单 位：
有机地球化学国家重点实验室;湖南省衡南县教师进修学校;南京大学环境学院;中国科学院广州地球化学研究所;污染控制与资源化研究国家重点实验室;有机地球化学国家重点实验室 广州510640
关键词：
吡虫啉;抑食肼;黑斑蛙;蝌蚪;微核;单细胞凝胶电泳试验(又名彗星试验)
摘 要：
应用青蛙红细胞微核试验和单细胞凝胶电泳试验研究了两种新型杀虫剂 -吡虫啉和抑食肼对青蛙蝌蚪和成体的遗传毒性 ,结果表明 :当吡虫啉为 2mg/L时 ,蝌蚪红细胞微核率与对照组相比 ,无显著性差异 (p >0 .0 5) ;浓度升高到 8mg/L时 ,微核率与对照组相比 ,有显著性差异 (p <0 .0 5) ;当浓度为 3 2mg/L时 ,微核率与对照组相比 ,有极显著性差异 (p <0 .0 1) ;并有明显的剂量 -效应关系 (r =0 .9843 )。而抑食肼在浓度为 2 .5mg/L和 10mg/L时 ,微核率与对照组相比 ,无显著性差异 (p >0 .0 5) ;当浓度增至 40mg/L时 ,微核与对照组相比 ,有极显著性差异 (p <0 .0 1) ;吡虫啉与抑食肼各浓度组对青蛙红细胞的DNA损伤与阴性对照组相比 ,都有极显著性差异 (p <0 .0 1) ,且具有明显的剂量 -效应关系 (r =0 .960 ,r=0 .990 )。
译 名：
GENOTOXICOLOGICAL STUDIES OF TWO PESTICIDES TO TADPOLES AND FROGS OF RANA NIGRINACULATA HALLOWEII BY MICRONUCLEI TEST AND SINGLE CELL GEL ELECTROPHORESIS ASSAY
作 者：
FENG Shao-Long 1,2 ,KONG Zhi-Ming 2, WANG Wu-Xiang 3,WANG Xin-Ming 1 and PENG Ping-An 1 (1.The State Key Laboratory of Organic Geochemistry Chinese Academy of Science, Guangzhou Institute of Geochemistry,Guangzhou510640; 2.The State Key Laboratory of Pollution Control and Resource Reuse,the School of Environment, Nanjing University, Nanjing210093; 3.The Refresher School of T eacher in Hennaan county, Hennan421141)
关键词：
Imidacloprid;RH-5849;Rana nigronaculata Ha ll owell; Tadpole; Micronuclei;Single cell gel electrophoresis assay (comet assay)
摘 要：
Imidacloprid [1-(6-chloro-3-pyridylmethyl)-N-n itro-imidazolidin-2-ylid eneamine] and RH-5849[2′-benzoyll′-tert-butylbenzoylhydrazinel]are two novel pesticides being used in China. Imidacloprid, which act as an agonist at t he nicotinic acetylcholine receptor,is highly effective against many sucking insec t s including ricehoppers, aphids, thrips and white flies. RH-5849, a nonsteroida l ecdysone agonist, which act similar to 20-hydroxyecdysone by binding to the e cdosone receptor, have been found to be very effective against lepidopteran pest s in vegetables, cotton, and cereals. To our knowledge, their effects on the aqu atic and agricultural ecosystems have not been fully investigated. Amphibians ar e important organisms in the aquatic and agricultural ecosystems; they are among the most important natural enemies of many agricultural pests. Because of their sensitivity to changes of their habiat and that their larvae live in the aquat ic environment, the amphibians were regarded as bio-indicators of aquatic and a gricultural ecosystems, and broadly used as typical test organisms in evaluating the effects of chemicals on the aquatic and agricultural ecosystems. This study was initiated to combine micronucleus test (MN) and comet assay to assess the g enotoxicity of the two pesticides on the amphibians from different endpoints. Th e objective was to achieve a more comprehensive understanding of the effects and potential risks of the pesticides on the aquatic and agricultural ecosystems. Amphibians,Rana nigronaculata Hallowellwas selected as the test organisms. T hey were acclimated to the conditions of the laboratory for 7 days before the te s ts. The tadpoles were one and half months old, with body lengths of 37.5±1.1mm and body weights of 461±60mg and the frogs with body weights of about 100 g we re chosen for the tests. After being tamed for 7 days in the laboratory, the tadpoles ofR. nigronacula t awere exposed to different levels of the two pesticides for 7 days. The concen tration of DO in the solutions was maintained at no less than 8.5 mg/L with bubb le aerator, and the temperature of solutions was controlled at 20±1℃, and the solutions were replaced with freshly prepared solution of the same concentration every 24hr. After the seven-day exposure, blood was taken from each of the tad poles by cardiac puncture and one smear was prepared per animal. Fixed in methan ol and stained with 5% of Giemsa in Sorensen buffer (pH 6.98),the smears were sc reened under a microscope. The erythrocytes with one or more micronuclei were co nuted for a total of at least 2000 erythrocytes per tadpole and seven random an imals were screened in each group. The procedure of comet assay was basically the same as that described by Singh e t al.Modifications due to the uniqueness of the biological material studied and due to the equipment available were relatively minor. Blood samples were collect ed fromR. nigronaculata frogs by decapitation followed by immediately placi ng the animals in a 10% solution of Hanks balanced salt solution. The survival r a te of erythrocytes after the isolation was higher than 95% as examined by trypan blue exclusion test. After 1hr of exposure at 20℃ in a 5% CO 2 atmosphere, t he cells were collected by centrifugation (10 min. 3000rpm, at 4℃)and washed tw ice with Hanks balanced salt solution (at 4℃) to minimize possible damage rep air. Samples were immediately placed on ice for comet assay. After lysing, elect rophoresis and neutralization, the slides were stained with ethidium bromide(EB ).For evaluation of DNA damage,100 cells per slide were analyzed at 400×ma gnif ication under a “TMD-EF" fluorescent microscope (Nikon, Japan). The cells were scored visually and given scores 0 (undamaged),1,2,3 or 4(maximally damaged) acc ording to tail intensity (size and shape).Thus, the total score for 100 comets r anges from 0 (all undamaged) to 400 (all maximally damage). The percentage of da maged cells was calculated and the results analyzed with theχ 2 test. The “AUs” was used to express t
