Sea Anemones Responding to Sex Hormones, Oxybenzone, and Benzyl Butyl Phthalate: Transcriptional Profiling and in Silico Modelling Provide Clues to Decipher Endocrine Disruption in Cnidarians

Endocrine disruption is suspected in cnidarians, but questions remain how occurs. Steroid sex hormones are detected in corals and sea anemones even though these animals do not have estrogen receptors and their repertoire of steroidogenic enzymes appears to be incomplete. Pathways associated with sex hormone biosynthesis and sterol signaling are an understudied area in cnidarian biology. The objective of this study was to identify a suite of genes that can be linked to exposure of endocrine disruptors. Exaiptasia diaphana were exposed to nominal 20ppb concentrations of estradiol (E2), testosterone (T), cholesterol, oxybenzone (BP-3), or benzyl butyl phthalate (BBP) for 4 h. Eleven genes of interest (GOIs) were chosen from a previously generated EST library. The GOIs are 17β-hydroxysteroid dehydrogenases type 14 (17β HSD14) and type 12 (17β HSD12), Niemann-Pick C type 2 (NPC2), Equistatin (EI), Complement component C3 (C3), Cathepsin L (CTSL), Patched domain-containing protein 3 (PTCH3), Smoothened (SMO), Desert Hedgehog (DHH), Zinc finger protein GLI2 (GLI2), and Vitellogenin (VTG). These GOIs were selected because of functional associations with steroid hormone biosynthesis; cholesterol binding/transport; immunity; phagocytosis; or Hedgehog signaling. Quantitative Real-Time PCR quantified expression of GOIs. In silico modelling utilized protein structures from Protein Data Bank as well as creating protein structures with SWISS-MODEL. Results show transcription of steroidogenic enzymes, and cholesterol binding/transport proteins have similar transcription profiles for E2, T, and cholesterol treatments, but different profiles when BP-3 or BBP is present. C3 expression can differentiate between exposures to BP-3 versus BBP as well as exposure to cholesterol versus sex hormones. In silico modelling revealed all ligands (E2, T, cholesterol, BBP, and BP-3) have favorable binding affinities with 17β HSD14, 17β HSD12, NPC2, SMO, and PTCH proteins. VTG expression was down-regulated in the sterol treatments but up-regulated in BP-3 and BBP treatments. In summary, these eleven GOIs collectively generate unique transcriptional profiles capable of discriminating between the five chemical exposures used in this investigation. This suite of GOIs are candidate biomarkers for detecting transcriptional changes in steroidogenesis, gametogenesis, sterol transport, and Hedgehog signaling. Detection of disruptions in these pathways offers new insight into endocrine disruption in cnidarians.

Application of Fungus Enzymes in Spent Mushroom Composts from Edible Mushroom Cultivation for Phthalate Removal

Spent mushroom composts (SMCs) are waste products of mushroom cultivation. The handling of large amounts of SMCs has become an important environmental issue. Phthalates are plasticizers which are widely distributed in the environment and urban wastewater, and cannot be effectively removed by conventional wastewater treatment methods. In this study, SMCs are tested for their ability to remove phthalates, including benzyl butyl phthalate (BBP), di-n-butyl phthalate (DBP), and diethyl phthalate (DEP). Batch experiments reveal that BBP, DBP, and DEP can be degraded by the SMC enzyme extracts of four edible mushrooms: Pleurotus eryngii, Pleurotus djamor, Pleurotus ostreatus, and Auricularia polytricha. Potential fungus enzymes associated with BBP, DBP, and DEP degradation in SMCs (i.e., esterases, oxygenases, and oxidases/dehydrogenases) are uncovered by metaproteomic analysis using mass spectrometry. Bioreactor experiments indicate that the direct application of SMCs can remove BBP, DBP, and DEP from wastewater, through adsorption and biodegradation. The results of this study extend the application of white-rot fungi without laccases (e.g., Auricularia sp.) for the removal of organic pollutants which are not degraded by laccases. The application of SMCs for phthalate removal can be developed into a mycoremediation-based green and sustainable technology.

Potential risk of BPA and phthalates in commercial water bottles: a minireview

The global water bottling market grows annually. Today, to ensure consumer safety, it is important to verify the possible migration of compounds from bottles into the water contained in them. Potential health risks due to the prevalence of bisphenol A (BPA) and phthalates (PAEs) exposure through water bottle consumption have become an important issue. BPA, benzyl butyl phthalate (BBP), di-n-butyl phthalate (DBP) and di (2-ethylhexyl) phthalate (DEHP) can cause adverse effects on human health. Papers of literature published in English, with BPA, BBP, DBP and DEHP detections during 2017, by 2019 by liquid chromatography and gas chromatography analysis methods were searched. The highest concentrations of BPA, BBP, DBP and DEHP in all the bottled waters studied were found to be 5.7, 12.11, 82.8 and 64.0 μg/L, respectively. DBP was the most compound detected and the main contributor by bottled water consumption with 23.7% of the TDI. Based on the risk assessment, BPA, BBP, DBP and DEHP in commercial water bottles do not pose a serious concern for humans. The average estrogen equivalente level revealed that BPA, BBP, DBP and DEHP in bottled waters may induce adverse estrogenic effects on human health.

AN ASSESSMENTOF PHTHALATES IN COSMETIC PRODUCTS BY DISPERSIVE LIQUID-LIQUID EXTRACTION METHOD USING HPLCAND LC-MS/MS

A Dispersive liquid-liquid microextraction (DLLME) based analytical method was developed and validated using HPLC-PDA and LC-MS/MS for quantitative determination of phthalates (Dimethyl phthalate (DMP), Diethyl phthalate (DEP), Benzyl Butyl Phthalate (BzBP), Dibutylphthalate(DBP), Diethylhexyl phthalate(DEHP), Di-n-octyl phthalate (DOP) in different cosmetic products (After Shave Lotion, Deodorants, Perfume and Liquid Body Lotion). The DLLME based developed and validated analytical method was found specific, sensitive, accurate and precise. The accuracy (% recovery) of the method at a spiking level of 0.1, 0.5 and 1.0 mg L-1 in the different cosmetic product was found in the range of 92-108. The interday and intraday precision (%RSD) of the method was found less than 15. Out of six analyzed phthalates, only four phthalates were detected in different cosmetic products. Dimethyl phthalate (DMP) and Di-n-octyl phthalate (DOP) were detected in aftershave lotion. Diethyl phthalate (DEP) was detected in deodorants. Dibutyl phthalate (DBP) and Diethyl phthalate (DEP) were detected in perfumes. None of the phthalates were detected in liquid body lotion.

Comparative Analysis of Neurotoxicity of Six Phthalates in Zebrafish Embryos

The effects and underlying mechanisms of phthalates on neurotoxicity remain unclear as compared with the potentials of these substances as endocrine disruptors. The locomotor activities of zebrafish embryos were investigated upon exposure to six phthalates: dimethyl phthalate (DMP), diethyl phthalate (DEP), benzyl butyl phthalate (BBzP), di-2-ethylhexyl phthalate (DEHP), di-n-octyl phthalate (DnOP), and diisononyl phthalate (DiNP). Moreover, changes in fluorescence intensity in the green fluorescent protein (GFP) transgenic (Tg) lines Tg(HuC:eGFP), Tg(sox10:eGFP), and Tg(mbp:GFP) were measured after exposure to six phthalates, and changes in the expression profiles of genes involved in the cholinergic (ache) and dopaminergic systems (dat, th, and drd1b) were assessed. Exposure to BBzP, DEHP, and DiNP affected larval behaviors, whereas exposure to DMP, DEP, and DnOP revealed no alterations. A reduced expression of Tg(HuC:eGFP) was observed upon exposure to BBzP, DEHP, and DiNP. The expression of Tg(sox10:eGFP) and Tg(mbp:GFP) was reduced only in response to BBzP and DiNP, respectively. Further, exposure to DiNP upregulated ache and drd1b. The upregulation of ache and downregulation of drd1b was observed in DEHP-exposed groups. Exposure to BBzP suppressed th expression. These observations indicate that exposure to phthalates impaired embryogenesis of the neurological system and neurochemicals in zebrafish embryos, although the detailed mechanisms varied among the individual phthalates. Further mechanistic studies are needed to better understand the causality between phthalate exposure and neurotoxicity.