戊唑醇在蚯蚓中手性选择性生物富集和生物可利用性研究
文献类型：学位论文
| 作者 | 于丁一 |
| 学位类别 | 硕士 |
| 答辩日期 | 2012-05 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 李建中 |
| 关键词 | 手性分析 手性选择性 生物富集 戊唑醇 enantioseparation enantioselectivity bioaccumulation tebuconazole |
| 其他题名 | Study on the enantioselective bioaccumulation and bioavailability of tebuconazole in earthworm |
| 学位专业 | 环境科学 |
| 中文摘要 | 手性农药的使用与环境安全是目前研究的热点问题。手性农药在某些生物和化学过程中常有较大的手性选择性差异，研究这些手性选择性差异对农药的合理生产使用、环境风险和生态毒理的评价有重要意义。 本文研究的目的在于利用固定相高效液相色谱和毛细管电泳的分析手段建立蚯蚓中戊唑醇手性对映体的分离分析方法；并利用所建立的方法研究蚯蚓对土壤中戊唑醇的手性选择性富集规律和手性对映体的生物可利用性评价。 1. 本文建立了蚯蚓中戊唑醇手性对映体的手性分析方法。利用液液分配（LLE）和固相萃取（SPE）前处理，以高效液相色谱（HPLC）和毛细管电泳（CE）串联紫外检测器为手段进行检测。实验探讨了SPE柱的选择、HPLC流动相的比例对对映体分离的影响、CE参数的优化等。最终优化的方法为——前处理方法：正己烷与乙腈提取液萃取除脂后，弗罗里硅土SPE柱净化；HPLC方法：Chiralpak IC手性色谱柱，流速1 mL/min，检测波长220 nm，流动相为正己烷：异丙醇=90:10（V/V）；CE方法：缓冲溶液为20 mmol/L KH2PO4，10 mmol/L H3PO4（pH 3.16），添加1% S-β-CD和2 mol/L尿素，分离电压为-25 kV，检测波长214 nm。HPLC和CE两种检测方法都有很好的分离度和回收率，RS均大于2，回收率均在70--120%之间。但HPLC方法有更低的检测限（LOD）和更宽的线性范围：HPLC方法对(−)-S-戊唑醇和(+)-R-戊唑醇的LOD分别为0.011 mg/kg和0.013 mg/kg，线性范围分别为0.087--1000 mg/kg和0.098--1000 mg/kg；CE方法对(−)-S-戊唑醇和(+)-R-戊唑醇的LOD分别为0.17 mg/kg和0.13 mg/kg，线性范围分别为1.79--102.4 mg/kg和1.90--99.6 mg/kg。 2. 利用所建立的固定相高效液相色谱方法研究了土壤中蚯蚓对戊唑醇的手性选择性富集，并用生物—土壤富集因子（BSAF）评价了土壤中戊唑醇手性对映体对蚯蚓的生物可利用性差异，利用平衡分配模型（EPT）研究了实验室条件下手性对映体富集动态规律的不同。戊唑醇手性对映体在土壤中的降解行为几乎没有手性差异，36天后，低剂量组和高剂量组中戊唑醇浓度分别下降约27%和17%；在蚯蚓体内，戊唑醇的富集出现手性选择性差异，(+)-R-戊唑醇浓度均高于(−)-S-戊唑醇，且对映体比率（ER）逐渐下降，暴露结束时低剂量组和高剂量组ER值分别为0.30和0.11；(+)-R-戊唑醇的富集动态符合平衡分配模型的一级动力学规律（r2分别为0.969和0.942），而(−)-S-戊唑醇则背离了一级动力学（r2分别为0.734和0.222）。蚯蚓对土壤中戊唑醇两种手性对映体的富集的BSAF值差异较大，反应了戊唑醇手性对映体对蚯蚓的生物可利用性差异较大；(+)-R-戊唑醇在土壤—蚯蚓传递时有生物富集作用（BSAF>1 kgOC/kglip），并有可能在进一步的食物传递中产生生物放大作用，(−)-S-戊唑醇则不会产生这种生物富集和放大作用。 |
| 英文摘要 | Chiral pesticides and their environmental safety is a hot issue for the present studChiral pesticides and their environmental safety is a hot issue for the present study. Chiral pesticides often have large enantioselective differences in some biological and chemical processes. To study these differences is very important for pesticides in their rational production and use, evaluation of environmental risk and ecotoxicology. The aim of this paper was to utilize high performance liquid chromatography (HPLC) with chiral stationary phases (CSPs) and capillary electrophoresis (CE) to develop methods for separation of tebuconazole enantiomers in earthworm samples and applied the methods to study the enantioselective bioaccumulation and bioavailability of tebuconazole to earthworm. 1. Enantioseparation methods of tebuconazole in earthworm were established. Based on liquid-liquid partition (LLE) and solid phase extraction (SPE) pre-treatment, earthworm samples could be determined by both HPLC and CE tandem UV detector. Selection of SPE columns, ratio of fluid phase in HPLC and optimization of CE parameters were discussed. 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 method, Chiralpak IC was employed as chromatographic column with cellulose tris (3,5-dichlorophenylcarbamate) stationary phase immobilised; flow rate was 1 mL/min; detection wavelength was 220 nm; fluid phase was hexanes:isopropanol=90:10(V/V). For CE method, buffer solution contained 20 mmol/L KH2PO4，10 mmol/L H3PO4（pH 3.16）, 1% S-β-CD and 2 mol/L urea; separation voltage was -25 kV; detection wavelength was 214 nm. Then HPLC method and CE method were compared. Both CE and HPLC methods had excellent resolution; RS were all above 2. However, HPLC method had lower limit of detection (LOD) and broader linearity range. LODs were 0.17 mg/kg and 0.13 mg/kg for (−)-S-tebuconazole and (+)-R-tebuconazole respectively in CE; 0.011 mg/kg and 0.013 mg/kg respectively in HPLC. Linearity ranges were 1.79--102.4 mg/kg and 1.90--99.6 mg/kg for (−)-S-tebuconazole and (+)-R-tebuconazole respectively respectively in CE; 0.087--1000 mg/kg and 0.098--1000 mg/kg respectively in HPLC. 2. HPLC method that developed above was employed to study the enantioselective bioaccumulation of tebuconazole in earthworm. Biota-soil-accumulation factor (BSAF) was employed to evaluate differences of bioavailability to earthworm between tebuconazole enantiomers. Equilibrium partitioning theory (EPT) model was employed to study dynamic laws of the bioaccumulation process. Enantioselective bioaccumulation in earthworm was investigated under laboratory condition at concentrations of 10 mg/kgdwt (dry weight) and 50 mg/kgdwt in soil. During the exposure period, there was no difference of the enantioselective degradation behavior of tebuconazole in test soil; tebuconazole concentrations were reduced by approximately 27% and 17% in low dose and high dose groups respectively. Bioaccumulation of tebuconazole in earthworm tissues was enantioselective with a preferential accumulation of (+)-R-tebuconazole. ER (enantiomeric ratio) values presented downward trends during 36 days' process in both low and high dose groups; ERs were 0.30 and 0.11 in low and high dose groups respectively after the exposure.The uptake kinetics of (+)-R-tebuconazole fitted the first-order kinetics of EPT model well (r2 were 0.969 and 0.942 under the exposure condition respectively), while (−)-S-tebuconazole obeyed (r2 were 0.734 and 0.222). BSAF values of tebuconazole enantiomers were of great difference, which means bioavailability to earthworm was different between tebuconazole enantiomers. (+)-tebuconzole was demonstrated to have bioaccumulation effect in earthworm from soil, and might have biomagnification effect in further earthworm food chain(BSAF>1 kgOC/kglip); (−)-S-tebuconzole was demonstrated not to bioaccmulate or biomagnify in earthworm food chain. |
| 公开日期 | 2014-05-13 |
| 源URL | [http://ir.rcees.ac.cn/handle/311016/7229]  |
| 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 | 于丁一. 戊唑醇在蚯蚓中手性选择性生物富集和生物可利用性研究[D]. 北京. 中国科学院研究生院. 2012. |
入库方式： OAI收割
来源：生态环境研究中心
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