Identification by volatolomics of hydrocarbon, oxygenated, sulfur and aromatic markers of livestock exposure to α-hexabromocyclododecane.

Volatile organic compounds (VOC)-based metabolomics, or volatolomics, was investigated for revealing livestock exposure to chemical contamination. Three farm animals, namely laying hens, broilers, and pigs, were experimentally exposed to 5 or 50 ng α-HBCDD g-1 feed. Liver and egg yolk for hens were analysed by headspace-SPME-GC-MS to reveal candidate markers of the livestock exposure to α-HBCDD. For hens, 2-butanol was found as marker in egg. In liver, twelve VOCs were highlighted as markers, with three aromatic VOCs - styrene, o-xylene, α-methylstyrene - highlighted for the two α-HBCDD doses. For broilers, six markers were revealed, with interestingly, styrene and phenol which were also found as markers in hen liver. For pigs, ten markers were revealed and the seven tentatively identified markers were oxygenated and sulfur VOCs. The candidate markers tentatively identified were discussed in light of previous volatolomics data, in particular from a γ-HBCDD exposure of laying hens.

Undernutrition combined with dietary mineral oil hastens depuration of stored dioxin and polychlorinated biphenyls in ewes. 1. Kinetics in blood, adipose tissue and faeces.

Food safety crises involving persistent organic pollutants [POPs, e.g. dioxins, polychlorinated biphenyls (PCBs), organochlorine pesticides] lead to systematic slaughter of livestock to prevent their entry into the food chain. Therefore, there is a need to develop strategies to depurate livestock moderately contaminated with POPs in order to reduce such economic and social damages. This study aimed to test a POPs depuration strategy based on undernutrition (37% of energy requirements) combined with mineral oil (10% in total dry matter intake) in nine non-lactating ewes contaminated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and PCBs 126 and 153. In order to better understand the underlying mechanisms of the depuration process, POPs kinetics and body lipids dynamics were followed concomitantly over 57-day of depuration in POPs storage (adipose tissue, AT), central distribution (blood) and excretion (faeces) compartments. Faecal POPs concentrations in underfed and mineral oil supplemented ewes increased by 2.0 to 2.6-fold, but not proportionally to lipids concentration which increased by 6-fold, compared to the control ewes. Nonetheless, after 57 days of depuration in undernutrition and mineral oil supplementation, AT POPs concentrations were 1.5 to 1.6-fold higher while serum concentrations remained unchanged compared to the control ewes. This was concomitant with a decrease by 2.7-fold of the AT estimated lipids weight along the depuration period. This reduction of the volume of the storage compartment combined with the increase of POPs faecal excretion in underfed and mineral oil supplemented ewes led to a reduction by 1.5-fold of the PCB 126 AT burden, while no changes were observed for TCDD and PCB 153 burdens (vs. no change for PCB 126 and increases for TCDD and PCB 153 AT burdens in control ewes). The original approach of this study combining the fine description at once of POPs kinetic and of body lipids dynamic improved our understanding of POPs fate in the ruminant.

Undernutrition combined with dietary mineral oil hastens depuration of stored dioxin and polychlorinated biphenyls in ewes. 2. Tissue distribution, mass balance and body burden.

Food safety crises involving persistent organic pollutants (POPs) lead to systematic slaughter of livestock to prevent contaminants from entering the food chain. Therefore, there is a need to develop strategies to depurate livestock moderately contaminated with POPs to reduce economic and social damage. This study aimed to test undernutrition (37% of energy requirements) combined with mineral oil (10% in total dry matter intake) in nine non-lactating ewes contaminated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and polychlorinated biphenyls (PCBs) 126 and 153 as a strategy to enhance the depuration of POPs through faecal excretion. To better understand the underlying mechanisms of the depuration process, lipophilic POPs and lipid fluxes were co-monitored in various body and excretion compartments. Body compartments (adipose tissues, muscle, liver and blood) and the total empty body were analyzed for lipids and POPs concentrations and burdens at slaughter, as well as excretion compartments (faeces and wool) collected during the depuration period. Decreases in empty body total and lipid weights were 6-fold higher in underfed and supplemented ewes compared to control ewes. In addition, over the depuration period undernutrition and supplementation treatment increased faecal TCDD, PCBs 126 and 153 excretions by 1.4- to 2.1-fold but tended to decrease wool PCB 153 excretion by 1.4-fold. This induced 2- to 3-fold higher decreases in the empty body POPs burdens for underfed and supplemented ewes. Nonetheless, when expressed relative to the calculated initial empty body burdens, burdens at slaughter decreased only slightly from 97%, 103% and 98% for control ewes to 92%, 97% and 94% for underfed and supplemented ones, for TCDD, PCBs 126 and 153, respectively. Fine descriptions at once of POPs kinetic (companion paper 1) and mass balance (companion paper 2), and of body lipid dynamics were very useful in improving our understanding of the fate of POPs in the ruminants.

A PBPK model to study the transfer of α-hexabromocyclododecane (α-HBCDD) to tissues of fast- and slow-growing broilers.

A physiologically based pharmacokinetic (PBPK) model was developed to investigate the production-specific factors involved in the transfer of α-hexabromocyclododecane (α-HBCDD) to broiler meat. The model describes growth and lipid deposition in tissues of fast- (FG) and slow- (SG) growing broilers from hatching to slaughter and simulates the exposure through the ingestion of contaminated feed or expanded polystyrene insulation material. Growth parameters were obtained from the literature while parameters relative to uptake, distribution, and elimination of α-HBCDD were adjusted using results of a previous experiment involving broilers exposed through feed throughout the rearing period or allowed to depurate before slaughter. The model was used to compare the two main edible tissues, breast and leg meat, as well as skin, and to investigate the variability within strain. Between strains and within strain, α-HBCDD assimilation efficiency (AE) is higher when the animals are slaughtered young or heavy. However, increasing slaughter age will lower α-HBCDD concentration in tissues, due to dilution. Based on fresh weight, the concentration of α-HBCDD in breast muscles and skin tends to be lower in SG than in FG broilers (-30 to +10%), while it is 10% to 80% higher in leg muscles. Compared to breast muscles, consuming leg muscles would elicit an exposure 9 and 16 times higher in FG and SG broilers, respectively. The consumption of skin together with muscles would multiply the exposure by up to 36 times compared to breast muscle alone. In case of acute exposure, the α-HBCDD concentration in tissues increased sharply, all the more since the animals are lighter in weight, and then decreased rapidly. In FG broilers, dilution through growth contributed for up to 37%, 28% and 97% to the decontamination of breast muscles, leg muscles and skin, respectively, depending on the duration of depuration before slaughter.

Comparison of common components analysis with principal components analysis and independent components analysis: Application to SPME-GC-MS volatolomic signatures.

The aim of this work is to compare a novel exploratory chemometrics method, Common Components Analysis (CCA), with Principal Components Analysis (PCA) and Independent Components Analysis (ICA). CCA consists in adapting the multi-block statistical method known as Common Components and Specific Weights Analysis (CCSWA or ComDim) by applying it to a single data matrix, with one variable per block. As an application, the three methods were applied to SPME-GC-MS volatolomic signatures of livers in an attempt to reveal volatile organic compounds (VOCs) markers of chicken exposure to different types of micropollutants. An application of CCA to the initial SPME-GC-MS data revealed a drift in the sample Scores along CC2, as a function of injection order, probably resulting from time-related evolution in the instrument. This drift was eliminated by orthogonalization of the data set with respect to CC2, and the resulting data are used as the orthogonalized data input into each of the three methods. Since the first step in CCA is to norm-scale all the variables, preliminary data scaling has no effect on the results, so that CCA was applied only to orthogonalized SPME-GC-MS data, while, PCA and ICA were applied to the "orthogonalized", "orthogonalized and Pareto-scaled", and "orthogonalized and autoscaled" data. The comparison showed that PCA results were highly dependent on the scaling of variables, contrary to ICA where the data scaling did not have a strong influence. Nevertheless, for both PCA and ICA the clearest separations of exposed groups were obtained after autoscaling of variables. The main part of this work was to compare the CCA results using the orthogonalized data with those obtained with PCA and ICA applied to orthogonalized and autoscaled variables. The clearest separations of exposed chicken groups were obtained by CCA. CCA Loadings also clearly identified the variables contributing most to the Common Components giving separations. The PCA Loadings did not highlight the most influencing variables for each separation, whereas the ICA Loadings highlighted the same variables as did CCA. This study shows the potential of CCA for the extraction of pertinent information from a data matrix, using a procedure based on an original optimisation criterion, to produce results that are complementary, and in some cases may be superior, to those of PCA and ICA.

Liver volatolomics to reveal poultry exposure to γ-hexabromocyclododecane (HBCD).

Hexabromocyclododecane (HBCD) is a critical emerging brominated flame retardant to which consumers can be exposed at high doses through a single food intake. Based on an animal experiment involving 3 groups of laying hens fed during 70 days with a control diet or γ-HBCD-contaminated diets at 0.1 or 10 µg γ-HBCD g-1 feed, this study aims to use the volatolome of biological samples for revealing markers of livestock exposure to HBCD. Liquid chromatography-tandem mass spectrometry was used to monitor the time-course of HBCD levels in bodily samples. Each liver was analyzed by solid-phase microextraction-gas chromatography-mass spectrometry for volatolome profiling. After 70 days, γ-HBCD concentrations in egg yolk, fat, liver and serum reached 54 ± 4, 85 ± 6, 31 ± 6, and 32 ± 4 ng g-1 lw, respectively, for the low exposure level and 4.6+/5.7, 7.8+/6.5, 3.9+/3.0 and 3.9+/6.1 µg g-1 lw, respectively, for the high exposure level. Isomerization of γ-HBCD into α- and ß-HBCD was observed in all tissues, at least for the high exposure level. Volatolome data allowed a significant discrimination between control and exposed animals whatever the feed contamination load, demonstrating a liver metabolic response to γ-HBCD exposure. The relevance of the twenty nine volatile exposure markers tentatively identified was discussed in light of literature data.

Hens can ingest extruded polystyrene in rearing buildings and lay eggs contaminated with hexabromocyclododecane.

The overall concentration of hexabromocyclododecane (HBCDD) in eggs is low although abnormally high concentrations exceeding 3000 ng g-1 lw have been reported. In order to test whether these contaminations may originate from the ingestion of insulating materials in rearing buildings, a group of 55 hens raised in a collective cage was provided with a 64-g piece of extruded polystyrene (XPS, 2.59% HBCDD of which 75, 15 and 10% as α-, ß- and γ-HBCDD, respectively). Hens entirely consumed the piece within 3 days, leading to a mean daily exposure of 4.7 mg HBCDD per kg body weight. Whole egg HBCDD concentration reached a maximum of 1037 ng HBCDD g-1 fresh weight (fw), recorded 2 days after the piece had disappeared, and decreased down to 86 ng g-1 fw within the 19 following days. In all these samples, HBCDD was made of 98.7 ± 0.7 and 1.3 ± 0.6% α- and ß-HBCDD, respectively, and 0.1% γ-HBCDD when quantified; it was enriched in (-)α- and (+)ß-HBCDD with enantiomeric fractions of 0.438 ± 0.009 and 0.579 ± 0.030, respectively. HBCDD was quantified in all the individual eggs collected the last day of experiment at concentrations ranging between 0.47 and 1361 ng g-1 fw, according to a lognormal distribution. The ingestion of XPS in degraded rearing buildings is thus a plausible cause of on-farm egg contamination by HBCDD which should be strictly avoided.

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.

Enantiomer-specific accumulation and depuration of α-hexabromocyclododecane (α-HBCDD) in chicken (Gallus gallus) as a tool to identify contamination sources.

A LC-ESI(-)-HRMS method dedicated to the analysis of 6 HBCDD enantiomers at trace levels in animal matrices was developed, using a cellulose based stationary phase with a particle size of 2.5 µm. This method was applied to a sample set derived from a kinetic study of α-HBCDD previously conducted in fast- and slow-growing chickens (Gallus gallus domesticus, n = 49, plus controls), in order to study the enantiomer specific accumulation and depuration of α-HBCDD in various tissues. Regarding abdominal adipose tissue, muscle and liver, the average enantiomeric fractions of α-HBCDD (EFα) for continuously exposed groups ranged between 0.434 and 0.467, with standard deviations below 0.014, showing a significant enrichment in (-)α enantiomer even accentuated for slow growing individuals during depuration with EFα reduced by about 0.020. Similar trends were observed for pooled plasma. Then, EFα of circulating plasma α-HBCDD appeared to closely reflect EFα in storage tissues and liver, suggesting some equilibrium. The racemic elimination of α enantiomer in excreta during the contamination phase indicated that no preferential gastrointestinal absorption took place. By contrast, preferential excretion of (-)α-HBCDD from the circulating compartment to the intestinal lumen occurred during the depuration. Finally, the method was applied to samples collected in three chicken farms, selected for total HBCDD levels in muscle in the ng/g range, as a tool to determine whether the contamination occurred ante- or post-mortem, according to the chiral signature. Ante-mortem contamination was hypothesised for 2 farms, with feed being excluded as contamination source.

Accumulation of α-hexabromocyclododecane (α-HBCDD) in tissues of fast- and slow-growing broilers (Gallus domesticus).

The aim of the current study was to describe the fate of ingested α-hexabromocyclododecane (α-HBCDD) in fast-growing (FG) and slow-growing (SG) broilers, through an exposure to a dietary concentration of 50 ng α-HBCDD g-1 feed during 42 and 84 days, respectively. Depuration parameters were assessed in SG broilers successively exposed during 42 days and depurated during 42 days. At market age, SG broilers had ingested 42% more feed than FG broilers, while their body weight gain per g of feed ingested was 34% lower. No isomerization of α- to ß- or γ-HBCDD forms occurred, while OH-HBCDD was identified as a product of α-HBCDD metabolism. Irrespective of the strain, abdominal fat displayed the highest α-HBCDD concentration on a lipid weight basis, followed leg muscles and then breast muscle, liver and plasma. The accumulation ratios of α-HBCDD were slightly higher in SG (6.7, 2.1, 2.6 and 9.9 in leg muscles, breast muscle, liver and abdominal fat, respectively) than in FG broilers (5.2, 2.2, 1.1 and 8.4, respectively). The elimination half-lives in SG broilers were 20, 12 and 19 d in leg muscles, breast muscle and abdominal fat, respectively, to which dilution through growth contributed for around 50%. The overall assimilation efficiency of α-HBCDD was estimated at 58 and 50% in FG and SG broilers, respectively, while 22 and 17% of α-HBCDD ingested were estimated to be eliminated in excreta as metabolites.

Tissue Distribution and Transfer to Eggs of Ingested α-Hexabromocyclododecane (α-HBCDD) in Laying Hens (Gallus domesticus).

The aim of the current study was to describe the fate of ingested α-hexabromocyclododecane (α-HBCDD) in laying hens. Individuals were exposed to two dietary concentrations of α-HBCDD (50 and 5 ng g(-1) feed) for 18 or 11 weeks followed by a 7-week decontamination period. The results show that no isomerization of α- to ß- or γ-HBCDD forms occurred, whereas OH-HBCDD was identified as a product of α-HBCDD metabolism. Irrespective of the level of feed contamination, estimates of steady-state accumulation ratios were 5.2, 3.6, and 9.2 and half-lives were estimated at 17.4, 22.8, and 35.3 days in egg yolk, liver tissue, and abdominal fat, respectively. The steady-state carry-over rate to eggs was 22.9%. Thus, α-HBCDD ingested by laying hens is readily transferred to eggs and significantly accumulates in adipose tissue.

Short-term effects of a perinatal exposure to the HBCDD α-isomer in rats: Assessment of early motor and sensory development, spontaneous locomotor activity and anxiety in pups.

The present study investigated the developmental neurotoxicity of an early exposure to α-HBCDD through the ingestion of contaminated hen's egg in pregnant and lactating Wistar female rats. Hens were given α-HBCDD-contaminated feed (40 ng/g fresh matter) for 5 and 10 days, which produced eggs with HBCDD content of 33 and 102 ng/glipid weight, respectively. Female rats were administered daily p.o. with an appropriate volume of the whole egg from the day of fertilization (GD0) to the weaning day for pups (PND21). Fetuses and pups were thus exposed continuously to α-HBCDD via the dam over a whole 42-day period that included both gestation and lactation. The administered egg volume was calculated on the basis of daily egg consumption in humans (0.7 egg/person/day) and duration of gestation and lactation in both species, which led animals to be exposed to α-HBCDD at levels of 22 and 66 ng/kg/day, respectively. Neurobehavioral development of pups was investigated from PND3 to PND25 using various tasks including the righting reflex (PND4), the grasping reflex (PND5), the negative geotaxis (PND9), the forelimb grip strength test (PND10) and the locomotor coordination test (PND20). Pup ultrasonic vocalizations were also recorded daily from PND4 to PND14. After weaning, behaviors related to spontaneous locomotor activity and anxiety were examined in the open-field (PND25) and in an elevated-plus maze (PND26), respectively. The results showed a significant decrease in body weight of pups exposed to the lower HBCDD level from PND3 to PND28, whereas the weight of rat pups given 66 ng/kg/day of HBCDD was not different from controls. During the first 3 weeks of life, impairments in motor maturation of pups were observed in a dose-dependent manner depending on the test, whereas no significant differences were reported between male and female pups. At PND26, the anxiety level of female rats exposed to the lowest dose of HBCDD (22 ng/kg/day) was significantly reduced whereas it remained unchanged in males. No significant variations were measured in rats exposed to the higher level of HBCDD (66 ng/kg/day). These results suggest the potent developmental neurotoxicity of an early chronic exposure to the HBCDD α-isomer through the ingestion of hen's eggs contaminated with this pollutant and question the long-lasting consequences of this exposure on behavior abilities and brain functioning in adulthood.

Modeling PCB transfer into hen eggs: influence of physiological characteristics of the animal.

Laying hens are likely to be exposed to a wide range of persistent organic pollutants (POPs), including polychlorinated biphenyls (PCBs). To improve the safety of poultry farming systems in terms of POPs, the present research focused on assessing the impact of physiological characteristics of the hen on the transfer of ingested PCBs to eggs. Modeling was used as a research tool to explore the impact of some physiological characteristics on the transfer of PCBs in the laying hen. The mathematical model simulates the dynamics of the size of the lipid compartments in the animal and the frequency of laying, with the PCB concentrations in egg yolk and adipose tissue being model outputs. Simulations were run to assess effects of animal characteristics on the transfer of PCBs to eggs. Laying rate proportionally influenced the PCB level of eggs and adipose tissue at steady state. Body fat diluted absorbed PCBs in the absence of laying and significantly influenced the decontamination rate of tissues during depuration after an exposure period. Application of the present model to actual exposure cases highlights its value in improving the support of risk management in livestock farming.

Kinetic study of chlordecone orally given to laying hens (Gallus domesticus).

The former use of chlordecone (CLD) in the French West Indies has resulted in long-term pollution of soils. In this area, CLD may be transferred into eggs of hens reared outdoors, through soil ingestion. In order to assess this risk, a kinetic study involving the contamination of laying hens (22 weeks of age) with a diet containing 500 µg CLD kg(-1) during 42 d, followed by a depuration period of 35 d was carried out. Forty-four hens were sequentially slaughtered all over the experimental period and their liver, egg, abdominal fat and serum were collected. Two additional edible tissues, pectoral and leg muscles, were collected in hens slaughtered at the end of the contamination period. The depuration half-life of CLD in liver, egg, abdominal fat and serum was estimated at 5.0 ± 0.38 (mean ± SE), 5.5 ± 0.29, 5.3 ± 0.37 and 5.1 ± 0.66 d, respectively. CLD concentration at the end of the contamination period reached 1640 ± 274, 460 ± 41, 331 ± 23, and 213 ± 8.5 µg kg(-1) fresh matter (FM), respectively. The corresponding concentrations in pectoral and leg muscles were 119 ± 8.4, 127 ± 11 µg kg(-1) FM, respectively. The steady state carry over rate of CLD in eggs reached 43 ± 7.6%. This experiment demonstrates the preferential accumulation of CLD in liver, its significant transfer to eggs and its quite short half-life. It is concluded that raising hens on even mildly contaminated areas would lead to products exceeding the regulatory maximum residue limit of 20 µg CLD kg(-1).

Contamination of free-range ducks by chlordecone in Martinique (French West Indies): a field study.

The former use of chlordecone (CLD) in the French West Indies has resulted in long-term pollution of soils and subsequently of food chains. In contaminated areas, free-range ducks used to control weeds in orchards may be exposed to CLD through polluted soil ingestion. The question arises whether they may be consumed. Muscovy ducks were raised on a guava orchard planted on a soil moderately contaminated (410 µg CLD/kg dry matter). Ducks were raised indoor up to 6 weeks of age and allowed to range freely outdoors thereafter. Twenty-nine females were sequentially slaughtered by groups of 2 to 5 ducks, after 4, 16, 19, 22 or 26 weeks spent in the orchard or after 16-17 weeks in the orchard followed by 3, 6 or 9 weeks in a closed shelter for depuration. CLD concentration increased from 258 to 1051, 96 to 278, 60 to 169 and 48 to 145 µg/kg fresh matter (FM) as the exposure through grazing increased from 4 to 22 weeks, in liver, abdominal fat and leg with and without skin, respectively. Eggs collected in the orchard contained up to 1001 µg CLD/kg FM. All these values exceeded the Maximum Residue Limit (MRL) of 20 µg/kg FM. CLD concentration in all tissues was divided by around 10 within the 9-week confinement period. Despite this quite rapid decontamination, it is estimated that 12-13 weeks would be required to achieve the MRL in liver and in eggs, and 5-6 weeks in leg muscle. Such durations would be too long in practice. Thus, the consumption of products from free-range ducks should be avoided, even in areas mildly contaminated with CLD.

Effects of condensed organic matter on PCBs bioavailability in juvenile swine, an animal model for young children.

The exposure assessment of polychlorinated biphenyls (PCBs) contaminated soils is a critical issue in terms of human health, especially since little reliable information on transfer of PCBs to humans via involuntary soil ingestion is available. Indeed, young children with their hand-to-mouth activity may be exposed to contaminated soils. The current study addresses the impact of soil organic matter (OM) condensation on bioavailability of sequestrated NDL-PCBs. Three artificial soils (ASs) were prepared according to OECD guideline 207. One standard soil (SS), devoid of OM, and two amended versions of this SS with fulvic acid (FA) or activated carbon (AC) were prepared to obtain 1% organic mass. This study involved fourteen juvenile male swine as a digestive physiology model of young children. Animals were randomly distributed into 4 contaminated groups (3 replicates) and a control one (2 replicates). During 10d, the piglets were fed AS or a corn oil spiked with 19200 ng of Aroclor 1254 per g of dry matter (6000 ng g(-1) of NDL-PCBs) to achieve an exposure dose of 1200 ng NDL-PCBskg(-1) of body weight per day. After 10d of oral exposure, NDL-PCBs in adipose tissue, liver and muscles were analyzed by GC-MS, after extraction and purification. Two distinct groups of treatments were found: on the one hand oil, SS and FA, on the other hand C and AC. This study highlights that condensed OM (AC) strongly reduces bioavailability whereas the less condensed one (FA) does not seem to have a significant effect. This result has to be considered as a first major step for further relative bioavailability studies involving mixture of different humic substances.

Relative bioavailability of tropical volcanic soil-bound chlordecone in piglets.

The application of chlordecone (CLD), a chlorinated polycyclic ketone pesticide, until 1993 in the French West Indies has resulted in long-term pollution of agricultural soils (10% of them exceed 1 mg kg(-1)). The aim of this study was to assess the impact of two tropical volcanic soils, an andosol and a nitisol, on CLD availability in piglets, using the relative bioavailability (RBA) approach. For both soils and relative to an oil matrix, RBA was close to 100%, indicating that CLD was not retained in the soil matrices during the piglet digestive process. Additionally, after a 14 day exposure period, liver and subcutaneous fat CLD concentrations exceeded the maximum residue limit (10 µg kg(-1) of fresh matter and 100 µg kg(-1) of fat for liver and subcutaneous fat, respectively) beyond a CLD ingestion of 2.1 and 6.8 µg CLD kg(-1) of body weight per day, respectively. Thus, rearing practices in CLD-contaminated areas should avoid involuntary soil ingestion by farm animals.

Relative bioavailability of soil-bound polychlorinated biphenyls in lactating goats.

Livestock may be exposed to organic pollutants via ingestion of contaminated matrices such as fodder or soil. The question on contribution of soil-bound polychlorinated biphenyls (PCB) to livestock exposure was not yet considered. The aim of this study was to assess the relative bioavailability of soil-bound PCB by assessing milk excretion of indicator PCB (I-PCB) after ingestion by goats of graded levels of PCB (mainly PCB forms 153, 180, and 138) in soil-contaminated feeds or in oil-contaminated feeds. Eight multiparous Alpine goats were grouped in 4 pairs on the basis of body weight and milk yield. In each pair, one goat was assigned to the soil feeds and the other one to the oil feeds. The experiment consisted of a 7-d adaptation period, followed by a 96-d exposure period. The exposure period was divided into 3 successive 32-d periods during which each goat received either 3 soil feeds or 3 oil feeds, distributed in increasing rank of contamination. During the last week of each 32-d period, milk from each goat was collected during 3 successive 24-h periods, stored at -20°C, and freeze dried before analysis (extraction by accelerated solvent extraction, followed by gas chromatography-mass spectrometry analysis). Bioavailability of I-PCB from soil or spiked oil feeds was estimated by means of the slope-ratio method from I-PCB concentration in milk in response to ingested I-PCB. Relative bioavailability was found to vary from 36 to 50% for PCB 118, 138, and 153 and it was 73% for PCB 180. When considered globally, the response obtained with the I-PCB was estimated to 51%. Relative bioavailability was not established for PCB 52 and 101, compounds known to be readily cleared and showing low concentrations in milk. For PCB 28, no significant interaction was found between matrix and dose. This experiment reveals that PCB bound to soil are potentially liberated from soil during the digestive process and may undergo absorption, distribution, metabolism, and excretion. Thus, soil has to be considered as a risk matrix for ruminants and rearing practices in contaminated areas should strictly reduce the risk of soil ingestion by the ruminants.

Relative bioavailability of tropical volcanic soil-bound chlordecone in laying hens (Gallus domesticus).

The former use of chlordecone (CLD) in the French West Indies has resulted in long-term pollution of soils and of food chains. CLD may be transferred into eggs of hens reared outdoors, through polluted soil ingestion. Tropical volcanic soils display variable capacities of pollutant retention: CLD is less available and more persistent in andosol than in nitisol. The impact of soil type on CLD bioavailability to hens was tested through a relative bioavailability study. The deposition of CLD in egg yolk and in abdominal fat was measured in 42 individually housed laying hens fed with diets containing graded levels of CLD from polluted andosol, nitisol, or spiked oil during 23 days. Within each ingested matrix, the concentration of CLD in yolk and in abdominal fat linearly increased with the amount of ingested CLD (P < 0.001). However, the response to andosol diets and to nitisol diets was not different from the response to oil diets (P > 0.1), indicating that CLD was equally bioavailable to laying hens, irrespective of the matrix. This suggests that the hen's gastrointestinal tract efficiently extracts CLD from the two tropical volcanic soils, regardless of their retention capacity. Thus, hens reared on polluted soils with CLD may lay contaminated eggs.

In vivo validation of the unified BARGE method to assess the bioaccessibility of arsenic, antimony, cadmium, and lead in soils.

The relative bioavailability of arsenic, antimony, cadmium, and lead for the ingestion pathway was measured in 16 soils contaminated by either smelting or mining activities using a juvenile swine model. The soils contained 18 to 25,000 mg kg(-1) As, 18 to 60,000 mg kg(-1) Sb, 20 to 184 mg kg(-1) Cd, and 1460 to 40,214 mg kg(-1) Pb. The bioavailability in the soils was measured in kidney, liver, bone, and urine relative to soluble salts of the four elements. The variety of soil types, the total concentrations of the elements, and the range of bioavailabilities found were considered to be suitable for calibrating the in vitro Unified BARGE bioaccessibility method. The bioaccessibility test has been developed by the BioAccessibility Research Group of Europe (BARGE) and is known as the Unified BARGE Method (UBM). The study looked at four end points from the in vivo measurements and two compartments in the in vitro study ("stomach" and "stomach and intestine"). Using benchmark criteria for assessing the "fitness for purpose" of the UBM bioaccessibility data to act as an analogue for bioavailability in risk assessment, the study shows that the UBM met criteria on repeatability (median relative standard deviation value <10%) and the regression statistics (slope 0.8 to 1.2 and r-square > 0.6) for As, Cd, and Pb. The data suggest a small bias in the UBM relative bioaccessibility of As and Pb compared to the relative bioavailability measurements of 3% and 5% respectively. Sb did not meet the criteria due to the small range of bioaccessibility values found in the samples.