臺灣博碩士論文加值系統

合成除蟲菊農藥(Pyrethroids)經常使用於農、林、園藝業及一般環境衛生用藥，除蟲菊農藥具有神經毒性，當中毒發生時可能會導致癱瘓甚至死亡，已有相關研究報導指出除蟲菊農藥可能具有基因毒性。隨著合成除蟲菊農藥在一般家庭中使用量及頻率的增加，除蟲菊農藥在一般人口的暴露型態及內在暴露劑量可能較過去增加，如何對人體除蟲菊農藥暴露進行有效的檢測評估已成一重要之課題。過去除蟲菊農藥尿中代謝物分析方法，僅針對部分代謝物進行探討，並未能針對全面性除蟲菊農藥之暴露進行量測，且樣本前處理方式較為繁瑣費時。
本研究利用薄膜型強陰離子交換樹脂 (strong anion exchange, SAX)暨氣相層析質譜儀 (GC/MS)建立一省時且操作簡單並能同時偵測尿中五種除蟲菊農藥生物指標之分析方法。前處理方式為利用薄膜型強陰離子交換樹脂對尿中五種常見除蟲菊代謝物3-(2,2-dimethyl)-2,2-dimethyl-(1-cyclopropane)-carboxylic acid (簡稱ChCA)、3-(2,2-dichlorovinyl)-2,2-dimethyl-(1-cyclopropane)-carboxylic acid (簡稱Cl2CA)、3-(2,2-dibromorovinyl)-2,2-dimethyl-(1-cyclopropane)-carboxylic acid (簡稱Br2CA)、3-phenoxybenzoic acid (簡稱3PBA)及3-(4-hydroxy)-phenoxybenzoic acid (簡稱4OHPBA)進行萃取、過濾、濃縮於一步驟，吹氮後加入CH3I加熱進行線上衍生化，前處理完畢之樣本以氣相層析質譜儀進行分析。
本分析方法可將樣本前處理時間由4-24小時減少為2.5小時，儀器分析時間則由1小時縮短為17分鐘，較以往文獻分析方法大幅減少。大鼠灌食約3mg cypermethrin後尿液檢體測試時，發現加入30μL 10N NaOH(aq)能有效去除代謝酸conjugation，並可降低尿液基值干擾。各代謝酸的方法偵測下限介於0.32~5.52μg/L。以此方法分析除蟲菊農藥噴灑員工(n=22)及無職 業暴露除蟲菊農藥一般民眾(n=20)尿中除蟲菊代謝物含量，經統計分析兩組尿中除蟲菊代謝物含量差異達統計上顯著差異(P<0.05)。
本分析方法不但能有效區別職業性或非職業性暴露除蟲菊農藥其尿中代謝物含量，並且可大幅縮短樣本前處理時間及儀器分析時間，為一經濟、省時且操作簡單並能同時偵測尿中五種除蟲菊農藥代謝物之分析方法。相較於文獻上其他合成除蟲菊農藥之分析方法，本方法更適於例行性大量樣本分析應用，對人體各種除蟲菊農藥暴露進行較為廣泛的檢測評估。
關鍵字：除蟲菊農藥、生物指標、強陰離子交換樹脂、分析方法、固相萃取

Pyrethroids have been extensively used as insecticides in agriculture, forestry, horticulture, and residential hygiene. Pyrethroids have been documented as a neurotoxicant as well as a possible genotoxicant in literature. The exposure profile and total body burden of pyrethroids in population draw more and more attention due to their increasing use. The analytical methods developed in the past, however, only targeted on some metabolites instead of full-spectrum application for widely diversified pyrethroids. Moreover, the procedures in previous methods were considerably complicated and time-consuming.
The purpose of this study is to establish a more timesaving and simpler analytical method incorporating strong anion exchange (SAX) / in-vail derivatization with gas chromatography/mass spectrometry (GC/MS) in the determination of five urinary pyrethroid biomarkers at the same time. SAX was used in extraction, percolation and enrichment in a single step for the following five urinary pyrethroid metabolites: 3-(2,2-dimethyl)-2,2-dimethyl- (1-cyclopropane)-carboxylic acid (ChCA), cis/trans 3-(2,2-dichlorovinyl)-2,2-dimethyl- (1-cyclopropane)-carboxylic acid (cis/trans Cl2CA), 3-(2,2-dibromorovinyl)-2,2-dimethyl- (1-cyclopropane)-carboxylic acid (Br2CA), 3-phenoxybenzoic acid (3PBA), and 3-(4-hydroxy)-phenoxybenzoic acid (4OHPBA). After nitrogen blow, CH3I was added and in-vial derivatization was achieved following by heating prior to GC/MS determination.
We found the method developed in this study reduced the sample preparation time to 2.5 hrs from 4-24 hrs and the chromatographic course to 17 min from 1 hr. The addition of 30μL of NaOH (aq) at 10N to the urine collected from a SD rat after oral exposure to cypermethrin at 3mg was able to effectively improve the recovery rates of biomarkers of interest by possibly breaking down the conjugated forms into free forms as well as eliminate the chromatographic interference. The limits of detection for five pyrethroid metabolites were ranged from 0.32 to 5.52 μg/L. Statistical significance was found in the determination of five pyrethroid metabolites for the urine samples collected from pesticide sprayers (n=22) and from unexposed population (n=20).
This method not only can greatly simplify sample preparation procedure but also significantly reduce the consumption of toxic solvents in sample preparation as well as covering more diversified pyrethroids. This method could be more suitable in routine analysis and for screening purpose in the determination of pyrethroid exposure for general population as well as for occupationally exposed groups.
Key words: Pyrethroids, biomarkers, SAX, analytical method, solid phase extraction

摘 要.............................................................I
Abstract............................................................II
誌 謝...........................................................III
目 錄............................................................IV
表 目 錄............................................................VI
圖 目 錄...........................................................VII
第一章 緒 論.....................................................1
1-1 前言.............................................................1
1-2 研究背景.........................................................2
1-2.1 合成除蟲菊農藥之演化...........................................2
1-2.2 合成除蟲菊農藥之使用...........................................2
1-2.3 合成除蟲菊農藥毒性.............................................3
1-2.4 合成除蟲菊農藥之暴露評估.......................................4
1-2.5 合成除蟲菊農藥的生物偵測.......................................4
1-2.6 合成除蟲菊農藥尿中代謝物.......................................5
1-3 研究目的.........................................................6
第二章 文獻回顧...................................................7
2-1 尿中合成除蟲菊農藥代謝物分析方法.................................7
2-2 生物指標的選擇...................................................7
2-3 樣本萃取方式.....................................................8
2-3.1 溶劑萃取 (Liquid-liquid extraction)............................8
2-3.2 固相萃取 (Solid phase extraction, SPE).........................9
2-4 衍生化方式......................................................10
2-5 儀器分析方析....................................................10
第三章 材料與方法................................................12
3-1 實驗材料與設備..................................................12
3-1.1 藥品與材料....................................................12
3-1.2 儀器及設備....................................................12
3-2 實驗室測試分析..................................................14
3-2.1 初步實驗流程..................................................14
3-2.2 前處理與分析條件最佳化測試....................................14
3-2.3 實驗室QA/QC測試...............................................16
3-3 尿液樣本分析....................................................20
3-3.1 研究對象選取..................................................20
3-3.2 尿液檢體收取..................................................20
3-3.3 問卷調查......................................................20
3-3.4 檢體儲存、運送及處理..........................................21
第四章 結果與討論................................................22
4-1 實驗室測試結果..................................................22
4-1.1 實驗流程測試..................................................22
4-1.2 實驗室QA/QC測試...............................................25
4-2 職業與非職業除蟲菊農藥暴露族群之尿液樣本分析結果................27
第五章 結 論....................................................31
第六章 未來工作..................................................32
參考文獻............................................................33
附 錄 一............................................................67

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