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研究生: Rajeshkumar Anbazhagan
Rajeshkumar Anbazhagan
論文名稱: Specific ligand chemically conjugated on exfoliated MoS2 nanomaterials for fluorescence, magnetic resonance and surface enhanced Raman scattering imaging.
Specific ligand chemically conjugated on exfoliated MoS2 nanomaterials for fluorescence, magnetic resonance and surface enhanced Raman scattering imaging.
指導教授: 蔡協致
Hsieh-Chih Tsai
口試委員: 朱智謙
陳瑞山
李奎毅
鄭如忠
蔡協致
學位類別: 博士
Doctor
系所名稱: 應用科技學院 - 應用科技研究所
Graduate Institute of Applied Science and Technology
論文出版年: 2017
畢業學年度: 105
語文別: 英文
論文頁數: 158
中文關鍵詞: 二硫化鉬殼核結構核磁共振影像
外文關鍵詞: MoS2, MoS2-QDs, TGA, Bioimaging, MRI, Core-shell, Enhanced relaxation, In Vivo MRI, Metallic 1T- MoS2, Semiconductor 2H- MoS2, SERS, Alkyne tag, SERS Imaging.
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  • 奈米科技的發展為生醫領域帶來新革命,自從石墨希成功剝落並且應用於生醫領域,現今有許多二維材料備受矚目,例如:氮化硼、二硫化鉬,特別是層狀二硫化鉬具有獨特的性質。二硫化鉬是由單層二硫化鉬晶體堆疊而成,且層與層之間由較弱的凡得瓦力堆疊,而單層由強共價鍵鍵結過渡金屬(Mo)和硫原子(S)鍵結。二硫化鉬為2H相結晶結構(金屬原子的配位是三棱柱形),此外二硫化鉬塊材具有能帶間隙為1.29eV(半導體),因此不適合應用在生醫領域,當二硫化鉬剝層為層狀材料時,能帶間隙提高至1.9 eV,此時有益於生醫應用,且確實已經有些研究將單層、多層和量子點(QD)的二硫化鉬應用於生醫領域。本文包含三個部分: 第一部分為利用巰基乙酸(TGA)作為輔助經由超聲波震盪剝落二硫化鉬,再經由水熱合成法獲得量子點(QD)應用於生醫影像。由TGA改質後的二硫化鉬利用穿透式電子顯微鏡(TEM)、原子力顯微鏡(AFM)、X-射線繞射分析(XRD)和拉曼光譜(Raman)鑑定特性,結果顯示層狀二硫化鉬的能帶間隙經過改質後大增,由於TGA分子鍵結至二硫化鉬並且減弱層層之間的凡得瓦力鍵。TGA分子不僅作為塊狀二硫化鉬的界面活性劑,而且同時改質表面和增加層狀材料的產量。此外,重金屬二硫化鉬量子點(QD)具有持續發螢光的特性可作為細胞追蹤劑。
    第二部分提到使用殼核結構磁性奈米材料作為體內MRI顯影劑,由二硫化鉬和Gd-DTPA chelate合成,並且藉由DTPA anhydride濃度調控控殼核結構的尺寸。gadolinium chelate離子緊密堆積在二硫化鉬殼核結構,所以生物毒性較低,並且可以增加T1鬆弛,由於合成的殼核結構分子相較於商業的gadolinium chelate(magnevist)尺寸較大。體內測試結果顯示奈米殼核結構具有潛在的對比劑特性,特別是在心臟、腎臟和膀胱組織。此外量化的數據中證實膀胱對於殼核結構的清除濾高,因此,所有的結果證實此殼核結構是種很好的顯影劑。
    第三部分研究,合成Li-MoS2和半導體單層結構(T-MoS2),並且進一步證實可增強拉曼光譜訊號。單層Li-MoS2經由掃描式顯微鏡(SEM)、穿透式顯微鏡(TEM)、X-射線繞射分析(XRD)鑑定,以及X射線光電子能譜學(XPS)確認化學成份。此外電化學阻抗譜(EIS)證實單層Li-MoS2相較於其他半導體單層材料具有較佳的電荷轉移性能。羅丹明6G(R6G)在Li-MoS2拉曼散射(SERS)訊號較強,且是在T-MoS2的兩倍,由於R6G均勻分布於Li-MoS2。alkyne與二硫化鉬的硫醇鍵結,然而Li-MoS2與alkyne鍵結程度相較於T-MoS2高,由於Li-MoS2存在較多的電子缺陷,更值得注意的是alkyne在Li-Mos2-Alk邊緣的表面增強拉曼散射(SERS)較中心位置強。Li-MoS2-Alk細胞毒性較低,最後將Li-MoS2-Alk於DiWr細胞測試Raman mapping,這項研究開起了二硫化鉬在生醫領域的應用價值,將來需要更進一步證實SERS的特性。


    The rapid development of nanobiotechnology and biomedicine provide some valuable efficient alternatives towards diagnosis and therapy. Layered two-dimensional (2D) nanomaterials have powerfully gained research attention due to their peculiar and singular properties after successful utilization of graphene into the biomedical application. To date, plenty of layered 2D nanomaterials have been reported, such as graphene and boron nitride (h-BN) and molybdenum disulfide (MoS2) etc. Particularly, two-dimensional MoS2 (2D MoS2) such as molybdenum disulfide (MoS2) have been investigated for use in a biomedical application. Notably, Transition metal (Mo) and chalcogen (S) atoms in a single-layer of MoS2 NSs interact with each other by strong covalent bonds, while the interlayer interaction is weak van der Waals forces. MoS2 has a 2H phase crystal structure (the coordination of its metal atoms is trigonal prismatic) with the bandgap of ∼1.29 eV, which renders it semi-insulating and therefore not immediately attractive as an imaging candidate for bioimaging application. However, exfoliating into single or few layers would enhance it band gap to 1.9 eV, which is good for energy and biomedical application. Indeed, only a few studies have described a biomedical application of single-, multi-layered MoS2 nanosheets, and quantum dots (QD).
    This dissertation is based on the three different research projects: The first projects deal with exfoliation of MoS2 nanosheets by thioglycolic acid (TGA) assisted sonication method followed by hydrothermal treatment to obtained QDs for bioimaging application. The successful functionalization of TGA confirmed by FT-IR, zeta potential, and the synthesized nanosheets and QDs confirmed by TEM, AFM, XRD, and Raman spectroscopy. The gaps of MoS2 layers were enlarged by the stirring process. Meanwhile, the Vander Waals force between the MoS2 layers was reduced by TGA molecules strongly binding to the defect sites of MoS2. The use of TGA molecules not only act as a surfactant for exfoliation of the bulk of MoS2 but also functionalization in a surface of MoS2 leading to highly concentrated MoS2 nanosheets was produced in water. Subsequently, heavy metal-free MoS2 quantum dots with sustained fluorescence emission, which were obtained from hydrothermal treatment of exfoliated MoS2, can be utilized as cell biomarker.
    The second part focused on core-shell magnetic nanomaterials composed of MoS2 and Gadolinium Diethylenetriaminepentaacetic dianhydride chelate (Gd-DTPA chelate) used as promising in vivo magnetic resonance imaging. The size of the synthesized core-shell magnetic nanomaterials could be controlled by varying the DTPA anhydride concentration. The gadolinium chelate functionalized core-shell MoS2 nanomaterials exhibit reduced toxicity due to the tight packing of gadolinium ion into it, and enhanced T1 relaxation owing to its macromolecular sized compared to the commercially available small gadolinium chelate such as magnevist. In vivo MRI of these nanomaterials revealed the potential contrasting property of these core-shell magnetic nanomaterials. Thereby, these core-shell magnetic nanomaterials show an enhanced contrast in heart, kidney, and bladder. In addition, the quantitative signal analysis in bladder clearly confirms the core-shell magnetic nanomaterials have enhanced clearance; it could be identified from its hyperintense signal in a bladder. Therefore, all these data revealed the potentials contrasting property of the synthesized core-shell magnetic nanomaterials could be used as better contrast agents in future.
    In the third project, alkyne conjugated metallic (Li- MoS2) and semiconductor (T- MoS2) monolayers were synthesized and investigated its potential surface enhanced Raman spectroscopy. The synthesized Li-MoS2 monolayers were confirmed through SEM, TEM Raman, XD, and its chemical composition confirmed by the XPS. The as per synthesized Li- MoS2 monolayers has enhanced charge transfer property than semiconductor monolayers. The enhanced charge transfer property confirmed by EIS. The SERS of R6G on Li-MoS2 two times higher than T- MoS2, the R6G uniformly distribution over Li- MoS2 than T- MoS2 followed by its Raman signal enhancement. The synthesized alkyne conjugated on both MoS2 through thiol group. Furthermore, Li- MoS2 has a high number of conjugation than T- MoS2 due to high defects sites presence in the Li- MoS2. Interestingly, highly enhanced alkyne SERS observed in the edge of the Li- MoS2-Alk than in the center of Li- MoS2-Alk. The cytotoxicity of synthesized Li- MoS2-Alk has less toxicity, finally, Raman mapping of synthesized Li- MoS2-Alk was executed in DiWr cells. These findings open a unique way to utilize MoS2 nanomaterials in biomedical application, the further study required to recheck its potential SERS and Raman imaging properties in the near future.

    中文摘要----------------------------------------------------------------------------------------------------viii Abstract------------------------------------------------------------------------------------------------------x Acknowledgement------------------------------------------------------------------------------------------xiii List of Figures-----------------------------------------------------------------------------------------------xv List of tables-------------------------------------------------------------------------------------------------xx List of schemes--------------------------------------------------------------------------------------------- xx List of abbreviations----------------------------------------------------------------------------------------xxi 1. Chapter 1: Backgrounds and its significance--------------------------------------------------------1 1.1. Imaging method------------------------------------------------------------------------------------1 1.2. Nanomaterials for biomedical imaging application-------------------------------------------2 1.3. Two dimensional nanomaterials for imaging application------------------------------------4 2. Chapter 2: Introduction--------------------------------------------------------------------------------6 2.1 Molybdenum disulphide (MoS2) ---------------------------------------------------------------6 2.2 Synthesis -------------------------------------------------------------------------------------------8 2.3 Top-down synthesis--------------------------------------------------------------------------------8 2.3.1 Mechanical exfoliation-------------------------------------------------------------------9 2.3.2 Chemical exfoliation---------------------------------------------------------------------9 2.3.3 Liquid base exfoliation------------------------------------------------------------------10 2.4 Bottom-up synthesis------------------------------------------------------------------------------12 2.5 Colloidal synthesis--------------------------------------------------------------------------------12 2.6 MoS2 QDs synthesis------------------------------------------------------------------------------13 2.7 Surface functionalization-------------------------------------------------------------------------13 2.8 Therapeutic application of 2D-MoS2-----------------------------------------------------------15 2.8.1. Biocompatibility-------------------------------------------------------------------------15 2.8.2. Phototherapy system--------------------------------------------------------------------16 2.8.3. Delivery system--------------------------------------------------------------------------17 2.8.4. Bioimaging system----------------------------------------------------------------------18 2.8.4.1. Fluorescence imaging--------------------------------------------------------------18 2.8.4.2. Multiphoton fluorescence imaging-----------------------------------------------20 2.8.4.3. MRI imaging------------------------------------------------------------------------20 2.8.4.4. SERS and Raman imaging--------------------------------------------------------23 3. Chapter 3: Research objectives--------------------------------------------------------------------25 4. Chapter 4: Highly concentrated MoS2 nanosheets in water achieved by thioglycolic acid as stabilizer and used as biomarkers-----------------------------------------------------------------26 4.1. Introduction----------------------------------------------------------------------------------26 4.2. Results and discussion----------------------------------------------------------------------28 4.2.1. Exfoliation of monolayers and its confirmation--------------------------28 4.2.2. Morphological identification------------------------------------------------34 4.2.3. Fluorescence property of MoS2-QDs and used as a biomarker---------36 4.3. Conclusion------------------------------------------------------------------------------------37 4.4. Experimental section------------------------------------------------------------------------37 4.4.1. Materials -----------------------------------------------------------------------37 4.4.2. Preparation of MoS2 monolayers-------------------------------------------37 4.4.3. Preparation of MoS2 quantum dot------------------------------------------38 4.4.4. Optimize the TGA concentrations------------------------------------------38 4.4.5. Measurement concentration of MoS2 monolayers -----------------------38 4.4.6. Characterization --------------------------------------------------------------39 4.4.7. Internalization-----------------------------------------------------------------40 5. Chapter 5: MoS2−Gd Chelate Magnetic Nanomaterials with Core−Shell Structure Used as Contrast Agents in in Vivo Magnetic Resonance Imaging------------------------------41 5.1. Introduction---------------------------------------------------------------------------41 5.2. Results and discussion---------------------------------------------------------------43 5.2.1. Synthesis and properties of core-shell MoS2-Gd chelates------------43 5.2.2. Mechanism of sphere formation------------------------------------------46 5.2.3. Binding energy and properties core shell magnetic chelate structure-----------------------------------------------------------------------------------48 5.2.4. Magnetic properties of core shell magnetic nanomaterials-----------54 5.2.5. T1 relaxation of core shell magnetic nanomaterials--------------------57 5.2.6. Gadolinium leaching and material cytotoxicity------------------------59 5.2.7. In vivo positive contrast imaging----------------------------------------61 5.3. Conclusion-----------------------------------------------------------------------------65 5.4. Experimental section-----------------------------------------------------------------66 5.4.1. Materials---------------------------------------------------------------------66 5.4.2. Synthesis of MoS2-cysteamine (M-Cys)--------------------------------66 5.4.3. Transfer of M-Cys from hydrated DMSO to anhydrous DMSO----66 5.4.4. Synthesis of DTPA-conjugated M-Cys (M-Cys-DTPA)-------------67 5.4.5. Preparation of M-Cys-DTPA-gadolinium complex (M-Cys-DTPA Gd)---------------------------------------------------------------------------67 5.4.6. Physicochemical property, morphology, and magnetic propert-----67 5.4.7. Cell culture experiment----------------------------------------------------68 5.4.8. Cytotoxicity test (MTT assay)--------------------------------------------69 5.4.9. Gadolinium leaching analysis---------------------------------------------69 5.4.10. In vivo positive contrast imaging-----------------------------------------70 5.4.11. Quantitative signal intensity measurement in organs------------------70 6. Chapter 6: Enhanced SERS using alkyne conjugated metallic MoS2. A comparative study between metallic and semiconductor phase -------------------------------------------------------71 6.1. Introduction.---------------------------------------------------------------------------71 6.2. Results and discussion---------------------------------------------------------------73 6.2.1. Exfoliation and its properties of exfoliated MoS2 monolayers-------73 6.2.2. SERS of Li-MoS2 and T-MoS2-------------------------------------------76 6.2.3. Binding energy investigation of Li-MoS2 and T-MoS2---------------80 6.2.4. Charge transfer property of Li-MoS2 and T-MoS2--------------------84 6.2.5. R6G mapping of Li-MoS2 and T-MoS2---------------------------------85 6.2.6. Alkyne conjugation and its SERS on Li-MoS2 and T-MoS2---------86 6.2.7. Alkyne of Li-MoS2 and T-MoS2-----------------------------------------93 6.3. Conclusion-----------------------------------------------------------------------------94 6.4. Experimental section-----------------------------------------------------------------94 6.4.1. Materials and methods-----------------------------------------------------94 6.4.2. Synthesis of Lithium exfoliated MoS2 (Li-MoS2)---------------------94 6.4.3. Synthesis of TGA exfoliated MoS2 (T-MoS2)-------------------------95 6.4.4. Characterization -----------------------------------------------------------95 6.4.5. R6G SERS in Li-MoS2 and T-MoS2------------------------------------96 6.5. Raman ligand synthesis-----------------------------------------------------------------96 6.5.1. Glycine methyl ester hydrochloride-------------------------------------97 6.5.2. N-(diphenylmethylene) glycine ester------------------------------------97 6.5.3. Methyl 2-(diphenylmethyleneamino)pent-4-ynoate-------------------97 6.5.4. Methyl-2-acetylamino-4-pentynoate------------------------------------98 6.5.5. Methyl 2-(3-(acetylthio)propanamido)pent-4-ynoate-----------------98 6.5.6. 2-(3-mercaptopropanamido) pent-4-ynoic acid------------------------99 6.5.7. Ligand conjugation on MoS2 nanosheets-------------------------------99 6.5.8. Alkyne SERS------------------------------------------------------------100 6.6. Cell culture experiment-----------------------------------------------------------------100 6.6.1. Cytotoxicity Test----------------------------------------------------------100 6.6.2. Raman imaging of DiWr cells-------------------------------------------100 7. Chapter 7: Conclusion-----------------------------------------------------------------------------102

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