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標題: | 奈米粒子對毛細管電泳分離胺及酸之影響 The impact of nanoparticles on the separation of DNA and amines by capillary electrophoresis |
作者: | Wan-Chi Yu 余婉琪 |
指導教授: | 張煥宗 |
關鍵字: | 奈米粒子,毛細管電泳,DNA分離,生物胺分離, nanoparticles,capillary electrophoresis,separation of DNA and biological amines, |
出版年 : | 2005 |
學位: | 碩士 |
摘要: | 本論文是在研究毛細管電泳對生物分子分離之應用,主要分成二個部分,第一個部分是探討毛細管電泳中,於聚合物溶液中加入添加物對DNA分離之影響。實驗所用的聚合物為乙基纖維素(2-hydroxyethyl cellulose,HEC),分子量為1,300 000,加入添加物thiourea之後,相較於單純以0.35% HEC分離DNA marker Ⅴ、Ⅵ,可大幅提高394-653 bp範圍的9個DNA片段之解析度,另外,我們也探討金奈米粒子對DNA分離之影響,金奈米粒子可以使DNA的分離在較短的時間內完成(整個分離時間可在五分鐘內完成),且能維持住相當的解析度,加入這些添加物可提高DNA之分離解析度,可能的原因為:添加物與聚合物作用因而改變聚合物的構型,此構型的改變有助於DNA分離解析度的提高。
第二部分則是研究毛細管表面修飾對生物胺及其代謝有機酸分離的影響,我們探討毛細管壁修飾聚離胺酸(poly-L-lysine,PLL)及二氧化矽奈米粒子(SiONP)對分析物分離的影響。毛細管壁修飾聚離胺酸(以(PLL-SiO2)n-PLL表示)時,最佳的條件是修飾(PLL-SiO2)2-PLL,並以添加3% acetonitrile (ACN) 之10 mM甲酸溶液(pH 3.7)進行分離。在此條件下,對酸的靈敏度較高,像是對5-HIAA及5-HT的偵測極限分別可達1 nM及35 nM;另一方面,毛細管壁修飾二氧化矽奈米粒子時(以(PLL-SiONP)n表示),是以修飾(PLL-SiONP)3的毛細管,並在10 mM甲酸溶液(pH 3.7)進行分離可得最佳的分離結果,在此條件下,對胺的靈敏度較高。像是對TA及5-HIAA的偵測極限為0.09 nM及19 nM。因此可依據欲分析的生物分子是胺或酸而選擇毛細管壁以聚離胺酸或二氧化矽奈米粒子修飾來進行分離。 In this thesis, we focus on the impact of nanoparticles on the separation of biological molecules by capillary electrophoresis (CE) in conjunction with laser-induced fluorescence (LIF) detection. In the first part of this thesis, the impact of additives in 0.35% 2-hydroxyethyl cellulose (HEC, Mw 1,300 000 g/mole) on DNA separation by CE-LIF is studied. By using 0.35% HEC prepared in 40 mM glycine buffer (pH 9.9) containing 1% thiourea, 32-nm gold nanoparticles, and 1.27μM ethidium bromide, 9 DNA fragments with their sizes ranging from 394 to 653 bp were well resolved within 5 min. The separation is fast, highly efficient, and reproducible, mainly because of the change in HEC morphologies as a result of the interaction among thiourea, HEC, and gold nanoparticles. In the second part of this thesis, we have investigated the separation of biological amines and acids by CE-LIF using capillaries coated with poly-L-lysine (PLL) and silica nanoparticles (SiONPs). The capillaries coated with (PLL-SiONP)3 and (PLL-SiONPs)2-PLL are suitable for the separations of amines and acids, respectively. By using a (PLL-SiONP)3 capillary filled with a solution consisted of 10 mM formic acid (pH 3.7) and 3% acetonitrile (ACN), the limits of detection (LOD) at signal-to-noise ratio (S/N) = 3 are 1.2 nM and 35.0 nM for 5-hydroxyindole-3-acetic acid (5-HIAA) and 5-hydroxytryptamine (5-HT), respectively. The LODs are 0.09 nM and 19.0 nM for TA and 5-HIAA, respectively, when using a (PLL-SiONPs)2-PLL capillary filled with 10 mM formic acid ( pH 3.7). The practical application of the method has been validated by the analysis of urine samples, with great sensitivity, high speed, efficiency, and good reproducibility. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/35253 |
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顯示於系所單位: | 化學系 |
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