Enzyme-linked immunosorbent assay for bisphenol A: Assay optimization and its application for surface water analysis

A range of ELISAs in indirect and direct formats for the determination of bisphenol A (BPA) was developed. Bisphenol A carboxymethyl ether (BPA-CME), (BPA-CPE) and 4,4-bis(4-hydroxyphenyl)valeric acid were coupled with bovine serum albumin (BSA) (immunogens for the production of polyclonal antibodies), ovalbumin (OVA) and horseradish peroxidase. In general, the indirect assay was more sensitive and specific than the direct one. Using heterologous combinations of immunoreagents in the indirect assay allowed to increase assay sensitivity and specificity. The highest sensitivity was obtained for the antibodies produced against a conjugate of BPA-CPE with BSA and a conjugate of 4,4-bis(4-hydroxyphenyl)valeric acid with OVA. The detection limit of BPA in phosphate buffer was 0.03?ng?mL^?1. The assay developed was also the most specific towards BPA. Maximum cross-reactivity values did not exceed 11% for 4-cumylphenol, 5% for bisphenol E and 2% for bisphenol S. Finally, the developed assay was used to analyze surface water samples spiked with known amount of BPA. The assay showed good recovery values (85–109%) for surface water with mineralization level lower than 500?mg?L^?1.     

Endocrine disruptors in the Leça River and nearby Porto Coast (NW Portugal): presence of estrogenic compounds and hypoxic conditions

The Leça River and two beaches close to the north and south margins of the estuary, nowadays an important seaport harbor, show signs of contamination. However, the chemical nature of that contamination is uncertain. Therefore, this study checked for the presence of 11 endocrine disrupting compounds (EDCs) of animal (17β-estradiol and estrone), pharmaceutical (17α-ethynylestradiol), and industrial (alkylphenols, alkylphenol ethoxylates, and bisphenol A) origins in water samples collected from six points at the river gradient and at two sites in the coastline (Atlantic Ocean). Along with the EDCs, evaluated by gas chromatography – mass spectroscopy (GC-MS) during spring, summer, and autumn, physico-chemical parameters were also assessed and particular attention was given to dissolved oxygen (DO) levels. Data showed the presence of high amounts of estrogens (up to 10?ng?L^?1 for estrone) and industrial compounds (up to 2?µg?L^?1 for the nonylphenol ethoxylates) at both river and seacoast. Along with this, clear signs of hypoxia were found in the river (DO??1 at several sampling sites). Taking into account these issues, it was concluded that there are local conditions for the occurrence of endocrine disruption in aquatic animals.     

环境中有机紫外防晒剂残留及其生态毒性研究进展

有机紫外防晒剂被广泛添加在个人护理品中,随着日常使用源源不断地进入环境中,成为一类新兴污染物。环境介质中有机紫外防晒剂的检测及其生态毒理是目前的研究热点。概述了个人护理品中常用的有机紫外防晒剂,总结了有机紫外防晒剂在各类地表水、沉积物和生物体等不同介质中的浓度水平,详细分析了有机紫外防晒剂对鱼类的内分泌干扰效应和对水生无脊椎动物的急性毒性,为今后深入研究和科学评价这类新兴污染物提供重要参考。     

内分泌干扰物的非单一剂量-效应研究进展

传统毒理学理论认为,有毒物质产生毒害作用的剂量-效应关系是线性单调的,线性剂量-效应关系也是目前管理毒理学中计算化学品安全剂量的理论基础。但越来越多的研究显示,内分泌干扰物的剂量-效应关系通常不是线性的,而是U型、倒U型或更复杂的非单调剂量-效应曲线形式。以典型案例的形式介绍了近年来生理激素和内分泌干扰物非单调剂量-效应的研究进展,总结了目前认为形成非单调剂量-效应关系的机制。在此基础上,讨论了内分泌干扰物的非单一剂量-效应给目前管理毒理学中的化学品风险评价带来的挑战,并提出该领域未来的研究方向。     

青春期双酚A结构类似物暴露对小鼠行为的影响

作为重要的环境内分泌干扰物,双酚A的神经发育毒性已经引起广泛关注,其结构类似物对健康影响相关研究少有报道。为探讨双酚A结构类似物长期暴露对个体抑郁行为的影响,选取60只小鼠随机分为5组,每组12只。小鼠分别灌胃给予0、5、50 mg·kg~(-1)的双酚F或双酚S,连续给药4周。给药结束后,利用强迫游泳测试和悬尾测试评价小鼠抑郁样行为,同时监测小鼠体质量增加和自发活动情况。结果表明,与溶媒对照组比较,持续4周的双酚F和双酚S给药组小鼠的体质量变化和自发活动均无显著差异;5mg·kg~(-1)的双酚F或双酚S处理均不显著改变小鼠的抑郁行为,50 mg·kg~(-1)双酚F显著增加小鼠悬尾和强迫游泳测试中的不动时间,50mg·kg~(-1)双酚S处理显著增加小鼠悬尾测试中的不动时间。研究结果提示高浓度双酚F和双酚S在青春期的暴露可导致成年后小鼠抑郁样行为异常,应当引起重视。     

In vitro assessment of corticosteroid effects of eight chiral herbicides

Abstract

Information regarding the enantioselective endocrine disruption of chiral herbicides is scarce. This study assessed the disrupting effects of eight typical chiral herbicides on corticosteroids (including glucocorticoids and mineralocorticoids). Enantioselectivity of eight chiral herbicides were evaluated for their agonistic/antagonistic effects on glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) with CHOK1 cell line using reporter gene assay. Their influence on the production of corticosteroids were further investigated in H295R cell line using enzyme-linked immunosorbent assay (ELISA). None of the racemates or enantiomers of eight chiral herbicides exhibited GR or MR agonistic activity at non-cytotoxic concentrations. However, rac-propisochlor and S-imazamox antagonized cortisol-induced transactivation of GR by 21.79% and 38.73% at the concentration of 1.0?×?10^?7 M and 1.0?×?10^?6 M, respectively, and R-napropamide remarkably attenuated aldosterone-induced MR transactivation by 68.78% at 1.0?×?10^?6 M. The secretion of cortisol was significantly restrained after treated with 1.0?×?10^?6 M rac-propisochlor and rac-/R-napropamide at the concentration of 1.0?×?10^?6 M by 26.49%, 30.10% and 35.27%, respectively, while this glucocorticoid was remarkably induced by 1.0?×?10^?5 M rac-diclofop-methyl and its two enantiomers at the concentration of 1.0?×?10^?5 M by 75.60%, 100.1% and 68.78%, respectively. Exposure to rac-propisochlor (1.0?×?10^?6 M), S-diclofop-methyl (1.0?×?10^?5 M) or rac-/S-/R- acetochlor (1.0?×?10^?6 M) and rac-/S-/R-lactofen (1.0?×?10^?6 M) inhibited the secretion of aldosterone by approximately 40%. Our findings suggested that chiral herbicides disrupted corticosteroid homeostasis in an enantioselective way. Therefore, more comprehensive screening is required to better understand the ecological and health risks of chiral pesticides.     

水中的环境内分泌干扰物

环境内分泌干扰物能通过生物富集作用进入生物体内,对生物和人体的内分泌机能造成影响。介绍了环境内分泌干扰物的定义、作用机理及特性。重点阐明了水中环境内分泌干扰物的检测,分布及去除。提出纳滤去除水中环境内分泌干扰物必将成为今后的研究方向。     

Selected endocrine disrupting compounds (Vinclozolin,Flutamide, Ketoconazole and Dicofol): Effects on survival,occurrence of males,growth, molting and reproduction of <Emphasis Type="Italic">Daphnia magna</Emphasis>

BACKGROUND, AIM AND SCOPE: Pollution-induced endocrine disruption in vertebrates and invertebrates is a worldwide environmental problem, but relatively little is known about effects of endocrine disrupting compounds (EDCs) in planktonic crustaceans (including Daphnia magna). Aims of the present study were to investigate acute 48 h toxicity and sub-chronic (4-6 days) and chronic (21 days) effects of selected EDCs in D. magna. We have investigated both traditional endpoints as well as other parameters such as sex determination, maturation, molting or embryogenesis in order to evaluate the sensitivity and possible use of these endpoints in ecological risk assessment. MATERIALS AND METHODS: We have studied effects of four model EDCs (vinclozolin, flutamide, ketoconazole and dicofol) on D. magna using (i) an acute 48 h immobilization assay, (ii) a sub-chronic, 4-6 day assay evaluating development and the sex ratio of neonates, and (iii) a chronic, 21 day assay studying number of neonates, sex of neonates, molting frequency, day of maturation and the growth of maternal organisms. RESULTS: Acute EC50 values in the 48 h immobilization test were as follows (mg/L): dicofol 0.2, ketoconazole 1.5, flutamide 2.7, vinclozolin >3. Short-term, 4-6 day assays with sublethal concentrations showed that the sex ratio in Daphnia was modulated by vinclozolin (decreased number of neonate males at 1 mg/L) and dicofol (increase in males at 0.1 mg/L). Flutamide (up to 1 mg/L) had no effect on the sex of neonates, but inhibited embryonic development at certain stages during chronic assay, resulting in abortions. Ketoconazole had no significant effects on the studied processes up to 1 mg/L. DISCUSSION: Sex ratio modulations by some chemicals (vinclozolin and dicofol) corresponded to the known action of these compounds in vertebrates (i.e. anti-androgenicity and anti-oestrogenicity, respectively). Our study revealed that some chemicals known to affect steroid-regulated processes in vertebrates can also affect sublethal endpoints (e.g. embryonic sex determination and/or reproduction) in invertebrates such as D. magna. CONCLUSIONS: A series of model vertebrate endocrine disrupters affected various sub-chronic and chronic parameters in D. magna including several endpoints that have not been previously studied in detail (such as sex determination in neonates, embryogenesis, molting and maturation). Evaluations of traditional reproduction parameters (obtained from the 21 day chronic assay). as well as the results from a rapid, 4-6 day, sub-chronic assay provide complementary information on non-lethal effects of suspected organic endocrine disrupters. RECOMMENDATIONS AND PERSPECTIVES: It seems that there are analogies between vertebrates and invertebrates in toxicity mechanisms and in vivo effects of endocrine disruptors. However, general physiological status of organisms may also indirectly affect endpoints that are traditionally considered 'hormone regulated' (especially at higher effective concentrations as observed in this study) and these factors should be carefully considered. Further research of D. magna physiology and comparative studies with various EDCs will help to understand mechanisms of action as well as ecological risks of EDCs in the environment.     

Estrogenic Compounds Downstream From Three Small Cities in Eastern Nebraska: Occurrence and Biological Effect1

Abstract: Recent studies have detected estrogenic compounds in surface waters in North America and Europe. Furthermore, the presence of estrogenic compounds in surface waters has been attributed, in some cases, to the discharge of wastewater treatment plant (WWTP) effluent. The primary objective of the current study was to determine if WWTP effluent contributes estrogens to the surface waters of Nebraska. A second objective of this study was to determine if estrogens were found in concentrations sufficient enough to manifest feminizing effects on fish. These objectives were satisfied by deploying polar organic chemical integrative samplers (POCIS) and caged fathead minnows at eight field sites. Deployment sites included: three reference sites (Pawnee Creek, the Little Blue River, and the Middle Loup River), two sites upstream of the WWTPs at Grand Island and Columbus, and three sites downstream of the WWTPs at Grand Island, Columbus, and Hastings. Following the seven day deployments, POCIS extracts were analyzed for estrone, 17β‐estradiol, estriol and 17α‐ethinylestradiol using liquid chromatography tandem mass spectrometry (LC/MS/MS). 17β‐estradiol was detected in POCIS from six of the eight field sites with the greatest quantities recovered in POCIS deployed downstream from the Grand Island and Hastings WWTPs. Estrone was detected only in the POCIS deployed downstream from the Grand Island and Hastings WWTPs. Estrogenic effects were detected in caged minnows analyzed for the hepatic mRNA expression of two estrogen‐responsive genes, vitellogenin (vg1) and estrogen receptor α (ERα). Fish deployed at the site where the greatest quantities of estrogens were recovered (Hastings) had significantly higher expression of both vg1 and ERα than fish deployed at any of the other sites. These results confirm that WWTP effluent contributes biologically significant levels of estrogens to Nebraska surface waters.     

无锡-常州地下水中内分泌干扰物的赋存特征和健康风险评价

采用固相萃取-超高效液相色谱串联质谱法（SPE-UPLC-MS/MS）分析了无锡-常州地区地下水中4类（性激素、孕激素、糖皮质激素和酚类激素）22种典型内分泌干扰物（EDCs）的赋存特征.结果表明,共有20种EDCs被检出,以双酚A（BPA）、雌酮（E1）、雌二醇（E2）和炔雌醇（EE2）为主,检出率均在90%以上,其次是雌三醇（E3）和甲基泼尼松龙（Meprednl）,检出率分别为67.7%和48.4%.BPA平均浓度最高,为4.95 ng·L^-1,其次是甲基泼尼松龙、雌二醇、泼尼松龙（Prednl）、丙酸睾酮（TES-pro）和醋酸甲地孕酮（MA）,平均浓度分别为2.84、0.71、0.64、0.58和0.53 ng·L^-1.各点位总浓度范围在0.38~147.35 ng·L^-1之间,高浓度点主要集中在常州市新北区、溧阳市和钟楼区.结合主成分分析（PCA）和采样点周边污染源对潜在来源进行解析,结果表明地下水中EDCs主要来源于生活污水、养殖和工业废水,其次为医疗废水.非致癌健康风险评价结果表明,研究区地下水中EDCs非致癌风险整体处于较低水平,饮水途径引起的风险水平大于皮肤暴露,EE2的非致癌风险最高,应作为重点监控对象.