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研究生：	陳靚蓉 Chen, Jing-Rong
論文名稱：	以單顆粒感應耦合電漿質譜儀結合前處理與奈米金標記來檢測魚體中的奈米塑膠 Using Single-Particle ICP-MS Combining with Pretreatment and Post-Au Nanoparticle Labeling to Detect Nanoplastics in Fish Samples
指導教授：	侯文哲 Hou, Wen-Che
學位類別：	碩士 Master
系所名稱：	工學院 - 環境工程學系 Department of Environmental Engineering
論文出版年：	2023
畢業學年度：	111
語文別：	英文
論文頁數：	76
中文關鍵詞：	奈米塑膠、單顆粒感應耦合電漿質譜儀、金標記
外文關鍵詞：	nanoplastic, spICP-MS, Au-labeling
相關次數：	點閱：86 下載：0
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奈米塑膠 (NP) ，一種被定義為粒徑小於 1 微米的塑膠碎片，隨著 NP 的存在被證實， NP 對環境及生物的可能危害也日漸受到關注。評估此類新興污染物的生物暴露和相關影響將取決於檢測及量化的分析方法，然而，檢測及量化複雜生物基質中的 NP 是一項具有挑戰性的任務。在本篇研究中我們使用金奈米顆粒 (AuNP) 對 NP 進行標記，以利用單顆粒感應耦合電漿質譜儀 (spICP-MS) 進行NP顆粒數計數。此外，我們研發從魚肉萃取 NP 的消化方法，並評估其對 AuNP 後標記NP的影響，開發可與 AuNP 後標記 NP 方法相容的魚樣本萃取消化程序，以使用 spICP-MS 對魚肉樣本中的 NP 進行量化計數。研究為初步結果顯示苯乙烯奈米塑膠顆粒(PSNP)數目濃度為 4.62 × 1012 #/L 時，該方法在水中回收率 99.29 ± 3.71% 。而此方法在應用於較低 PSNP 數目濃度標記時，需調整金前驅物及還原劑添加濃度，以避免過多的游離 AuNPs 產生或造成標記不均等干擾檢測的問題。而現階段發現蛋白酶搭配雙氧水的雙重消化程序，可有效消解魚肉且不干擾標記程序，而目前此消化程序及標記方法可從魚肉樣本回收 94.29 ± 48.98% 的 PSNPs 。

Nanoplastics (NPs) are plastic fragments with particle sizes smaller than 1 μm. Their presence in the environment has prompted great concerns regarding their potential hazards and accumulation in biota. The availability of analytical methods to detect and quantify NPs in complex environmental and biological matrices significantly affects our ability to assess the ecological exposure and impacts of this emerging class of pollutants. However, detecting and quantifying NPs in complex biological matrices is a challenging task. In this study, we first developed a method to label NPs with gold nanoparticles (AuNPs) to enable NP counting using single-particle inductively coupled plasma mass spectrometry (spICP-MS). We explored various fish tissue digestion methods to extract NPs from fish tissue and evaluate their impacts on the AuNP post-labeling of extracted NPs for counting by spICP-MS. Preliminary results demonstrated that the recoveries of polystyrene nanoparticles (PSNPs) at a concentration of 4.62 × 1012 #/L from water were 99.29 ± 3.71%. Furthermore, when the method was applied to lower PSNP concentrations, adjustments in the concentration of gold precursors and reductants were required to prevent excessive generation of free AuNPs or uneven attachment of AuNPs on PSNPs during the labeling process. It was discovered that a dual digestion process using proteinase K and hydrogen peroxide effectively digested fish tissue without interfering with the labeling process. Currently, this digestion process and labeling method could recover 94.29 ± 48.98% of PSNPs from fish tissue samples.

摘要 I
ABSTRACT II
CONTENTS III
LIST OF TABLE VI
LIST OF FIGURE VII
Chapter 1 INTRODUCTION 1
1 Background and motivation 1
2 Objective 3
Chapter 2 LITERATURE REVIEWS 4
1 Occurrences of plastic waste, MPs, and NPs 4
2 MPs and NPs in the environment 5
3 Potential health hazards of MPs and NPs 6
4 Characterization and analysis technologies of MPs and NPs 7
4.1 Morphology analysis of MPs and NPs 7
4.2 Qualitative and quantitative analyses of MPs 8
4.3 Challenges and recent development in NP analysis 9
5 Extraction of NPs from organisms 10
Chapter 3 MATERIALS AND METHODS 12
1 Materials and chemicals 12
2 Synthesis of AuNP precursor 13
3 Development and evaluation of AuNP labeling methods for PSNPs counting 14
3.1 Labeling PSNPs with AuNPs according to an existing study 16
3.2 Different reductant NaBH4 addition speed during labeling 17
3.3 Different standing times during labeling 18
3.4 Different AuNP coverages during labeling 18
3.5 Counting performance of the labeling method 20
4 Development and evaluation of fish sample extraction methods for PSNP counting using AuNP post-labeling 22
4.1 Fish sample preparation 23
4.2 Evaluation of fish tissue digestion methods 23
4.3 Labeling PSNPs incubated at high temperature 24
4.4 Labeling PSNPs after different digestion method 25
4.5 Labeling PSNPs spiked in the fish tissue 26
5 Analytical methods 27
5.1 NMR characterization 27
5.2 ICP-MS analysis 27
5.3 spICP-MS analysis 28
5.4 TEM characterization 32
Chapter 4 RESULTS AND DISCUSSION 33
1 Characterization of PSNPs 34
2 Characterization of home-made Au(en)2Cl3 35
3 Development and evaluation of AuNP labeling methods for PSNP counting in water 37
3.1 Preliminary results of comparing unstirred and stirred sample 37
3.2 Effect of different reductant NaBH4 addition speed on labeling 40
3.3 Effect of different standing time on labeling 42
3.4 Effect of different AuNP coverage on labeling 43
3.5 Counting performance of the labeling method 45
4 Development and evaluation of fish sample extraction methods for PSNP counting using AuNP post-labeling 50
4.1 The efficiency of the digestion method 51
4.2 Effect of PSNPs incubated at high temperature on labeling 53
4.3 Effect of different digestion method on labeling 54
4.4 The result of the AuNP labeled PSNPs in fish tissue 58
Chapter 5 CONCLUSION AND OUTLOOK 61
REFERENCES 63
                                    

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