran (nifurpirinol) were simultaneously detected. Furthermore, 21 banned substances were quantified by LC-MS/MS with ESI using one single injection. To evaluate and validate the performance of the method, the criteria of the Decision (EC) no 2002/657 were applied. Decision limit (CCα) of target analytes ranged 0.067-1.655 µg/kg in aquaculture products. The recovery ranged 77.2%-125.6%, and the relative standard deviations of inter-day analyses (RSD) were less than 25%.

A non-targeted LC-HRMS approach for detecting exposure to illegal veterinary treatments: The case of cephalosporins in commercial laying Hens.

Cephalosporins are of particular importance in human medicine and should be reserved for second-line curative treatment in the veterinary field to avoid any emerging antimicrobial resistance. Due to misuse of ceftiofur in the poultry sector in France, it is now recommended to completely stop using cephalosporins in this sector. Methods currently used for the control of veterinary practices are mostly based on liquid chromatography coupled to mass spectrometry in a targeted mode, including parent compounds and any major metabolites. The aim of the present study was to evaluate the relevance of untargeted metabolomic approaches to highlight a possible exposure of laying hens to cephalosporins using a predictive model including selected treatment biomarkers. An experimentation carried out on living animals involved the administration of cefquinome and ceftiofur. Three biological matrices-droppings, eggs and liver-were investigated. Metabolites were extracted and analysed by liquid chromatography coupled to high resolution mass spectrometry in a full scan mode. Metabolites impacted by the treatment were selected by using univariate and multivariate statistical analyses. Predictive models built from the potential biomarkers selected in the "droppings" matrix were validated and able to classify "treated" and "control" hens. PLS-DA and logistic regression models were compared and both models gave satisfactory results in terms of prediction. Results were of less interest for other matrices in which only biomarkers of exposure to cefquinome were detected.

Multiplex immunoassay based on biochip technology for the screening of antibiotic residues in milk: validation according to the European guideline.

The Infiniplex for milk® (IPM) kit is a quick method for the simultaneous and qualitative detection of more than 100 molecules including antibiotic residues, mycotoxins, anti-inflammatories and antiparasitic drugs into a single test that does not require milk treatment. The IPM® kit was validated according to the European decision EC/2002/657 and according to the European guideline for the validation of screening methods (2010). Our validation was focused only on antibiotic residues. The washing step was identified as the most critical step of the assay. Insufficient washes could cause a significant background noise that prevents imaging. Positive controls have to be freshly prepared each day (insufficient stability). The method was specific with a low false-positive rate of 1.7% on 5 discrete test regions (DTR) ((beta-lactams, lincomycin, virginiamycin, quinolones and sulphonamides)) and a false-positive rate of 0% on the 26 other DTR. During our validation, the 42 determined detection capabilities CCß for 12 antibiotic families (aminoglycosides, cephalosporins, lincosamides, macrolides, miscellaneous antibiotics, penicillins, phenolated polymixins, polypeptide antibiotics, quinolones, sulphonamides, tetracyclines) were at between once and twice the decision levels stated by the manufacturer. Forty CCß determined were lower than the respective regulatory limits (i.e. MRL, RC, MRPL) in milk, except for tilmicosin (1.5 times the MRL) and neospiramycin (>1.25 times the MRL). The estimated CCß of thiamphenicol, cloxacillin, danofloxacin, sulphathiazol, ceftiofur and sulphamonomethoxine were lower than or at the MRL. However, it was difficult to approach an accurate CCß with only qualitative results. It is impossible to know whether or not we were close to the cut-off value. The software could be improved by differentiating between low-positive and high-positive results. The results of our participation in three qualitative proficiency tests in 2016 and 2017 for the detection of quinolones, tetracyclines and sulphonamides in cows' milk were very satisfactory.

Impact of ion-pairs for the determination of multiclass antimicrobials residues in honey by LC-MS/MS.

This study examines the behaviour of volatile perfluorinated carboxylic acids used as ion-pairing reagents for the separation of multiclass antimicrobial residues in honey (tetracyclines, sulphonamides, macrolides, and aminoglycosides). Heptafluorobutanoic acid and pentafluoropentanoic acid were compared as mobile phase additives at concentrations of 0-20 mmol.L-1 using reversed-phase chromatography. Our objective was to find optimal conditions that enable the separation of antimicrobials with a minimum loss in sensitivity and efficiency. The influence of ion pairing on chromatographic performance was examined. Results showed that heptafluorobutanoic acid was able to separate aminoglycosides faster and more efficiently than pentafluoropentanoic acid, but at low concentrations of heptafluorobutanoic acid between 2.5 and 5 mM, tetracyclines showed poor chromatographic efficiency. A concentration of 10 mM was found to be optimal for all studied veterinary drugs. The method was then applied to determine multiclass antimicrobial residues in honey. Based on validation data, the mean recoveries of analytes ranged between 93% and 104%, and the intermediate precisions were <21%. The decision limit (CCα) and detection capability (CCß) were in the ranges of 5-25 and 7-33 µg kg-1, respectively. The method was found to be suitable for use in analysis of veterinary drugs with a broader group of compounds found in food-producing animals.

Strategies for the screening of antibiotic residues in eggs: comparison of the validation of the classical microbiological method with an immunobiosensor method.

Efficient screening methods are needed to control antibiotic residues in eggs. A microbiological kit (Explorer® 2.0 test (Zeu Inmunotech, Spain)) and an immunobiosensor kit (Microarray II (AM® II) on Evidence Investigator™ system (Randox, UK)) have been evaluated and validated for screening of antibiotic residues in eggs, according to the European decision EC/2002/657 and to the European guideline for the validation of screening methods. The e-reader™ system, a new automatic incubator/reading system, was coupled to the Explorer 2.0 test. The AM II kit can detect residues of six different families of antibiotics in different matrices including eggs. For both tests, a different liquid/liquid extraction of eggs had to be developed. Specificities of the Explorer 2.0 and AM II kit were equal to 8% and 0% respectively. The detection capabilities were determined for 19 antibiotics, with representatives from different families, for Explorer 2.0 and 12 antibiotics for the AM II kit. For the nine antibiotics having a maximum residue limit (MRL) in eggs, the detection capabilities CCß of Explorer 2.0 were below the MRL for four antibiotics, equal to the MRL for two antibiotics and between 1 and 1.5 MRLs for the three remaining antibiotics (tetracyclines). For the antibiotics from other families, the detection capabilities were low for beta-lactams and sulfonamides and satisfactory for dihydrostreptomycin (DHS) and fluoroquinolones, which are usually difficult to detect with microbiological tests. The CCß values of the AM II kit were much lower than the respective MRLs for three detected antibiotics (tetracycline, oxytetracycline, tylosin). Concerning the nine other antibiotics, the detection capabilities determined were low. The highest CCß was obtained for streptomycin (100 µg kg-1).

Design for the transfer of a validated liquid chromatography/tandem mass spectrometry analytical method for the determination of antimicrobial residues in honey from low-resolution to high-resolution mass spectrometry.

RATIONALE: This paper investigates the validity of the transfer of a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the determination of veterinary medicinal products in honey and compares it with an LC/linear ion trap/Orbitrap mass spectrometry method. A descriptive statistical approach was used in order to assess whether such a transfer would succeed or fail. This approach is based on the simultaneous evaluation of the trueness and of the intermediate precision for each compound at a 95% interval of confidence of both analytical techniques. METHODS: Two grams of honey were placed in a centrifuge tube and diluted with 2.5 mL of ultra-pure water and 2.5 mL of acidified methanol with hydrochloric acid at 2 mol.mL-1 . The extract was purified with 50 mg of primary secondary amine and then analyzed using LC/MS/MS in multiple reaction monitoring (MRM) mode and LC/orbitrap high-resolution mass spectrometry in full scan mode. Both analytical techniques were compared by using the descriptive statistical approach for the determination of antimicrobial residues in honey. RESULTS: The transfer of the method showed that the Orbitrap system provides the same accurate analytical results compared with the LC/MS/MS method except for 4-epitetracycline, anhydroerythromycin A, erythromycin A enol ether, and dihydrostreptomycin. Furthermore, the LC/LTQ-Orbitrap system is capable of successfully competing with the LC/MS/MS method by additional provision of high mass resolution and mass accuracy even though it shows less sensitivity compared with the LC/MS/MS instrument. CCα levels for most analytes were 1.3 times higher by LC/MS/MS than those observed by LC/LTQ-Orbitrap. The method was assessed in terms of relative bias through analysis of a reference material provided by FAPAS (Food Analysis Performance and Assessment Scheme) and also through the control of several contaminated honey samples from local Lebanese markets. Satisfactory relative bias was below 22% except for tetracycline found in one sample that showed a higher bias at 29%. CONCLUSIONS: The LC/LTQ-Orbitrap method offers adequate performance in comparison with previously validated method on a LC/MS/MS system resulting in acceptance of the transfer of the method from LC/MS/MS to LC/LTQ-Orbitrap. Copyright © 2017 John Wiley & Sons, Ltd.

Micropollutants and chemical residues in organic and conventional meat.

The chemical contamination levels of both conventional and organic meats were assessed. The objective was to provide occurrence data in a context of chronic exposure. Environmental contaminants (17 polychlorinated dibenzodioxins/dibenzofurans, 18 polychlorinated biphenyls (PCBs), 3 hexabromocyclododecane (HBCD) isomers, 6 mycotoxins, 6 inorganic compounds) together with chemical residues arising from production inputs (75 antimicrobials, 10 coccidiostats and 121 pesticides) have been selected as relevant compounds. A dedicated sampling strategy, representative of the French production allowed quantification of a large sample set (n=266) including both conventional (n=139) and organic (n=127) raw meat from three animal species (bovine, porcine, poultry). While contamination levels below regulatory limits were measured in all the samples, significant differences were observed between both species and types of farming. Several environmental contaminants (Dioxins, PCBs, HBCD, Zn, Cu, Cd, Pb, As) were measured at significantly higher levels in organic samples.

Solid-phase microextraction set-up for the analysis of liver volatolome to detect livestock exposure to micropollutants.

Starting from a critical analysis of a first "proof of concept" study on the utility of the liver volatolome for detecting livestock exposure to environmental micropollutants (Berge et al., 2011), the primary aim of this paper is to improve extraction conditions so as to obtain more representative extracts by using an extraction temperature closer to livestock physiological conditions while minimizing analytical variability and maximizing Volatile Organic Compound (VOC) abundancies. Levers related to extraction conditions and sample preparation were assessed in the light of both abundance and coefficient of variation of 22 candidate VOC markers identified in earlier volatolomic studies. Starting with a CAR/PDMS fiber and a 30min extraction, the reduction of SPME temperature to 40°C resulted in a significant decrease in the area of 14 candidate VOC markers (p<0.05), mainly carbonyls and alcohols but also a reduction in the coefficient of variation for 17 of them. In order to restore VOC abundances and to minimize variability, two approaches dealing with sample preparation were investigated. By increasing sample defrosting time at 4°C from 0 to 24h yielded higher abundances and lower variabilities for 15 and 13 compounds, respectively. Lastly, by using additives favouring the release of VOCs (1.2g of NaCl) the sensitivity of the analysis was improved with a significant increase in VOC abundances of more than 50% for 13 out of the 22 candidate markers. The modified SPME parameters significantly enhanced the abundances while decreasing the analytical variability for most candidate VOC markers. The second step was to validate the ability of the revised SPME protocol to discriminate intentionally contaminated broiler chickens from controls, under case/control animal testing conditions. After verification of the contamination levels of the animals by national reference laboratories, data analysis by a multivariate chemometric method (Common Components and Specific Weights Analysis - ComDim) showed that the liver volatolome could reveal dietary exposure of broilers to a group of environmental pollutants (PCBs), a veterinary treatment (monensin), and a pesticide (deltamethrin), thus confirming the usefulness of this analytical set-up.