【作者】 骆爱兰；

【导师】 祝树德； 刘贤进；

【作者基本信息】 扬州大学 ， 农业昆虫与害虫防治， 2004， 硕士

【摘要】 拟除虫菊酯杀虫剂是目前使用最广泛的杀虫剂之一，亦是农产品中残留农药检出率较高的药种之一。因其种类多，含量低，目前尚缺乏高效、经济、快速的筛选检测方法。另外，其污染以表面残留为主，实用消解去除相对较可行。本论文采用以广谱性抗体（generic antibody）检测多种农药残留的原理，建立了针对4种拟除虫菊酯类农药ELISA检测技术，并进行蔬菜样本检测应用。同时，论文还对青菜中残留高效氯氰菊酯农药的去除方法开展了探索性研究，取得结果如下： 1．以间苯氧基苯甲酸（PBA）作为半抗原，与牛血清蛋白偶联后免疫新西兰大白兔，获得对多种菊酯类农药有特异性的广谱性抗体。以6-氨基己酸法合成的偶联物作包被原，建立并优化了间接竞争ELISA（IC-ELISA）检测方法，对PBA最低检出限达0.2172μg·mL^-1，抑制中浓度为3.8613μg·mL^-1，且对氯菊酯、甲氰菊酯、氯氰菊酯、三氟氯氰菊酯、溴氰菊酯均有特异性识别，对氰戊菊酯识别弱，抑制中浓度分别为4.0665、4.2324、4.2867、4.8749、8.9791和77.3600μg·mL^-1。结果表明该抗体可用于上述菊酯类农药残留筛选检测。 2．以辣根过氧化物酶标记抗体，建立拟除虫菊酯的直接竞争抑制ELISA（DC-ELISA）检测方法，进一步优化分析条件并建立了一种适用于蔬菜样品中菊酯残留快速检测的规程（protocol）。该规程的最低检测限、精密度以及青菜样品的添加回收率分别为0.2462～0.7422μg·mL^-1、6.05％～10.87％和68.46％～96.10％，并与气相色谱仪（GC）分析方法有良好的相关性。采用该规程在对南京市场收取的107个样品检测中，测得样品阳性检出率为8.46％，经GC确证的假阳性率为4.72％。表明该方法对含甲氰菊酯、氯氰菊酯、三氟氯氰菊酯、溴氰菊酯等残留农药菜样的快速初筛具有良好的实用性。农药残留下降情况。结果表明:各种处理对去除青菜中残留高效氯氰菊醋均有一定作用。随着存储和紫外照射时间的延长，青菜中的高效氯氰菊酷残留逐渐减少，存储72h的消解率为28.01%，紫外照射lh的消解率为27.15%。水温的升高或碱性的增强，青菜中的高效氯氰菊酷残留降解加快，60℃的水溶液处理smin，消解率为77.99%，pHg水溶液处理lh的消解率为40.15%。洗涤剂水溶液浸泡，其残留下降亦加快，2%的洗涤剂溶液处理lh的去除率为49.04%。臭氧处理对高效氯氰菊醋残留去除作用效果较明显，处理30min去除率可达50.27%。这对农产品产后的残留去除有重要参考价值。更多还原

【Abstract】 Pyrethroids are a group of pesticides widely used in agriculture and are found easily and commonly in agricultural products. It is necessary to develop a kind of economical, effective, and rapid screening method to monitor levels of pyrethroids residue to ensure the safety of human and environment. It is feasible to remove the residue of beta-cypermethrin Which is on the leaves of vegetables. The generic antibody which was raised against 3-phenoxybenzoic acid (PBA) conjugate with Bovine Serum Albamin(BSA) was applied to detect pyrethroids simultaneously. The enzyme-linked immunosorbent assay (ELISA) protocol was developed to screen the residue of pyrethroids on the vegetables. Moreover, removing techniques for pyrethroids residueon vegetables were preliminary probed. The results were reported as follows:1. PBA was used as generic hapten in order to generate broad specificity antibody against a group of pyrethroid insecticides. An indirect competitive ELISA(IC-ELISA) was developed. Assay performance was improved by the use of a coating antigen which possessed a four carbon spacer arm between the hapten and BSA. Typical detection limits for PBA was 0.2172ng-mL-1 with an I50 value of 3.8623 g-mL-1. Different cross-reactivities with the closely related pyrethroids,including permethrin, fenpropathrin, cypermetnrin, cyhalothrin and deltamethrin, were conducted, with I50 value of 4.0665, 4.2324, 4.2867, 4.8749 and 8.9791 g-mL-1,respectivity. There was a low cross reactivity for esfenvalerate with an I50 value of 77.3600ug-mL-1. The resultindicated the antibody was practicable to determinate the residues of pyrethroids.2.A direct competitive ELISA (DC-ELISA) was established by using the highly generic antibody against PBA, which was labeled with Horseradish Peroxidase(HRP). TheDC-ELISA protocol was developed for screening pyrethroids on vegetables with the values of detection limit 0.2462~0.7422ug-mL-1, the values of precisioa6.05%~10.87%, and the recovery rate 68.46%~96.10%. The samples of 107 vegetables from Nanjing region were detected with the protocol. Compared the results with Gas Chromatogrphy(GC), the positive rate of the protocol was 8.46%, the false-positive rate was 4.72%. The preliminary application showed the protocol was a time-saving and easily-operating to screen the pyrethrbid residue on vegetables.3.The residue of beta-cypermethrin on vegetables could be removed through different approaches including storage, washing, heating, irradiating with ultraviolet wave, and treating with base or ozone. The results showed that all of them were effective. Thedegradation of the residue by storage and UV-light was a slow process, the removingrate was 28.01% with storage for 72 hour and 27.15% under UV-light for Ihour. The removal of beta-cypermethrin was quickened with the increase of temperature and value of pH, the rate was 77.99% through soaking in 60 C water for 5 minutes and 40.15% under pH11 for Ihour. The removing rate was over 30% through washing with 0.05%detergent. The rate of 50.27% was obtained with ozone for 30 minutes.更多还原