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研究生:張家耀
研究生(外文):Jia-Yaw Chang
論文名稱:超臨界水對奈米碳管之處理暨生物錯合性奈米粒子之製備與在生物上應用
論文名稱(外文):Processing of carbon nanotubes in supercritical water and synthesis of bioconjugated nanoparticles for biological applications
指導教授:凌永健凌永健引用關係
指導教授(外文):Yong-Chien Ling
學位類別:博士
校院名稱:國立清華大學
系所名稱:化學系
學門:自然科學學門
學類:化學學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:156
中文關鍵詞:超臨界水奈米碳管肥胖性蛋白質分子信標金奈米棒螢光二氧化矽奈米粒子
外文關鍵詞:supercritical watercarbon nanotubesobesitymolecular beacongold nanorodfluorescent silica nanoparticle
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  • 收藏至我的研究室書目清單書目收藏:0
使用超臨界水對於多層奈米碳管進行將兩端變細及層與層之間的剝蝕。超臨界水中以不同的條件,例如: 溫度,壓力,時間 進行實驗。當超臨界水搭配氧氣(∼ 2mmol)的存在之下特別能對於將變細的多層奈米碳管外層結構變成崩塌結構。除此之外,超臨界水還能改變多層奈米碳管的外觀形狀,並且搭配電子損失能譜觀察此種外觀上變化會引起電子能階的分布情況。同時在4種假設之下搭配簡單的數學模式解釋在超臨界水實驗中所觀察的多層奈米碳管的外觀形狀變化。本文中更以超臨界水的高度破壞性的環境下將銀的凝聚物脆裂成2~20 nm奈米粒子而將其帶入奈米碳管的奈米孔洞。實驗中發現超臨界水除了將銀奈米粒子帶入奈米碳管的奈米孔洞,並且隨著時間增長銀奈米粒子會堆積在一起,並因為高溫燒結使其成為奈米棒子。不同的超臨界水實驗條件可以得到不同粒徑分布的銀奈米粒子,更隨著壓力增大銀奈粒子會形成銀奈米線,和 少許類似三角形的銀奈米板。
除了銀奈米粒子的製備,更將其使用在生物上應用例如分子信標。分子信標示一種髮夾形狀的核甘酸,並在兩端分別接上螢光團和螢光抑制物(quencher)。螢光團和螢光被抑制物會由於螢光共振能量傳遞(fluorescence resonance energy transfer)而不發光,所以分子信標會是關閉的狀態。而當遇到欲偵測的互補性核甘酸,分子信標與互補性核甘酸雜合(hybridization)會呈現打開狀態而發出螢光,藉以檢測目標核甘酸。一般文獻上都以有機物DABCYL當成螢光抑制物,本實驗中則是將金銀合金當成螢光抑制物,探討不同金銀比例對於螢光抑制的效率影響。
發螢光的二氧化矽奈米粒子和金奈米棒也分別應用在肥胖性相關的蛋白質檢測和免疫分析。發螢光的二氧化矽奈米粒子是以反微胞方法將水溶性的螢光染料包覆在二氧化矽奈米粒子之中,同時二氧化矽奈米粒子的表面更可以修飾上不同的官能基例如酸基(carboxylic acid)、胺基(amine) 、硫基(thiol)藉以進一步能與生物分子產生錯合(biojugation),因此可以將發螢光的二氧化矽奈米粒子當成肥胖性相關的蛋白質的檢測探針(probe)。同時在金奈米棒的兩端接上帶有酸基的官能基後,利用Carbodimmide 方法將其與抗-老鼠的免疫球蛋白(抗體)產生錯合,此時的金奈米棒便具有免疫分析的能力可以專一性辨識 老鼠的免疫球蛋白(抗原)以及自組裝的功能。
Abstract
Supercritical water (SCW) is used for the first time for the opening and thinning of multiwall carbon nanotubes (MWNTs). The influence of variation of pressure, temperature and time on the opening and thinning of MWNTs is examined. In SCW, opening and thinning of MWNTs is observed. The presence of oxygen (~2 mmol) shows improved thinning of MWNTs with the collapsed outer graphene layers tending towards the inner layers. The morphologies of MWNTs are critically analyzed using transmission electron microscopy (TEM). MWNTs with different morphology were prepared using SCW oxidation and investigated by TEM and electron energy-loss spectroscopy (EELS). TEM results indicate that the peeling and sharpening of MWNTs are influenced by the etching process in SCW oxidation, of which oxidation time and amount of oxygen used is crucial. A simplified etching model is proposed, which indicates that the difference of mean etching rate between two adjoining blocks causes the morphological variation of MWNTs. The EELS results show change in characteristic energy-loss peaks as a function of total shell numbers along longitudinal axis of individual peeled-tube.

SCW medium also served as a highly destructive environment, which broke the silver aggregates into nanoparticles (diameter 2~20 nm). Water was drawn into open-ended MWNTs by capillary suction which caused silver (Ag) nanoparticles being pulled into the ends of MWNTs. The Ag nanoparticles presumably transported in nanochannels of MWNTs by the fluidity of SCW, stacked and fused to form nanorods, suggesting SCW associated with MWNTs might be exploited as a nanoreactor. Furthermore, SCW used to be a medium alone for fragmenting Ag aggregates and organized the resulting Ag nanoparticles into chain-like nanowires and nanobanners. A variety of 1D and 2D nanostructured assemblies were formed from the nanoparticles by variations in pressure, temperature, and time. The size distribution of Ag nanoparticles is controllable in the range of 2–20 nm. Under appropriate conditions, the SCW medium allows the merging of Ag nanoparticles to form Ag nanowires with diameters of ~60 nm, and nanobanners of triangle-shaped morphology with lengths of several hundred �慆.

Rather than just preparation of silver nanoparticles, the bioapplication of these metal nanoparticles were also concerned. Since fluorescent probes for biomolecular recognition are of great importance in the fields of chemistry, biology, and medical sciences, as well as in biotechnology. These probes have been used for mechanism studies of biological functions and in ultra sensitive detection of biological species responsible for many diseases. In the post-genome era, quantitative studies of genomic information for disease diagnosis and prevention and drug discovery become fast growing areas of research and development. This has led to a continued demand for advanced biomolecular recognition probes with high sensitivity and high specificity. The molecular beacon (MB), a recently developed single-stranded DNA molecule, appears to be a very promising probe for quantitative genomic studies. MBs are hairpin-shaped oligonucleotides that contain both fluorophore and quencher moieties and act like switches that are normally closed to bring the fluorophore/quencher pair together to turn fluorescence “off”. When prompted to undergo conformational changes that open the hairpin structure, the fluorophore and the quencher are separated, and fluorescence is turned “on”. A hybrided system composed of a ssDNA molecule and a 20 nm diameter gold/silver alloy nanoparticles. The quenching efficiency was varied by the mole ratio of gold and silver. The pure silver nanoparticles as quencher shows the best quenching efficiency and the regain efficiency was increased by gold mole fraction.

Except metal nanoparticles used as the quencher of MBs for reorganizing target DNA, fluorescent silica nanoparticles and gold nanorods were also applied in the obesity-related protein assay and immunosensing, respectively. Fluorescent silica NPs were prepared using reverse microemussion method and can be functionalized with carboxylate, amine, and thiol group for conjugated with biomolecules. The phosphonate group plays an important role in the dispersion of fluorescent silica NPs with amine and thiol groups. Obesity is a common nutritional disorder associated with diabetes, hypertension, hyperlipidemia, cancer and the other health related problems. Fluorescent silica NPs with carboxylate group were further used as the probe in the obesity-related protein assay. The oriented assembly of gold nanorods as biorecognition system was useful for the immune sensing events. Self-assembled thioctic acid containing a terminal carboxylate group at the end of gold nanorod can facilitate conjugation with anti-mouse IgG, which provides an anchoring site. Driven by the biorecognition with mouse IgG, nanorods assemble to form extended nanorod chains.
Table of Contents
Page
CHAPTER 1 BACKGROUND AND INTRODUCTION 1
1.1 CARBON NANOTUBES 1
1.1.1 PROPERTIES OF CARBON NANOTUBES 1
1.1.1.1 Chemical reactivity 1
1.1.1.2 Electronic properties 3
1.1.1.3 Mechanical strength 4
1.1.2 SYNTHESIS METHODS 5
1.1.2.1 Arc discharge method 6
1.1.2.2 CVD method 7
1.1.2.3 Laser ablation method 8
1.1.3 POTENTIAL APPLICATIONS 9
1.1.3.1 Field-effect transistors 9
1.1.3.2 Electron field emission 10
1.1.3.3 Sensors 11
1.1.3.4 Composite materials 12
1.1.3.5 Hydrogen storage 13
1.2 MOLECULAR BEACON 14
1.2.1 PRINCIPLE OF MBS 14
1.2.2 ADVANTAGES OF MBS 15
1.2.3 APPLICATIONS OF MBS 16
1.3 NANOMATERIALS 17
1.3.1 METAL NANORODS AND NANOWIRES 17
1.3.1.1 Gold nanorods 18
1.3.1.2 Silver nanowires 18
1.3.2 FLUORESCENT-DOPED SILICA NANOPARTICLES 19
1.3.3 BIOCONJUGATED NANOPARTICLES 21
1.3.3.1 Carboxylic acid-amine cross-linking 22
1.3.3.2 Thiol-amine cross-linking 22
1.3.3.3 Amine–amine cross-linking 23
1.3.3.4 Hydroxy-amine cross-linking 23
1.4 OBESITY-RELATED PROTEIN 24
1.5 FIGURES 27
1.6 REFERENCES 29
CHAPTER 2 OPENING AND THINNING OF MULTIWALL CARBON NANOTUBES IN SUPERCRITICAL WATER 38
2.1 INTRODUCTION 38
2.2 EXPERIMENTAL SECTION 40
2.3 RESULTS AND DISCUSSION 41
2.3.1 OXIDATION 41
2.3.2 OPENING AND THINNING OF MWNTS 44
2.3.3 RAMAN SPECTROSCOPY 48
2.4 CONCLUSION 49
2.5 FIGURES 50
2.6 REFERENCES 53
CHAPTER 3 MORPHOLOGICAL VARIATION OF MULTIWALL CARBON NANOTUBES IN SUPERCRITICAL WATER OXIDATION 55
3.1 INTRODUCTION 55
3.2 EXPERIMENTAL SECTION 57
3.3 RESULT AND DISCUSSION 57
3.3.1 ASSUMPTION FOR THE MORPHOLOGICAL VARIATION OF CNTS 57
3.3.2 MORPHOLOGICAL VARIATION BETWEEN TWO ADJOINING BLOCKS 59
3.3.3 EELS SPECTRA OF PEELED-TUBES 61
3.4 CONCLUSION 63
3.5 FIGURES 64
CHAPTER 4 TRANSPORTATION OF SILVER NANOPATICLES IN NANOCHANNELS OF CARBON NANOTUBES WITH SUPERCRITICAL WATER 68
4.1 INTRODUCTION 68
4.2 EXPERIMENTAL SECTION 70
4.3 RESULTS AND DISCUSSION 71
4.4 CONCLUSION 73
4.5 FIGURES 74
4.6 REFERENCES 76
CHAPTER 5 SILVER NANOPARTICLES SPONTANEOUS ORGANIZE INTO NANOWIRES AND NANOBANNERS IN SUPERCRITICAL WATER 78
5.1 INTRODUCTION 78
5.2 EXPERIMENTAL SECTION 79
5.3 RESULTS AND DISCUSSION 80
5.3.1 SILVER NANOPARTICLES PREPARED IN SCW 80
5.3.2 SILVER NANOWIRES AND NANOBANNERS PREPARED IN SCW 81
5.3.3 SILVER NANOSTRUCTURES FORMATION MECHANISM 83
5.4 CONCLUSION 84
5.5 FIGURES 86
5.6 REFERENCES 89
CHAPTER 6 GOLD/SILVER ALLOY NANOPARTICLES AS THE QUENCHER IN THE MOLECULAR BEACON 91
6.1 INTRODUCTION 91
6.2 EXPERIMENTAL SECTION 95
6.2.1 BUFFER 95
6.2.2 COUPLING OF DABCYL 95
6.2.3 COUPLING OF FLUOROPHORE 96
6.2.4 AUTOMATED SYNTHESIS 97
6.2.6 MB WITH CY3 FLUOROPHORE AND QUENCHER OF GOLD/SILVER NANOPARTICLE 98
6.3 RESULTS AND DISCUSSION 99
6.3.1 SYMTHESIS OF MB WITH CY3 FLUOROPHORE AND DABCYL QUENCHER 99
6.3.2 SYNTHESIS OF MB WITH TRI-THIOL AT 3’ TERMINUS 101
6.3.3 SILVER, GOLD, AND GOLD/SILVER ALLOY NANOPARTICLES 102
6.3.4 FRET EFFICIENCY 103
6.4 CONCLUSION 105
6.5 FIGURES 106
6.6 REFERENCES 116
CHAPTER 7 FLUORESCENT BIOCONJUGATED SILICA NANOPARTICLES FOR THE OBESITY-RELATED PROTEIN ASSAY 118
7.1 INTRODUCTION 118
7.2 EXPERIMENTAL SECTION 121
7.2.1 CHEMICALS AND REAGENTS 121
7.2.2 PREPARATION OF SAMPLE AND BUFFER SOLUTION 122
7.2.3 PREPARATION OF FLUORESCENT SILICA NPS 122
7.2.4 SURFACE MODIFICATION OF FLUORESCENT SILICA NPS 122
7.2.5 DETERMINATION OF CARBOXYLATE, AMINE, AND THIOL FUNCTIONAL GROUP ON THE NPS 123
7.2.6 PREPARATION OF FLUORESCENT SILICA NPS COATING WITH STV 124
7.2.7 CONSTRUCTION OF PROTEIN SANDWICH ASSAY 124
7.2.8 CONFOCAL FLUORESCENCE MICROSCOPY AND IMAGE ANALYSIS 125
7.3 RESULTS AND DISCUSSION 125
7.3.1 SURFACE MODIFICATION OF NPS 125
7.3.2 CHARACTERIZATION OF FLUORESCENT SILICA NPS WITH DIFFERENT FUNCTION GROUP 127
7.3.3 CHARACTERIZATION OF FLUORESCENT SILICA NPS COATING WITH STV 127
7.3.4 CONSTRUCTION OF PROTEIN SANDWICH ASSAY 128
7.4 CONCLUSION 131
7.5 FIGURES 132
7.6 REFERENCES 141
CHAPTER 8 ORIENTED ASSEMBLY OF GOLD NANORODS AS BIORECOGNITION SYSTEM FOR IMMUNOSENSING 143
8.1 INTRODUCTION 143
8.2 EXPERIMENTAL SECTION 144
8.3 RESULTS AND DISCUSSION 145
8.4 CONCLUSION 147
8.5 FIGURES 148
8.6 REFERENCES 152
LIST OF PUBLICATIONS 153
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