Determination of organic fluorinated compounds content in complex samples through combustion ion chromatography methods: a way to define a "Total Per- and Polyfluoroalkyl Substances (PFAS)" parameter?

Emerging contaminants are a growing concern for scientists and public authorities. The group of per-polyfluoroalkyl substances (PFAS), known as 'forever chemicals', in complex environmental liquid and solid matrices was analysed in this study. The development of global analytical methods based on combustion ion chromatography (CIC) is expected to provide accurate picture of the overall PFAS contamination level via the determination of extractable organic fluorine (EOF) and adsorbable organic fluorine (AOF). The obtained results may be put into perspective with other methods such as targeted analyses (LC-MS/MS). The impact of pH, the presence of dissolved organic carbon and suspended particles on AOF measurements were explored. The effectiveness of the washing step to remove adsorbed inorganic fluorine (IF) has been proven for samples containing up to 8 mgF.L-1. CIC-based methods showed good repeatability and reproducibility for the complex matrices studied. Environmental applications of these methods have been tested. AOF and EOF analyses could explain between 1 % and 23 % and 0.1 % to 2 % of total organic fluorine (TOF), respectively. The sum of PFAS compounds expressed as fluorine could explain from 0.2 % to 11 % and from 0.003 % to 5 % for AOF and EOF, respectively. These results also suggest that some fluorinated compounds are not adsorbed or extractable and/or lost by volatilisation during the application of AOF and EOF analytical procedure. These findings highlight that AOF and EOF are not entirely efficient as proxy to assess "total PFAS" for assessing environmental contamination by PFAS. However, these methods could still be applied to gain a better understanding of the sources and fate of PFAS in the environment.

Low concentrations of ethylene bisdithiocarbamate pesticides maneb and mancozeb impair manganese and zinc homeostasis to induce oxidative stress and caspase-dependent apoptosis in human hepatocytes.

The worldwide and intensive use of phytosanitary compounds results in environmental and food contamination by chemical residues. Human exposure to multiple pesticide residues is a major health issue. Considering that the liver is not only the main organ for metabolizing pesticides but also a major target of toxicities induced by xenobiotics, we studied the effects of a mixture of 7 pesticides (chlorpyrifos-ethyl, dimethoate, diazinon, iprodione, imazalil, maneb, mancozeb) often detected in food samples. Effects of the mixture was investigated using metabolically competent HepaRG cells and human hepatocytes in primary culture. We report the strong cytotoxicity of the pesticide mixture towards hepatocytes-like HepaRG cells and human hepatocytes upon acute and chronic exposures at low concentrations extrapolated from the Acceptable Daily Intake (ADI) of each compound. Unexpectedly, we demonstrated that the manganese (Mn)-containing dithiocarbamates (DTCs) maneb and mancozeb were solely responsible for the cytotoxicity induced by the mixture. The mechanism of cell death involved the induction of oxidative stress, which led to cell death by intrinsic apoptosis involving caspases 3 and 9. Importantly, this cytotoxic effect was found only in cells metabolizing these pesticides. Herein, we unveil a novel mechanism of toxicity of the Mn-containing DTCs maneb and mancozeb through their metabolization in hepatocytes generating the main metabolite ethylene thiourea (ETU) and the release of Mn leading to intracellular Mn overload and depletion in zinc (Zn). Alteration of the Mn and Zn homeostasis provokes the oxidative stress and the induction of apoptosis, which can be prevented by Zn supplementation. Our data demonstrate the hepatotoxicity of Mn-containing fungicides at very low doses and unveil their adverse effect in disrupting Mn and Zn homeostasis and triggering oxidative stress in human hepatocytes.

Microbial Transformation of Chlordecone and Two Transformation Products Formed During in situ Chemical Reduction.

Chlordecone (CLD) is a very persistent synthetic organochlorine pesticide found in the French West Indies. Recently published work has demonstrated the potential of zero-valent iron to dechlorinate CLD by in situ chemical reduction (ISCR) in soils under water-saturated conditions, forming mono- to penta-dechlorinated CLD transformation products. These transformation products are more mobile than CLD and less toxic; however, nothing is known about their further degradation, although increasing evidence of CLD biodegradation by bacteria is being found. The present study began with the enrichment from wastewater sludge of a CLD-transforming community which was then inoculated into fresh media in the presence of either CLD or two of the main ISCR transformation products, 10-monohydroCLD (-1Cl-CLD) and tri-hydroCLD (-3Cl-CLD). Carried out in triplicate batches and incubated at 38°C under anoxic conditions and in the dark, the cultures were sampled regularly during 3 months and analyzed for CLD, -1Cl-CLD, -3Cl-CLD, and possible transformation products by gas chromatography coupled to mass spectrometry. All batches showed a decrease in the amended substrates (CLD or hydroCLD). CLD degradation occurred with concomitant formation of a nine-carbon compound (pentachloroindene) and two sulfur-containing transformation products (chlordecthiol, CLD-SH; methyl chlordecsulfide, CLD-SCH3), demonstrating competing transformation pathways. In contrast, -1Cl-CLD and -3Cl-CLD only underwent a sequential reductive sulfidation/S-methylation process resulting in -1Cl-CLD-SH and -1Cl-CLD-SCH3 on the one hand, and -3Cl-CLD-SH, -3Cl-CLD-SCH3 on the other hand. Some sulfur-containing transformation products have been reported previously with single bacterial strains, but never in the presence of a complex microbial community. At the end of the experiment, bacterial and archaeal populations were investigated by 16S rRNA gene amplicon sequencing. The observed diversity was mostly similar in the CLD and -1Cl-CLD conditions to the inoculum with a dominant archaea genus, Methanobacterium, and four OTU affiliated to bacteria, identified at the family (Spirochaetaceae) or genus level (Desulfovibrio, Aminobacterium, and Soehngenia). On the other hand, in the -3Cl-CLD condition, although the same OTU were found, Clostridium sensu stricto 7, Candidatus Cloacimonas, and Proteiniphilum were also present at > 2% sequences. Presence of methanogens and sulfate-reducing bacteria could contribute to sulfidation and S-methylation biotransformations. Overall, these results contribute to increasing our knowledge on the biodegradability of CLD and its transformation products, helping to progress toward effective remediation solutions.

Removal efficiency of emerging micropollutants in biofilter wastewater treatment plants in tropical areas.

We studied the removal of 61 emerging micropollutants, including illicit drugs, in a biofilter wastewater treatment plant located in the French Indies (Martinique). Raw wastewater concentrations were the highest for paracetamol followed by caffeine, naproxen, ibuprofen, its metabolite 2-hydroxyibuprofen, atenolol, ketoprofen, furosemide, methylparaben, cocaine, benzoylecgonine, and 11-nor-delta-9-carboxytetrahydrocannabinol (THC-COOH). The calculated removals were better than those reported in the literature, while the cumulative removal efficacy (i.e., removal of the total mass load) was estimated to be 92 ± 4%. However, this good performance may be partly explained by the removal of paracetamol (also named acetaminophen) and caffeine, which represented 86.4% of the total mass load. Our results point to the adsorption of some molecules on sludge, thus raising the question about local soil pollution from sludge spreading.

Laboratory study on the mobility of chlordecone and seven of its transformation products formed by chemical reduction in nitisol lysimeters of a banana plantation in Martinique (French Caribbean).

The contamination by chlordecone (CLD) of soils and water in the French Caribbean (FC) has major environmental and human health impacts. In Situ Chemical Reduction (ISCR) is a promising method to degrade CLD but it generates transformation products (TPs). Here, the fate and transport of CLD and its TPs have been studied using three lysimeters, 65-70 cm-long and 20 cm in diameter, collected from a CLD contaminated nitisol in the FC. A simulated ISCR remediation process (Sim-ISCR) was applied to the top 15 cm layer. An equivalent of 9.8 years of effective rainfall was simulated during the 451 days of the experiments. CLD and seven TPs were analyzed in soils, soil pore waters and outflow waters of the lysimeters before and after the Sim-ISCR. CLD concentration in the soil pore waters increases with depth. In the Sim-ISCR treated layer, the CLD contamination was lowered by 41 to 47% in the soil and by 48 to 73% in the soil pore water. In the lysimeters outflow, however, the CLD concentration was lowered by only 13 to 25%, the flux of CLD from the untreated 50-55 cm of the profile concealing much of the beneficial impact of treating the top 15 cm. Remediating by ISCR the topsoil only will therefore not be sufficient for preventing further CLD contamination of the underlying groundwater. Sim-ISCR generated 5-hydroCLD in soils and waters and, to a much lesser extent, a trihydroCLD, a tetrahydroCLD, a pentahydroCLD and a heptahydroCLD. 5-hydroCLD is more mobile than CLD, but it still interacts strongly with the soil. The 5-hydroCLD values measured in the outflow were up to a factor of 4.4 lower than in the treated soil pore waters, indicating some natural attenuation.

Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France).

Soils, surface and groundwater in Martinique (French West Indies) are contaminated by chlordecone (CLD), a highly persistent organochlorine pesticide. In Situ Chemical Reduction (ISCR) using zero valent iron has been tested as a remediation technique to lower CLD levels in soils but it produces derivatives whose fate in environment may differ from the parent molecule. Here, the transfer of CLD and two of its main derivatives resulting from ISCR, CLD5aH and a CLD-3Cl, have been investigated in untreated and treated nitisol from a banana plantation using column experiments (20 cm long and 2.5 cm in diameter) under saturated conditions. The circulation of CaCl2 10-2M solution, simulating the ionic strength of soil water, in untreated nitisol results in CLD concentrations in solutions that remain for decades above the threshold limit for drinking water. ISCR treatment lowers the CLD concentration by ~50% in soil and by a factor 3 in waters but they remain above the threshold values. CLD derivatives, CLD5aH and a CLD-3Cl and, to a lesser extent, a CLD-2Cl and a CLD-5Cl, are found in waters after treatment. Dechlorination increases the mobility of the derivatives with respect to the parent molecule, which is likely to induce their transfer to deeper soil layers than those treated by ISCR: CLD-3Cl is more mobile than CLD5aH which is more mobile than CLD. When the water is in contact with the contaminated soil, a period of fast desorption kinetic of CLD and its derivatives, followed by a period of slow kinetics are found. This attests the high risk for water contamination and the potential influence of rainfall events on the concentrations likely to be encountered in soil waters or in waters accumulated on the soil surface.

Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordecone-contaminated nitisol at plot scale in a French Caribbean banana plantation.

The In Situ Chemical Reduction (ISCR) process was tested in a nitisol in a French Caribbean banana plantation using five different soil amendments. The addition of 2.8% or 4.0% of Zero Valent Iron (ZVI; dw/dw, 2 different trial plots) in the 0-40-cm soil layer lowered the initial chlordecone (CLD) concentration by up to 74% or 69% in 37 days or 94 days, with 75% of the decrease achieved after only 21 or 24 days of treatment depending on the trial plot. The addition of commercially available Daramend® was also tested by applying the 6% dose (dw/dw) recommended by the manufacturer and using either the regular alfalfa-based product or a bagasse-based product specifically formulated for the study. Both significantly lowered CLD concentrations, but to a lesser extent than with the ZVI-only amendment. A bagasse-ZVI mixture prepared on site produced results slightly better than the two Daramend®. The percentage decreases in CLD concentrations were correlated with the negative redox potentials achieved. In all the trial plots, dechlorinated transformation products appeared in the soil and soil water as the CLD concentrations decreased, with H atoms replacing up to 4 and 7 of the 10 Cl atoms, respectively. None of these degradation products appeared to accumulate in the soil or soil water during the treatment. Instead, the reverse occurred, with an overall downward trend in their concentrations over time. The effects of ISCR treatment on agronomic and human health-related parameters were measured in three different crops. The radishes produced with some treatments were visually of lower quality or smaller in size than those grown in the control plots. Lower yields were observed for the cucumbers and sweet potatoes grown after applying the bagasse-based amendments. Mortality among cucumber seedlings was observed after treatment with ZVI only. Simple operational solutions should suffice to remedy these negative agronomic effects. As regards human health-related effects, the CLD concentrations in radishes grown with three of the amendments were significantly lower than in the two control plots and well below the maximum residue level (MRL), which was substantially exceeded in the radishes grown on untreated soil. For cucumbers, the treatments with regular Daramend® and with a local bagasse-ZVI mixture produced fruits with CLD below the MRL and also below the concentrations in one of the two control plots. As for the sweet potatoes, adding a bagasse-ZVI mixture had a significant positive effect by decreasing contamination below the levels in the two control plots and below the MRL.

In vivo comparison of the proangiogenic properties of chlordecone and three of its dechlorinated derivatives formed by in situ chemical reduction.

In situ chemical reduction (ISCR) has been identified as a possible way for the remediation of soils contaminated by chlordecone (CLD). Evidences provided by the literature indicate an association between the development of prostate cancer and CLD exposure (Multigner et al. 2010). In a previous in vitro study, we demonstrated that the two main dechlorinated CLD derivatives formed by ISCR, CLD-1Cl, and CLD-3Cl have lower cytotoxicity and proangiogenic properties than CLD itself (Legeay et al. 2017). By contrast, nothing is known on the in vivo proangiogenic effect of these dechlorinated derivatives. Based on in vitro data, the aims of this study were therefore to evaluate the in vivo influence of CLD and three of its dechlorinated metabolites in the control of neovascularization in a mice model of prostate cancer. The proangiogenic effect of CLD and three of its dechlorinated derivatives, CLD-1Cl, CLD-3Cl, and CLD-4Cl, was evaluated on a murine model of human prostate tumor (PC-3) treated, at two exposure levels: 33 µg/kg and 1.7 µg/kg respectively reflecting acute and chronic toxic exposure in human. The results of serum measurements show that, for the same ingested dose, the three metabolite concentrations were significantly lower than that of CLD. Dechlorination of CLD lead therefore to molecules that are biologically absorbed or metabolized, or both, faster than the parent molecule. Prostate tumor growth was lower in the groups treated by the three metabolites compared to the one treated by CLD. The vascularization measured on the tumor sections was inversely proportional to the rate of dechlorination, the treatment with CLD-4Cl showing no difference with control animals treated with only the vehicle oil used for all substances tested. We can therefore conclude that the proangiogenic effect of CLD is significantly decreased following the ISCR-resulting dechlorination. Further investigations are needed to elucidate the molecular mechanisms by which dechlorination of CLD reduces proangiogenic effects in prostate tumor.

Chlordecone consumption estimated by sewage epidemiology approach for health policy assessment.

Chlordecone is an organochlorine insecticide that was intensively used in the French West Indies to control the black weevil Cosmopolites sordidus in bananas. Its usage, however, resulted in the widespread pollution of the environment with heavy sanitary and social consequences, leading to population exposure mainly through food. Time-consuming and costly programs have been used to tackle this problem, and this study proposes to use the emerging sewage epidemiology approach to evaluate the current situation and the effect of such programs. The results determine the maximal value of chlordecone consumption, and considering the detection limit of the analytical protocol, the wastewater was found to have undetectable amounts of chlordecone. This value confirms the efficiency of the population protection strategy provided by French sanitary and environmental authorities. It also bolsters the usage of sewage epidemiology in pesticide assessment and relativizes the chlordecone risk compared to other chemicals of lesser concern.

Two dechlorinated chlordecone derivatives formed by in situ chemical reduction are devoid of genotoxicity and mutagenicity and have lower proangiogenic properties compared to the parent compound.

Chlordecone (CLD) is a chlorinated hydrocarbon insecticide, now classified as a persistent organic pollutant. Several studies have previously reported that chronic exposure to CLD leads to hepatotoxicity, neurotoxicity, raises early child development and pregnancy complications, and increases the risk of liver and prostate cancer. In situ chemical reduction (ISCR) has been identified as a possible way for the remediation of soils contaminated by CLD. In the present study, the objectives were (i) to evaluate the genotoxicity and the mutagenicity of two CLD metabolites formed by ISCR, CLD-5a-hydro, or CLD-5-hydro (5a- or 5- according to CAS nomenclature; CLD-1Cl) and tri-hydroCLD (CLD-3Cl), and (ii) to explore the angiogenic properties of these molecules. Mutagenicity and genotoxicity were investigated using the Ames's technique on Salmonella typhimurium and the in vitro micronucleus micromethod with TK6 human lymphoblastoid cells. The proangiogenic properties were evaluated on the in vitro capillary network formation of human primary endothelial cells. Like CLD, the dechlorinated derivatives of CLD studied were devoid of genotoxic and mutagenic activity. In the assay targeting angiogenic properties, significantly lower microvessel lengths formed by endothelial cells were observed for the CLD-3Cl-treated cells compared to the CLD-treated cells for two of the three tested concentrations. These results suggest that dechlorinated CLD derivatives are devoid of mutagenicity and genotoxicity and have lower proangiogenic properties than CLD.

Remediation by chemical reduction in laboratory mesocosms of three chlordecone-contaminated tropical soils.

Chlordecone (CLD), a highly persistent organochlorine pesticide commonly encountered in French West Indies (FWI) agricultural soils, represents a major source of contamination of FWI ecosystems. The potential of chemical reduction for remediation of CLD-contaminated soil has been investigated in laboratory pilot-scale 80 kg mesocosms for andosol, ferralsol, and nitisol from FWI banana plantations. Six cycles consisting of a 3-week reducing phase followed by a 1-week oxidizing phase were applied, with 2 % (dw/dw) Daramend® (organic plant matter fortified with zero valent iron) added at the start of each cycle. Complementary amendments of zero valent iron and zinc (total of 3 % dw/dw) were added at the start of the first three cycles. After the 6-month treatment, the CLD soil concentration was lowered by 74 % in nitisol, 71 % in ferralsol, and 22 % in andosol. Eleven CLD-dechlorinated transformation products, from mono- to penta-dechlorinated, were identified. None of them accumulated over the duration of the experiment. Six of the seven ecotoxicological tests applied showed no difference between the control and treated soils. The treatment applied in this study may offer a means to remediate CLD-contaminated soils, especially nitisol and ferralsol.

Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants.

Chlordecone (CLD) was an organochlorine insecticide whose previous use resulted in an extensive pollution of the environment with severe health effects and social consequences. A closely related compound, 5b-hydrochlordecone (5b-hydroCLD), has been searched for and often detected in environmental matrices from the geographical area where CLD was applied. The current consensus considered that its presence was not the result of a biotic or abiotic dechlorination of CLD in these matrices but rather the consequence of its presence as impurity (synthesis by-product) in the CLD released into the environment. The aim of the present study was to determine if and to what extent degradation of CLD into 5b-hydroCLD occurred in the field. To test this hypothesis, the ratios of 5b-hydroCLD and CLD concentrations in a dataset of 810 soils collected between 2006 and 2012 in Martinique were compared to the ratios measured in 3 samples of the CLD dust commercial formulations applied in the banana fields of French West Indies (FWI) and 1 sample of the technical-grade CLD corresponding to the active ingredient used in such formulations. Soil data were processed with a hierarchical Bayesian model to account for random measurement errors and data censoring. Any pathway of CLD transformation into 5b-hydroCLD occurring over the long term in FWI soils would indeed change the ratio of 5b-hydroCLD/CLD compared to what it was in the initially applied formulations. Results showed a significant increase of the 5b-hydroCLD/CLD ratio in the soils-25 times greater in soil than in commercial formulations-which suggested that natural CLD transformation into 5b-hydroCLD over the long term occurred in these soils. Results from this study may impact future decisions for the remediation of the polluted areas.