In Silico Identification of Potential Thyroid Hormone System Disruptors among Chemicals in Human Serum and Chemicals with a High Exposure Index.

Data on toxic effects are at large missing the prevailing understanding of the risks of industrial chemicals. Thyroid hormone (TH) system disruption includes interferences of the life cycle of the thyroid hormones and may occur in various organs. In the current study, high-throughput screening data available for 14 putative molecular initiating events of adverse outcome pathways, related to disruption of the TH system, were used to develop 19 in silico models for identification of potential thyroid hormone system-disrupting chemicals. The conformal prediction framework with the underlying Random Forest was used as a wrapper for the models allowing for setting the desired confidence level and controlling the error rate of predictions. The trained models were then applied to two different databases: (i) an in-house database comprising xenobiotics identified in human blood and ii) currently used chemicals registered in the Swedish Product Register, which have been predicted to have a high exposure index to consumers. The application of these models showed that among currently used chemicals, fewer were overall predicted as active compared to chemicals identified in human blood. Chemicals of specific concern for TH disruption were identified from both databases based on their predicted activity.

Fate of Per- and Polyfluoroalkyl Substances from Durable Water-Repellent Clothing during Use.

To make outdoor clothing water- or dirt-repellent, durable water-repellent (DWR) coatings based on side-chain fluorinated polymers (SFPs) are used. During use of outdoor clothing, per- and polyfluoroalkyl substances (PFASs) can be emitted from the DWR to the environment. In this study, the effects of aging, washing, and tumble drying on the concentration of extractable PFASs in the DWR of perfluorohexane-based short-chain SFPs (FC-6 chemistry) and of perfluorooctane-based long-chain SFPs (FC-8 chemistry) were assessed. For this purpose, polyamide (PA) and polyester (PES) fabrics were coated with FC-6- and FC-8-based DWRs. Results show that aging of the coated fabrics causes an increase in concentration and formation of perfluoroalkyl acids (PFAAs). The effect of aging on the volatile PFASs depends on the type of fabric. Washing causes a decrease in PFAA concentrations, and in general, volatile PFASs are partly washed out of the textiles. However, washing can also increase the extractable concentration of volatile PFASs in the fabrics. This effect becomes stronger by a combination of aging and washing. Tumble drying does not affect the PFAS concentrations in textiles. In conclusion, aging and washing of fabrics coated with the DWR based on SFPs release PFASs to the environment.

Estimation of the concentrations of hydroxylated polychlorinated biphenyls in human serum using ionization efficiency prediction for electrospray.

Hydroxylated PCBs are an important class of metabolites of the widely distributed environmental contaminants polychlorinated biphenyls (PCBs). However, the absence of authentic standards is often a limitation when subject to detection, identification, and quantification. Recently, new strategies to quantify compounds detected with non-targeted LC/ESI/HRMS based on predicted ionization efficiency values have emerged. Here, we evaluate the impact of chemical space coverage and sample matrix on the accuracy of ionization efficiency-based quantification. We show that extending the chemical space of interest is crucial in improving the performance of quantification. Therefore, we extend the ionization efficiency-based quantification approach to hydroxylated PCBs in serum samples with a retraining approach that involves 14 OH-PCBs and validate it with an additional four OH-PCBs. The predicted and measured ionization efficiency values of the OH-PCBs agreed within the mean error of 2.1 × and enabled quantification with the mean error of 4.4 × or better. We observed that the error mostly arose from the ionization efficiency predictions and the impact of matrix effects was of less importance, varying from 37 to 165%. The results show that there is potential for predictive machine learning models for quantification even in very complex matrices such as serum. Further, retraining the already developed models provides a timely and cost-effective solution for extending the chemical space of the application area.

Microglial CD74 Expression Is Regulated by TGFß Signaling.

Microglia play important roles during physiological and pathological situations in the CNS. Several reports have described the expression of Cd74 in disease-associated and aged microglia. Here, we demonstrated that TGFß1 controled the expression of Cd74 in microglia in vitro and in vivo. Using BV2 cells, primary microglia cultures as well as Cx3cr1CreERT2:R26-YFP:Tgfbr2fl/fl in combination with qPCR, flow cytometry, and immunohistochemistry, we were able to provide evidence that TGFß1 inhibited LPS-induced upregulation of Cd74 in microglia. Interestingly, TGFß1 alone was able to mediate downregulation of CD74 in vitro. Moreover, silencing of TGFß signaling in vivo resulted in marked upregulation of CD74, further underlining the importance of microglial TGFß signaling during regulation of microglia activation. Taken together, our data indicated that CD74 is a marker for activated microglia and further demonstrated that microglial TGFß signaling is important for regulation of Cd74 expression during microglia activation.

Carbamazepine Ozonation Byproducts: Toxicity in Zebrafish (Danio rerio) Embryos and Chemical Stability.

Carbamazepine (CBZ) is an anticonvulsant medication with highly persistent properties in the aquatic environment, where it has the potential to affect nontarget biota. Because CBZ and many other pharmaceuticals are not readily removed in conventional sewage treatment plants (STP), additional STP effluent treatment technologies are being evaluated and implemented. Whole effluent ozonation is a prospective method to remove pharmaceuticals such as CBZ, yet knowledge on the toxicity of CBZ ozonation byproducts (OBPs) is lacking. This study presents, for the first time, in vivo individual and mixture toxicity of four putative OBPs, that is, carbamazepine 10,11-epoxide, 10,11-Dihydrocarbamazepine, 1-(2-benzaldehyde)-4-hydro-(1H,3H)-quinazoline-2-one (BQM), and 1-(2-benzaldehyde)-(1H,3H)-quinazoline-2,4-dione (BQD) in developing zebrafish (Danio rerio) embryos. BQM and BQD were isolated from the ozonated solution as they were not commercially available. The study confirmed that the OBP mixture caused embryotoxic responses comparable to that of ozonated CBZ. Individual compound embryotoxicity assessment further revealed that BQM and BQD were the drivers of embryotoxicity. OBP chemical stability in ozonated CBZ water solution during 2 week dark storage at 22 °C was also assessed. The OBP concentrations remained over time, except for BQD which decreased by 94%. Meanwhile, ozonated CBZ persistently induced embryotoxicity over 2 week storage, potentially illustrating environmental concern.

Thyroid disruption properties of three indoor dust chemicals tested in Silurana tropicalis tadpoles.

Indoor dust contains a multitude of industrial chemicals, and ingestion of dust is considered an important exposure route to organic contaminants. Some of these contaminants have been shown to interfere with the thyroid system, which may result in significant consequences on public health. The amphibian metamorphosis is a thyroid hormone-dependent process, which can be used as an in vivo model for studies on thyroid hormone-disrupting potency. Three contaminants of indoor dust were tested on metamorphosing Silurana (Xenopus) tropicalis tadpoles. The tested chemicals were Tris (1,3-dichloroisopropyl) phosphate (TDCiPP), tetrabromobisphenol-A (TBBPA) and propylparaben (PrP). Measurements reflecting general growth, development progress and thyroid epithelial cell height were performed on the exposed tadpoles as well as chemical analyses of the exposure water. It was shown that TDCiPP acts as a thyroid hormone-disrupting chemical in metamorphosing tadpoles by causing increased epithelial cell height in thyroid glands after exposure to a nominal concentration of 0.010 mg/L and in higher concentrations. TBBPA caused reductions in general growth of tadpoles at the nominal concentration 0.125 mg/L, and PrP caused acute toxicity at the nominal concentration 12.5 mg/L. However, no evident indications of specific thyroid-disrupting effects caused by TBBPA or PrP were observed.

Source characterisation and distribution of selected PCBs, PAHs and alkyl PAHs in sediments from the Klip and Jukskei Rivers, South Africa.

A study of the distribution of polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) utilising 16 priority PAHs, benzo(e)pyrene, perylene, 19 alkylated PAHs and 31 ortho substituted PCBs in South Africa is presented. It was aimed to (a) deduce characteristic contamination patterns for both PCBs and PAHs and (b) provide the first comprehensive dataset for establishment of source characterisation of PCBs and PAHs. This is in line with new South African legislation on mandatory monitoring of PCB and PAH emissions. Bar charts, principal component analysis (PCA) and biplots were utilised to identify signature contamination patterns and distribution of PCBs and PAHs within the Jukskei and Klip Rivers. Sediments from the Jukskei and Klip River catchments both showed distinct contamination signatures for hexa to nonachlorinated PCBs, characteristic of contamination by Aroclor 1254 and 1260 technical mixtures. PCB signature patterns in order of abundance were 138 > 180 > 206 > 153 > 187 > 149 and 138 > 153 > 180 > 149 > 187 > 110 > 170 for the Jukskei and Klip River sediments, respectively. The upstream Alberton point had the highest Σ31 PCB and Σ (parent+alkyl) PAH concentrations in the Klip River of 61 and 6000 µg kg-1 dry weight (dw), respectively. In the Jukskei River, the upstream Marlboro point had the highest Σ31 PCB concentration of 19 µg kg-1 dw and the N14 site recorded the highest Σ (parent+alkyl) PAH concentration of 2750 µg kg-1 dw. PAH concentrations in both the Jukskei and Klip Rivers were significantly higher than the PCB concentrations. Fluoranthene, phenanthrene and pyrene were found in the highest concentrations in both the Jukskei and Klip River sediments. Both the Jukskei and Klip River sediments showed trends of a mixed pyrogenic-petrogenic PAH source contamination.

In Silico Approach To Identify Potential Thyroid Hormone Disruptors among Currently Known Dust Contaminants and Their Metabolites.

Thyroid hormone disrupting chemicals (THDCs) interfere with the thyroid hormone system and may induce multiple severe physiological disorders. Indoor dust ingestion is a major route of THDCs exposure in humans, and one of the molecular targets of these chemicals is the hormone transporter transthyretin (TTR). To virtually screen indoor dust contaminants and their metabolites for THDCs targeting TTR, we developed a quantitative structure-activity relationship (QSAR) classification model. The QSAR model was applied to an in-house database including 485 organic dust contaminants reported from literature data and their 433 in silico derived metabolites. The model predicted 37 (7.6%) dust contaminants and 230 (53.1%) metabolites as potential TTR binders. Four new THDCs were identified after testing 23 selected parent dust contaminants in a radio-ligand TTR binding assay; 2,2',4,4'-tetrahydroxybenzophenone, perfluoroheptanesulfonic acid, 3,5,6-trichloro-2-pyridinol, and 2,4,5-trichlorophenoxyacetic acid. These chemicals competitively bind to TTR with 50% inhibition (IC50) values at or below 10 µM. Molecular docking studies suggested that these THDCs interacted similarly with TTR via the residue Ser117A, but their binding poses were dissimilar to the endogenous ligand T4. This study identified new THDCs using an in silico approach in combination with bioassay testing and highlighted the importance of metabolic activation for TTR binding.

Tracing thyroid hormone-disrupting compounds: database compilation and structure-activity evaluation for an effect-directed analysis of sediment.

A variety of anthropogenic compounds has been found to be capable of disrupting the endocrine systems of organisms, in laboratory studies as well as in wildlife. The most widely described endpoint is estrogenicity, but other hormonal disturbances, e.g., thyroid hormone disruption, are gaining more and more attention. Here, we present a review and chemical characterization, using principal component analysis, of organic compounds that have been tested for their capacity to bind competitively to the thyroid hormone transport protein transthyretin (TTR). The database contains 250 individual compounds and technical mixtures, of which 144 compounds are defined as TTR binders. Almost one third of these compounds (n = 52) were even more potent than the natural hormone thyroxine (T4). The database was used as a tool to assist in the identification of thyroid hormone-disrupting compounds (THDCs) in an effect-directed analysis (EDA) study of a sediment sample. Two compounds could be confirmed to contribute to the detected TTR-binding potency in the sediment sample, i.e., triclosan and nonylphenol technical mixture. They constituted less than 1% of the TTR-binding potency of the unfractionated extract. The low rate of explained activity may be attributed to the challenges related to identification of unknown contaminants in combination with the limited knowledge about THDCs in general. This study demonstrates the need for databases containing compound-specific toxicological properties. In the framework of EDA, such a database could be used to assist in the identification and confirmation of causative compounds focusing on thyroid hormone disruption.

Exploring a mechanism-based approach for the identification of endocrine disruptors using Adverse Outcome Pathways (AOPs) and New Approach Methodologies (NAMs) : A perfluorooctane sulfonic acid case study.

Endocrine disruptors (EDs) pose a serious threat to human health and the environment and require a comprehensive evaluation to be identified. The identification of EDs require a substantial amount of data, both in vitro and in vivo, due to the current scientific criteria in the EU. At the same time, the EU strives to reduce animal testing due to concerns regarding animal welfare and sensitivity of animal studies to adequately detect adverse effects relevant for human health. Perfluorooctane sulfonic acid (PFOS) is a persistent organic pollutant that is suspected to be an ED based on academic research, however it is not identified as such from a regulatory perspective. It has previously been shown that PFOS has the potential to cause neurotoxicity as well as affect the thyroid system, and it is known that specific thyroid hormone levels are critical in the development of the brain during. In this work, the aim was to evaluate a mechanism-based approach to identify ED properties of PFOS based on the Adverse Outcome Pathway (AOP) framework and using New Approach Methods (NAMs), by comparing this approach to an ED assessment based on the currently available guidance document. An AOP network (AOPN) was generated for the thyroid modality, and AOPs leading to developmental neurotoxicity (DNT) were identified. A literature search and screening process based on the AOPN, and systematic review methodology, was performed, followed by a rigorous Weight-of-Evidence (WoE) assessment. Evidence was mapped back onto the AOPN used for the literature search, to identify possible endocrine Modes-of-Action (MoAs) for PFOS and data gaps in the two assessments. It could be concluded that PFOS fulfils the criteria for ED classification in the standard ED assessment, but not in the mechanism-based assessment. The need for quantitative information, such as quantitative AOPs, for the mechanism-based approach is discussed. The possibility of a directly neurotoxic alternative MoA was also highlighted based on available in vitro data. Opportunities and challenges with implementing AOPs and NAMs into the regulatory assessment of EDs, and assessing hazard in the Next Generation Risk Assessment, is discussed. This case study exploring the mechanism-based approach to ED identification represents an important step toward more accurate and predictive assessment of EDs based on AOPs and NAMs, and to the Next Generation Risk Assessment (NGRA) concept.

Gas Phase Reactivity of Isomeric Hydroxylated Polychlorinated Biphenyls.

Identification of stereo- and positional isomers detected with high-resolution mass spectrometry (HRMS) is often challenging due to near-identical fragmentation spectra (MS2), similar retention times, and collision cross-section values (CCS). Here we address this challenge on the example of hydroxylated polychlorinated biphenyls (OH-PCBs) with the aim to (1) distinguish between isomers of OH-PCBs using two-dimensional ion mobility spectrometry (2D-IMS) and (2) investigate the structure of the fragments of OH-PCBs and their fragmentation mechanisms by ion mobility spectrometry coupled to high-resolution mass spectrometry (IMS-HRMS). The MS2 spectra as well as CCS values of the deprotonated molecule and fragment ions were measured for 18 OH-PCBs using flow injections coupled to a cyclic IMS-HRMS. The MS2 spectra as well as the CCS values of the parent and fragment ions were similar between parent compound isomers; however, ion mobility separation of the fragment ions is hinting at the formation of isomeric fragments. Different parent compound isomers also yielded different numbers of isomeric fragment mobilogram peaks giving new insights into the fragmentation of these compounds and indicating new possibilities for identification. For spectral interpretation, Gibbs free energies and CCS values for the fragment ions of 4'-OH-CB35, 4'-OH-CB79, 2-OH-CB77 and 4-OH-CB107 were calculated and enabled assignment of structures to the isomeric mobilogram peaks of [M-H-HCl]- fragments. Finally, further fragmentation of the isomeric fragments revealed different fragmentation pathways depending on the isomeric fragment ions.

Metabolic activation of nonpolar sediment extracts results in enhanced thyroid hormone disrupting potency.

Traditional sediment risk assessment predominantly considers the hazard derived from legacy contaminants that are present in nonpolar sediment extracts, such as polychlorinated biphenyls (PCBs), dioxins, furans (PCDD/Fs), and polyaromatic hydrocarbons (PAHs). Although in vivo experiments with these compounds have shown to be thyroid hormone disrupting (THD), in vitro their THD potency is not observed in nonpolar sediment extracts. This is hypothesized to be due to the absence of in vitro biotransformation which will result in bioactivation of the lipophilic compounds into THD hydroxyl metabolites. This study reveals that indeed metabolically activated nonpolar contaminants in sediments can competitively bind to thyroid hormone transport proteins. Sediment fractions were incubated with S9 rat microsomes, and the metabolites were extracted with a newly developed method that excludes most of the lipids to avoid interference in the applied nonradioactive 96-well plate TTR competitive binding assay. Metabolic activation increased the TTR binding potency of nonpolar fractions of POP-polluted sediments up to 100 times, resulting in potencies up to 240 nmol T4 equivalents/g sediment equivalent (nmol T4-Eq/g SEQ). This demonstrates that a more realistic in vitro sediment THD risk characterization should also include testing of both polar and medium polar sediment extracts for THD, as well as bioactivated nonpolar sediment fractions to prevent underestimation of its toxic potency.

In vivo effect confirmation of anti-androgenic compounds in sediment contact tests with Potamopyrgus antipodarum.

In order to investigate the sensitivity of Potamopyrgus antipodarum to anti-androgenic compounds, three spiked sediment tests were performed. The substances benzanthrone (7H-benz[de]anthracen-7-one), traseolide (ATII) and androstenone (5α-Androst-16-en-3-one) were previously identified in an effect-directed analysis study of the river Schijn in the north of Belgium. Although, in previous studies, all of the three compounds exhibited anti-androgenic activities in vitro, only the oxy-PAH benzanthrone had significant stimulating effects on the snails' reproduction. The reproduction of P. antipodarum was significantly stimulated, following a sigmoidal dose response curve, whereby an EC(50) of 10 ng/g dry sediment was calculated. Mortality was significantly increased at the highest concentration (69 ng/g dry sediment). The results indicate different relative potencies for the in vivo test with P. antipodarum and the in vitro anti-AR-CALUX assay, performed in a previous study. This highlights the importance of combined in vitro and in vivo assays for the effect assessment of field sediments.

Estimated daily intake of per- and polyfluoroalkyl substances related to different particle size fractions of house dust.

Indoor environmental pollutants are a threat to human health. In the current study, we analysed 25 per- and polyfluoroalkyl substances (PFASs) in seven different size fraction of house dust including the two relevant for exposure via ingestion and inhalation. The highest PFAS concentration is found in the inhalable particulate fraction which is explained by the increased surface area as the particulate's sizes decrease. The estimated daily intake (EDI) of the individual PFAS and exposure pathways were calculated for children and adults. In addition, the total EDI for PFOA and its precursors was estimated. The polyfluoroalkyl phosphoric acid diesters (diPAP), followed by PFOA and PFHxA fluortelomer, showed the highest concentrations of PFAS analysed. The cumulative EDI of PFAS for children was 3.0 ng/kg bw per day, a worst-case scenario, which is 17 times higher than the calculated EDI for adults. For children, ingestion of dust was found to result in 800 times higher PFOA exposure than via inhalation. The contribution from PFOA precursors corresponded to only 1% of the EDI from dust indicating PFOA as the main source of exposure. The EDI's of PFOA and PFOS from dust were lower than the calculated EDI's from food ingestion reported by the Swedish Food Agency. Our data indicate that the EDI for the sum of four PFASs: PFOA, PFNA, PFHxS and PFOS from dust intake alone is close to the established tolerable weakly intake of 4.4 ng/kg bw in children, set by European Food Safety Authority (EFSA) in 2020. The combined EDI levels PFOA and PFOS from both dust and food exceeded the EFSA TWI for both children and adults. This study demonstrates that dust is a relevant exposure pathway for PFAS intake and that analysis of relevant particle size fractions is important for evaluation of dust as an exposure pathway.

Polar compounds dominate in vitro effects of sediment extracts.

Sediment extracts from three polluted sites of the river Elbe basin were fractionated using a novel online fractionation procedure. Resulting fractions were screened for mutagenic, aryl hydrocarbon receptor (AhR)-mediated, transthyretin (TTR)-binding, and estrogenic activities and their potency to inhibit gap junctional intercellular communication (GJIC) to compare toxicity patterns and identify priority fractions. Additionally, more than 200 compounds and compound classes were identified using GC-MS/MS, LC-MS/MS, and HPLC-DAD methods. For all investigated end points, major activities were found in polar fractions, which are defined here as fractions containing dominantly compounds with at least one polar functional group. Nonpolar PAH fractions contributed to mutagenic and AhR-mediated activities while inhibition of GJIC and estrogenic and TTR-binding activities were exclusively observed in the polar fractions. Known mutagens in polar fractions included nitro- and dinitro-PAHs, azaarenes, and keto-PAHs, while parent and monomethylated PAHs such as benzo[a]pyrene and benzofluoranthenes were identified in nonpolar fractions. Additionally, for one sample, high AhR-mediated activities were determined in one fraction characterized by PCDD/Fs, PCBs, and PCNs. Estrone, 17ß-estradiol, 9H-benz[de]anthracen-7-one, and 4-nonylphenol were identified as possible estrogenic and TTR-binding compounds. Thus, not only nonpolar compounds such as PAHs, PCBs, and PCDD/Fs but also the less characterized and investigated more polar substances should be considered as potent mutagenic, estrogenic, AhR-inducing, TTR-binding, and GJIC-inhibiting components for future studies.

Identification strategy for unknown pollutants using high-resolution mass spectrometry: androgen-disrupting compounds identified through effect-directed analysis.

Effect-directed analysis has been applied to a river sediment sample of concern to identify the compounds responsible for the observed effects in an in vitro (anti-)androgenicity assay. For identification after non-target analysis performed on a high-resolution LTQ-Orbitrap, we developed a de novo identification strategy including physico-chemical parameters derived from the effect-directed analysis approach. With this identification strategy, we were able to handle the immense amount of data produced by non-target accurate mass analysis. The effect-directed analysis approach, together with the identification strategy, led to the successful identification of eight androgen-disrupting compounds belonging to very diverse compound classes: an oxygenated polyaromatic hydrocarbon, organophosphates, musks, and steroids. This is one of the first studies in the field of environmental analysis dealing with the difficult task of handling the large amount of data produced from non-target analysis. The combination of bioassay activity assessment, accurate mass measurement, and the identification and confirmation strategy is a promising approach for future identification of environmental key toxicants that are not included as priority pollutants in monitoring programs.

Per- and polyfluoroalkyl substances (PFASs) in Swedish household dust and exposure of pet cats.

Per- and polyfluoroalkyl substances (PFASs) are used in a wide range of products and have been found ubiquitously in our indoor environment, and there is evidence that exposure to PFAS can lead to adverse endocrine effects, such as thyroid hormone disruption. Pet cats have a high dust intake due to their grooming behavior and have been shown to be a suitable sentinel species for assessment of toddler's exposure. Here we used paired household dust (n=46) and cat serum (n=27) samples to establish whether dust is a relevant exposure pathway to PFASs. An analytical method for PFAS analysis was optimized using a low volume of cat serum samples, combining solid-phase extraction and online sample cleanup. Dust was extracted with methanol by sonication and cleaned up by addition of active carbon. In total, 27 PFASs were analyzed by liquid chromatography/mass spectrometry analysis. The correlation between PFAS levels in dust and serum, serum lipids and thyroid hormone levels, and PFAS levels in dust between different rooms were statistically evaluated. PFOS and PFDA could be quantified in all cat serum samples (median 2300 pg/mL and 430 pg/mL, respectively), followed by PFOA (median 1100 pg/mL), quantified in 96% of the samples. The levels of 6:2 and 8:2 diPAPs were determined in 65% and 92% of the serum samples, respectively, and were an order of magnitude lower (1.4-160 pg/mL). Household dust on the other hand was dominated by 6:2 and 8:2 diPAPs, with a median of 65 ng/g dust and 49 ng/g dust, respectively. PFOS (median 13 ng/g dust) and PFOA (median 9 ng/g dust) were quantified in 93% of the dust samples. Only eight PFASs were detected (>LOD) in at least 50% of the samples of both matrices and could be paired. Significant correlations between cat serum and dust were found for PFOA (rS=0.32, p<0.049) and PFUnDA (rS=0.55, p<0.001). Significant positive correlations were found between serum total thyroxine (rS=0.11, p<0.05) and PFNA and between serum cholesterol and PFHpA (rS=0.46, p<0.01), PFUnDA (rS=0.40, p<0.05), PFDoDA (rS=0.44, p<0.01), and sum PFAS (rS=0.48, p<0.01). In conclusion, this study confirmed that dust is a relevant exposure pathway for the ingestion of some PFASs for cats, and the serum levels of PFASs could be of relevance for the cat's health.

Testing endocrine disruption in biota samples: a method to remove interfering lipids and natural hormones.

A cleanup method was developed to remove coextracted lipids and natural hormones from biota samples in order to test the endocrine-disrupting (ED) capacity of their extracts in in vitro bioassays. Unspiked and spiked fish tissues were cleaned with a combination of dialysis, gel permeation chromatography (GPC), and normal-phase liquid chromatography (NP-HPLC). The spiking mixture consisted of a broad range of environmental pollutants (endocrine disruptors and genotoxic compounds). Chemical recoveries of each test compound, and thyroid-hormone-like and (anti)androgenic activities of the cleaned extracts were investigated. Despite the chemical and toxicological complexity of the spiking mixture and the sequential sample treatment, chemical analysis revealed acceptable recoveries on average: 89 ± 8% after each cleanup step separately and 75 ± 3% after the whole extraction and cleanup procedure in the extracts. In addition, recovered activities in the bioassays were in good agreement with the spiking levels. The developed cleanup method proved to be capable of lipid and natural hormone removal from fish extracts, enabling the measurement of selected endocrine-hormone-like activities in T(4)*-TTR and AR-CALUX bioassays. The method can be used as a sample preparation method of biota samples for toxicity profiling and effect-directed analysis (EDA).

Transthyretin-Binding Activity of Complex Mixtures Representing the Composition of Thyroid-Hormone Disrupting Contaminants in House Dust and Human Serum.

BACKGROUND: House dust contains many organic contaminants that can compete with the thyroid hormone (TH) thyroxine (T4) for binding to transthyretin (TTR). How these contaminants work together at levels found in humans and how displacement from TTR in vitro relates to in vivo T4-TTR binding is unknown. OBJECTIVES: Our aims were to determine the TTR-binding potency for contaminant mixtures as found in house dust, maternal serum, and infant serum; to study whether the TTR-binding potency of the mixtures follows the principle of concentration addition; and to extrapolate the in vitro TTR-binding potency to in vivo inhibition levels of T4-TTR binding in maternal and infant serum. METHODS: Twenty-five contaminants were tested for their in vitro capacity to compete for TTR-binding with a fluorescent FITC-T4 probe. Three mixtures were reconstituted proportionally to median concentrations for these chemicals in house dust, maternal serum, or infant serum from Nordic countries. Measured concentration-response curves were compared with concentration-response curves predicted by concentration addition. For each reconstituted serum mixture, its inhibitor-TTR dissociation constant (Ki) was used to estimate inhibition levels of T4-TTR binding in human blood. RESULTS: The TTR-binding potency of the mixtures was well predicted by concentration addition. The ∼20% inhibition in FITC-T4 binding observed for the mixtures reflecting median concentrations in maternal and infant serum was extrapolated to 1.3% inhibition of T4-TTR binding in maternal and 1.5% in infant blood. For nontested mixtures reflecting high-end serum concentrations, these estimates were 6.2% and 4.9%, respectively. DISCUSSION: The relatively low estimated inhibition levels at median exposure levels may explain why no relationship between exposure to TTR-binding compounds and circulating T4 levels in humans has been reported, so far. We hypothesize, however, that 1.3% inhibition of T4-TTR binding may ultimately be decisive for reaching a status of maternal hypothyroidism or hypothyroxinemia associated with impaired neurodevelopment in children. https://doi.org/10.1289/EHP5911.

Short-, medium-, and long-chain chlorinated paraffins in South African indoor dust and cat hair.

Polychlorinated n-alkanes or chlorinated paraffins (CPs) contain a magnitude of structural isomers and are categorized as short-chain (SCCPs), medium-chain (MCCPs), and long-chain (LCCPs) CPs, according to the carbon chain lengths. In this study the Æ©SCCPs, Æ©MCCPs, and Æ©LCCP concentrations are reported for South African indoor dust and pet cat hair. The median concentrations of the Æ©CPs (C9-C37) ranged from 33 to 663 µg/g for freshly collected dust (FD), 36-488 µg/g for dust collected from household vacuum cleaner bags (VD), and 1.2-15 µg/g for cat hair (CH) samples. MCCPs were the dominant CP group, followed by SCCPs and LCCPs. The Æ©MCCP concentration ranged from 13 to 498 µg/g in dust and 0.6-6.5 µg/g in cat hair. SCCPs with shorter carbon chains and lower chlorine substitution were observed in cat hair. LCCPs with carbon chains > C20 were detected in dust and hair samples, possibly indicating the use of wax grade LCCP formulations. Non-traditional Kendrick mass defect plots were used to obtain information on the magnitude of CPs and provide evidence of possible interfering compounds. This is the first report on the occurrence of SCCPs, MCCPs, and LCCPs in the South African indoor environment.