甲醛诱导Hela细胞凋亡的研究

细胞凋亡是当今环境毒理学领域的研究热点之一.为了探讨甲醛能否诱导细胞凋亡,以Hela细胞为实验材料,通过不同浓度的甲醛处理24h后,应用形态学观察、四甲基偶氮唑蓝(MTT)实验和流式细胞仪检测了Hela细胞凋亡情况.实验结果表明低浓度甲醛(≤0.2mmol·L-1)处理Hela细胞24h后,对细胞生长有显著的抑制作用(p<0.05),但对细胞形态改变不大,且对细胞核DNA没有损伤;而高浓度甲醛(≥0.4mmol·L-1)处理Hela细胞24h后,对细胞生长有极显著的抑制作用(p<0.01),细胞形态逐渐变的模糊不清,并有少量细胞萎缩成圆形,且会造成细胞核DNA有规律的断裂,细胞凋亡率随甲醛浓度的增加而增大.以上实验结果显示,高浓度甲醛处理Hela细胞24h后,会诱导细胞凋亡;并且随甲醛浓度的升高凋亡率逐渐增大,呈一定的剂量-效应关系.     

酶联免疫吸附分析法测定苯并(a)芘和多氯联苯

对苯并(a)芘作了五种不同的化学修饰,并与载体蛋白交联,制备出十四种单克隆抗体.经单克隆抗体的筛选和实验条件的优化,建立了测定苯并(a)芘的酶联免疫吸附分析法.以4,4'-二氯联苯为测定多氯联苯的酶联免疫吸附分析法的起始物,作了二种化学修饰,制备出九种多克隆抗体,建立了以羊抗多氯联苯抗体为基础的酶联免疫吸附分析法,并用于土壤中多氯联苯的测定.     

己酸二乙氨基乙醇酯的致突变性研究

采用鼠伤寒沙门氏菌回复突变试验(Ames试验),用TA97,TA98,TA100和TA102菌株,加与不加S9,剂量分别设为每皿1000μg,500μg和250μg;小鼠骨髓多染红细胞微核试验,剂量设为雄性、雌性小鼠给药剂量均为250,500和1000mg*kg-1;小鼠睾丸精母细胞染色体畸变试验,剂量设为625mg*kg-1,1250mg*kg-1和2500mg*kg-1.结果表明,Ames试验中,各测试浓度的诱发回变菌落数均未超过自发回变菌落数的2倍;小鼠骨髓多染红细胞微核试验和小鼠睾丸精母细胞染色体畸变试验,各剂量组和溶剂对照组的微核率进行统计学处理,未见有显着性差异(P＞0.05),表明各项试验结果均为阴性.因此,己酸二乙氨基乙醇酯无致突变作用.     

单克隆抗体酶免疫分析法测定大米中杀虫脒残留量

用抗杀虫脒单克隆抗体为试剂,建立了间接竞争法,直接竞争法和标记抗原的竞争法等３种测定大米中杀生脒残留量的ＥＬＩＳＡ法,标准曲线上抑制率为５０％时对应的杀虫脒浓度分别为１．３、１．７和５．３ｎｇ／ｍｌ,大米样品用甲醇提到,提取液经吸附剂柱简单净化后即可用上述ＥＬＩＳＡ对痕量杀虫脒进行定量,使用直接竞争法时,样品加标回收率为９０％－１１６％,平行测定的相对标准偏差为５．１％－１６％,方法检出限为０．５     

Polyclonal antibody-based indirect competitive enzyme-linked immunosorbent assay for screening of paclobutrazol in fruits

Abstract

Paclobutrazol (PBZ) is a plant growth regulator (PGR) widely used in fruit and vegetable cultivation. However, due to the severe toxicity of PBZ, a sub-ppm level maximum residue limit (MRL) was established worldwide. Therefore, it is significant to propose a rapid, sensitive and high throughput screening method for monitoring the PBZ residues in foods. In this study, a simple and sensitive indirect competitive Enzyme-linked immunosorbent assay (icELISA) was established for PBZ detection in fruits basing polyclonal antibody. For both economy and pollution prevention, a microwave-solvent-free method was used to synthesize the PBZ hapten with high efficiency. The detection conditions, such as coating antigen concentration, antibody concentration, organic reagent concentration, ionic strength and pH, were optimized. Under the optimized conditions, this method showed high sensitivity and specificity. The detection range is 1.27-138.23?ng/mL, half-maximum inhibition concentration (IC₅₀) is 13.26?ng/mL, and the IC₂₀ was lower than the reported ELISAs for PBZ. Additionally, this method had high accuracy and precision. The recoveries were ranged from 88.78% to 96.80% in PBZ spiked apple samples with RSD below 4%. All the results showed that the polyclonal antibody based icELISA could be useful for PBZ screening in fruit samples.     

Genotoxicity evaluation and a primary risk assessment of organic pollutants in the drinking water sources of Nanjing, China

An increasing number of industrial, agricultural, and commercial chemicals in the aquatic environment leads to various deleterious effects on organisms, which is becoming an increasingly serious problem in China. In this study, the comet assay was conducted to investigate the genotoxicity to human body caused by organic concentrates in the drinking water sources of Nanjing City from Yangtze River of China, and health and ecology risk due to expose to these organic pollutants were evaluated with the multimedia environmental assessment system （MEAS）. For all the water samples, they were collected from four different locations in the drinking water sourcr samples, es of Nanjing City. The results of the comet assay showed that all the organic concentrates from the water samples could induce different levels DNA damages on human peripheral blood lymphocytes, and a statistically significant difference （p〈0.01） was observed compared with the solvent control, which demonstrated the genotoxicity was in existence. According to the ambient severity （AS） of individual compound, we had sorted out the main organic pollutants in the drinking water source of the four waterworks, and the results showed that there was some potential hazard to human body for all the source water, namely the total ambient severity （TAS） of health for each water source was more than 1. However, the TAS of ecology for each water source was less than 1, which indicated that it was safe to ecology. The results of this investigation demonstrate the application of the comet assay and the MEAS in aquatic environmental monitoring studies, and the comet assay found to be fast, sensitive, and suitable for genotoxicity monitoring programs of drinking water source.     

彗星试验检测地下水有机污染物的遗传毒性效应

应用人外周血淋巴细胞和小鼠淋巴细胞彗星试验,对长治市5个样点的地下水有机浓集物进行了遗传毒性研究。结果表明,各水样中有机浓集物在3个染毒剂量50,200,800 m L·管-1条件下均可对人外周血淋巴细胞和小鼠淋巴细胞DNA产生不同程度的遗传损伤,并且随着浓度的升高,DNA损伤程度明显增加,各实验组与溶剂对照组相比有P<0.05或P<0.01显著差异。研究表明,上述2种淋巴细胞彗星试验可有效地检测地下水体有机污染物的遗传毒性。     

Characterization of the pro-inflammatory potencies of purified Jatropha phorbol esters by a gene expression-based in vitro bioassay

The oil extracted from Jatropha seeds is an emerging biodiesel feedstock that also contains several pro-inflammatory phorbol esters. These phorbol esters can elicit adverse inflammatory responses through activation of the protein kinase C, as previously described for the prototypical phorbol ester 12-O-tetradecanoylphorbol-13-acetate. We extracted and purified the six phorbol esters identified in Jatropha oil and assessed their pro-inflammatory activities using a recently developed gene expression-based bioassay. Borrowing from an approach used for the assessment of dioxin toxicity, we expressed their pro-inflammatory potencies in relation to the potency of 12-O-tetradecanoylphorbol-13-acetate (in terms of 12-O-tetradecanoylphorbol-13-acetate toxic equivalency factors). The pro-inflammatory potencies of Jatropha phorbol esters were orders of magnitudes below the potency of 12-O-tetradecanoylphorbol-13-acetate. Interestingly, the cytotoxicity of phorbol esters did not appear to be directly related to their pro-inflammatory potencies. Calculations based on phorbol ester potencies and concentrations led to overestimation of the pro-inflammatory activity of Jatropha oil, as measured by the same gene expression-based bioassay. The preliminary results presented here suggest that further work on the described approach may lead to the development of valuable tools and metrics to quantify and predict the pro-inflammatory activities of complex phorbol ester mixtures.     

Transcriptional responses of heat shock protein 70 (Hsp70) to thermal, bisphenol A, and copper stresses in the dinoflagellate Prorocentrum minimum

The designation of biodiesel as an environmental-friendly alternative to diesel oil has improved its commercialization and use. However, most biodiesel environmental safety studies refer to air pollution and so far there have been very few literature data about its impacts upon other biotic systems, e.g. water, and exposed organisms. Spill simulations in water were carried out with neat diesel and biodiesel and their blends aiming at assessing their genotoxic potentials should there be contaminations of water systems. The water soluble fractions (WSF) from the spill simulations were submitted to solid phase extraction with C-18 cartridge and the extracts obtained were evaluated carrying out genotoxic and mutagenic bioassays [the Salmonella assay and the in vitro MicroFlow® kit (Litron) assay]. Mutagenic and genotoxic effects were observed, respectively, in the Salmonella/microsome preincubation assay and the in vitro MN test carried out with the biodiesel WSF. This interesting result may be related to the presence of pollutants in biodiesel derived from the raw material source used in its production chain. The data showed that care while using biodiesel should be taken to avoid harmful effects on living organisms in cases of water pollution.     

Doramectin induced cytotoxic and genotoxic effects on bovine peripheral lymphocytes and cumulus cells in vitro

The effect of doramectin (DOR) was tested on two experimental somatic bovine cells in vitro: peripheral lymphocytes (PL) and cumulus cells (CC). The cytotoxicity and genotoxicity of DOR were assessed using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, single cell gel electrophoresis assay (SCGE) and cytokinesis-block micronucleus cytome (CBMN Cyt) assay. Both cells were treated with three concentrations of DOR (20, 40, 60?ng?mL^–1) for 24?h. The results obtained from PL demonstrated that DOR was able to induce cytotoxic effect and DNA damage with all concentrations tested. Additionally, DOR increased micronuclei (MNi) frequency and nuclear buds (NBuds) with 20, 40, 60?ng?mL^–1, and nucleoplasmic bridges (NPBs) only with 40?ng?mL^–1. On the other hand, the three concentrations of DOR were not able to induce cytotoxic effect and DNA damage using SCGE in the bovine CC. Nevertheless, the two higher concentrations of DOR (20, 40?µg mL^–1) significantly increased the frequency of micronucleus formation in bovine CC. These results represent the first experimental evidence of genotoxic and cytotoxic effects exerted by DOR on bovine PL and CC.