Bruno Le Bizec

Exposure assessment of French women and their newborn to brominated flame retardants : Determination of tri-to deca-polybromodiphenylethers (PBDE) in maternal adipose tissue, serum, breast milk and cord serum

In the frame of a French monitoring program, tri- to deca- polybromodiphenylethers (PBDE) have been measured in maternal and cord serum, adipose tissue, and breast milk samples, collected from 93 volunteer women during caesarean deliveries. The seven major tri- to heptaBDE (BDE-28, 47, 99, 100, 153, 154, and 183) were detected in adipose tissue and breast milk with cumulated median values of 2.59 and 2.51 ng g(-1) l w. Nine highly brominated octa- to decaBDE (BDE-196, 197, 201, 202, 203, 206, 207, 208 and 209) was performed in the same samples, with cumulated median values of 2.73 and 3.39 ng g(-1) l w in adipose tissue and breast milk, respectively. At this opposite, median levels of octa- to decaBDE in maternal and cord serum appeared significantly higher than the levels of tri- to heptaBDE in the same matrices, i.e. 8.85 and 12.34 versus 0.98 and 0.69 ng g(-1) l w, respectively.

Exposure assessment of French women and their newborns to tetrabromobisphenol-A: Occurrence measurements in maternal adipose tissue, serum, breast milk and cord serum

A French monitoring study was initiated to evaluate the exposure of fetus and newborn to brominated flame retardants (BFR). A previously developed multi-residue analytical method was used for measuring the main classes of BFR (tetrabromobisphenol-A, and tri- to decabomodiphenyl ethers) in various human biological matrices. Analyzed samples (maternal and cord serum, adipose tissue and breast milk) were collected from 93 volunteer women during caesarean deliveries. TBBPA was detected in 44% of the analyzed breast milk samples, at levels varying from 0.06 to 37.34 ng g(-1) lipid weight, but was not detected in adipose tissue. This compound was also detected in 30% of the analyzed serum samples, with similar average values in maternal and cord serum (154 pg g(-1) fresh weight versus 199 pg g(-1) fresh weight, respectively). The interpretation of the collected data permitted the demonstration of (1) a significant exposure to TBBPA both for mothers and fetuses and (2) a possible risk of overexposure of newborns through breastfeeding.

Fate and Complex Pathogenic Effects of Dioxins and Polychlorinated Biphenyls in Obese Subjects before and after Drastic Weight Loss

Background In humans, persistent organic pollutants (POPs) are stored primarily in adipose tissue. Their total body burden and their contribution to obesity-associated diseases remain unclear. Objectives We characterized POP total body burden and their redistribution in obese individuals before and after drastic weight loss and compared these values with a variety of molecular, biological, and clinical parameters. Methods Seventy-one obese subjects were enrolled and underwent bariatric surgery. Blood and adipose tissue samples were obtained at different times from these individuals as well as from 18 lean women. Results POP content (17 dioxins/furans and 18 polychlorinated biphenyl congeners) in different adipose tissue territories was similar, allowing us to assess total POP body burden from a single biopsy. Total POP body burden was 2 to 3 times higher in obese than in lean individuals. We also found increased expression of some POP target genes in obese adipose tissue. Drastic weight loss led to increased serum POPs and, within 6–12 months, to a significant 15% decrease in total polychlorinated biphenyl body burden. Importantly, serum POP levels were positively correlated with liver toxicity markers and lipid parameters, independently of age and body mass index. Conclusions POP content in adipose tissue and serum correlate with biological markers of obesity-related dysfunctions. Drastic weight loss leads to a redistribution of POPs and to a moderate decrease of their total body burden.

Toxicological Function of Adipose Tissue: Focus on Persistent Organic Pollutants

Background: Adipose tissue (AT) is involved in several physiological functions, including metabolic regulation, energy storage, and endocrine functions. Objectives: In this review we examined the evidence that an additional function of AT is to modulate persistent organic pollutant (POP) toxicity through several mechanisms. Methods: We reviewed the literature on the interaction of AT with POPs to provide a comprehensive model for this additional function of AT. Discussion: As a storage compartment for lipophilic POPs, AT plays a critical role in the toxicokinetics of a variety of drugs and pollutants, in particular, POPs. By sequestering POPs, AT can protect other organs and tissues from POPs overload. However, this protective function could prove to be a threat in the long run. The accumulation of lipophilic POPs will increase total body burden. These accumulated POPs are slowly released into the bloodstream, and more so during weight loss. Thus, AT constitutes a continual source of internal exposure to POPs. In addition to its buffering function, AT is also a target of POPs and may mediate part of their metabolic effects. This is particularly relevant because many POPs induce obesogenic effects that may lead to quantitative and qualitative alterations of AT. Some POPs also induce a proinflammatory state in AT, which may lead to detrimental metabolic effects. Conclusion: AT appears to play diverse functions both as a modulator and as a target of POPs toxicity.

Validation of analytical methods based on mass spectrometric detection according to the “2002/657/EC” European decision: guideline and application

Abstract The purpose of the present paper is to present an interpretation of the concepts introduced in the new 2002/657/EC European decision and to propose a practical guideline dedicated to the validation of analytical methods based on mass spectrometry. Considering both the statistical significance of the results and practical aspects, the minimal number of assays permitting a satisfying validation to be achieved appeared to be 45 for qualitative methods and 55 for quantitative methods. The parameters validated with this protocol are specificity, sensitivity, linearity, decision limit (CC α ), repeatability, detection capability (CC β ) and recovery. It is proposed to estimate these parameters on the basis of the most intense (or unique) ion for screening methods and on the basis of the “critical ion” (less intense ion permitting the unambiguous identification of the analyte according to the required number of identification points) for confirmatory methods. An application of this guideline is presented and discussed, through the validation of a liquid chromatography-tandem mass spectrometric (LC–MS/MS) method dedicated to the determination of the corticosteroid triamcinolone acetonide (Tri Acn) in meat samples.

Assessment of circulating sex steroid levels in prepubertal and pubertal boys and girls by a novel ultrasensitive gas chromatography-tandem mass spectrometry method.

CONTEXT Estrogens and androgens play key roles for pubertal onset and sexual maturation. Most currently used immunoassays are not sensitive enough to accurately measure the low circulating levels of sex steroids in children without any signs of puberty. However, this does not exclude that sex steroids have important biological roles in prepubertal children. OBJECTIVES To accurately determine levels of sex steroid hormones and their metabolites in serum of healthy children before any physical signs of puberty and to evaluate possible sex differences. MAIN OUTCOME MEASURES Total (unconjugated plus conjugated) serum levels of 17beta-testosterone, 17alpha-testosterone, 5alpha-dihydrotestosterone, 5beta-dihydrotestosterone, androsterone, etiocholanolone, estradiol, and estrone measured by an ultrasensitive method based on gas chromatography-tandem mass spectrometry in samples from 81 healthy schoolchildren (42 boys) without any signs of puberty. For comparison, 48 pubertal children were studied. RESULTS 17beta-Estradiol levels in prepubertal boys were undetectable or extremely low (median < 3.7 pmol/liter), whereas levels in prepubertal girls were significantly higher (median 9.6 pmol/liter, P < 0.001). Among the older prepubertal children (>8 yr), girls had significantly higher androsterone (4.07 vs. 1.45 nmol/liter, P < 0.05), etiocholanolone (5.45 vs. 1.95 nmol/liter, P < 0.0001), 5alpha-dihydrotestosterone (0.11 vs. <0.10 nmol/liter, P < 0.01), and 17beta-testosterone concentrations (0.69 vs. 0.47 nmol/liter, P < 0.05) compared with similarly aged prepubertal boys. CONCLUSION Using an accurate and sensitive method, we found significantly higher levels of estrogens as well as androgen metabolites in prepubertal girls compared with age-matched boys. The higher prepubertal sex steroid levels in girls may contribute to their earlier onset of puberty including pubic hair development.

Collision‐induced dissociation of corticosteroids in electrospray tandem mass spectrometry and development of a screening method by high performance liquid chromatography/tandem mass spectrometry

A screening method based on liquid chromatography/electrospray tandem mass spectrometry was developed in order to control the illegal use of corticosteroids as growth promoters in cattle. The objective was the detection of low residue levels of corticosteroids or metabolites in biological matrices. Relative to other studies published on this subject, the present work focused on enhancing specificity and sensitivity. Firstly, fragmentation of corticosteroids by collision-induced dissociation was studied. In positive mode, the losses of H(2)O for each hydroxyl group fixed on the molecule, as well as the loss of HF or HCl for halogenated compounds, were observed. For higher collision energy, fragmentations in the B, C and D rings were induced. The negative mode was found to be more specific, inducing a cleavage of the C(20)-C(21) bond with concomitant loss of formaldehyde (CH(2)O). Secondly, three acquisition methods in the negative mode were studied and evaluated, recorded signals being the parent ion [M + acetate](-) and the two daughter ions, [M - H](-) and [M - H - CH(2)O](-). For dexamethasone, MS/MS instrumental detection limits of fragment ion and neutral loss scans, and of multiple reaction monitoring (MRM), were 250, 20 and 5 pg injected, respectively. The MRM method was then evaluated with the objective of use for the detection of corticosteroid residues in biological samples (urine, hair, muscle) and for a metabolism study.

PFOS (perfluorooctanesulfonate) in serum is negatively associated with testosterone levels, but not with semen quality, in healthy men

STUDY QUESTION Is exposure to perfluorinated compounds (PFCs) associated with testicular function (reproductive hormone levels and semen quality) in healthy men? SUMMARY ANSWER PFOS levels were significantly negatively associated with serum testosterone (total and calculated free), but not with any other reproductive hormones or semen quality. WHAT IS KNOWN ALREADY In animals, some PFCs have endocrine disrupting potential, but few studies have investigated PFCs in relation to human testicular function. Previously, we and others have observed a negative association between serum PFC levels and sperm morphology. The potential associations with reproductive hormones remain largely unresolved. STUDY DESIGN, SIZE, DURATION A cross-sectional study of 247 men was conducted during 2008-2009. PARTICIPANTS/MATERIALS, SETTING, METHODS Healthy men from the general population, median age of 19 years, gave serum and semen samples. Serum samples were analysed for total testosterone (T), estradiol (E), sex hormone-binding globulin (SHBG), luteinizing hormone (LH), follicle-stimulating hormone (FSH) and inhibin-B and 14 PFCs, including perfluorooctanesulfonate (PFOS). Semen samples were analysed according to the WHO criteria. MAIN RESULTS AND THE ROLE OF CHANCE PFOS levels were negatively associated with testosterone (T), calculated free testosterone (FT), free androgen index (FAI) and ratios of T/LH, FAI/LH and FT/LH. Other PFCs were found at lower levels than PFOS and did not exhibit the same associations. PFC levels were not significantly associated with semen quality. PFOS levels in these samples collected in 2008-2009 were lower than in our previous study of men participating in 2003. LIMITATIONS, REASONS FOR CAUTION Results were robust to adjustment for relevant confounders; however, the possibility of chance associations due to multiple testing or effects of uncontrolled confounding cannot be ruled out. WIDER IMPLICATIONS OF THE FINDINGS Our previous findings of decreased sperm morphology in the most highly PFC exposed men were not replicated, possibly due to a lack of highly exposed individuals; however, a recent independent study also did corroborate such an inverse association. The negative association between serum PFOS and testosterone indicates that testosterone production may be compromised in individuals with high PFOS exposure. STUDY FUNDING/COMPETING INTEREST(S) The authors received financial support from the European Commission (DEER, FP7-2007-212844), the Danish Agency for Science, Technology and Innovation (grant nos. 27107068 and 09-067180), Rigshospitalet (grant no. 961506336), the University of Copenhagen, the Danish Ministry of Health and the Danish Environmental Protection Agency (MST-621-00013), and Kirsten and Freddy Johansen Foundation (grant no. 95-103-72087). The funding organizations played no role in the design and conduct of the study, in collection, management, analysis and interpretation of the data; or in the presentation, review or approval of the manuscript. The authors declare that they have no competing financial interests.

Exposure assessment of fetus and newborn to brominated flame retardants in France: preliminary data

Brominated flame retardants (BFR) are chemicals extensively used in many manufactured products to reduce the risk of fire, but also environmental pollutants. In order to assess the potential risk linked to these compounds in human, a French monitoring study was initiated to evaluate the exposure of fetus and newborn. A previously described multi-residue analytical method was used, for measuring the main classes of BFR (hexabromocyclododecane, tetrabromobisphenol-A, and tri- to deca-polybromodiphenylethers) in various biological matrices. These analyzed samples (maternal and umbilical serum, adipose tissue and breast milk) were collected on volunteer women during caesarean deliveries. Preliminary results obtained on 26 individuals (mother/newborn pairs) mainly demonstrated the presence of polybromodiphenylethers (PBDE) and tetrabromobisphenol A both in maternal and fetal matrices, and a possible risk of overexposure of newborns through breastfeeding. Contaminations levels were found globally in the ng/g lipid weight range, consistent with other published European data. Exposure results regarding highly brominated PBDE congeners (octa- to deca-BDE) appeared particularly informative and non-commonly reported, these compounds accounting for around 50% of the total PBDE load. Additional data collection and metabolism investigations are now on-going. A more complete statistical analysis related to this BFR exposition study will be provided in a next future.

Multi-residue extraction–purification procedure for corticosteroids in biological samples for efficient control of their misuse in livestock production

A fast and efficient multi-residue extraction-purification procedure was developed for 12 corticosteroids in biological matrices (hair, urine and meat), in order to control their illegal use as growth promoters in cattle. Detection and identification of the analytes were achieved using a previously described LC-MS-MS method based on negative electrospray ionisation and a triple quadrupole analyser. The presented procedures included acid (hair) or enzymatic (urine and meat) hydrolysis, C18 reversed-phase SPE, Na2CO3 liquid-liquid clean-up and SiOH normal-phase SPE. The detection limits of the developed methods were between 2.9 and 9.3 pg/mg (ppb) for hair samples and in the 40 - 70 pg/g (ppt) range for the urine or meat samples. The acid hydrolysis used for corticosteroid extraction in hair was optimised using an experimental design and response surface methodology. Achieved performances were linked to a physico-chemical approach based on the corticosteroids specific C17 side-chain. This original multi-residue and multi-matrices analytical methodology will be used for further metabolism studies.