Potential Endocrine Disruptors Research Articles

Thresholds of adversity for endocrine disrupting substances: a conceptual case study.

For endocrine disrupting chemicals (EDC) the existence of "safe exposure levels", that is exposure levels that do not present an appreciable risk to human health is most controversially discussed, as is the existence of health-based reference values. Concerns have been especially raised that EDCs might not possess a threshold level such that no exposure level to EDCs can be considered safe. To explore whether or not threshold levels can be identified, we performed a screening exercise on 14 pesticidal and biocidal active substances previously identified as EDCs in the European Union. The respective substances are ideal subjects for case studies to review for endocrine activity and disruptive potential following well-defined regulatory assessment based on solid data to effectually establish adversity as consequence of endocrine disruption. Dimethomorph, metiram and propiconazole for which the weight of evidence demonstrating endocrine disruption was the strongest were used as subjects for further study. Epoxiconazole was additionally selected as its effects on the endocrine system are extensive. For all four substances, analysis of the toxicological data clearly indicated thresholds of adversity below which no adverse effects mediated through an endocrine mechanism were observed. Particular emphasis was placed on mechanistic considerations including homeostasis and the concept of adversity. As a proof of concept this study provides evidence that like other substances of toxicological concern EDCs have threshold levels for adversity. While for some EDCs the respective thresholds might indeed be very low this shows that, data allowing, for other EDCs sufficiently protective reference values can be derived.

Toxicity threshold and ecological risk of nitrate in rivers based on endocrine-disrupting effects: A case study in the Luan River basin, China

Nitrate, as a crucial nutrient, is consistently targeted for controlling water eutrophication globally. However, there is considerable evidence suggesting that nitrate has endocrine-disrupting potential on aquatic organisms. In this study, the sensitivity of various adverse effects to nitrate nitrogen (nitrate-N) was compared, and a toxicity threshold based on endocrine-disrupting effects was derived. The spatiotemporal variations of nitrate-N concentrations in the Luan River basin were investigated, and the associated aquatic ecological risks were evaluated using a comprehensive approach. The results showed that reproduction and development were the most sensitive endpoints to nitrate, and their distribution exhibited significant differences compared to behavior. The derived threshold based on endocrine-disrupting effects was 0.65 mgL−1, providing adequate protection for the aquatic ecosystem. In the Luan River basin, the mean nitrate-N concentrations during winter (4.4 mgL−1) were significantly higher than those observed in spring (0.7 mgL−1) and summer (1.2 mgL−1). Tributary inputs had an important influence on the spatial characteristics of nitrate-N in the mainstream, primarily due to agricultural and population-related contamination. The risk quotients (RQ) during winter, summer, and spring were evaluated as 6.7, 1.8, and 1.1, respectively, and the frequency of exposure concentrations exceeding the threshold was 100 %, 64.3 %, and 42.5 %, respectively. At the ecosystem level, nitrate posed intermediate risks to aquatic organisms during winter and summer in the Luan River basin and at the national scale in China. We suggest that nitrate pollution control should not solely focus on water eutrophication but also consider the endocrine disruptive effect on aquatic animals.

Estetrol/drospirenone versus 17α-ethinylestradiol/drospirenone: An extended one generation test to evaluate the endocrine disruption potential on zebrafish (Danio rerio)

Combined oral contraceptives, comprising of both an oestrogen and a progestin component, are released in aquatic environments and potentially pose a risk to aquatic wildlife by their capacity to disrupt physiological mechanisms.In this study, the endocrine disruptive potential of two mixtures, 17α-ethinylestradiol (EE2), a synthetic oestrogen, or estetrol (E4), a natural oestrogen, with the progestin drospirenone (DRSP) have been characterised in three generations of zebrafish, according to an adapted Medaka Extended One Generation Reproduction Test. Zebrafish (Danio rerio) were exposed to a range of concentrations of EE2/DRSP and E4/DRSP (∼1×, ∼3×, ∼10× and ∼30× predicted environmental concentration, PEC). Survival, growth, hatching success, fecundity, fertilisation success, vitellogenin (VTG), gonad histopathology, sex differentiation, and transcriptional analysis of genes related to gonadal sex steroid hormones synthesis were assessed.In the F0 generation, exposure to EE2/DRSP at ∼10 and ∼30× PEC decreased fecundity and increased male VTG concentrations. The highest concentration of EE2/DRSP also affected VTG concentrations in female zebrafish and the expression of genes implicated in steroid hormones synthesis. In the F1 generation, sex determination was impaired in fish exposed to EE2/DRSP at concentrations as low as ∼3× PEC. Decreased fecundity and fertility, and abnormal gonadal histopathology were also observed. No effects were observed in the F2 generation. In contrast, E4/DRSP induced only minor histopathological changes and an increase in the proportion of males, at the highest concentration tested (∼30× PEC) in the F1 generation and had no effect on hatching success of F2 generation.Overall, this study suggests that the combination E4/DRSP has a more favourable environmental profile than EE2/DRSP.

Cocktail effects of tire wear particles leachates on diverse biological models: A multilevel analysis

Tire wear particles (TWP) stand out as a major contributor to microplastic pollution, yet their environmental impact remains inadequately understood. This study delves into the cocktail effects of TWP leachates, employing molecular, cellular, and organismal assessments on diverse biological models. Extracted in artificial seawater and analyzed for metals and organic compounds, TWP leachates revealed the presence of polyaromatic hydrocarbons and 4-tert-octylphenol. Exposure to TWP leachates (1.5 to 1000 mg peq L−1) inhibited algae growth and induced zebrafish embryotoxicity, pigment alterations, and behavioral changes. Cell painting uncovered pro-apoptotic changes, while mechanism-specific gene-reporter assays highlighted endocrine-disrupting potential, particularly antiandrogenic effects. Although heavy metals like zinc have been suggested as major players in TWP leachate toxicity, this study emphasizes water-leachable organic compounds as the primary causative agents of observed acute toxicity. The findings underscore the need to reduce TWP pollution in aquatic systems and enhance regulations governing highly toxic tire additives.

Predicting the Activity of Unidentified Chemicals in Complementary Bioassays from the HRMS Data to Pinpoint Potential Endocrine Disruptors.

The majority of chemicals detected via nontarget liquid chromatography high-resolution mass spectrometry (HRMS) in environmental samples remain unidentified, challenging the capability of existing machine learning models to pinpoint potential endocrine disruptors (EDs). Here, we predict the activity of unidentified chemicals across 12 bioassays related to EDs within the Tox21 10K dataset. Single- and multi-output models, utilizing various machine learning algorithms and molecular fingerprint features as an input, were trained for this purpose. To evaluate the models under near real-world conditions, Monte Carlo sampling was implemented for the first time. This technique enables the use of probabilistic fingerprint features derived from the experimental HRMS data with SIRIUS+CSI:FingerID as an input for models trained on true binary fingerprint features. Depending on the bioassay, the lowest false-positive rate at 90% recall ranged from 0.251 (sr.mmp, mitochondrial membrane potential) to 0.824 (nr.ar, androgen receptor), which is consistent with the trends observed in the models' performances submitted for the Tox21 Data Challenge. These findings underscore the informativeness of fingerprint features that can be compiled from HRMS in predicting the endocrine-disrupting activity. Moreover, an in-depth SHapley Additive exPlanations analysis unveiled the models' ability to pinpoint structural patterns linked to the modes of action of active chemicals. Despite the superior performance of the single-output models compared to that of the multi-output models, the latter's potential cannot be disregarded for similar tasks in the field of in silico toxicology. This study presents a significant advancement in identifying potentially toxic chemicals within complex mixtures without unambiguous identification and effectively reducing the workload for postprocessing by up to 75% in nontarget HRMS.

Indications of Endocrine Disruptor Effects of JP-5 Jet Fuel Using a Rat-Model Reproductive Study and an In Vitro Human Hormone Receptor Assay.

Recent events concerning jet fuel contamination of drinking water have shown that we need a better understanding of the effects of ingested jet fuel. To this end, a reproductive study with ingested jet fuel in rats was undertaken with relatively high concentrations of Jet Propellant (JP)-5 along with a human estrogen receptor activation in vitro assay using JP-5, JP-8, and an alternative jet fuel derived from the camelina plant referred to as HydroRenewable Jet (HRJ) fuel, to help evaluate potential effects of ingested jet fuel. The results of the in vivo study provide evidence that JP-5 can act as an endocrine disruptor, with specific observations including altered hormone levels with JP-5 exposure (significantly lower estradiol levels in male rats and significantly increased Dehydroepiandrosterone levels in females), and a decreased male/female offspring ratio. The in vitro hormone receptor activation assay indicated that JP-5 and JP-8 are capable of upregulating human estrogen receptor (ER) activity, while HRJ was not active in the ER assay. The jet fuels were not able to activate androgen or glucocorticoid receptors in further in vitro assays. These results infer potential endocrine disruption associated with JP-5, with activation of the estrogen receptor as one potential mechanism of action.

Nondestructive Quantification of Isoflavones in Cotyledons by Near-Infrared Spectroscopy and Potential and Limits for Sustainable Soybean Breeding

The isoflavones in the cotyledon of soybean seeds mimic human estrogen in structure, conferring them complex effects on health. Their regulation represents a major challenge for the sustainable breeding of new varieties with lower levels of potential endocrine disruptors. To develop a rapid, nondestructive, and eco-friendly analysis method, this study explores how sample grinding affects the results of near-infrared spectroscopy (NIRS) and the preprocessing methods. The prediction of the daidzein and genistein content would help the specific reduction in isoflavones in the cotyledon without harming seed development. The potential of a nonlinear approach (artificial neural network) is also compared with the more conventional partial least squares (PLS) regression. The isoflavone content of cotyledons from 529 soybean samples (65 genotypes) was quantified by HPLC, and the NIR spectra of these samples were collected using a Brucker multi-purpose analyzer. The spectra of whole and ground cotyledons were also collected for 155 samples. The results show that grain fragmentation improves the model calibration, although spectral preprocessing can harmonize this effect. Although the best PLS regression in cross-validation did not suffice to quantify the daidzein and genistein percentages, the artificial neural network (ANN) approach allowed us to develop much more reliable models than PLS. The performance of ANNs in external validation is remarkable in terms of both precision and applicability (R2 = 0.89 and a ratio of prediction to deviation of 2.92), making ANNs suitable in the breeding context for screening soybean grains regarding their isoflavone content.

Advancing Endocrine Disruptors via In Vitro Evaluation: Recognizing the Significance of the Organization for Economic Co-Operation and Development and United States Environmental Protection Agency Guidelines, Embracing New Assessment Methods, and the Urgent Need for a Comprehensive Battery of Tests.

Efforts are being made globally to improve the evaluation and understanding of endocrine-disrupting chemicals. Recognition of their impact on human health and the environment has stimulated attention and research in this field. Various stakeholders, including scientists, regulatory agencies, policymakers, and industry representatives, are collaborating to develop robust methodologies and guidelines for assessing these disruptors. A key aspect of these efforts is the development of standardized testing protocols and guidelines that aim to provide consistent and reliable methods for identifying and characterizing endocrine disruptors. When evaluating the potential endocrine-disrupting activity of chemicals, no single test is capable of detecting all relevant endocrine-disrupting agents. The test battery approach is designed to reduce the risk of false negative results for compounds with toxic potential. A weight-of-evidence approach is therefore necessary for endocrine disruptor evaluation. This approach considers various types of data from multiple sources, assessing the overall strength, consistency, and reliability of the evidence. OECD guidelines are highly regarded for their scientific rigor, transparency, and consensus-based development process. It is crucial to explore and develop new methodologies that can effectively evaluate the risks associated with potential endocrine disruptors. Integrating these methods into a comprehensive weight-of-evidence framework will enhance risk assessments and facilitate informed decisions regarding the regulation and management of these substances, ensuring the protection of human health and the environment from their adverse effects.

Bioaccumulation and Potential Endocrine Disruption Risk of Legacy and Emerging Organophosphate Esters in Cetaceans from the Northern South China Sea.

Despite the increasing health risks shown by the continuous detection of organophosphate esters (OPEs) in biota in recent years, information on the occurrence and potential risks of OPEs in marine mammals remains limited. This study conducted the first investigation into the body burdens and potential risks of 10 traditional OPEs (tOPEs) and five emerging OPEs (eOPEs) in 10 cetacean species (n = 84) from the northern South China Sea (NSCS) during 2005-2021. All OPEs, except for 2-ethylhexyl diphenyl phosphate (EHDPHP), were detected in these cetaceans, indicating their widespread occurrence in the NSCS. Although the levels of the ∑10tOPEs in humpback dolphins remained stable from 2005 to 2021, the concentrations of the ∑5eOPEs showed a significant increase, suggesting a growing demand for these new-generation OPEs in South China. Dolphins in proximity to urban regions generally exhibited higher OPE concentrations than those from rural areas, mirroring the environmental trends of OPEs occurring in this area. All OPE congeners, except for EHDPHP, in humpback dolphins exhibited a maternal transfer ratio >1, indicating that the dolphin placenta may not be an efficient barrier for OPEs. The observed significant correlations between levels of OPEs and hormones (triiodothyronine, thyroxine, and testosterone) in humpback dolphins indicated that OPE exposures might have endocrine disruption effects on the dolphin population.

Chlorothalonil as a potential endocrine disruptor in male zebrafish (Danio rerio): Impact on the hypothalamus-pituitary-gonad axis and sperm quality

Chlorothalonil is a broad-spectrum organochlorine fungicide widely employed in agriculture to control fungal foliar diseases. This fungicide enters aquatic environments through the leaching process, leading to toxicity in non-target organisms. Organic contaminants can impact organism reproduction as they have the potential to interact with the neuroendocrine system. Although there are reports of toxic effects of chlorothalonil, information regarding its impact on reproduction is limited. The aim of the present study was to evaluate the influence of chlorothalonil on male reproductive physiology using the zebrafish (Danio rerio) as ecotoxicological model. Zebrafish were exposed for 7 days to two concentrations of chlorothalonil (0.1 and 10 μg/L) along with a control group (with DMSO – 0.001%). Gene expression of hypothalamus-pituitary-gonad axis components (gnrh2, gnrh3, lhr, fshr, star, hsd17b1, hsd17b3, and cyp19a1), as well as hepatic vitellogenin concentration were assessed. In sperm cells, reactive oxygen species (ROS) content, lipid peroxidation (LPO), mitochondrial functionality, and membrane integrity and fluidity were evaluated. Results indicate that exposure to the higher concentration of chlorothalonil led to a reduction in brain gnr2 expression. In gonads, mRNA levels of lhr, star, and hsd17b1 were decreased at both chlorothalonil concentrations tested. Similarly, hepatic vitellogenin concentration was reduced. Regarding sperm cells, a decreased ROS level was observed, without significant difference in LPO level. Additionally, a higher mitochondrial potential and lower membrane fluidity were observed in zebrafish exposed to chlorothalonil. These findings demonstrate that chlorothalonil acts as an endocrine disruptor, influencing reproductive control mechanisms, as evidenced by changes in expression of genes HPG axis, as well as hepatic vitellogenin concentration. Furthermore, our findings reveal that exposure to this contaminant may compromise the reproductive success of the species, as it affected sperm quality parameters.

Spermatogenic Inhibition Properties of a Phenolic Phytoestrogen Isolated from Momordica charantia (Bitter Guard) Seeds

Mechanisms and potencies of phytoestrogens are not completely clarified and they may be considered potential endocrine disruptors, and therefore caution should be exercised while taking them. Eating very high levels of some phytoestrogens may pose some health risks. Reproductive problems have been documented in laboratory animals, farm animals and wildlife that ate very high (up to 100% of their diet) amounts of phytoestrogen-rich plants. Sheep consuming large amounts of clover showed infertility and reproductive disorders. Cheetahs in captivity also had reduced fertility rates when consuming a feline diet composed of a soybean product, which was reversed when it was removed from the diet. Toxicities associated with herbal medicines that include phytoestrogens have also been reported in the literature.
 Phenolphthalein a phenolic phytoestrogen has been isolated from the crude ethanol extract of Momordica charantia Linn. seeds. After preparative HPLC whitish amorphous compound was obtained. Its structural elucidation using IR, NMR and Mass spectral data revealed that the molecule isolated from the ethanol extract of M. charantia seeds was surprisingly, phenolphthalein. In order to clarify testicular influence of ethanol extract, fractions and isolated phenolphthalein were treated for sixty days to adult male albino rats. All the treated groups showed statistically significant reduction in testis weight. On histological examinations of testis showed spermatogenic inhibition effect, as the number of spermatogonia, spermatocytes, spermatids and spermatozoa were significantly decreased.

Unveiling the connections: Chlorpyrifos and its association with breast cancer

Chlorpyrifos, a broad-spectrum insecticide categorized within the organophosphate family, is recognized for its potent inhibition of the enzyme Acetylcholinesterase (AChE), resulting in the manifestation of cholinergic syndrome in humans. Beyond its well-established toxicity in the central nervous system, recent studies have explored additional pathways through which this pesticide may adversely impact human health. Breast cancer, characterized by the uncontrolled proliferation of epithelial cells in the mammary gland, stands as the most diagnosed cancer in women and is a leading global cause of female cancer-related deaths. Chlorpyrifos, extensively employed worldwide for pest control in agriculture, domestic settings, and industries, has notably faced recent bans in the European Union, marking a significant regulatory shift. This bibliographical review aims to unravel the intricate mechanisms by which chlorpyrifos may contribute to the development of breast cancer. Collating findings from human studies, as well as in vitro and in vivo research spanning the past decade, the review sheds light on chlorpyrifos as a potent endocrine disruptor. It influences female sex hormones, exhibits estrogenic effects on breast cancer cells, and induces alterations in breast tissue. Additionally, chlorpyrifos acts as an agonist of Estrogen Receptor α(ERα) and Aryl hydrocarbon Receptor (AhR), contributing to cell proliferation, oxidative stress, and engaging epigenetic and angiogenic mechanisms. This comprehensive review underscores the compelling association between chlorpyrifos exposure and mammary cancer. It emphasizes the urgent need for further research on pesticide usage to mitigate potential adverse health consequences.

Applications of Optical Method As an Effective Platform for Progesterone Sensing Using Carbon Dots and Graphene Oxide

Today, ‘Hormonal imbalance’ is a significant concern worldwide that lead to various types of complications in human body. For instance, abnormal levels of steroid hormones, such as progesterone are associated with irregular or absent menstrual periods, reproductive disorders, infertility, cardiovascular diseases, and cancer. Nowadays, progesterone is being excessively consumed as oral contraceptives, menopausal hormone therapy and treating infertility, resulting into the deleterious effects due to its endocrine disrupting activity. Furthermore, its presence as a potent endocrine disruptor in surface or environmental waters and various daily use products is another concern. Therefore, monitoring progesterone hormone is vital in environmental samples because of the adverse effects on reproductive health and causing hormonal imbalance. Herein, we have developed optical photoluminescence (PL) method implementing carbon dots (CDs) and graphene oxide (GO) based fluorescence resonance energy transfer (FRET) mechanism for progesterone hormone sensing. In this method, firstly progesterone-specific antibodies are covalently functionalized to CDs whose fluorescence has been quenched (turn off) by adding GO. However, the presence of progesterone increases the distance between donor and acceptor moieties and turns-on the fluorescence by displacing GO off the surface of CDs which in turn substantially reduces the FRET efficiency. Various optimization steps have been performed under control experiments to tune the fluorescence of CDs and the concentration of graphene oxide for maximum efficiency. In conclusion, a linear relationship is achieved in between PL intensity and different progesterone concentrations (10-900 nM) with 13.8 nM detection limits (R2=0.974). The structural and morphological analysis (XPS, and FT-IR) revealed the structural and surface functionalities of CDs. Finally, the potential of the developed biosensor has been explored in water matrices.

Computational Risk Assessment of Persistence, Bioaccumulation, and Toxicity of Novel Flame-Retardant Chemicals.

Novel brominated flame retardants (NBFRs) have emerged as chemicals of environmental concern, as they have been widely used as an alternative to polybrominated diphenyl ethers (PBDEs). Considering the similar structural features of NBFRs and PBDEs necessitates a comprehensive investigation to understand the physicochemical relationships of these compounds and their ability to alter biological functions. In this study, we investigated the persistent nature of NBFRs in terms of thyroid-disrupting potential by understanding the structure-stability aspects using density functional theory (DFT)-based reactivity parameters and interactions via molecular docking and molecular dynamics (MD) simulations. The results indicate that the DFT-based stability descriptor (chemical hardness) is associated with the persistent nature of NBFRs. The computed molecular interaction profile revealed prominent interactions between thyroid receptor-β (TR-β) and NBFRs. Stable trajectory and interactions with TR-β were obtained with ATE, p-TBX, PBT, PBEB, and TBBPA-DBPE during 100 ns of MD simulation. The results of these studies have suggested that the presence of a higher number of halogenated atoms increases the stability vis-à-vis the persistence and endocrine disruption potential of NBFRs.

BDE-47-mediated cytotoxicity via autophagy blockade in 3D HepaRG spheroids cultured in alginate microcapsules

Polybrominated Diphenyl Ethers (PBDEs) are a major class of brominated flame retardants, and their widespread use has led them to be considered contaminants with emerging concern. PBDEs have been detected in the indoor air, house dust, food, and all environmental compartments. The congener BDE-47 (2,2′,4,4′-tetrabromodiphenyl ether) is the most prevalent, and hepatotoxicity, neurotoxicity, immunological changes, endocrine disruption, and genotoxic potential have been related to its exposure. Although the BDE-47 molecular toxicity pathway is directly related to intrinsic apoptotic cell death, the role of autophagy in BDE-47 toxicity remains unclear. In this context, three-dimensional cell culture has emerged as a good strategy for the replacement of animals in toxicological testing. Here, we used HepaRG spheroids cultured in alginate microcapsules to investigate the role of autophagy in BDE-47-mediated hepatotoxicity. We developed mature and functional HepaRG spheroids by culturing them in alginate microcapsules. Histological analysis revealed that HepaRG spheroids formed an extracellular matrix and stored glycogen. No apoptotic and/or necrotic cores were observed. BDE-47 showed concentration- and time-dependent cytotoxicity in HepaRG spheroids. In the early exposure period, BDE-47 initially disrupted mitochondrial activity and increased the formation of acid compartments that promoted the increase in autophagic activity; however, this autophagy was blocked, and long-term exposure to BDE-47 promoted efficient apoptotic cell death through autophagy blockade, as evidenced by an increased number of fragmented/condensed nuclei. Therefore, for the first time, we demonstrated BDE-47 toxicity and its cell pathway induces cell death using a three-dimensional liver cell culture, the HepaRG cell line.

The ROS/SIRT1/STAR axis as a target for melatonin ameliorating atrazine-induced mitochondrial dysfunction and steroid disorders in granulosa cells

The granulosa cells (GCs) of birds are essential for the reproduction and maintenance of populations in nature. Atrazine (ATR) is a potent endocrine disruptor that can interfere with reproductive function in females and Diaminochlorotriazine (DACT) is the primary metabolite of ATR in the organism. Melatonin (MT) is an endogenous hormone with antioxidant properties that plays a crucial role in development of animal germ cells. However, how ATR causes mitochondrial dysfunction, abnormal secretion of steroid hormones, and whether MT prevents ATR-induced female reproductive toxicity remains unclear. Thus, the purpose of this study is to investigate the protective effect of MT against ATR-induced female reproduction. In the present study, the GCs of quail were divided into 6 groups, as follows: C (Serum-free medium), MT (10 µM MT), A250 (250 µM ATR), MA250 (10 µM MT+250 µM ATR), D200 (200 µM DACT) and MD200 (10 µM MT+200 µM DACT), and were cultured for 24 h. The results revealed that ATR prevented GCs proliferation and decreased cell differentiation. ATR caused oxidative damage and mitochondrial dysfunction, leading to disruption of steroid synthesis, which posed a severe risk to GC's function. However, MT supplements reversed these changes. Mechanistically, our study exhibited that the ROS/SIRT1/STAR axis as a target for MT to ameliorate ATR-induced mitochondrial dysfunction and steroid disorders in GCs, which provides new insights into the role of MT in ATR-induced reproductive capacity and species conservation in birds.

Ethylparaben Toxicity on Cladocerans Daphnia Similis and Ceriodaphnia Silvestrii and Species Sensitivity Analysis.

Parabens, a group of preservatives with a wide industrial range, threaten human and aquatic biota health due to their toxicity and endocrine disruption potential. As conventional wastewater treatment may not be enough to keep natural environments safe, toxicity studies are useful tools for supporting ecological risk assessments. Here, we focused on assessing ethylparaben's, one of the most common kinds of paraben, toxicity in the cladocerans Daphnia similis and Ceriodaphnia silvestrii. The EC50 sensitivity for D. similis and C. silvestrii was 24 (21-28) mg L- 1 and 25 (19-33) mg L- 1, respectively. Inhibition of reproduction and late development of females were observed in C. silvestrii exposed to 8mg L- 1. Furthermore, species sensitivity distribution was used to assess ecological risk, and ethylparaben demonstrated low potential risk for aquatic biota.

Toxicological investigation of lilial

Lilial (also called lysmeral) is a fragrance ingredient presented in many everyday cosmetics and household products. The concentrations of lilial in the final products is rather low. Its maximum concentration in cosmetics was limited and recently, its use in cosmetics products was prohibited in the EU due to the classification as reproductive toxicant. Additionally, according to the European Chemicals Agency, it was under assessment as one of the potential endocrine disruptors, i.e. a substance that may alter the function of the endocrine system and, as a result, cause health problems. Its ability to act as an androgen receptor agonist and the estrogenic and androgenic activity of its metabolites, to the best of our knowledge, have not yet been tested. The aim of this work was to determine the intestinal absorption, cytotoxicity, nephrotoxicity, mutagenicity, activation of cellular stress-related signal pathways and, most importantly, to test the ability to disrupt the endocrine system of lilial and its Phase I metabolites. This was tested using set of in vitro assays including resazurin assay, the CHO/HPRT mutation assay, γH2AX biomarker-based genotoxicity assay, qPCR and in vitro reporter assays based on luminescence of luciferase for estrogen, androgen, NF-κB and NRF2 signalling pathway. It was determined that neither lilial nor its metabolites have a negative effect on cell viability in the concentration range from 1 nM to 100 µM. Using human cell lines HeLa9903 and MDA-kb2, it was verified that this substance did not have agonistic activity towards estrogen or androgen receptor, respectively. Lilial metabolites, generated by incubation with the rat liver S9 fraction, did not show the ability to bind to estrogen or androgen receptors. Neither lilial nor its metabolites showed a nephrotoxic effect on human renal tubular cells (RPTEC/TERT1 line) and at the same time they were unable to activate the NF-κB and NRF2 signalling pathway at a concentration of 50 µM (HEK 293/pGL4.32 or pGL4.37). Neither lilial nor its metabolites showed mutagenic activity in the HPRT gene mutation test in CHO-K1 cells, nor were they able to cause double-strand breaks in DNA (γH2AX biomarker) in CHO-K1 and HeLa cells. In our study, no negative effects of lilial or its in vitro metabolites were observed up to 100 µM using different in vitro tests.

Examination of the endocrine-disrupting properties of “active chlorine generated from seawater by electrolysis” in response to the European Biocidal Products Regulation: current knowledge and methodological challenges

BackgroundCurrently, active chlorine is considered the most effective treatment for preventing biofouling of structures in contact with seawater. This compound falls under the scope of the EU Biocidal Products Regulation, which includes since 2018 a requirement to assess all active ingredients for their potential endocrine-disrupting properties on humans and non-target organisms. Therefore, this study examines the endocrine-disrupting (ED) potential of active chlorine based on the European Chemicals Agency and European Food Safety Authority guidance (ED TGD). It includes two approaches: (i) a systematic literature review using appropriate search terms and (ii) an in silico assessment, both supported by expert judgement. Finally, the feasibility and relevance of in vitro tests were examined by considering the stability of chlorine and the applicability domain of the recommended in vitro assays.ResultsNo significant adversity or endocrine activity based on EATS (estrogen, androgen, thyroid, and steroidogenesis)-modalities were evidenced based on the literature data. However, these modalities remain understudied and further datasets are needed for a comprehensive assessment. The in silico approach revealed a low probability of binding between active chlorine and a set of 14 human nuclear receptors, for both agonist and antagonist effects. This is not surprising given the great structural difference between active chlorine and natural ligands. The in vitro investigation of the ED potential of active chlorine raises several operational limits, including: (i) its instability (t1/2 < 48 h) which is incompatible with a reasonable time window between collection and ex situ analysis; (ii) its rapid and complete reaction with several essential nutrients in cell culture media; (iii) its documented cytotoxicity on various cell lines; and (iv) its exclusion from the scope of certain OECD guidelines.ConclusionsOverall, neither the in silico evaluation nor the systematic literature review performed indicates a significant adversity based on EATS-mediated parameters or EATS-related endocrine activities. This study highlights the challenges of performing a comprehensive ED assessment for a data-poor chemical and questions the relevance of transposing generic methodologies to the case of unstable and inorganic molecules.

Identification of Transformation Products of Organic UV Filters by Photooxidation and Their Differential Estrogenicity Assessment.

Organic ultraviolet filters (OUVFs) are extensively released into aquatic environments, where they undergo complex phototransformation. However, there is little knowledge regarding their transformation products (TPs) and associated endocrine disruption potentials. In the present study, we characterized the chemical and toxicological profiles of TPs for two common OUVFs, oxybenzone (BP3) and ethylhexyl methoxycinnamate (EHMC), by photooxidation under environmentally relevant conditions. It is hypothesized that TPs of the tested OUVFs will show varied estrogenicity at different reaction times. High-resolution liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) identified 17 TPs of 7 m/z for BP-3 and 13 TPs of 8 m/z for EHMC at confidence levels ≤2. Five novel TPs of 2 m/z were reported for the first time with structure-diagnostic MS/MS spectra. Estrogenicity assessment using the MCF-7-luc cell line showed discrepant estrogenic activities exhibited by OUVF-TPs over time. Specifically, BP3-TPs exhibited significantly greater estrogenicity than the parent at several reaction times, whereas EHMC-TPs displayed fluctuating estrogenicity with a declining trend. Correlation analysis coupled with molecular docking simulations further suggested several TPs of BP3 as potential endocrine disruptive compounds. These findings underscore the necessity of considering mixtures during chemical testing and risk assessment and highlight the potentially greater risks associated with post-transformation cocktails.