以雙重震盪搭配超音波輔助基質固相分散萃取法快速檢測魚體中 BTRs 與 BTs 的殘留

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陳建豪(Chien-Hao Chen) 查詢紙本館藏

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以雙重震盪搭配超音波輔助基質固相分散萃取法快速檢測魚體中 BTRs 與 BTs 的殘留

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摘要(中)

苯并三唑類 (Benzotriazoles, BTRs) 與苯并噻唑類 (Benzothiazoles, BTs) 被歸納為高產量的新興汙染物，近年來廣泛使用於工業生產與家庭用品中，其對環境有高暴露的風險。當進入水環境體系，有可能透過食物鏈生物放大效應累積於動物體內，進而危害人類健康。因此有必要開發出一套快速又方便的檢驗方法來檢測生物檢體中 BTRs 與 BTs 的殘留。
本研究前處理方法使用新開發的雙重震盪搭配超音波輔助基質固相分散 (Double vortex-ultrasound assisted matrix solid-phase dispersion ，簡稱 DVUA-MSPD) 萃取法，分析儀器使用超高效液相層析搭配電噴灑串聯飛行時間質譜儀 (UHPLC-ESI-qTOF-MS) ，進行定性與定量檢測。
本研究使用實驗設計中的 Multilevel categoric design 與 Box-Behnken design 搭配變異數分析 (Analysis of variance, ANOVA) 進行 DVUA-MSPD 萃取法的最佳化條件探討。萃取條件篩選與最佳化的結果為：取 0.5 g 金目鱸魚魚體粉末與 1 g 矽酸鎂震盪 1 分鐘，加入 10 mL 超純水，在 63 ℃ 下超音波 10 分鐘，接著加入 10 mL 乙酸乙酯與 1.3 g 無水硫酸鈉，震盪 10 分鐘後，以 2000 rpm 離心 10 分鐘。取出 5 mL 上層清液，以 13 mm 0.22 µm PTFE 疏水過濾頭進行過濾，吹氮濃縮到乾，並且以 50 μL 的甲醇回溶，最後進樣 2 µL 到 UHPLC-ESI-qTOF-MS 進行分析。
5 種 BTRs 與 2 種 BTs 的方法定量極限介於 0.15-2 ng/g ；在 5-500 ng/g 的濃度範圍內，檢量線 R2 值都在 0.9964 以上，呈現良好的線性關係；在 Intra-day 與 Inter-day 的萃取回收率介於 70 到 93 % ，相對標準偏差都小於 10 % ，表示此方法有良好的準確度與精密度。在四種市售魚體樣品中都有檢測到 BTRs 與 BTs 不同濃度的殘留，其中 BTRs 殘留的濃度介於 n.d.-71.43 ng/g ， BTs 殘留的濃度則介於 n.d.-26.07 ng/g 。

摘要(英)

Benzotriazoles (BTRs) and benzothiazoles (BTs) are classified as high production volume contaminants of emerging concern with a broad range of applications in industry and household products in recent years. High production may indicate high exposure to environment. When they enter the aquatic ecosystems, they have potential to be accumulated in animals via biomagnification in the food chain and further jeopardize human health. Hence it is necessary to develop a fast and convenient method to determine BTRs and BTs in biota samples.
This study used a newly developed pretreatment method, called double vortex-ultrasound assisted matrix solid-phase dispersion (DVUA-MSPD) extraction, coupled with UHPLC-ESI-qTOF-MS for qualitative and quantitative detection.
The parameters of DVUA-MSPD extraction were screened by multilevel categoric design firstly, and then optimized by Box-Behnken design relied on analysis of variance (ANOVA). The screening and optimal conditions of DVUA-MSPD were: weighting 0.5 g fish powder and 1 g florisil into a glass centrifuge tube, homogenized by mechanical vortex-blending for 1 minute. Ultrapure water (10 mL) was added and sonicated in an ultrasonic bath for 10 minutes at 63 ℃. Then, 10 mL ethyl acetate and 1.3 g sodium sulfate anhydrate were added and vortexed for 10 minutes followed by centrifugation at 2000 rpm for 10 minutes. After that, a 5 mL aliquot of the supernatant was collected and filtered through a 0.22 μm PTFE hydrophobic syringe filter. Dried under nitrogen and then reconstituted with 50 μL methanol. Finally, a 2 μL aliquot was injected into the UHPLC-ESI- qTOF-MS for analysis.
The method quantification limits of five BTRs and two BTs ranged from 0.15 to 2 ng/g. Excellent linearities (coefficient of determination (R2) greater than 0.9964) were found for each target analyte in the six-level calibration standard curves, and the ranges were 5-500 ng/g. High precisions for both intra- and inter-day analysis were all below 10 %. High trueness (or mean extraction recovery) varied from 70 to 93%. The DVUA-MSPD combined with UHPLC-qTOF-MS was successfully applied to determine BTRs and BTs in marketed fish samples, and the total concentrations ranged from n.d. to 71.43 ng/g for BTRs, and from n.d. to 26.07 ng/g for BTs.

關鍵字(中)

★ 苯并三唑類
★ 苯并噻唑類
★ 生物檢體分析
★ 雙重震盪搭配超音波輔助基質固相分散萃取法
★ 超高效液相層析搭配電噴灑串聯飛行時間質譜儀

關鍵字(英)

★ Benzotriazoles
★ Benzothiazoles
★ Biota analysis
★ DVUA-MSPD
★ UHPLC-ESI-qTOF-MS

論文目次

摘要 i
Abstract iii
謝誌 v
目錄 vii
圖目錄 xi
表目錄 xiii
第一章前言 1
1-1 新興汙染物 1
1-2 研究目標 3
第二章文獻回顧 5
2-1 苯并三唑類 (Benzotriazoles, BTRs) 5
2-1-1 結構與性質 5
2-1-2 用途 7
2-1-3 毒性探討 7
2-1-4 環流分布 11
2-2 苯并噻唑類 (Benzothiazoles, BTs) 11
2-2-1 結構與性質 11
2-2-2 用途 12
2-2-3 毒性探討 13
2-2-4 環流分布 15
2-3 相關法規與文獻 16
2-4 分析儀器 18
2-4-1 超高效液相層析儀 18
2-4-1-1 Poroshell 120 管柱 20
2-4-2 四極桿飛行時間串聯式質譜儀 20
2-5 基質固相分散萃取法 21
2-5-1 前言 21
2-5-2 原理與流程 21
2-6 雙重震盪搭配超音波輔助基質固相分散萃取法 23
2-6-1 原理與流程 23
2-6-2 超音波萃取法 24
2-6-3 固體支持物 26
2-7 實驗設計 27
2-7-1 簡介 27
2-7-2 應用 28
2-8 脂質定量分析 29
第三章實驗步驟與樣品分析 31
3-1 實驗藥品與設備 31
3-1-1 實驗藥品 31
3-1-2 儀器設備 34
3-2 實驗步驟 35
3-2-1 標準品的配製 35
3-2-2 UHPLC 參數設定 36
3-2-3 qTOF-MS 參數設定 37
3-2-4 質荷比校正 38
3-2-5 DVUA-MSPD 萃取法實驗步驟 39
3-2-6 空白魚體粉末的淨化 40
3-2-7 生物檢體採集 41
3-2-8 脂質定量分析實驗步驟 42
第四章結果與討論 43
4-1 分析儀器對待測物標準品與內標準品的測定 43
4-1-1 待測物標準品與內標準品的層析圖 43
4-1-2 待測物標準品的質譜圖 45
4-2 DVUA-MSPD 萃取法條件探討 47
4-2-1 Multilevel categoric design 47
4-2-1-1 分析 49
4-2-1-2 殘差分布圖 51
4-2-1-3 種類探討 53
4-2-2 Box-Behnken design 55
4-2-2-1 分析 57
4-2-2-2 殘差分布圖 59
4-2-2-3 數量探討 61
4-2-3 實驗設計條件探討結果 62
4-3 方法偵測極限與檢量線 63
4-3-1 Mandel’s fitting test 64
4-3-2 Lack-of-Fit test 66
4-4 方法準確度與精密度 68
4-5 真實樣品的檢測 69
4-6 前處理方法比較 74
第五章結論 77
第六章參考文獻 79
附錄 89

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指導教授

丁望賢(Wang-Hsien Ding)

審核日期

2019-7-23

推文