A protocol for the evaluation of genotoxicity in bile of carp (Cyprinus carpio) exposed to lake water treated with different disinfectants

A sensitive and rapid method to evaluate toxic and genotoxic properties of drinking water supplied from Lake Trasimeno (Umbria, Central Italy) was worked out analysing bile in Cyprinus carpio exposed for 20 d to lake water treated with 3 different disinfectants, sodium hypochlorite (NaClO), chlorine dioxide (ClO₂) and peracetic acid (PAA). Fish were sacrificed at 0, 10 and 20 d in order to investigate the time course of these endpoints. An aliquot of bile samples was fractionated by adsorption on C₁₈ silica cartridges and the genotoxic potential of whole bile and of bile fractions was evaluated by the single-cell microgel-electrophoresis (comet) assay on human colonic adenocarcinoma cells (Caco-2). Bile (both whole and fractionated) from specimens exposed to the three disinfectants always showed a genotoxic activity as compared to the control group. The results of this study provide evidence that all three disinfectants cause an increase in bile genotoxicity of chronically exposed fish.     

Bioconcentration and Phytotoxicity of Cd in Eichhornia crassipes

Plants of Eichhornia crassipes grown at various levels of cadmium ranging from 0.1 to 100 μg ml⁻¹ accumulated Cd in a concentration and duration dependent manner. At all levels, Cd accumulation by various plant tissues followed the order roots shoot leaves. Approximately 80% of total Cd was accumulated by plant at highest concentration (100 μg ml⁻¹) used in the experiment. Cadmium induced phytotoxicity appears at 25.0 μg ml⁻¹ resulting into reduced levels of chlorophyll, protein and in vivo nitrate reductase activity of the plant. However, a slight induction of these physiological variables was obtained at lowest Cd (0.1 μg ml⁻¹) concentration. In contrast, carotenoid content increased at highest Cd concentration i.e., 100 μg ml⁻¹. Similar effects at low and high levels of Cd was obtained with respect to mitotic index and micronuclei in root meristem of the plant. It could be inferred that Cd toxicity in plant is differential depending upon the low and high concentration of Cd in the medium.     

Perfluorooctane Sulfonate Increases the Genotoxicity of Cyclophosphamide in the Micronucleus Assay with V79 Cells: Further Proof of Alterations in Cell Membrane Properties Caused by PFOS (3 pp)

Background, Aim and Scope Perfluorooctane sulfonate (PFOS; C8F17SO3-) is a fully fluorinated organic compound which has been manufactured for decades and was used widely in industrial and commercial products. The recent toxicological knowledge of PFOS mainly concerns mono-substance exposures of PFOS to biological systems, leaving the potential interactive effects of PFOS with other compounds as an area where understanding is significantly lacking. However, a recent study, reported the potential of PFOS to enhance the toxicity of two compounds by increasing cell membrane permeability. This is of particular concern since PFOS has been reported to be widely distributed in the environment where contaminants are known to occur in complex mixtures. In this study, PFOS was evaluated alone and in combination with cyclophosphamide (CPP) to investigate whether a presence of PFOS leads to an increased genotoxic potential of CPP towards hamster lung V79 cells. Genotoxicity was investigated using the micronucleus (MN) assay according to the recent draft ISO/DIS 21427-2 method. PFOS alone demonstrated no genotoxicity up to a concentration of 12.5 mg/L. However, PFOS combined with two different concentrations of CPP, with metabolic activation, caused a significant increase in the number of micronucleated cells compared to treatments with CPP only. These results provide a first indication that PFOS has the potential to enhance the genotoxic action of CPP towards V79 cells, suggesting that together with the alterations in cell membrane properties shown previously, that genotoxicity of complex mixtures may be increased significantly by changes in chemical uptake. Together with an earlier study performed by the own working group it can be concluded that PFOS alone is not genotoxic in this bioassay using V79 cells up to 12.5 mg/L, but that further investigations are needed to assess the potential interaction between PFOS and other substances, in particular regarding the impact of membrane alterations on the uptake of toxic substances. Materials and Methods: - Results: - Discussion: - Conclusions: - Recommendations and Perspectives: -     

15N Metabolic test for the determination of phytotoxic effects of chemicals and contaminated environmental samples

A stable isotope¹⁵N-nitrogen test (ESIMA = Ecotoxicological Stable Isotope Metabolic Assay) was developed to assess biological effects and the potential toxicological hazard of chemicals and contaminated environmental samples on plant metabolism. The assay measures the effect of toxicants on the incorporation of a¹⁵N labelled tracer into the total nitrogen fraction (both the nonprotein and protein fraction) of plants. Segments ofPisum arvense epicotyls are used as test substrates because of their high metabolic activity. The plant material is incubated under standardised conditions for two hours; subsequently¹⁵N incorporation is analysed by determining the¹⁵N abundance (¹⁵N atom-%) in the epicotyl segments. The effects of toxicants are evaluated by comparing the¹⁵N incorporation rates of control tissue and epicotyl segments exposed to individual chemicals or complex environmental samples. The specificity and sensitivity of effects as indicated by ESIMA were compared with effects as measured by two established ecotoxicological bioassays, the pollen tube growth test using pollen ofNicotiana sylvestris and the bacterial luminescence inhibition test using pollen ofPhotobacterium phosphoreum. The results of the study clearly indicate the suitability of ESIMA for assessing toxic impacts on plant nitrogen metabolism. Prof. Dr. habil. Hans Faust dedicated to his 70^th birthday.     

镉对小麦叶片DNA伤害的彗星实验研究

为了建立植物基因毒性的研究方法 ,以小麦为对象 ,进行了植物彗星实验方法的研究 .以 0 1mg·L- 1 的Cd2 + 溶液处理小麦叶片后 ,采用机械方法分离细胞核 ,制备玻片后变性 0 ,5 ,15 ,3 0min ,在 10 0 ,2 0 0 ,3 0 0mA下电泳 5 ,15 ,3 0min .实验结果表明 ,采用机械分离的方法可以获得大量的、能够进行彗星实验的细胞核 ,增加DNA的变性时间、电泳电流大小和电泳时间可以增加DNA片段在电场中的迁移 ,提高实验的灵敏度 ,但变性时间和电泳时间过长增加了对照处理DNA片段的迁移 .因此 ,本实验提出了小麦叶片彗星实验的最佳条件是DNA变性 15min ,3 0 0mA下电泳 15min ,并在该条件下研究了重金属镉对小麦的基因损伤 .结果表明 ,重金属镉能引起小麦基因的损伤 ,并随镉浓度的增加伤害程度加大     

不同粒径大气颗粒物几种重金属成分分析及其致突变性研究

采集冬季太原市一采样点不同粒径大气颗粒物,经分析,总悬浮颗粒物日均浓度为1.04mg/m~3,严重超标。其中粒径小于7.0μm的占49.6％,小于3.3μm的占33.5％。颗粒物无机提取液中5种金属元素的浓度由高到低依次为Pb、Mn、Cr、Ni和Cd。每一种元素均呈随颗粒物粒径减小浓度增高的趋势。以SOS显色法和小鼠体内骨髓细胞染色体畸变试验检测颗粒物的无机提取液和模拟肺泡液溶出液的遗传毒性,表明小粒径颗粒物遗传毒性较强;小于1.1μm的颗粒物,仅用相当于5m~3或10m~3空气量的样品液,即可诱发SOS反应或致染色体损伤。     

硝基甲苯对小鼠睾丸生殖细胞DNA的损伤作用

为提供硝基甲苯的环境遗传毒理学依据和建立鱼类生殖细胞的培养方法,采用昆明小鼠睾丸支持细胞/生殖细胞共培养法以及彗星实验,研究了2,4-二硝基甲苯(2,4-DNT)、2,6-二硝基甲苯(2,6-DNT)、对硝基甲苯(4-NT)对小鼠睾丸生殖细胞DNA的损伤作用.结果表明,3种受试硝基甲苯化合物均能够诱导小鼠睾丸生殖细胞DNA单链断裂,而且其受损率与剂量对数具有明显的剂量-效应关系.2,4-DNT、2,6-DNT以及4-NT的各个剂量浓度引起细胞DNA损伤的程度,与对照组相比,均具有显着性差异(p<0.01,p<0.05).受试化合物的毒性顺序为2,6-DNT>2,4-DNT>4-NT,DNA的损伤作用二硝基甲苯大于单硝基甲苯.提示在体外条件下,2,4-DNT、2,6-DNT和4-NT具有生殖毒性,可以引起小鼠睾丸生殖细胞DNA损伤.     

Preparation of hapten-specific monoclonal antibody for cadmium and its ELISA application to aqueous samples

High-affinity and specific monoclonal antibodies against cadmium-ethylene diamine tetraacetic acid (EDTA) complex have been produced using the hybridoma technique. A hapten was synthesized and characterized by Fourier Transform Infrared Spectroscopy (FT-IR) and UV-Vis. Competitive enzyme-linked immunosorbent assay (ELISA) for quantitative detection of cadmium in aqueous sample was developed. The monoclonal antibody with high level of binding affinity for Cd-IEDTA-BSA and high specificity for soluble Cd-EDTA complex showed less than 0.99% cross-reactivity with other 11 metals. The limit of detection was 0.10 μg·L^-1, and the effective linear range was 10^-1–10³ μg·L^-1. The intra- and inter-assay coefficient variations were 1.5%–6.3% and 3.2%–7.4%, respectively. The spike recovery in different water samples were between 98.5% and 110.3%. The detection limit of this assay was well below the allowable concentration of cadmium (3 μg·L^-1), and the working range was wider than that of other methods which showed the range of 2.19–86.38 and 0–10³ μg·L^-1. The competitive ELISA established in this paper was sensitive and accurate in the screening of cadmium in aqueous samples. The results will lay a solid foundation for construction of an immunoassay kit for cadmium.     

盐酸小檗碱对小鼠脾细胞的DNA损伤和氧化性损伤研究

为了探讨盐酸小檗碱对小鼠的DNA损伤和氧化性损伤。随机选取30只小鼠分成对照组以及7.5,15,30,60与120 mg?kg^-1实验组,处理后,应用小鼠脾细胞进行彗星实验与抗氧化酶实验。测定DNA损伤情况以及超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、过氧化物酶（POD）活性以及丙二醛（MDA）含量变化。对盐酸小檗碱的DNA损伤与氧化性损伤作用进行比较研究。研究结果表明：彗星实验中,随着盐酸小檗碱浓度的增加,尾部DNA含量、尾长与尾矩均增加,与阴性对照组相比差异有统计学意义（p<0.05或p<0.01）,且呈剂量-效应关系;超氧化物歧化酶(SOD)与过氧化氢酶(CAT)活性随盐酸小檗碱剂量增加逐渐降低,丙二醛(MDA)含量明显下降,过氧化物酶(POD)活性在7.5 mg?kg^-1时上升,而后逐渐下降。在60 mg?kg^-1和120 mg?kg^-1时,有极显著性差异（p<0.01）产生。由此可见,盐酸小檗碱对小鼠脾细胞有一定的损伤作用,能够引起小鼠脾细胞的DNA损伤和氧化性损伤。     

Evaluation of environmental stress by comet assay on freshwater snail Lymnea luteola L. exposed to titanium dioxide nanoparticles

The use of aquatic organisms to monitor for contamination is well-established. Therefore, this study was designed to assess the adverse effects of titanium dioxide nanoparticles (TiO₂NP) in freshwater snail Lymnea luteola L. (L. luteola). For TiO₂NPs ecotoxicity tests, snails were exposed for seven days. A dose and time-response relationship was observed for TiO₂NP-induced genotoxicity. Induction of oxidative stress in digestive gland was observed by a decrease in glutathione and gluthathions-S-transferase levels accompanied by elevated malondialdehyde levels at TiO₂NP (9 and 28 µg/mL). Superoxide dismutase activities were markedly reduced at TiO₂NP (9 and 28 µg/mL) at days 1 and 3, but not at day 7. Catalase activities were decreased at days 1 and 3 but increased at higher concentration of TiO₂NP at day 7. DNA fragmentation occurring in L. luteola due to ecotoxic impact TiO₂NP was further substantiated by alkaline single-cell gel electrophoresis assay and expressed in terms of percent tail DNA and olive tail moment. The results indicate that the interaction of these TiO₂NP with snail influences the toxicity, which is mediated by oxidative stress in a dose- and time-dependent manner. The measurement of DNA integrity in L. luteola thus provides an early warning signal of contamination of the aquatic ecosystem by TiO₂NP. Data suggest the freshwater snail L. luteola is a potential biomonitor organism.