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研究生： 鄭丞勛
Chen-Hsun Cheng
論文名稱： 蛋白質溶液在黏彈性特徵與動態侷限下之液-液相分離行為研究
A Study on Liquid-liquid Phase Separation Behavior of Protein Solutions with Viscoelastic Characteristics and Dynamical Arrest
指導教授： 洪伯達
Po-Da Hong
口試委員: 洪伯達
Po-Da Hong
白孟宜
Meng-Yi Bai
江少華
Adhimoorthy Prasannan
學位類別： 碩士
Master
系所名稱： 工程學院 - 材料科學與工程系
Department of Materials Science and Engineering
論文出版年： 2023
畢業學年度： 112
語文別： 英文
論文頁數： 74
中文關鍵詞： 溶菌酶雞蛋白 、液-液相分離 、動態侷限 、分形維度 、小角度光散射
外文關鍵詞： lysozyme, liquid-liquid phase separation, dynamical arrest, fractal dimension, small-angle light scattering
相關次數： 點閱：69  下載：0 
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    • 在以短程作用力主導的蛋白質溶液相分離行為中，蛋白質稠密相（protein-rich phase）的液態-固態轉變會導致相分離動力學發生動態侷限（dynamical arrest）的現象。本研究利用小角光散射儀、雷射掃描共軛焦顯微鏡，以及相差顯微鏡，追蹤溶菌酶球蛋白溶液相分離的動力學過程以及形態學變化。
    透過在不同的熱力學條件下觀察相分離過程，我們發現蛋白質稠密相的黏彈性特徵，顯示蛋白質稠密相從液態到固態的轉變之間存在平滑的過渡區間。除此之外，在非臨界淬火的顯微影像中，我們觀察到稠密相液滴形成類似於膠體凝膠的結構，並在液滴內發現球蛋白的分形聚集現象。綜合以上結果，我們推測溶菌酶球蛋白由微觀到介觀的跨尺度凝膠化現象，將賦予蛋白質稠密相黏彈性特徵，從而影響蛋白質溶液相分離的動力學機制。

    In protein solutions where short-range forces dominate the phase separation behavior, the transition from liquid to solid of the protein-rich phase leads to dynamical arrest in the phase separation kinetics. We employed apparatus of small-angle light scattering, laser scanning confocal microscope, and phase contrast microscope to investigate the kinetic process and morphology changes of phase separation in lysozyme solution.
    We found the viscoelastic characteristics of the protein-rich phase by observing the phase separation process under different conditions. We noted a smooth transition between the liquid and solid phases of the protein-rich phase. In addition, in microscopy images of off-critical quench, we observed that protein-rich droplets formed structures similar to those in colloidal gels. We also found fractal aggregation of globulin within the droplets. Based on our results, we suggested that the gelation phenomenon of lysozyme globulin gives viscoelastic characteristics to the protein-rich phase, thereby influencing the kinetic mechanism of protein solution phase separation.

    Abstract IV Contents V Chart Catalogues VII Chapter 1. Introduction 11 1.1. Application of Protein Aggregation 11 1.1.1. Fluorescence modification by Eosin Y 13 1.2. Protein Structure and Colloidal System 14 1.2.1. Amino Acids and Primary Structure 14 1.2.2. Secondary Structure 15 1.2.3. Tertiary Structure 15 1.2.4. Quaternary Structure 16 1.2.5. Colloidal System 16 1.2.6. Van der Waals Force 17 1.2.7. Repulsive Electrical Double-layer Force 18 1.2.8. Ion Dispersion Forces 19 1.3. Protein Phase Diagram 20 1.3.1. Phase Transition 20 1.3.2. Phase Diagram 21 1.4. Classical Spinodal Decomposition 25 1.4.1. Cahn-Hilliard Theory 25 1.4.2. Binder-Stauffer-Siggia (BSS) Theory 27 1.5. Viscoelastic Phase Separation 29 1.5.1. The Moving Droplet Phase 30 1.6. Fractal Aggregation Model 32 1.7. The Purpose of This Thesis 33 Chapter 2. Experimental Section 34 2.1. Materials 34 2.2. Sample Preparation 35 2.3. Experimental Methods 35 2.3.1. UV-Visible/NIR Spectrophotometer (UV-Vis) 35 2.3.2. Micro Refrigerated High-Speed Centrifuge 35 2.3.3. Phase Contrast Microscopy (PCM) 36 2.3.4. Laser Scanning Confocal Microscopy (LSCM) 36 2.4. Small-angle Light Scattering 36 2.4.1. Data Analysis 38 2.4.2. Calibration of the Scattering Vector 39 Chapter 3. Results and Discussion 41 3.1. Determination of Protein Concentration 41 3.2. Phase Diagram of Lysozyme Solution 43 3.2.1. Glass Line and Arrest Tie Line 47 3.2.2. Investigate Arrest Tie Line by PCM Measurement 50 3.3. Morphology of Lysozyme Solutions by PCM and LSCM Measurement 52 3.4. Phase Separation Dynamics of Lysozyme Solutions 55 Chapter 4. Conclusion 66 References 69

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