| 摘要: | 近年來在環境檢測方面，科學家對於新興污染物(emerging contaminants, ECs)已產生 濃厚的興趣與關注。新興污染物定義為“新認定”或“之前未定確認”的污染物，包含日常 生活用品如：界面活性劑及其殘留物、個人護理用品、汽油添加劑、塑化劑及各種工業 添加劑等。由於新興污染物的蹤跡、風險評估及其生物毒性資訊均難以獲得，對於人體 及水中生物體所造成的影響亦是難以預估的。而造成資訊難以獲得的主要因素之一則是 因為欠缺適當的微量方析分法，以致無法對於環境中新興污染物進行有效的監測與評 估。由於環境樣品中基質複雜且濃度微量，偵測此類新興污染物需以最適當分析方法檢 測，並搭配有效的前處理方式將待測物濃縮淨化，以避免在偵測時產生干擾。常用檢測 方式是以層析方法搭配質譜儀以達到分離及質量與結構鑑定之定性與定量目的。本研究 將以液相層析串聯質譜技術(liquid chromatography/tandem mass spectrometry, LC-MS-MS) 在微量分析上的方法開法與應用，並根據待測物的極性程度及酸鹼性質找尋適當的游離 源，結合串聯質譜技術 (MS-MS) 對待測物進行結構鑑定並有效提升靈敏度。 本研究首先針對四個苯二乙烯類及一個二苯乙烯聯苯類之強離子性螢光增白劑 (fluorescent whitening agents, FWAs)，發展出一套穩定且可靠的微量檢測技術，藉由探討 兩種不同固相萃取管柱對回收率及離子抑制效應之影響，尋找出一套較佳之樣品前處理 方法，偵測水環境中之螢光增白劑含量。利用揮發性離子配對-液相層析，以電灑游離 法(electrospray ionization, ESI)結合離子阱二次質譜技術對待測物在複雜基質中進行定 性以確定分析物結構，並選定特徵斷裂離子作為定量，藉此亦可提高靈敏度及降低偵測 極限。在最佳之樣品前處理方法搭配層析分離方法與最佳之游離條件下，偵測極限可達 4 – 18 ng/L，檢量線r2值均大於0.995。在分析方法之精密度與準確度分別小於13%及 11%，且平均回收率為68% – 97%。將此方法應用至廢污水處理廠放流水與河流水樣中， 皆可偵測到五種螢光增白劑的存在，濃度為n.d. – 1580 ng/L。 藉由上述方法之開發，進而將樣品類別延伸至餐巾紙、嬰幼兒衣物等商業化產品 上。樣品先經由熱水萃取法(hot-water extraction)萃取固體樣品中之螢光增白劑，將萃取 濾液以Oasis WAX固相萃取管柱萃取，再以揮發性離子配對LC-MS-MS 檢測之。回收 率為42 – 95%，相對標準偏差均小於11%。初步結果顯示螢光增白劑在餐巾紙中的含量 為n.d. – 4.2 mg/g、在衛生紙中含量為n.d. – 7.0 ng/g、在嬰幼兒衣物中之含量為n.d. – 118 mg/g。檢測出之主要物質為兩種苯二乙烯類螢光增白劑。 由於研究強陰離子型化合物之概念已成型，因此在研究的第三部份，則是探討弱酸 鹼性等極性化合物的檢測方法開發，在此方法中選擇新興污染物中漸受重視的藥物殘留 物(pharmaceuticals residues)作為研究對象。此部份主要是發展一套快速、可靠且穩定的 分析方法，研究這類藥物殘留物在台灣水環境中的流布與含量。藉由最佳的層析分離與 游離化條件，搭配良好的固相萃取方法(solid-phase extraction, SPE)，可達到濃縮及移除 干擾物質之目的。實驗結果顯示LC流速設定為0.2 mL/min，搭配ESI並結合MS-MS 可得到較佳的感度。其偵測極限可達0.5 – 20 ng/L。利用此方法針對民生廢污水及河流 水樣進行檢測，發現在民生廢污水及河流水樣均可測得藥物殘留物的存在，且最高濃度 可達960 ng/L。 在本研究的最後部份則是針對非離子性的雌激素化合物(estrogenic compounds)以三 種不同離子源游離偵測，探討游離效率及基質干擾在分析含有複雜基質的真實樣品過程 中所造成的影響，試以較適當的離子源搭配MS-MS，降低基質所造成的離子抑制現象。 研究結果顯示，ESI及摻雜劑輔助之大氣壓力光游離法(dopant-assisted atmospheric pressure photoionzation, DA-APPI)對雌激素化合物具較好的游離效率及感度。以去鹽萃 取法(de-salting extraction)結合SPE，針對含有複雜基質的廢污水樣進行樣品前處理，再 以LC-MS-MS搭配ESI及DA-APPI，探討基質干擾所造成之離子抑制現象，並利用此 兩種游離法之互補的特性，以補足單一游離法在複雜基質水樣中雌激素化合物的定量。 ESI的平均離子抑制因子為52 ± 5%；而DA-APPI之平均離子抑制因子為27 ± 7%。在 LC-ESI-MS-MS 的系統中，當水樣為含有複雜基質之廢污水進流水(influent)時，其感度 與游離效率均明顯降低。廢污水進流水與放流水中均可測得雌激素化合物的存在，在進 流水中之濃度為n.d. – 230 ng/L，而在放流水中之濃度為 n.d. – 28.5 ng/L。此部份的研究 結果顯示，利用具有較低偵測極限之ESI方法輔以具有較高基質容忍度之DA-APPI游 離技術，對於複雜基質中之雌激素化合物之定量可達相輔相成的作用。 由本論文中之結果可得知新興污染物的檢測不易，尤其在利用LC-MS 時，具有不 同官能基性質之新興污染物對質譜游離方式亦有不同的感度與限制，因此檢測方法的開 發與最佳化需要投入更多的研究才能獲得高品質的結果。新興污染物除了存在於環境 中，亦存在於我們的日常生活。此類新興污染物之存在主要都是經由不當的使用及排放 所造成，因此希望藉由此研究結果，有助於提升國人的環保意識，建立適當的使用及排 放標準，以減輕人類對環境所造成的負擔。 In recent years, the development of new sensitive methods for detecting chemicals and determining biological effects of emerging contaminants (ECs) has shifted attention of the scientific community toward new undetected unregulated contaminants or those that had not been considered as a risk. Notably, ECs, which are defined as newly identified or previously unrecognized pollutants, primarily comprise products utilized in large quantities in everyday life such as fluorescent whitening agents (FWAs), human and veterinary pharmaceuticals, personal care products, surfactants and surfactant residues, plasticizers and various industrial additives. However, little information of these chemicals is available, and it is difficult to predict the impacts on ecobiological environment, public health and safety. The reason of a lack of information for ECs is a lack of analytical methods for determining trace concentrations. Analyzing ECs is extremely challenging, not only due to the diversity of chemical properties of ECs, but also because of the typically low concentrations and matrix complexities. Novel analytical methodologies for ECs containing different polarities and functional groups are presented in this thesis that detected EC content in environmental samples. The first part of this thesis developed and validated a comprehensive method for determining trace amounts of five ionic FWAs in environmental water samples. Various solid-phase extraction cartridges were evaluated regarding recoveries, abundance and ion suppression effect. The new Oasis WAX-SPE cartridge generates optimal sample extraction results. Analytes were then identified and quantified by liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS) in negative ionization mode, applying di-n-hexylammonium acetate (DHAA) as the ion-pairing reagent in mobile phases. Qualification with the MS-MS spectral comparison would confirm the presence of FWAs, and selectivity and sensitivity can be improved using product ion scan technique. Limits of quantification (LOQs) were established at 4 – 18 ng/L in 50 mL water samples. Intrabatch and interbatch precision at two concentration levels was also investigated. Precision for these five FWAs, as indicated by relative standard deviation (RSD), was <13 and 11% for the intra- and interbatch, respectively. Accuracy, expressed as mean recovery, was 68 – 97%. The proposed method was finally applied to environmental water samples. The concentrations of selected FWAs in water samples were n.d. – 1580 ng/L. Preliminary results demonstrated that five FWAs existed in river water and wastewater treatment plant (WWTP) effluent samples. In the second part, the proposed method for trace determination of five FWAs in environmental water samples was further applied to paper materials and infant clothing. The FWAs were extracted from paper material and cloth samples using hot-water extraction; aqueous extracts were then effectively extracted with the Oasis WAX-SPE cartridge. The analytes were accurately determined by ion-pair chromatography combined with negative LC-ESI-MS-MS. The LOQs were 0.2 – 0.9 ng/g in 2g samples. Recovery of the five FWAs in spiked commercial samples was 42 – 95% and %RSD (n = 3) was 2 – 11%. The proposed method was finally applied to commercial samples, and the preliminary results revealed that concentration of FWAs was n.d. – 4.2 mg/g in napkins, n.d. –7.0 ng/g in tissues, and n.d. – 118 mg/g in infant clothes. The results also indicated that two stilbene-type disulfonates were predominant in FWAs detected in napkin and infant clothing samples. Because the analyses of ionic substances were well-developed in the 1st and 2nd parts, the investigation of the third part focused on the occurrence and levels of selected acidic and neutral pharmaceutical residues in Taiwanese river and wastewater samples. These pharmaceutical residues were extracted from water samples using Oasis HLB-SPE. The analytes were then identified and quantified using LC-MS-MS at a flow-rate of 0.2 mL/min with dual-polarity ESI. The LOQs were 0.5 – 20 ng/L for 250 mL water samples. The selected analytes were detected at concentrations of <0.5 – 960 ng/L in WWTP effluent and river water samples. Finally, three complementary LC-MS ionization methods, ESI, atmospheric pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI) were evaluated regarding the ion suppression effect in real samples containing complex matrix and optimized to identify five neutral estrogens in influent and effluent of WWTPs. Wastewater samples were first subjected to SPE coupled with desalting extraction to eliminate matrix interference. As the abundances of ESI and dopant-assisted APPI (DA-APPI) were better than APCI, both ionization methods with LC-MS-MS were utilized to detect five estrogenic compounds to evaluate the ion suppression effect and complement detection and quantification of estrogenic compounds in complex wastewater samples. Average ion suppression factors for extracts from the WWTP influent—analyzed using ESI and APPI—were 52 ± 5% and 27 ± 7%, respectively. Sensitivity and ionization efficiency of the LC-ESI-MS-MS system decreased dramatically when a complicated matrix existed in the WWTP influent sample. Estrogenic compounds were detected in WWTP influent and effluent samples at concentrations of n.d. – 230 ng/L and n.d. – 28.5 ng/L, respectively. The lower detection limits obtained when using ESI and the high matrix tolerance of the APPI method facilitated complete quantification of estrogenic compounds in extremely complex samples in a complementary manner. The dissertation reveals that the ionization efficiency and limitations vary due to diverse chemical properties and different functional groups of ECs as LC-MS is utilized. As difficulties are encountered during method development, more investigations are needed to ensure the quality of developed and optimized analyzing methods. The preliminary results of this dissertation also demonstrate that these ECs present not only in environment, but also in our daily life because of improper use and discharge. Since little information exists regarding the occurrence of FWAs, pharmaceuticals and estrogenic compounds in aquatic environments in Taiwan, this study is significant in that it provides information regarding the contents and fate of selected ECs in Taiwanese environments. Moreover, this study promotes environmental protection and conservation, and support pollution-control policies and sustainable development in Taiwan. |
