Determination of estrogenic activity in landfill leachate by simplified yeast two-hybrid assay

We previously developed a simplified yeast two-hybrid assay of estrogenic activity. In the present study, the optimal conditions for sensitivity and stability of this assay were decided. The assay could determine the estrogenic activity of 4 x 10(-11) mol dm(-1) 17beta-estradiol and also anti-estrogenic activity by using 4-hydroxytamoxifen as a control. The assay was used to test various chemicals suspected of estrogenicity. Many kinds of chemical, including alkylphenols, naphthols, xylenols, methoxychlor, phthalates, and bisphenol-A, showed estrogenic activity, and several, such as 2,5-xylenol and di-iso-octyl phthalate, showed anti-estrogenic but no estrogenic activity. Copresence of two estrogenic chemicals produced additive activity. The assay was also used to test leachate and water samples from the sea-based landfill of Osaka North Port in Japan. Estrogenic activity was detected in leachate from a municipal waste-dumping site, but not in that from a dredged soil-dumping site, which did however cause inhibition of cell growth.     

Bioluminescent yeast estrogen assay (BLYES) as a sensitive tool to monitor surface and drinking water for estrogenicity

Estrogenic Endocrine Disrupting Chemicals (EDCs) are a concern due to their ubiquity and recognized adverse effects to humans and wildlife. Methods to assess exposure to and associated risks of their presence in aquatic environment are still under development. The aim of this work is to assess estrogenicity of raw and treated waters with different degrees of pollution. Chemical analyses of selected EDCs were performed by liquid chromatography-tandem mass spectrometry, and estrogenic activity was evaluated using in vitro bioluminescent yeast estrogen assay (BLYES). Most raw water samples (18/20) presented at least one EDC and 16 rendered positive in BLYES. When EDCs were detected, the bioassay usually provided a positive response, except when only bisphenol A was detected at low concentrations. The highest values of estrogenic activity were detected in the most polluted sites. The maximum estrogenic activity observed was 8.7 ng equiv. of E2 L(-1). We compared potencies observed in the bioassay to the relative potency of target compounds and their concentrations failed to fully explain the biological response. This indicates that bioassay is more sensitive than the chemical approach either detecting estrogenic target compounds at lower concentrations, other non-target compounds or even synergistic effects, which should be considered on further investigations. We have not detected either estrogenic activity or estrogenic compounds in drinking water. BLYES showed good sensitivity with a detection limit of 0.1 ng equiv. E2 L(-1) and it seems to be a suitable tool for water monitoring.     

Spatial distribution and quantification of endocrine-disrupting chemicals in Sado River estuary, Portugal

The important Portuguese Sado River estuary has never been investigated for the presence of potentially endocrine-disrupting chemicals (EDCs), such as natural estrogens (estradiol, estrone), pharmaceutical estrogens (17α-ethynylestradiol), phytoestrogens (daidzein, genistein and biochanin A), or industrial chemicals (4-octylphenol, 4-nonylphenol, and bisphenol A). Thus, the main objective of this study was to evaluate their presence at 13 sampling points distributed between both the industrial and the natural reserve areas of the estuary, zones 1 and 2, respectively. For that, water samples collected in summer and winter were processed by solid phase extraction and analyzed by high-performance liquid chromatography with photodiode array detection and gas chromatography–mass spectroscopy. Results showed that estrone, ethynylestradiol, all the aforementioned phytoestrogens as well as bisphenol A and 4-octylphenol were found in zone 1. In zone 2, neither estrogens nor 4-OP were found. However, in the same zone, daidzein (500 ng/L) and genistein (320 ng/L) attained their highest levels in summer, whereas biochanin A peaked in winter (170 ng/L). Furthermore, bisphenol A was also found in some areas of zone 2, but showed similar concentrations in both surveys (about 220 ng/L). This study demonstrated that the Sado River estuary had low EDCs levels, suggesting that the Sado’s high hydrodynamic activity may be involved in the dilution of local pollution. It was suggested that at the current levels of concentrations, all assayed EDCs are unlikely to individually cause endocrine disruption in local animals. However, under a continuous exposure scenario, an additive and/or synergistic action of the estrogenic chemicals load can not be excluded, and so, continuous monitoring is advisable.     

Occurrence of estrogenic chemicals in South Korean surface waters and municipal wastewaters

Broad scale monitoring of estrogenic compounds was performed at 19 sampling points throughout the Yeongsan and Seomjin river basins and 5 wastewater treatment plants (WWTPs) adjacent to the Gwangju area, Korea, from December 2005 to August 2007. The concentrations of estrogenic compounds, including estrone (E1), 17β-estradiol (E2), 17α-ethynylestradiol (EE2), bisphenol-A, nonylphenol (NP) and 4-octylphenol (OP), in the samples was measured with gas chromatography/mass spectrometry (GC-MS). In addition, the estrogenic activities throughout the river were investigated using the E-screen assay. Of the six estrogenic chemicals, NP (114.6-336.1 ng L(-1)) and EE2 (0.23-1.90 ng L(-1)) were detected at the highest and lowest levels, respectively in both the river waters and the WWTP effluents. Bisphenol-A showed the largest concentration range, from 7.5 to 335 ng L(-1). The concentrations of E1, E2 and octylphenol ranges were 3.6-69.1, 1.2-10.7, and 2.2-16.9 ng L(-1), respectively. According to the calculated estradiol equivalent concentration (EEQ); however, no estrogenic contribution was observed due to the phenolic compounds in the river waters and effluents. E1 and E2 dominated in both the river water and effluent samples, with contributions to the calculated EEQ of over 79 and 77%, respectively. Conversely, EE2 was rarely detected in the river waters (21%) and effluents (0%). The largest contribution of EE2 to the calculated EEQ was 21% in the river water at S-7. The levels of E1, E2, and EE2 were remarkably decreased in the effluents, indicating that the 5 WWTPs did not contribute to the estrogenic effect of the receiving streams. Overall, the WWTPs did not contributed to the estrogenic activity of the receiving waters, but the livestock industry or wildlife may play an important role in the estrogenic contribution to river water.     

Improved application of recombinant yeast assays on environmental samples by size exclusion chromatography

The utility of HPLC with a size exclusion chromatography (SEC) column for the fractionation of environmental samples was examined to expand the applicability of recombinant yeast assays (RYAs) for the measurement of estrogenic activity in environmental samples. The elution of steroidal hormone standards through SEC was tested, and river water and sediment samples were subjected to SEC fractionation. The retention times of all estrogen standards were longer than those of the major components with absorbance at 230 nm of the environmental sample matrix. The estrogenic activity of SEC fractions of environmental samples from highly polluted sites was measured using RYA and revealed the existence of estrogenic compounds that could not be detected or quantified in extracted samples before fractionation. The fractions from environmental samples that corresponded to the retention time of estrone, 17[small beta]-estradiol, and estriol were analyzed with LC/MS/MS. These three estrogens were separated into three different fractions, and the concentration of estrone coincided with the estrogenic activity of the fraction in which it was detected. The profile of estrogenic activity of SEC fractions indicated the molecular size of the estrogenic compounds and the yeast growth inhibitors in samples.     

水中多种酚类化合物同步提取方法研究

对水中14种酚类化合物的液液萃取过程进行研究,选取不同萃取溶剂体系、水样pH和盐析条件等多个对萃取效率有决定性影响的参数进行优化。通过上述条件的优化选择,实现了同步对包括低沸点的苯酚、一氯代酚及较难提取的甲基酚和硝基酚等14种不同类型酚类化合物最优的萃取,提取效率达到75.2%~110.3%。     

紫外分光光度法测定水中挥发酚的研究

目前,测定水中挥发酚一般采用4-氨基安替比林直接比色法和萃取法.为了克服4-氨基安替比林比色法测定挥发酚时需蒸馏、试剂不稳定等缺点或不足,提高挥发酚的检测上限,本文提出用紫外光度法测定水中挥发酚.用浓磷酸固定水样,用三氯甲烷、乙醚萃取水样,使挥发酚转移到乙醚中,排除多种离子的干扰;加入无水硫酸钠脱水,用乙醚定容,269 nm处测定吸光度.此方法不仅使测定范围增大到0.409～120 mg/L,且简便、快速、准确.本方法与标准方法有很好的可比性,同时,有很好的重现性.     

Investigation of arsenic speciation on drinking water treatment media utilizing automated sequential continuous flow extraction with IC-ICP-MS detection

Three treatment media, used for the removal of arsenic from drinking water, were sequentially extracted using 10 mM MgCl2(pH 8), 10 mM NaH2PO4(pH 7) followed by 10 mM (NH4)2C2O4(pH 3). The media were extracted using an on-line automated continuous extraction system which allowed the arsenic in each of the extraction fluids to be speciated on-line using IC-ICP-MS. The 10 mM MgCl2 preferentially extracted As(III) from each of the media. The percentage of the arsenic extracted by the MgCl2, relative to a HNO3/H2O2 digestion of the media, ranged from 0.1-2.3% for the three solids. The next sequential extraction fluid, 10 mM NaH2PO4, extracted some of the residual As(III) remaining on each of the media but the predominant species extracted was As(V). The 10 mM NaH2PO4 extracted 15.3 to 42.8% of the total arsenic relative to a total digested concentration for each of the media. The As(III) and As(V) stability studies conducted in these two extraction fluids indicated that conversion between As(III) and As(V) was not significant for the short extraction fluid sample contact time associated with the on-line continuous flow extraction cell. Finally, the 10 mM (NH4)2C2O4 extraction fluid was utilized in an off-line analysis mode because the Fe and As concentrations extracted from the media were not compatible with direct ICP-MS detection. The (NH4)2C2O4 extracted 2.9-29% As(III) for all three media and caused an oxidation of As(III) to As(V) during the extraction period for one of the three media. The sum of the arsenic from each of the three extraction fluids represented 92%, 44% and 53% of the available total arsenic for the three media, respectively. The speciation results for each media were obtained by adding all the speciation results from all three extraction fluids together and the resulting distribution of As(III)/As(V) compared well with the speciation results obtained via XANES.     

Artemia salina as test organism for assessment of acute toxicity of leachate water from landfills

Artemia salina has, for the first time, been used as test organism for acute toxicity of leachate water from three landfills (the municipal landfills at Kristianstad, Sweden and Siauliai, Lithuania, and an industrial landfill at Stena fragmenting AB, Halmstad, as well as for leachate from Kristianstad treated in different ways in a pilot plan). Artemia can tolerate the high concentrations of chloride ions found in such waters. Large differences in toxicities were found, the leachate from Siauliai being the most toxic one. To increase the selectivity in the measurements, a fractionation was done by using ion exchange to separate ammonium/ammonia and metal ions from the leachate, and activated carbon adsorbents for organic pollutants. The influence of some metals and phenol compounds on the toxicity was investigated separately. It was found that most of the toxicity emanated from the ammonium/ammonia components in the leachate. However, there was also a significant contribution from organic pollutants, other than phenol compounds, since separate experiments had in this latter case indicated negligible impact. The concentrations of metals were at a level, shown by separate experiments, where only small contribution to the toxicity could be expected.     

Dietary exposure to xenoestrogens in New Zealand

Continuing evidence of the feminising effects of xenoestrogens on a range of wildlife species increases the need to assess the human health risk of these estrogen mimics. We have estimated the exposure of New Zealand males, females and young men to a range of naturally occurring and synthetic xenoestrogens found in food. Only estrogenic compounds that act by interaction with the estrogen receptor have been included. Theoretical plasma estrogen activity levels were derived from estrogen exposure estimates and estrogenic potency data. Theoretical plasma levels were compared with published data for specific xenoestrogens. There was surprisingly close agreement. Xenoestrogenicity from dietary intake was almost equally attributed to naturally occurring and synthetic xenoestrogens. Relative contributions for a male, for example were isoflavones (genistein and daidzein) (36%) and bisphenol A (34%) with smaller contributions from alkyl phenols (18%) and the flavonoids (phloretin and kaempferol) (12%). It is suggested that dietary xenoestrogens might have a pharmacological effect on New Zealand males and postmenopausal women, but are unlikely to be significant for pre-menopausal women.     

Labile pools of Pb in vegetable-growing soils investigated by an isotope dilution method and its influence on soil pH

Pollution of Pb in the surface of agricultural soils is of increasing concern due to its serious impact on the plant growth and the human health through the food chain. However, the mobility, activity and bioavailability of Pb rely mainly on its various chemical species in soils. In the present study, E and L values, the labile pools of isotopically exchangeable Pb, were estimated using the method of isotope dilution in three vegetable-growing soils. The experiments involved adding a stable enriched isotope ((206)Pb > 96%) to a soil suspension and to soils in which plants are subsequently grown, the labile pools of Pb were then estimated by measuring the isotopic composition of Pb in soil solutions and in the plant tissues, respectively. In addition, the correlation of E values and soil pH was investigated at the ranges of pH 4.5-7.0. The amount of labile Pb in soils was also estimated using different single chemical extractants and a modified BCR approach. The results showed that after spiking the enriched isotopes of (206)Pb (>96%) for 24 hours an equilibration of isotopic exchanges in soil suspensions was achieved, and the isotope ratios of (208)Pb/(206)Pb measured at that time was used for calculating the E(24 h) values. The labile pools of Pb by %E(24 h) values, ranging from 53.2% to 61.7% with an average 57%, were found to be significantly higher (p < 0.05) than the values estimated with L values, single chemical extractants and the Σ(BCR) values obtained with the BCR approach, respectively. A strong negative correlation (R(2) = 0.984) between E(24 h) values and soil pH was found in the tested soil sample. The results indicate that the %E(24 h) value can more rapidly and easily predict the labile pools of Pb in soils compared with L values, but it might be readily overestimated because of the artificial soil acidity derived from the spiked isotopic tracer and the excess of spiked enriched isotopes. The results also suggest that the amounts of Pb extracted with EDTA and the Σ(BCR) values extracted with the modified BCR approach are helpful to detect the labile pools of Pb in soils. In addition, the negative correlation between soil pH and the labile pools of Pb in soils may be useful for further remediation to reduce the bioavailability of Pb in contaminated soils.     

垃圾渗出液中酚类物质的气相色谱分析

建立了一种用气相色谱法准确、快速测定垃圾渗出液中酚类物质的新方法。外标法定量分析 4 -甲基苯酚及苯酚。采用填装了 80～ 1 0 0目 Chromosorb WAW DCMS的玻璃填充柱 ,担体上涂渍了 2 % OV-1 7,2 .5% QF-1的固定液 ,分离酚类物质效果较好 ,线性范围为 1 0 0～ 1 50 0 mg/ L,相关系数为 0 .9996,方法回收率为 94 .7%。     

Estrogenic activity profiles and risks in surface waters and sediments of the Pearl River system in South China assessed by chemical analysis and in vitro bioassay

Estrogenic activity risks in the Pearl River system (Liuxi River, Zhujiang River and Shijing River) in South China were assessed by combined chemical analysis and recombinant yeast estrogen screen (YES) bioassay for surface waters and sediments collected in both dry and wet seasons. The xenoestrogens 4-tert-octylphenol, 4-nonylphenol and bisphenol A were detected at almost every sampling site at concentrations of several ng L(-1) (ng g(-1)) to tens of μg L(-1) (μg g(-1)) in surface waters (and sediments). The estrogens estrone and 17β-estradiol were also detected in most of the samples with concentrations from several ng L(-1) (ng g(-1)) to tens of ng L(-1) (ng g(-1)) in surface waters (and sediments). However, synthetic estrogens diethylstilbestrol and 17α-ethinylestradiol were only detected at a few sites. The 17β-estradiol equivalents (EEQ) screened by the YES bioassay were in the range of 0.23-324 ng L(-1) in surface waters and from not detected to 101 ng g(-1) in sediments. Shijing River displayed one to two orders of magnitude higher levels for both measured chemical concentrations and estrogenic activities than the Zhujiang River and the Liuxi River. A risk assessment for the surface waters showed high risks for the downstream reaches of the Liuxi River and the upstream to midstream reaches of the Zhujiang River and the Shijing River. Higher estrogenic risks were observed in the wet season than in the dry season for surface waters, probably due to the input of runoff and direct overflow of small urban streams during heavy rain events. Only small variations in estrogenic risk were found for the sediments between the two seasons, suggesting that sediments are a sink for these estrogenic compounds in the rivers.     

水样中痕量十溴联苯醚的分散液-液微萃取条件的优化

采用正交设计和Box-Penhnken响应面设计,对分散液-液微萃取技术萃取水样中痕量十溴联苯醚的条件进行了筛选和优化,得到最佳条件:四氯乙烯为萃取剂(10μl)、丙酮作分散剂(1ml)、pH范围5~9、离子强度,2%NaCl及萃取时间10min。此优化条件下分散液-液微萃取技术的萃取回收率可达92.37%~104.38%,富集倍数为508~611。优化条件下方法的线性范围为0.01~100ng/ml,检出限(S/N=2)为3.0pg/ml,加标回收率为96.25%~102.16%,精密度为5.44%~6.34%。     

Estrogenicity of pyrethroid insecticide metabolites

There is concern that insecticides are able to mimic the action of 17beta-estradiol by interaction with the human estrogen receptor. Pyrethroids are commonly used insecticides and several have been assessed for potential endocrine disrupting activity by various methods. It has been noted that some metabolites of pyrethroids, in particular, permethrin and cypermethrin, have chemical structures that are more likely to interact with the cellular estrogen receptor than the parent pyrethroid. For this study permethrin and cypermethrin metabolites 3-(4-hydroxy-3-phenoxy)benzyl alcohol, 3-(4-hydroxy-3-phenoxy)benzoic acid, and N-3-(phenoxybenzoyl)glycine were synthesised, and together with the commercially available 3-phenoxybenzyl alcohol, 3-phenoxybenzaldehyde, and 3-phenoxybenzoic acid, were studied in a recombinant yeast assay expressing human estrogen receptors (YES). Three metabolites, 3-phenoxybenzyl alcohol, 3-(4-hydroxy-3-phenoxy)benzyl alcohol, and 3-phenoxybenzaldehyde, showed estrogenic activity of approximately 10(5) less than that of 17beta-estradiol. No activity was observed in the yeast assay for 3-phenoxybenzoic acid, 3-(4-hydroxy-3-phenoxy)benzoic acid, and N-3-(phenoxybenzoyl)glycine. The results from this study show that pyrethroid metabolites are capable of interacting with the human estrogen receptor, and so might present a risk to human health and environmental well being. The impact would be expected to be small, but still add to the overall environmental xenoestrogen load.     

Performance optimization of a membrane assisted passive sampler for monitoring of ionizable organic compounds in water

A thin-walled silicone rubber hollow fibre membrane has been developed as a passive sampler. The inside of the tube is filled with an aqueous solution at an appropriate pH. The tube is sealed at both ends and then immersed in a water sample. In order for the ionizable permeating compounds to be trapped in the aqueous receiving phase, the pH is adjusted such that the compounds are ionized and trapped. The major advantages are its simplicity, low cost and high selectivity, since only ionizable organic compounds are trapped. Additionally, the sampler uses no organic solvent. By adjusting the pH of the acceptor phase, it is possible to control the extraction process and whether the sampler is used in the kinetic or equilibrium regime. Since it is very selective, no further clean-up of the extract is required. The membrane assisted passive sampler has been tested for extraction of chlorophenols under laboratory conditions. The extraction process was found to be linear over a 72 h sampling period. Selectivity of the passive sampler in river water was demonstrated and the extraction process was independent of sample concentration, even at lower concentration levels of analytes. However, the sample matrix in some river water samples led to incomplete trapping, thereby reducing the amount trapped in the acceptor phase. Detection limits (three times signal to noise ratio) were dependant on sample matrix and type of detection system and ranged from 0.05 microg L(-1) to about 1 microg L(-1) with a UV photodiode detector in water samples from one river and 1.0 microg L(-1) to 20 microg L(-1) in another but with an ordinary UV detector. The enrichment factors in river water were 28 for 2-chlorophenol and 44 for 2,4-dichlorophenol over a 72 h sampling period. 4-chlorophenol was poorly extracted and its enrichment factor was 3.     

Occurrence and removal efficiencies of eight EDCs and estrogenicity in a STP

The occurrence and removal of eight endocrine disrupting compounds (EDCs), including estrone (E(1)), 17β-estradiol (E(2)), estriol (E(3)), 17α-ethinylestradiol (EE(2)), diethylstilbestrol (DES), bisphenol A (BPA), nonylphenol (NP) and octylphenol (OP), and their estrogenicities were investigated in a sewage treatment plant in Harbin city, China. The EDCs were extracted from wastewater samples by solid phase extraction (SPE) method and analyzed with gas chromatography coupled with mass spectrometry (GC-MS). The average concentrations in the influents and effluents ranged from 6.3 (EE(2)) to 1725.8 ng L(-1) (NP) and from 相似文献   

The continuous monitoring of field water samples with a novel multi-channel two-stage mini-bioreactor system

Toxicity monitoring of field water samples was performed using a novel multi-channel two-stage mini-bioreactor system and genetically engineered bioluminescent bacteria for the continuous monitoring and classification of the toxicity present in the samples. The toxicity of various samples spiked with known endocrine disrupting chemicals and phenol was also investigated for system characterization. The field samples used in this study were obtained from two different sites on a monthly basis--from a drinking water treatment plant, referred to as site N, and from a stream near a dam which is currently being constructed, referred to as site T. These samples were either pumped or injected into the second mini-bioreactors to initiate the toxicity test. Most of the samples did not show any specific toxicity. However, one sample showed to have, based upon the detection results, and was deemed toxic. The samples spiked with phenol showed possible responses in the DPD2540 and TV1061 channels, indicating the occurrence of both membrane and protein damage due to phenol. In the tests using an endocrine disrupting chemical, bisphenol A, DNA damage was detected in the DPD2794 channel with a concentration of 2 ppm. Finally, a simple but novel early warning protocol that can be used in a drinking water reservoir and a suspected place where effluents of toxic materials enter the water sourse was suggested with a schematic diagram. In conclusion, this system showed good feasibility for use as a toxicity monitoring system in the field and as an early warning system, indicating if effluents are toxic.     

Carbonate induced dissolution of uranium containing precipitates under cement leachate conditions

The effects of carbonate on uranium (VI) solubility under aerobic and cementitious conditions have been investigated. The information is of relevance to low level nuclear waste disposal. Aqueous NaOH, KOH, Ca(OH)2 and a cement leachate solution were added to uranyl nitrate solution. Afterwards, increasing amounts of ammonium carbonate were added to re-dissolve the precipitates. The precipitates were characterised by means of X-ray powder diffraction (XRPD) measurements and modelling studies. The model calculations were performed using the MINTEQA2 speciation code, with an expanded database incorporating uranium stability constants taken from the HATCHES database. The measured and predicted amounts of CO3(2-) needed to dissolve the precipitates were compared. The knowledge gained from the 'pure' systems was used to rationalise the precipitation and re-dissolution behaviour observed in the leachate system. The lack of uranium solubility at low carbonate levels brought into question literature formation constants for UO2(OH)3- and UO2(OH)4(2-). An approximate log K value of 26.8 for K2U2O7 formation was estimated from the KOH results at pH 12. Generally, uranium solubilities are expected to be insignificant at low level nuclear waste sites because anaerobic conditions should persist. However, this study has demonstrated that solubility in leachate could rise at high pH and high carbonate levels, if aerobic conditions were to develop. Soluble U(VI) species would be formed. The novel part of the study has been to reproduce the effects and to correlate enhanced solubilities with model predictions. Consequently, the study has re-emphasised the need for appropriate environmental monitoring of such sites.