An enzyme-linked immunosorbent assay for rare minnow (Gobiocypris rarus) vitellogenin and comparison of vitellogenin responses in rare minnow and zebrafish (Danio rerio)

A competitive enzyme-linked immunosorbent assay (ELISA) was developed to determine vitellogenin (Vtg) in rare minnow (Gobiocypris rarus) based on the separation and purification of rare minnow Vtg (r-Vtg) as well as the production of polyclonal antibody against r-Vtg in rabbits. Three different ELISAs for measuring r-Vtg were then compared: (1) indirect ELISA with the antibody against carp (Cyprinus carpio) Vtg (c-Vtg) (IC-ELISA); (2) competitive ELISA with the antibody against c-Vtg, and using r-Vtg for coating the plates and preparing standard curve (CC-ELISA); (3) competitive ELISA with the antibody against r-Vtg, and using r-Vtg for coating the plates and preparing standard curve (CR-ELISA). The result showed that the homologous CR-ELISA was the most sensitive among the three assays for quantifying r-Vtg. The sensitivities to 17 alpha-ethinylestradiol (EE2) Of rare minnow and zebrafish (Danio rerio) were compared upon the establishment of homologous competitive ELISA. The lowest observed effect concentrations (LOECs) to induce Vtg were found to be 0.8 ng EE2 l(-1) for rare minnow and 4 ng EE2 l(-1) for zebrafish respectively. Afterwards, CR-ELISA was applied to measure Vtg concentration in whole body homogenate (WBH) of juvenile rare minnow fed by three diets (tubifex from wastewater treatment plant, Artemia nauplii and commercial pellet food), and the agreements between bioassay and GC-MS analysis demonstrated that rare minnow was a sensitive fish model for assessing estrogenic effects of endocrine disrupting compounds in aquatic environment. (c) 2006 Elsevier B.V. All rights reserved.

Assessment of estrogenic activity of leachate from automobile tires with two in vitro bioassays

In this study, a combination of enzyme-linked receptor assay (ELRA) and yeast estrogen screen (YES) assay was firstly applied to determine whether automobile tires immersed in fresh water can leach chemicals, which display estrogenic activity. We optimized ELRA substituting the chromogene substrate by a luminescent one, and found that luminescent ELBA was more sensitive to 17 beta-estradiol (17 beta-E2) with a detection limit of 0.016 mu g/l, compared to 0.088 mu g/l in the chromogene version. In ELRA, all tire leachates obviously showed estrogenic activity, which was increased with duration of immersion. Moreover, the leachate from hackled tires showed more potent estrogenicity than that from the whole ones. In comparison to ELRA, no detectable estrogenic activity was found in all tire leachates with YES assay. The results from YES assay further evidenced that antiestrogenic compounds can be leached from tires. As tire leachates contain estrogenic compounds, they could be important pollution sources, potentially harmful to wildlife and human health. Thus, use of shredded tires as road fill or in landfill sites should arouse our attention.

Environmental signals: Synthetic humic substances act as xeno-estrogen and affect the thyroid system of Xenopus laevis

According to outdated paradigms humic substances (HS) are considered to be refractory or inert that do not directly interact with aquatic organisms. However, they are taken up and induce biotransformation activities and may act as hormone-like substances. In the present study, we tested whether HS can interfere with endocrine regulation in the amphibian Xenopus laevis. In order to exclude contamination with phyto-hormones, which may occur in environmental isolates, the artificial HS 1500 was applied. The in vivo results showed that HS 1500 causes significant estrogenic effects on X. laevis during its larval development and results of semi-quantitative RT-PCR revealed a marked increase of the estrogenic biomarker estrogen receptor mRNA (ER-mRNA). Furthermore, preliminary RT-PCR results showed that the thyroid-stimulating hormone (TSH beta-mRNA) is enhanced after exposure to HS1500, indicating a weak adverse effect on T3/T4 availability. Hence, HS may have estrogenic and anti-thyroidal effects on aquatic animals, and therefore may influence the structure of aquatic communities and they may be considered environmental signaling chemicals. (c) 2005 Elsevier Ltd. All rights reserved.

Effects of polyaluminum chloride and copper sulfate on phosphorus and UV254 under different anoxic levels

Experimental sediments and water from shallow, eutrophic Dianchi Lakes were treated in a controlled laboratory microcosm using different chemicals under different anoxic levels. This study revealed that the polyaluminum chloride (PAC) was able to inhibit the phosphorus release and decrease the UV254 value at any anoxic level. When the DO concentrations were between 0.76-0.95 mg(.) L-1, the UV(254)value, total phosphorus (TP), and total dissolved phosphorus (TDP) in the water column were decreased by 71.93%, 87.12% and 64.24% respectively. The UV254, TP, and TDP were also decreased by 72.94%, 70.87% and 50.76% respectively at the levels of 4.56-5.32mg(.)L(-1) of DO concentrations. The treatment effects of TP and TDP in the water column using copper sulfate however were not as efficient as the PAC treatment. The UV254 value was increased with the addition of copper sulfate at every anoxic level tested but the chlorophyll-a (Chl-a) content was decreased rapidly and efficiently by copper sulfate more than the treatment by PAC. When the DO concentrations were 0.76-0.86mg(.)L(-1) and 4.75-5.14mg(.)L(-1), the Chl-a concentrations were decreased by 84.87% and 75.07% respectively through copper sulfate treatment. With additions of PAC and copper sulfate, the phosphorus fractions in sediments were shifted forward to the favorable shapes that have little ability of release. The TP concentrations in sediments were increased after treatment via PAC and copper sulfate. Under anoxic conditions, most of the BD-P (Fe-P) to NaOH-P (Al-P) was converted using the recommended PAC dose in BD-P rich sediment. Similar to the PAC, the copper sulfate also could flocculate the exchange phosphorus from sediment to overlying water. Overall though, the effects of copper sulfate treatment were not better than that of the PAC.

Effects of nitrogen forms on the production of cyanobacterial toxin microcystin-LR by an isolated Microcystis aeruginosa

A cyanobacterial strain, which produced high content of microcystin-LR (MC-LR) but no rnicrocystin-RR (MC-RR), was isolated from the hypertrophic Dianchi Lake in China and identified as Microcystis aeruginosa DC-1. Effects of nitrogen containing chemicals and trace elements on the growth and the production of MC-LR by this strain were Studied. In the presence of bicine, compared with urea and ammonium, nitrate greatly promoted the growth and the production of MC-LR. However, leucine and arginine, which were the constitutional components in the molecular structure of MC-LR or RR, inhibited the production of MC-LR. Iron and silicon up to 10mg/L had little effects on the growth of M. aeruginosa DC-1, but the production of MC-LR was apparently enhanced. Under all conditions studied here, only MC-LR but no RR was detected within the cells of M. aeruginosa DC-1. Thus, chemical forms of nitrogen, rather than the usually concerned the total nitrogen, Lind trace elements played important roles in the production of MC toxins during cyanobacterial blooms.

Comparative study of estrogenic potencies of estradiol, tamoxifen, bisphenol-A and resveratrol with two in vitro bioassays

This study was undertaken to compare the sensitivity of two in vitro screening test methods and to determine the accuracy of predicted response to spiked laboratory water samples. A newly developed enzyme-linked receptor assay (ELRA) and a widely used yeast estrogen screen (YES) assay were selected to evaluate the estrogenic responses. Four natural, pharmaceutical, xenobiotic or phytobiotic chemicals: 17beta-estradiol (E2), tamoxifen, bisphenol-A and resveratrol were examined, and 17beta-E2 was used as a positive control. 17beta-E2 can strongly induce estrogenic response in both test systems, however, ELRA was found to be more sensitive to 17beta-E2 with a detection limit of 0.07 mug/l compared to 0.88 mug/l in YES assay. Similar results were obtained for bisphenol-A and resveratrol, and their estrogen potencies relative to E2 (100%) determined by ELRA were at least 5.6 times greater than produced by YES assay. ELRA was unable to distinguish the anti-estrogen tamoxifen and YES assay is also poor at distinguishing. Comparison of response to spiked laboratory water samples show that ELRA can give accurate determination to all four chemicals with recoveries among 70-120%, while YES can only give accurate determination to 17beta-E2 and bisphenol-A with recoveries among 69-112%. The comparative results provide evidence that ELRA is more suitable for rapid screening estrogenic potency of the environmental samples. Combination of ELRA and mammalian cellular assay will constitute an advantageous test to specify agonistic or antagonistic effects. (C) 2003 Elsevier Ltd. All rights reserved.

Quantitative structure-property relationships for octanol-air partition coefficients of polychlorinated naphthalenes, chlorobenzenes and p,p '-DDT

The octanol-air partition coefficient (K-OA) is a key descriptor of chemicals partitioning between the atmosphere and environmental organic phases. Quantitative structure-property relationships (QSPR) are necessary to model and predict KOA from molecular structures. Based on 12 quantum chemical descriptors computed by the PM3 Hamiltonian, using partial least squares (PLS) analysis, a QSPR model for logarithms of K-OA to base 10 (log K-OA) for polychlorinated naphthalenes (PCNs), chlorobenzenes and p,p'-DDT was obtained. The cross-validated Q(cum)(2) value of the model is 0.973, indicating a good predictive ability of the model. The main factors governing log K-OA of the PCNs, chlorobenzenes, and p,p'-DDT are, in order of decreasing importance, molecular size and molecular ability of donating/accepting electrons to participate in intermolecular interactions. The intermolecular dispersive interactions play a leading role in governing log K-OA. The more chlorines in PCN and chlorobenzene molecules, the greater the log K-OA values. Increasing E-LUMO (the energy of the lowest unoccupied molecular orbital) of the molecules leads to decreasing log K-OA values, implying possible intermolecular interactions between the molecules under study and octanol molecules. (C) 2002 Elsevier Science Ltd. All rights reserved.

Persistent organic pollutants in water and surface sediments of Taihu Lake, China and risk assessment

Organic pollutants, especially persistent organic pollutants were examined in the water and surface sediments of Taihu Lake, China. Both 12 water and 12 sediment samples were collected over the lake. C-18 solid-phase extraction technique was applied to extract organic pollutants in collected water samples. Soxhlet extraction procedure was used to extract organic pollutants in sediment samples. The analysis was performed by GC-MS controlled by a Hewlett Packard chemstation. Two hundred and seventy-three kinds of organic chemicals in water were examined, 200 more than that detected in 1985; 188 kinds of chemicals in sediments were detected as well. Among them 21 kinds of chemicals belong to priority pollutants as well as 17 kinds to be the endocrine disruptors. The concentrations of the pollutants were more than 2 times higher than that in 1985. The possible source and relation to anthropogenic activity were discussed. (C) 2002 Elsevier Science Ltd. All rights reserved.

Cytological and biochemical alterations in Carassius auratus hepatocytes from exposure to sediment containing dioxins and related compounds

Cytological and biochemical alterations of crucial carp (Carassius auratus) hepatocytes were characterized after exposure to sediments from a lake contaminated with dioxins and other industrial chemicals. Carp were exposed in 20 L water containing 25, 50, or 100 g of contaminated sediment for 2 and 4 weeks. Ultrastructural changes in the liver were characterized by severe enlargement of hepatocytes. Alterations in the cell. included formation of condensed and irregular cell nucleus, polynuclei, dispersed heterochromatin, enlargement of the nucleolus, and degeneration of the nucleus. Mitochondrial numbers were reduced and cristae were deformed. Myelin figures and lysosomes were increased, and sometimes cell organelles and cell matrix were totally lost after 4 weeks of exposure. The ultrastructural alterations were correlated with exposure time and sediment concentrations. Hepatosometic index was significantly increased in experimental groups at 2 and 4 weeks as compared with the control group. EROD enzyme activities were strongly induced in liver. A trend from rough endoplasmic reticulum (RER) to SER was observed. Our results suggest that the dioxin-like compounds bound by sediment were bioavailable to C. auratus and cause sublethal effects.

黄土沟壑区小流域土壤pH值的空间分布及条件模拟

土壤pH值是影响土壤养分有效性和化学物质在土壤中行为的主要因素,研究土壤pH值的空间分布特征对于土壤养分管理和土壤污染预测具有重要意义。该文用地统计学方法研究了环境因素复杂的黄土高原小流域土壤pH值空间分布特征。结果表明,黄土沟壑区小流域土壤pH值具有球形—指数套合模型的空间结构特征,其空间异质性主要来源于流域内土地利用和土壤侵蚀等随机因素。与有机质协同的Kriging法能较好地对土壤pH值进行估值,其估值范围小于实测数据,估值误差来源于复杂的环境因素。序贯高斯条件模拟的土壤pH值范围与实测数据接近,模拟的平均值低于实测数据,模拟误差来源于模拟过程中独特的Kriging算法及高斯特性。

Waterborne exposure to PFOS causes disruption of the hypothalamus-pituitary-thyroid axis in zebrafish larvae

Thyroid hormones (THs) play an important role in the normal development and physiological functions in fish. Environmental chemicals may adversely affect thyroid function by disturbing gene transcription. Perfluorooctane sulfonate (PFOS), a persistent compound, is widely distributed in the aquatic environment and wildlife. In the present study, we investigated whether PFOS could disrupt the hypothalamic– pituitary–thyroid (HPT) axis. Zebrafish embryos were exposed to various concentrations of PFOS (0, 100, 200 and 400 lg L 1) and gene expression patterns were examined 15 d post-fertilization. The expression of several genes in the HPT system, i.e., corticotropin-releasing factor (CRF), thyroid-stimulating hormone (TSH), sodium/iodide symporter (NIS), thyroglobulin (TG), thyroid peroxidase (TPO), transthyretin (TTR), iodothyronine deiodinases (Dio1 and Dio2) and thyroid receptor (TRa and TRb), was quantitatively measured using real-time PCR. The gene expression levels of CRF and TSH were significantly up-regulated and down-regulated, respectively, upon exposure to 200 and 400 lg L 1 PFOS. A significant increase in NIS and Dio1 gene expression was observed at 200 lg L 1 PFOS exposure, while TG gene expression was down-regulated at 200 and 400 lg L 1 PFOS exposure. TTR gene expression was down-regulated in a concentration-dependent manner. Up-regulation and down-regulation of TRa and TRb gene expression, respectively, was observed upon exposure to PFOS. The whole body thyroxine (T4) content remained unchanged, whereas triiodothyronine (T3) levels were significantly increased, which could directly reflect disrupted thyroid hormone status after PFOS exposure. The overall results indicated that PFOS exposure could alter gene expression in the HPT axis and that mechanisms of disruption of thyroid status by PFOS could occur at several steps in the synthesis, regulation, and action of thyroid hormones.

生物催化的Baeyer-Villiger氧化反应研究

Baeyer-Villiger氧化反应是一种很重要的化学反应，产生的许多中间体或产物可以被用来生产多种化学产品和药物。此反应具有多功能性，可以氧化多种羰基化合物，但是化学方法中的必需反应物——氧化剂在生产、储存、运输、反应的过程中都存在很多的不安全因素，反应的立体选择性也不强，而生物转化则具有底物选择性、立构选择性、化学选择性、对映选择性等一般化学反应中不具备的优点，在精细化工中占有很大的优势。在工业生物催化中有很好的应用前景。 为了研究生物催化的Baeyer-Villiger反应，我们从本实验室保藏菌种中分离筛选出一株能够以环己酮作为唯一碳源的菌株，进行初步研究并对其产物进行GC/MS定性，探讨了pH，装液量，底物浓度，培养时间，温度以及转速等条件对细菌生长的影响，并进一步研究了细菌的底物广谱性。 此菌株经鉴定属于邻单胞菌属Plesiomonas sp.)， 根据正交试验，确定了菌的最佳生长条件：底物浓度为1mL/L，底物浓度过高对菌株生长有抑制作用，转速为150 rpm ，温度为30℃ ，pH为7.0； 此菌株转化环己酮的产物通过GC/MS检测含有内酯，表明此菌株能够催化Baeyer-Villiger氧化反应；此菌株还能够以与环己酮有相似结构的环己烷，环戊酮等作为唯一碳源生长，说明此菌株底物利用范围比较广，用途比较广泛。 Baeyer-Villiger oxidation is an important chemical conversion, its products and intermediates can be used to produce a lot of medicine and fine chemicals. Its success is largely due to its versatility: a variety of carbonyl compounds can be oxidized, a large number of functional groups are tolerated, the regiochemistry is highly predictable and so on, but the oxidants that the traditional chemistry way needs have a number of problem in their production, storage, transportation and reaction, Chemistry way has not a high stereochemistry yet. However, biotransformations have many attractive characters, such as substrate-, stereo-, chemo- and enantioselectivity, so it has a great advantage in the fine chemical industry and has a bright prospect in the industrial biological catalysis. In order to study Baeyer-Villiger oxidation, we isolated a strain which can utilize cyclohexanone as sole carbon source and had a primary research on it. Its product was identified by GC/MS. Effects of pH, volume, concentration of cyclohexanone, cultivating time, temperature and rotate speed on the growth of bacteria were discussed, and the other organic substrates were also studied. The strain was identified as Plesiomonas sp.. The result of orthogonal test made it sure that the best growth condition of the strain is: rotate speed 150 rpm, temperature 30℃, pH7.0, concentration of cyclohexanone1ml/L. There is caprolactone in the product of the fermentation with cyclohexanone as substrate by GC/MS,which indicated that the strain can catalyse Baeyer-Villiger oxidation.In addition,the strain can utilize other organic substrates having the similar structure with cyclohexanone such as cyclohexane, cyclopentanone, Swertiamarin as sole carbon source.So the strain can be applied extentively.