利用恆溫吸附以及恆溫滴定卡計探討二氧化矽表面吸附DNA之吸附機制

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姓名

王瑋豪(Wei-Hao Wang) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

利用恆溫吸附以及恆溫滴定卡計探討二氧化矽表面吸附DNA之吸附機制
(Studies of the Interaction Mechanism between DNA with Silica Surface by Microcalorimetry and Isotherm Measurements)

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摘要(中)

微小核醣核酸 (miRNAs)在許多疾病的診斷以以及治療上為一個非常具有潛力的生物標靶分子。目前市售萃取miRNA的萃取套組大多是結合液相萃取及固相萃取以達到萃取miRNA之最大效率。但目前大多的研究專注於萃取出更大量及更高純度的miRNA，而缺乏有關核酸吸附機制之研究，進而限制了萃取核酸的應用及發展。
在固相萃取中，最常以二氧化矽當作是吸附核酸的材料，而二氧化矽和核酸之間的吸附作用力依據不同溶液條件，大致可以分成以下四種(1)核酸和二氧化矽表面之氫鍵鍵結 (2)核酸和二氧化矽表面水分子之輸水效應 (3)分子間的靜電作用力 (4)核酸二氧化矽間之鹽橋效應。此外，我們觀察在Hofmeister series中將鹽離子之水合能力，由較強的kosmotrope至較弱的chaotrope對二氧化矽吸附核酸之吸附行為之影響。我的研究主要是利用恆溫吸附實驗和微熱量滴定卡計(ITC)所得出之熱力學參數分析吸附行為，並以熱力學的角度去觀察上述四種作用力，在不同溶液條件下主導作用力之轉換。
實驗中我們使用不同pH值(靜電作用力、氫鍵)不同鹽濃度以及五種不同種類的鹽(Na2SO4、CaCl2、NaCl、GuHCl、GuSCN)，從恆溫吸附曲線及ITC的數據中，可以發現在低鹽濃度時，雖然在焓(∆H)的部分是些微放熱的，不過自由能(∆G)的貢獻主要來自於熵(∆S)的變化，隨著鹽濃度增加，可以從焓的變化發現Na+和Gu+在高pH值時所扮演的吸附作用力的不同。最後，我們可以從焓(∆H)以及熵(∆S)在不同條件的溶液中發現，上述四種作用力的轉換。

摘要(英)

MicroRNAs (miRNAs) are potential biomarkers that could be applied on the diagnosis and treatment of different disease. However, the lack of knowledge of the binding mechanism of using silica as a material for adsorbing nucleic acids limit the improvement and application.
The driving forces of the adsorption process can be divided into four parts based on different solution conditions (1) shielded intermolecular electrostatic forces (2) dehydration of the DNA and silica surfaces, (3) intermolecular hydrogen bond formation in the DNA–silica contact layer and (4)salt bridge effect between silica and DNA.
This work demonstrates the mechanistic aspects underlying the adsorption behaviors of DNA with mesoporous silica particles in aqueous solution. We changing pH value, ionic strength, three types of salt (NaCl, GuHCl, GuSCN). Using the isotherm adsorption experiment combined with isotherm titration calorimetry (ITC), we observe adsorption mechanism from the viewpoint of thermodynamics. From the ITC data, Na+ and Gu+ play different roles in the adsorption process from the change in reaction enthalpy at high pH value. Finally, we observe the relationship between these four driven forces by the change of enthalpy (∆H) and entropy (∆S).

關鍵字(中)

★ 二氧化矽
★ 無氧核醣核酸
★ 恆溫滴定卡計

關鍵字(英)

★ silica
★ DNA
★ Isothermal Titration Calorimetry

論文目次

摘要 i
ABSTRACT iii
圖目錄 vi
表目錄 xi
一、緒論 1
二、文獻回顧 3
2.1核酸介紹 3
2.1.1核酸分子 3
2.1.2去氧核醣核酸 5
2.1.3核糖核酸結構 7
2.2核酸萃取 8
2.2.1液相核酸萃取 8
2.2.2固相核酸萃取 8
2.3核酸吸附於二氧化矽表面之吸附機制 10
2.3.1吸附作用力 10
2.4Hofmeister series 22
2.5Langmuir adsorption isotherm model 27
2.6恆溫滴定微卡計 28
2.6.1恆溫滴定微卡計介紹 28
2.6.2吸附熱計算 31
三、實驗藥品、儀器及方法 33
3.1實驗藥品 33
3.2儀器設備 34
3.3實驗方法 35
3.3.1恆溫吸附實驗 35
3.3.2恆溫滴定微卡計實驗 36
四、結果與討論 38
4.1pH值對吸附行為之影響 38
4.2鹽類差異 42
4.2.1離子水合能力對吸附行為之影響 42
4.2.2鹽橋效應 50
五、結論 61
六、未來展望 64
七、參考文獻 65
八、附錄 69
恆溫吸附實驗數據 69
ITC數據 70

參考文獻

[1] R. Dahm, "Friedrich Miescher and the discovery of DNA," Developmental biology, vol. 278, no. 2, pp. 274-288, 2005.
[2] J. Watson and F. Crick, "Molecular structure of nucleic acids: a structure for deoxyribose nucleic acid," Jama, vol. 269, no. 15, pp. 1966-1967, 1993.
[3] F. S. Collins, A. Patrinos, E. Jordan, A. Chakravarti, R. Gesteland, and L. Walters, "New goals for the US human genome project: 1998-2003," science, vol. 282, no. 5389, pp. 682-689, 1998.
[4] M. Mandelkern, J. G. Elias, D. Eden, and D. M. Crothers, "The dimensions of DNA in solution," Journal of molecular biology, vol. 152, no. 1, pp. 153-161, 1981.
[5] R. Wing et al., "Crystal structure analysis of a complete turn of B-DNA," Nature, vol. 287, no. 5784, p. 755, 1980.
[6] A. Leslie, S. Arnott, R. Chandrasekaran, and R. Ratliff, "Polymorphism of DNA double helices," Journal of molecular biology, vol. 143, no. 1, pp. 49-72, 1980.
[7] M. C. Wahl and M. Sundaralingam, "Crystal structures of A‐DNA duplexes," Biopolymers: Original Research on Biomolecules, vol. 44, no. 1, pp. 45-63, 1997.
[8] X.-J. Lu, Z. Shakked, and W. K. Olson, "A-form conformational motifs in ligand-bound DNA structures," Journal of molecular biology, vol. 300, no. 4, pp. 819-840, 2000.
[9] R. Boom, C. Sol, M. Salimans, C. Jansen, P. Wertheim-van Dillen, and J. Van der Noordaa, "Rapid and simple method for purification of nucleic acids," Journal of clinical microbiology, vol. 28, no. 3, pp. 495-503, 1990.
[10] S. Jiang, J. Zhuang, C. Wang, J. Li, and W. Yang, "Highly efficient adsorption of DNA on Fe3+–iminodiacetic acid modified silica particles," Colloids and Surfaces A: Physicochemical and Engineering Aspects, vol. 409, pp. 143-148, 2012.
[11] X. Li, J. Zhang, and H. Gu, "Adsorption and desorption behaviors of DNA with magnetic mesoporous silica nanoparticles," Langmuir, vol. 27, no. 10, pp. 6099-6106, 2011.
[12] K. Kang et al., "Preparation and characterization of chemically functionalized silica-coated magnetic nanoparticles as a DNA separator," The Journal of Physical Chemistry B, vol. 113, no. 2, pp. 536-543, 2008.
[13] S. Berensmeier, "Magnetic particles for the separation and purification of nucleic acids," Applied microbiology and biotechnology, vol. 73, no. 3, pp. 495-504, 2006.
[14] Z. Zhang, L. Zhang, L. Chen, L. Chen, and Q. H. Wan, "Synthesis of novel porous magnetic silica microspheres as adsorbents for isolation of genomic DNA," Biotechnology progress, vol. 22, no. 2, pp. 514-518, 2006.
[15] T. Sen, A. Sebastianelli, and I. J. Bruce, "Mesoporous silica− magnetite nanocomposite: fabrication and applications in magnetic bioseparations," Journal of the American Chemical Society, vol. 128, no. 22, pp. 7130-7131, 2006.
[16] L. Liu, Z. Guo, Z. Huang, J. Zhuang, and W. Yang, "Size-selective separation of DNA fragments by using lysine-functionalized silica particles," Scientific reports, vol. 6, p. 22029, 2016.
[17] A. A. Vertegel, R. W. Siegel, and J. S. Dordick, "Silica nanoparticle size influences the structure and enzymatic activity of adsorbed lysozyme," Langmuir, vol. 20, no. 16, pp. 6800-6807, 2004.
[18] L. Han, J. Ruan, Y. Li, O. Terasaki, and S. Che, "Synthesis and characterization of the amphoteric amino acid bifunctional mesoporous silica," Chemistry of materials, vol. 19, no. 11, pp. 2860-2867, 2007.
[19] A. Salis et al., "Ion specific surface charge density of SBA-15 mesoporous silica," Langmuir, vol. 26, no. 4, pp. 2484-2490, 2009.
[20] M. Barisik, S. Atalay, A. Beskok, and S. Qian, "Size dependent surface charge properties of silica nanoparticles," The Journal of Physical Chemistry C, vol. 118, no. 4, pp. 1836-1842, 2014.
[21] A. H. Jalil and U. Pyell, "Quantification of zeta-potential and electrokinetic surface charge density for colloidal silica nanoparticles dependent on type and concentration of the counterion: probing the outer Helmholtz plane," The Journal of Physical Chemistry C, vol. 122, no. 8, pp. 4437-4453, 2018.
[22] J. Nawrocki, "The silanol group and its role in liquid chromatography," Journal of Chromatography A, vol. 779, no. 1-2, pp. 29-71, 1997.
[23] R. Van Wagenen, J. Andrade, and J. Hibbs, "Streaming potential measurements of biosurfaces," Journal of the Electrochemical Society, vol. 123, no. 10, pp. 1438-1444, 1976.
[24] M. L. Hair and W. Hertl, "Acidity of surface hydroxyl groups," The Journal of Physical Chemistry, vol. 74, no. 1, pp. 91-94, 1970.
[25] J. Fripiat, "Silanol groups and properties of silica surfaces," in ACS Symposium Series, 1982, vol. 194, pp. 165-184: AMER CHEMICAL SOC 1155 16TH ST, NW, WASHINGTON, DC 20036 USA.
[26] K. A. Melzak, C. S. Sherwood, R. F. Turner, and C. A. Haynes, "Driving forces for DNA adsorption to silica in perchlorate solutions," Journal of colloid and interface science, vol. 181, no. 2, pp. 635-644, 1996.
[27] M. G. Lorenz and W. Wackernagel, "Adsorption of DNA to sand and variable degradation rates of adsorbed DNA," Applied and environmental microbiology, vol. 53, no. 12, pp. 2948-2952, 1987.
[28] T. H. Nguyen and M. Elimelech, "Plasmid DNA adsorption on silica: kinetics and conformational changes in monovalent and divalent salts," Biomacromolecules, vol. 8, no. 1, pp. 24-32, 2007.
[29] Y. Shen, H. Kim, M. Tong, and Q. Li, "Influence of solution chemistry on the deposition and detachment kinetics of RNA on silica surfaces," Colloids and Surfaces B: Biointerfaces, vol. 82, no. 2, pp. 443-449, 2011.
[30] L. Xu et al., "Altered nucleic acid partitioning during phenol extraction or silica adsorption by guanidinium and potassium salts," Analytical biochemistry, vol. 419, no. 2, pp. 309-316, 2011.
[31] N. Sun et al., "Optimization of influencing factors of nucleic acid adsorption onto silica-coated magnetic particles: application to viral nucleic acid extraction from serum," Journal of Chromatography A, vol. 1325, pp. 31-39, 2014.
[32] Y. Mao, L. N. Daniel, N. Whittaker, and U. Saffiotti, "DNA binding to crystalline silica characterized by Fourier-transform infrared spectroscopy," Environmental Health Perspectives, vol. 102, no. Suppl 10, p. 165, 1994.
[33] W. J. Xie and Y. Q. Gao, "A simple theory for the Hofmeister series," The journal of physical chemistry letters, vol. 4, no. 24, pp. 4247-4252, 2013.
[34] M. Cacace, E. Landau, and J. Ramsden, "The Hofmeister series: salt and solvent effects on interfacial phenomena," Quarterly reviews of biophysics, vol. 30, no. 3, pp. 241-277, 1997.
[35] Y. Zhang and P. S. Cremer, "Interactions between macromolecules and ions: the Hofmeister series," Current opinion in chemical biology, vol. 10, no. 6, pp. 658-663, 2006.
[36] F. Hofmeister, "Archiv für experimentelle pathologie und pharmakologie," Archiv fűr Experimentelle Pathologie und Pharmakologie, vol. 24, p. 247, 1888.
[37] L. M. Pegram and M. T. Record, "Hofmeister salt effects on surface tension arise from partitioning of anions and cations between bulk water and the air− water interface," The journal of physical chemistry B, vol. 111, no. 19, pp. 5411-5417, 2007.
[38] X. Li, J. Zhang, and H. Gu, "Study on the adsorption mechanism of DNA with mesoporous silica nanoparticles in aqueous solution," Langmuir, vol. 28, no. 5, pp. 2827-2834, 2012.
[39] S. L. Sahoo and C.-H. Liu, "Adsorption behaviors of DNA by modified magnetic nanoparticles: effect of spacer and salt," Colloids and Surfaces A: Physicochemical and Engineering Aspects, vol. 482, pp. 184-194, 2015.
[40] K. C. Duong-Ly and S. B. Gabelli, "Salting out of proteins using ammonium sulfate precipitation," in Methods in enzymology, vol. 541: Elsevier, 2014, pp. 85-94.
[41] 陳昱圻, "Improved the process for miRNA isolation from the culture cells by using silica membrane column," 中央大學化材所碩士論文, 2018.

指導教授

陳文逸(Wen-Yih Chen)

審核日期

2019-7-25

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