题名 | 氟环唑和腈菌唑手性对映体在黄粉虫幼虫体内生物富集和环境风险评价 |
作者 | 吕笑天 |
学位类别 | 硕士 |
答辩日期 | 2014-05 |
授予单位 | 中国科学院研究生院 |
授予地点 | 北京 |
导师 | 郭宝元 |
关键词 | 手性农药 立体选择性 生物富集 动力学 风险评价 Chiral pesticides stereoselectivity bioaccumulation dynamics risk assessment |
其他题名 | Enantioselective Bioaccumulation and Environment risk assessment of Epoxiconazole and Myclobutanil in Tenebrio molitor Larvae |
学位专业 | 环境工程 |
中文摘要 | 随着立体化学的发展,农药的研究也已经深入到分子立体异构领域。生产、利用和研究单一的或者经过富集纯化的具有靶标活性的农药手性对映体成为目前绿色农药化学发展的重要机遇和发展方向。因此,为完善手性农药的风险评价体系,需要在对映体水平上研究手性农药通过食物链富集和排泄的相关规律。本文研究的目的在于利用高效液相色谱-质谱联用的分析手段建立黄粉虫幼虫中氟环唑和腈菌唑手性对映体的分离分析方法,并利用所建立的方法研究,黄粉虫对食物中氟环唑和腈菌唑的手性选择性富集和排泄规律,评价其环境风险。 1.本文在实验室内喂养驯化黄粉虫幼虫后,建立了幼虫体内腈菌唑和氟环唑手性对映体的分析方法。利用液液分配(LLE)和固相萃取(SPE)前处理,以高效液相色谱(HPLC)和质谱(MS)串联为手段检测。优化得到的方法为——前处理方法:样品低温冷冻,乙腈提取,乙腈和正己烷液液分配,SPE预淋洗、上样、洗杂和洗脱,定容待测;HPLC-MS方法:手性分离:固定相 ChiralcelOD-3R,流动相:A(0.05%甲酸超纯水),B乙腈40:60(V/V), 流速 0.25 mL·min-1 MS 检测:正离子模式该方法回收率在 70-110%之间,对氟环唑和腈菌唑的对映体的检测限(LOD)分别为 0.001 mg/kg和 0.002 mg/kg。 2.利用所建立的分析方法研究了黄粉虫分别从麦麸中富集和排泄氟环唑与腈菌唑的规律,并用生物富集系数(BAF)评价它们对黄粉虫幼虫的生物富集和生物可利用能力,以对映体比值( ER)评价对映体的差异。在实验室条件下,对 2 mg/kg和 20 mg/kg两组暴露浓度下,21天内幼虫体内氟环唑和腈菌唑手性对映体的变化进行测定。实验结果表明,在此暴露期间内,氟环唑和腈菌唑手性对映体在麦麸中均未发生明显降解,而在幼虫体内,高浓度组中出现了选择性富集的现象,低浓度组中则未有明显选择性趋势出现。同时,幼虫排泄物中浓度大大高于幼虫体内测得浓度,表现出对药物迅速而强烈的排泄,且排泄物中 ER值基本维持为 1,说明药物排泄为被动运输过程,不表现对映体选择性。BAF值计算值均远小于 1,幼虫对氟环唑和腈菌唑的富集能力较为微弱,与排泄物中数据相互吻合。 3.对腈菌唑对映体在黄粉虫体内浓度变化建立一级动力学模型进行刻画,通过模型的数学求解与软件拟合结果发现,高浓度组中腈菌唑对映体对模型吻合程度较高(r2分别为 0.702和 0.729),而低浓度组的浓度数据吻合度较低。通过对高浓度组中模型相关速率常数的进一步计算得出,两个对映体置换转化的速率常数不同(k12/k21≠1),可能是由于酶促反应的优先选择引起。 |
英文摘要 | With the development of stereochemistry, pesticide research has also been widen into the field of molecular stereoisomerism. The production, utilization and research of single or enriched and purificated chiral enantiomers with target activity of pesticides have become the important direction of the development of green chemical pesticides. Chiral pesticides not only present selective differences when they act with the target and non-target organisms, they also have potential chiral selectivity when they are involved in biological behaviors such as soil degradation, transformation and bioaccumulation in the food chain. Therefore, in order to improve the chiral pesticide risk assessment system, study of accumulation and excretion of .chiral pesticides on the level of enantiomers is needed. In this work, the differences in bioaccumulation behavior of stereoisomers of epoxiconazole and myclobutanil in Tenebrio molitor L at two dose levels through dietary exposure under laboratory conditions by HPLC-MS/MS were investigated. A method for extraction, cleaning and chiral determination of the residues of epoxiconazole in the tissue of the larvae was developed and the primary aim of this experiment was to study the stereoselectivity in the bioaccumulation and excretion of epoxiconazole and myclobutanil in Tenebrio molitor L. 1. In this paper, an analysis method was built to detect the enantiomers in the larvae after the acclimation of them under laboratory conditions. Liquid-liquid distribution (LLE) and solid phase extraction (SPE) were used with high performance liquid chromatography (HPLC) and mass spectrometry (MS) as detection instruments. The final optimized conditions were as follows. For pre-treatment methods, acetonitril extracted solution was defatted with hexane through LLE followed by Florisil SPE clearup. For HPLC-MS/MS method, ChiralcelOD-3R [cellulosetris-tris-(3,5-dichlorophenyl-carbamate)] was employed as chromatographic column; The separation used solvent A (0.05% formal acid in ultrapure water) and solvent B(acetonitrile) in a 40:60 (V/V) ratio and 0.25 mL/min flow rate; TSQ QUANTUM ACCESS MAX was used for LC-MS/MS analysis (Thermo Electron Corporation, Hopkinson, MA). Quantification was achieved in positive-ion mode (ESI+). 2. HPLC-MS/MS analytical method that developed above was employed to study the enantioselective bioaccumulation and excretion of epoxiconazole and myclobutanil in the larvae. Bioaccumulation factor (BAF) was employed to evaluate differences of bioavailability to the larvae between enantiomers and enantiomer ratio (ER) was employed to evaluation the differences of enantiomers.Under laboratory conditions, the wheat bran fed to Tenebrio molitor larvae was spiked with racemic myclobutanil at two dose levels of 20 mg/kg and 2 mg/kg (dry weight). The results showed that there was a significant trend of enantioselective bioaccumulation in the larvae with a preferential accumulation of (-)- myclobutanil and (-)-epoxiconazole in 20 mg/kg dosage exposure, but it was not obviously observed in 2 mg/kg dosage group. Results of excretion indicated an active excretion is an important pathway for the larvae to eliminate epoxiconazole which was a passive transport process with non stereoselectivity. The faster elimination might be the reason for the low accumulation of epoxiconazole, as measured by bioaccumulation factor (BAF). 3. A kinetic model considering enantiomerization between the two enantiomers of myclobutanil based on first-order reactions was built and the rate constants were estimated to discuss the kinetic reason for the different concentrations of individual enantiomers in the larvae. The data of 2 mg/kg treatment group did not obey the model well with the low correlation coefficients and just the rate constants in 20 mg/kg treatment group were calculated applying the estimated values obtained from the regression results (r2=0.702 and 0.729 respectively). The approximations implied an inversion between the two enantiomers with a relative higher rate of the inversion from (-)- myclobutanil to (+)-myclobutanil and the differences in inversion rates might be caused by enzyme-catalyzed reactions. |
公开日期 | 2015-07-08 |
内容类型 | 学位论文 |
源URL | [http://ir.rcees.ac.cn/handle/311016/15626] |
专题 | 生态环境研究中心_中国科学院环境生物技术重点实验室 |
推荐引用方式 GB/T 7714 | 吕笑天. 氟环唑和腈菌唑手性对映体在黄粉虫幼虫体内生物富集和环境风险评价[D]. 北京. 中国科学院研究生院. 2014. |
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