本研究開發一套具有快速、綠色化學的萃取方式檢測苯並三唑類化合物 (Benzotriazoles，簡稱BTRs) 與苯並?唑類化合物 (Benzothiazoles，簡稱BTHs) 在人體尿液中所含濃度。由於BTRs及BTHs在工業製造、農業用品及家用產品的各個方面應用越來越廣泛，但近期研究指出，BTRs及BTHs在生物系統中不僅含有潛在的危害性，且藉由生物累積的方式，易對環境及人類造成負面影響。 本研究萃取方法是以超音波輔助深共熔溶劑液液微萃取法 (Ultrasonic?assisted DES liquid?liquid microextraction，簡稱USA?DES?LLME) 萃取人體尿液中共九種BTRs及BTHs衍生物，接著利用超高效液相層析串聯電灑游離 (+) ?四極桿飛行時間質譜儀 (UHPLC?ESI(+)?QTOF?MS) 進行後續檢測。本方法中所利用的萃取劑為新興綠色化學溶劑，深共熔溶劑 (Deep Eutectic Solvents，簡稱DES)，因其具有低毒性、低成本、易回收且組合多樣性，取代以往有機溶劑的萃取方式，達到綠色分析化學的宗旨。而本研究以氯化膽鹼 (Choline Chloride) 為氫鍵接受者搭配苯酚 (Phenol) 為氫鍵供應者，以莫耳比1:2比例合成深共熔溶劑。 透過Multilevel Categoric Design 選擇實驗因子種類，以及使用Box?Behnken Design (BBD) 中的變異數分析 (Analysis of variance, ANOVA) 優化各項因子選擇實驗的最佳條件。本實驗方法最佳條件為：將1 mL的尿液經由酵素水解後，加入 0.25 mL 的萃取劑DES，經過6分鐘的超音波震盪後，加入氯化鈉 0.25 g，再以3000 rpm離心 7分鐘，取出上層的萃取液，加入甲醇至 1 mL，最後以2 ?L進樣至UHPLC?QTOF?MS中進行檢測。 利用USA?DES?LLME方式，檢測九種BTRs及BTHs待測物的定量極限 (LOQ) 為 0.4?9.0 ng/mL；Inter?day 和Inter?day 的相對標準偏差 (RSD) 介於4 %?17 % 之間，而萃取回收率則介在 90 %?106 % 之間，表示本方法呈現良好的準確性及穩定度。在人體尿液檢測結果顯示，BTRs殘留濃度介於n.d?13.5 ng/mL之間，BTHs殘留濃度介於n.d.?23 ng/mL之間。 ;Benzotriazole (BTRs) and benzothiazole (BTHs), a group of polar heterocyclic aromatic derivatives, have been classified as high production volume contaminants of emerging concern, especially since they are widely applied in household products as well as for industrial usages. High volume production may be indicative of a high level of exposure to humans and as having widespread effects on the environment. This study describes a simple and environmental?friendly method for the rapid determination of five BTRs and four BTHs derivatives in human urine. The target analytes were effectively extracted from urine samples using a deep eutectic solvent-based ultrasonic-assisted liquid-liquid microextraction (DES?USA?LLME), and their determination were performed by ultrahigh?performance liquid chromatography and electrospray ionization (+) ?quadrupole time?of?flight mass spectrometry. DESs are a group of novel “green” solvents, and their applications in sample pretreatment are appropriate for the requirements for green chemistry, environmental protection and sustainable development. To overcome the challenges related to different experimental conditions, multivariate experimental design approaches conducted by means of a multilevel categorical design and a Box-Behnken Design were applied to screen and optimize parameters that have significant influences on the extraction efficiency of DES?USA?LLME. The developed method was validated, and provided low limits of quantitation (LOQs) ranging from 0.4 to 9 ng/mL; high precisions for both repeatability and reproducibility ranging from 3 to 12 %; and high accuracy (mean extraction recovery) ranging from 90 to 106 % at two spiked concentration levels. The developed method was then successfully applied for the analysis of BTRs and BTHs in human urine samples.
