Discovery of a series of portimine-A fatty acid esters in mussels.

Vulcanodinium rugosum is a benthic dinoflagellate known for producing pinnatoxins, pteriatoxins, portimines and kabirimine. In this study, we aimed to identify unknown analogs of these emerging toxins in mussels collected in the Ingril lagoon, France. First, untargeted data acquisitions were conducted by means of liquid chromatography coupled to hybrid quadrupole-orbitrap mass spectrometry. Data processing involved a molecular networking approach, and a workflow dedicated to the identification of biotransformed metabolites. Additionally, targeted analyses by liquid chromatography coupled to triple quadrupole mass spectrometry were also implemented to further investigate and confirm the identification of new compounds. For the first time, a series of 13-O-acyl esters of portimine-A (n = 13) were identified, with fatty acid chains ranging between C12:0 and C22:6. The profile was dominated by the palmitic acid conjugation. This discovery was supported by fractionation experiments combined with the implementation of a hydrolysis reaction, providing further evidence of the metabolite identities. Furthermore, several analogs were semi-synthesized, definitively confirming the discovery of these metabolization products. A new analog of pinnatoxin, with a molecular formula of C42H65NO9, was also identified across the year 2018, with the highest concentration observed in August (4.5 µg/kg). The MS/MS data collected for this compound exhibited strong structural similarities with PnTX-A and PnTX-G, likely indicating a substituent C2H5O2 in the side chain at C33. The discovery of these new analogs will contribute to deeper knowledge of the chemodiversity of toxins produced by V. rugosum or resulting from shellfish metabolism, thereby improving our ability to characterize the risks associated with these emerging toxins.

Phosphinoquinoline supported CoII, NiII, and FeII complexes: divergent behaviour upon reduction.

The reduction of [CoLBr2], a CoII complex supported by a diisopropylphosphinoquinoline (L) ligand, induced a ligand coupling giving access to a (PNNP) supported CoII complex which was isolated in 70% yield. This complex was formed using a minimum of 2 equivalents of a reductant (either Mn or KC8). The fate of [CoLBr2] in the presence of 1 equivalent of a reductant was more difficult to study; nevertheless, a CoI complex was characterised in the solid state. In order to determine whether this ligand coupling could occur with other 3d metals, L supported FeII and NiII complexes were synthesised. While no compound could be identified upon reduction of [FeLBr2], both [NiLBr2] and [NiL2Br](Br) led to the reduction at the metal center allowing the isolation of an original Ni0 trimer in a satisfactory yield. This study shows the different behaviours of these 3d metal complexes in the presence of a reductant.

Tandem InCl3-Promoted Hydroperoxide Rearrangements and Nucleophilic Additions: A Straightforward Entry to Benzoxacycles.

The acid-catalyzed rearrangement of organic peroxides is generally associated with C-C-bond cleavages (Hock and Criegee rearrangements), with the concomitant formation of an oxocarbenium intermediate. This article describes the tandem process between a Hock or Criegee oxidative cleavage and a nucleophilic addition onto the oxocarbenium species (in particular a Hosomi-Sakurai-type allylation), under InCl3 catalysis. It was applied to the synthesis of 2-substituted benzoxacycles (chromanes and benzoxepanes), including a synthesis of the 2-(aminomethyl)chromane part of sarizotan, and a total synthesis of erythrococcamide B.

Characterization of photoproducts and global ecotoxicity of chlorphenesin: A preservative used in skin care products.

Photolysis experiments of chlorphenesin, used as a preservative in cosmetic products, were performed in aqueous solution and on a cream used in cosmetics. Three by-products resulting from the direct UV-visible photodegradation of chlorphenesin were characterized by chromatography (gas and liquid) coupled with tandem mass spectrometry (GC-MS/MS and LC-HR MS/MS) and found in both solutions. In vitro tests on Vibrio fischeri bacteria showed that the overall ecotoxicity of chlorphenesin increased with increasing irradiation time in both samples. In silico QSAR (Quantitative Structure Activity Relationship) tests were performed using T.E.S.T. (Toxicity Estimation Software Tool). Among the degradation compounds identified, 4-chlorophenol must contribute to the increased ecotoxicity of the photolyzed solution since the in silico LC50 estimated for all tests performed are always lower than those obtained for chlorphenesin.

Les expériences de photolyse de la chlorphénésine, qui s'utilise comme conservateur dans les produits cosmétiques, ont été réalisées en solution aqueuse et sur une crème utilisée dans les cosmétiques. Trois sous-produits résultant de la photodégradation directe UV-visible de la chlorphénésine ont été caractérisés par chromatographie (gaz et liquide) couplée à une spectrométrie de masse en tandem (GC-MS/MS et LC-HR MS/MS) et trouvés dans les deux solutions. Les tests in vitro sur les bactéries Vibrio fischeri ont montré que l'écotoxicité globale de la chlorphénésine augmentait avec l'augmentation du temps d'irradiation dans les deux échantillons. Des tests QSAR (Quantitative Structure Activity Relationship, relation d'activité de structure quantitative) in silico ont été réalisés à l'aide du logiciel T.E.S.T. (Toxicity Estimation Software Tool, outil logiciel d'estimation de la toxicité). Parmi les composés de dégradation identifiés, le 4-chlorophénol doit contribuer à l'augmentation de l'écotoxicité de la solution photolysée puisque la LC50 in silico estimée pour l'ensemble des essais réalisés est toujours inférieure à celle obtenue pour la chlorphénésine.

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.

Reaction of Phosphines with 1-Azido-(2-halogenomethyl)benzene Giving Aminophosphonium-Substituted Indazoles.

The reaction between a 1-azido-(2-halogenomethyl)benzene and a phosphine gives different products depending on the nature of the halogen, the phosphine itself, and the solvent employed. While PPh3 (2 equiv) reacts with the chloro reagent in toluene to give the expected iminophosphorane-phosphonium adduct, trialkylphosphines (PCy3 and PEt3) surprisingly furnish an aminophosphonium substituted by a zwitterionic indazole. The bicyclic product can also form from PPh3 using the bromo reagent in acetonitrile. A mechanism is proposed for this cyclization based on DFT calculations.

FeIII and FeII Phosphasalen Complexes: Synthesis, Characterization, and Catalytic Application for 2-Naphthol Oxidative Coupling.

We report on the synthesis and characterization of three iron(III) phosphasalen complexes, [FeIII (Psalen)(X)] differing in the nature of the counter-anion/exogenous ligand (X- =Cl- , NO3 - , OTf- ), as well as the neutral iron(II) analogue, [FeII (Psalen)]. Phosphasalen (Psalen) differs from salen by the presence of iminophosphorane (P=N) functions in place of the imines. All the complexes were characterized by single-crystal X-ray diffraction, UV/Vis, EPR, and cyclic voltammetry. The [FeII (Psalen)] complex was shown to remain tetracoordinated even in coordinating solvent but surprisingly exhibits a magnetic moment in line with a FeII high-spin ground state. For the FeIII complexes, the higher lability of triflate anion compared to nitrate was demonstrated. As they exhibit lower reduction potentials compared to their salen analogues, these complexes were tested for the coupling of 2-naphthol using O2 from air as oxidant. In order to shed light on this reaction, the interaction between 2-naphthol and the FeIII (Psalen) complexes was studied by cyclic voltammetry as well as UV/Vis spectroscopy.

Cryo-EM study of an archaeal 30S initiation complex gives insights into evolution of translation initiation.

Archaeal translation initiation occurs within a macromolecular complex containing the small ribosomal subunit (30S) bound to mRNA, initiation factors aIF1, aIF1A and the ternary complex aIF2:GDPNP:Met-tRNAiMet. Here, we determine the cryo-EM structure of a 30S:mRNA:aIF1A:aIF2:GTP:Met-tRNAiMet complex from Pyrococcus abyssi at 3.2 Å resolution. It highlights archaeal features in ribosomal proteins and rRNA modifications. We find an aS21 protein, at the location of eS21 in eukaryotic ribosomes. Moreover, we identify an N-terminal extension of archaeal eL41 contacting the P site. We characterize 34 N4-acetylcytidines distributed throughout 16S rRNA, likely contributing to hyperthermostability. Without aIF1, the 30S head is stabilized and initiator tRNA is tightly bound to the P site. A network of interactions involving tRNA, mRNA, rRNA modified nucleotides and C-terminal tails of uS9, uS13 and uS19 is observed. Universal features and domain-specific idiosyncrasies of translation initiation are discussed in light of ribosomal structures from representatives of each domain of life.

Use of ß3-methionine as an amino acid substrate of Escherichia coli methionyl-tRNA synthetase.

Polypeptides containing ß-amino acids are attractive tools for the design of novel proteins having unique properties of medical or industrial interest. Incorporation of ß-amino acids in vivo requires the development of efficient aminoacyl-tRNA synthetases specific of these non-canonical amino acids. Here, we have performed a detailed structural and biochemical study of the recognition and use of ß3-Met by Escherichia coli methionyl-tRNA synthetase (MetRS). We show that MetRS binds ß3-Met with a 24-fold lower affinity but catalyzes the esterification of the non-canonical amino acid onto tRNA with a rate lowered by three orders of magnitude. Accurate measurements of the catalytic parameters required careful consideration of the presence of contaminating α-Met in ß3-Met commercial samples. The 1.45 Å crystal structure of the MetRS: ß3-Met complex shows that ß3-Met binds the enzyme essentially like α-Met, but the carboxylate moiety is mobile and not adequately positioned to react with ATP for aminoacyl adenylate formation. This study provides structural and biochemical bases for engineering MetRS with improved ß3-Met aminoacylation capabilities.

Ultraviolet-visible phototransformation of dehydroacetic acid - Structural characterization of photoproducts and global ecotoxicity.

RATIONALE: The present work is devoted to the structural elucidation of by-products issued from the direct ultraviolet-visible (UV-vis) irradiation of dehydroacetic acid (DHA) in solution and in cosmetic emulsion. METHODS: Analyses were carried out using gas chromatography coupled with ion trap mass spectrometry and by high-performance liquid chromatography coupled with ultrahigh-resolution Fourier transform ion cyclotron resonance mass spectrometry (LC/UHRMS). The potential toxicities of by-products were estimated by in silico calculations based on a QSAR (Quantitative Structure-Activity Relationship) approach and by in vitro bioassays conducted on Vibrio fischeri bacteria. RESULTS: Three photoproducts were detected by LC/MS while one photoproduct was detected by GC/MS. The first photoproduct (PP1) corresponds to an isomer of DHA while two isomeric compounds correspond to dimeric structures. The oral rat LD50 of PP1 was evaluated to be 4.5 times lower than that of the parent molecule which classes it in the category 'moderately toxic' on the Hodge and Sterne toxicity classification. In vitro assays on Vibrio fischeri bacteria showed that the global ecotoxicity of the DHA solution increases with irradiation time. CONCLUSIONS: With the exception of one photoproduct, the structures proposed for the photoproducts on the basis of mass spectral interpretation have not been reported in previous studies. All photoproducts, with the exception of dimers, were detected after irradiation in the cosmetic emulsion. This result shows that personal care products containing DHA must be protected from direct sunlight to prevent photodegradation.

UV-vis degradation of α-tocopherol in a model system and in a cosmetic emulsion-Structural elucidation of photoproducts and toxicological consequences.

The UV-vis photodegradation of α-tocopherol was investigated in a model system and in a cosmetic emulsion. Both gas chromatography coupled with tandem mass spectrometry (GC-MS/MS) and high performance liquid chromatography coupled with ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry (LC-UHR-MS) were used for photoproducts structural identification. Nine photoproduct families were detected and identified based on their mass spectra and additional experiments with α-tocopherol-d9; phototransformation mechanisms were postulated to rationalize their formation under irradiation. In silico QSAR (Quantitative Structure Activity Relationship) toxicity predictions were conducted with the Toxicity Estimation Software Tool (T.E.S.T.). Low oral rat LD50 values of 466.78mgkg-1 and 467.9mgkg-1 were predicted for some photoproducts, indicating a potential toxicity more than 10 times greater that of α-tocopherol (5742.54mgkg-1). In vitro assays on Vibrio fischeri bacteria showed that the global ecotoxicity of the α-tocopherol solution significantly increases with irradiation time. One identified product should contribute to this ecotoxicity enhancement since in silico estimations for D. magna provide a LC50 value 4 times lower than that of the parent molecule.

Nanostructure Changes upon Polymerization of Aqueous and Organic Phases in Organized Mixtures.

The nanostructure of a microemulsion can be strongly affected by the liquid-to-solid transition during polymerization. Here, we examined the evolution of nanostructures of different ternary mixtures, including two microemulsions and a single lamellar phase that upon polymerization are quantitatively studied by SAXS/WAXS and DSC experiments systematically performed before and after the polymerization of both aqueous and organic phases. Samples are mixtures of the poly(2-acrylamido-2-methylpropanesulfonic acid) network as the aqueous phase and poly(hexyl methacrylate) as the organic phase stabilized by Brij35 surfactant. Upon polymerization, the surfactant is excluded from the water/oil interface and crystallizes, strongly changing the nanostructure of samples where it is the main component. In samples where the aqueous phase is the main component, only a few changes in structure are observed upon polymerization. This study demonstrates quantitatively the possibility to preserve nanostructures during polymerization, thus inducing a templating effect.

Photodegradation of cyprodinil under UV-visible irradiation - chemical and toxicological approaches.

RATIONALE: Cyprodinil is a fungicide active on grapes, strawberries, tomatoes, and many other fruits. Under UV-visible irradiation, it undergoes photodegradation through various processes to form transformation products (TPs) whose structures and potential toxicities are unknown. The structures of the TPs were elucidated by comparing the photodegradation of cyprodinil and cyprodinil-D5 . The potential toxicities of these compounds were compared with that of cyprodinil. METHODS: Aqueous solutions of cyprodinil were irradiated in a reactor equipped with a mercury vapor lamp. Analyses were carried out using high-performance liquid chromatography coupled to a quadrupole time-of-flight (QTOF) mass spectrometer or to a SolarixXR 9.4 Tesla Fourier transform (FT) mass spectrometer. High-resolution mass measurements, MS/MS and isotopic labeling experiments allowed structural elucidation of the cyprodinil TPs. The toxicities were estimated by three tests in silico using the TEST software and in vitro bioassays using Vibrio fischeri bacteria. These bioassays were carried out on irradiated solution for several exposure times and non-irradiated solutions. RESULTS: The structures of 19 photoproducts were characterized by LC/HRMS/MS after 4 h of irradiation of a cyprodinil aqueous solution. The use of cyprodinil-D5 allowed the TPs to be characterized with more confidence. Knowing the structure of the TPs allows the estimation of their potential toxicities by in silico tests. Most of the photoproducts are potentially more toxic than the parent compound, based on the oral rat LD50 values, and most of them might induce more developmental and mutagenic toxicities. In vitro assays on Vibrio fischeri bacteria showed that the global ecotoxicity of the cyprodinil solution significantly increases with irradiation time. CONCLUSIONS: Structures of photoproducts were characterized after irradiation of a cyprodinil aqueous solution combining LC/HRMS, LC/HRMS/MS and the use of a labeled compound. Their formations imply several photodegradation reactions, namely direct bond cleavages, cyclization, isomerization and hydroxylation. Most of the TPs exhibit a toxicity significantly higher than that of the parent molecule. Copyright © 2016 John Wiley & Sons, Ltd.

Characterization of the ultraviolet-visible photoproducts of thiophanate-methyl using high performance liquid chromatography coupled with high resolution tandem mass spectrometry-Detection in grapes and tomatoes.

UV-visible irradiation of thiophanate-methyl (TM) led to the formation of nine photoproducts that were characterized by high performance liquid chromatography coupled with high resolution Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). Although carbendazime has been reported in the literature to be the major metabolite and photoproduct of thiophanate-methyl, it was not detected in this study. However, an isomer of carbendazime referred as PP2, which was unambiguously characterized owing to CID experiments, was found in great abundance. Grape berries and cherry tomatoes treated with aqueous solutions of thiophanate-methyl were submitted to irradiation under laboratory conditions. TM and PP2 were detected in both peel and flesh of berries. The ability of TM and PP2 to pass through the fruit skin has been shown to be highly compound and matrix dependent. In vitro bioassays on Vibrio fischeri bacteria showed that the global ecotoxicity of the TM solution increases significantly with the irradiation time. PP2 should likely contribute to this ecotoxicity enhancement since in silico estimations for Daphnia magna provide a LC50 value seven times lower for PP2 than for the parent molecule.

Structural elucidation of metolachlor photoproducts by liquid chromatography/high-resolution tandem mass spectrometry.

RATIONALE: Metolachlor is one of the most intensively used chloroacetanilide herbicides in agriculture. It has been detected in water; consequently, under UV-visible irradiation, it can be transformed in degradation products (TPs). The structures of TPs were elucidated by liquid chromatography/high-resolution tandem mass spectrometry (LC/HR-MS/MS). The potential toxicities of these TPs were estimated by in silico tests. METHODS: Aqueous solutions of metolachlor were irradiated in a self-made reactor equipped with a mercury vapor lamp. Analyses were carried out using high-performance liquid chromatography coupled to quadrupole time-of-flight (QTOF) mass spectrometer. High-resolution m/z measurements, MS/MS and isotopic labeling experiments allowed structural elucidation of metolachlor TPs. Their toxicities were estimated in silico, using the T.E.S.T. RESULTS: Ten major metolachlor photoproducts were characterized by LC/MS/MS after irradiation of metolachlor in aqueous solution. Elucidation of their chemical structures was identified using high-resolution measurements and MS/MS experiments. They resulted from the combination of dehalogenation, hydroxylation and cyclisation processes. The potential oral rat lethal dose (LD50) was assessed with QSAR tests for metolachlor and each photoproduct. Results indicate that most of the TPs are much more toxic than metolachlor. CONCLUSIONS: UV-vis irradiation of metolachlor in aqueous solution leads to the formation of ten photoproducts. QSAR estimations show that the location of added hydroxyl group(s) is of key relevance as regards to biological activity and that routine water analysis should take into account the TPs are more toxic than the parent molecule.

Isomerization of fenbuconazole under UV-visible irradiation - chemical and toxicological approaches.

RATIONALE: Fenbuconazole is a fungicide commonly used for the protection of vineyards, vegetables and grain crops. Under UV-visible irradiation, it undergoes isomerization through various cyclization processes. Isomeric structures were elucidated by liquid chromatography/high-resolution multiple-stage mass spectrometry (LC/HR-MS(n) ) coupling. The potential toxicities of these isomers were estimated by in silico tests. METHODS: Aqueous solutions of fenbuconazole and grapes treated with this fungicide were irradiated in a self-made reactor equipped with a mercury vapor lamp. Analyses were carried out using high-performance liquid chromatography (HPLC) coupled with Fourier transform ion cyclotron resonance mass spectrometry (FT-ICRMS). High-resolution m/z measurements, multiple-stage mass spectrometry and isotopic labeling experiments allowed structural elucidation of the isomers of fenbuconazole. In silico toxicity estimations were carried out using the T.E.S.T. RESULTS: Seven isomers of fenbuconazole were detected after irradiation of the fungicide in aqueous solution; the major ones were also detected in the flesh of treated grapes irradiated under laboratory conditions. Elucidation of their chemical structures owing to high resolution measurements and multi-stage collision induced dissociation experiments allowed confirmation of photo-transformation pathways mainly dominated by cyclization processes. Photo-induced isomers exhibited higher potential toxicities than fenbuconazole for Daphnia magna and fathead minnow species. CONCLUSIONS: UV-visible irradiation of fenbuconazole in aqueous solution and on grapes leads to the formation of isomers, all of which being potentially much more toxic than the parent fungicide.

Gas-phase lithium cation affinity of glycine.

The gas-phase lithium cation binding thermochemistry of glycine has been determined theoretically by quantum chemical calculations at the G4 level and experimentally by the extended kinetic method using electrospray ionization quadrupole time-of-flight tandem mass spectrometry. The lithium cation affinity of glycine, ∆(Li)H°(298)(GLY), i.e. the∆(Li)H°(298) of the reaction GlyLi(+)→ Gly + Li(+)) given by the G4 method is equal to 241.4 kJ.mol(-1) if only the most stable conformer of glycine is considered or to 242.3 kJ.mol(-1) if the 298K equilibrium mixture of neutral conformers is included in the calculation. The ∆(Li)H°(298)(GLY) deduced from the extended kinetic method is obviously dependent on the choice of the Li(+) affinity scale, thus∆(Li)H°(298)(GLY) is equal to 228.7±0.9(2.0) kJ.mol(- 1) if anchored to the recently re-evaluated lithium cation affinity scale but shifted to 235.4±1.0 kJ.mol(-1) if G4 computed lithium cation affinities of the reference molecules is used. This difference of 6.3 kJ.mol(-1) may originate from a compression of the experimental lithium affinity scale in the high ∆(Li)H°(298) region. The entropy change associated with the reaction GlyLi(+)→Gly + Li(+) reveals a gain of approximately 15 J.mol(-) 1.K(-1) with respect to monodentate Li(+) acceptors. The origin of this excess entropy is attributed to the bidentate interaction between the Li(+) cation and both the carbonyl oxygen and the nitrogen atoms of glycine. The computed G4 Gibbs free energy,∆(Li)G°(298)(GLY) is equal to 205.3 kJ.mol(-1), a similar result, 201.0±3.4 kJ.mol(-1), is obtained from the experiment if the∆(Li)G°(298) of the reference molecules is anchored on the G4 results.

Structural elucidation and estimation of the acute toxicity of the major UV-visible photoproduct of fludioxonil - detection in both skin and flesh samples of grape.

Ultraviolet (UV)-visible irradiation of fludioxonil was investigated with two photoreactors using either a mercury or xenon vapor lamp. In both cases, it led to the formation of only one photoproduct in significant amount: 2-(2,2-difluorobenzo[d][1,3]dioxol-4-yl)-2-(nitrosomethylene)-4-oxobutanenitrile, which has been characterized using Liquid Chromatography - High Resolution - Tandem Mass Spectrometry (LC-HR-MS/MS) coupling. A photolysis pathway has been proposed to rationalize its formation in degassed water. In vitro bioassays on Vibrio fischeri bacteria showed that UV-vis irradiation of an aqueous solution of fludioxonil significantly increases its toxicity. Because no other by-product was detected in significant amount, the photoproduct mentioned above may be considered mainly responsible for this increase in toxicity. Grape berries treated with a 50 ppm aqueous solution of fludioxonil were submitted to UV-visible irradiation under laboratory conditions. The fungicide and photoproduct were detected in both skin and flesh of berries, even after they have been rinsed with water. The ability of the photoproduct to pass through the fruit skin is comparable with that of fludioxonil. These results are of concern for consumers because they mean that water tap rinsing does not lead to efficient removing of both compounds.

6-Iso-chlortetracycline or keto form of chlortetracycline? Need for clarification for relevant monitoring of chlortetracycline residues in food.

Chlortetracycline (CTC) is a broad-spectrum antibiotic used in veterinary medicine for pulmonary or digestive infections and having a regulatory maximum residue limit (MRL) necessitating an official analytical control method. The purpose of this study was to clarify the identification of different forms of CTC observed in standard solution, in spiked muscle samples and in naturally incurred muscle samples of pigs analysed by LC-MS/MS and to demonstrate the in vivo formation of 6-iso-chlortetracycline and 4-epi-6-iso-CTC as a metabolite of CTC and 4-epi-CTC in muscle. The six following forms were identified, all being isobaric with a protonated molecule at m/z 479 (precursor ion): the keto-enol forms of CTC and the keto-enol forms of 4-epi-chlortetracycline (4-epi-CTC), 6-iso-chlortetracycline (6-iso-CTC) and 4-epi-6-iso-chlortetracycline (4-epi-6-iso-CTC). The 6-iso-CTC and 4-epi-6-iso-CTC were observed only in incurred pig samples so were identified for the first time as metabolites of CTC and 4-epi-CTC. Identification of the different forms was obtained by comparing incurred muscle samples with standard solutions and with spiked samples. Then the differences between the features of the chromatograms obtained by LC-TQ-MS and the fragmentation study of the different forms of CTC obtained by LC-Q-TOF-MS helped us to support this identification. The extraction steps and the LC-MS/MS conditions developed to analyse muscle tissue samples are described. This clarification concerning the rigorous identification of chromatographic peaks allowed us to evaluate the relevance of our monitoring method with regard to the regulations in place in the European Union and could be of help to laboratories involved in official control of antibiotic residues in food of animal origin. Additional results are also presented highlighting the transformation of the CTC when prepared in a mixture with other antibiotics.

Ultraviolet-vis degradation of iprodione and estimation of the acute toxicity of its photodegradation products.

The UV-vis photodegradation of iprodione in water was investigated with a high pressure mercury lamp photoreactor. Five photoproducts of iprodione were characterized by LC-HR-MS/MS and isotopic labeling; none of them has been reported in previous studies. Three of them result from the elimination of one or two chlorine atoms followed by hydroxy or hydrogen addition while the two others are cyclic isomers of iprodione. An ICR mass spectrometer was used for by-products identification; concentrations of photoproducts were estimated with a triple quadrupole instrument, using iprodione-D5 as an internal standard. Phototransformation mechanisms were postulated to rationalize photoproducts formation. In silico QSAR toxicity predictions were conducted with the Toxicity Estimation Software Tool (T.E.S.T.) considering oral rat LD50, mutagenicity and developmental toxicity. Low oral rat LD50 values of 350 mg/kg and 759 mg/kg were predicted for cyclic isomers of iprodione, compared to that of the parent molecule (2776 mg/kg). Toxicity estimations exhibited that all the iprodione photoproducts could be mutagenic while the parent compound is not. In vitro assays on Vibrio fischeri were achieved on both irradiated and non-irradiated aqueous solutions of iprodione and on HPLC fractions containing isolated photoproducts. Phenolic photoproducts were shown to be mainly responsible for toxicity enhancement with EC50 values of 0.3 and 0.5 ppm, for the bi- and mono-phenolic compounds issued from chlorine elimination.