臺灣博碩士論文加值系統

基質輔助雷射脫附游離質譜法(MALDI-TOF MS)可以快速偵測樣品以測定生物分子的質量。然而，MALDI-TOF MS在低分子量範圍嚴重的基質干擾限制了其對小分子的偵測。近來，功能化奈米材料的發展被應用在小分子無背景雜訊的分析上。利用實驗室建立的基質功能化磁奈米粒子(DHB@MNP)作為無背景雜訊測量，我們開發出可調式游離碎裂方法，透過調整基質功能化磁奈米粒子的不同濃度，可以在一級質譜模式下產生分析物碎片，作為快速結構鑑定的工具。
在生物分子之中，醣具有低游離效率以及容易解離產生碎片的特性造成其分析較為困難。在這裡我們使用醣異構物作為模型，在低濃度奈米粒子的情況下可以得到無背景雜訊且較強的完整分子量訊號。另一方面，使用高濃度的奈米粒子作為基質則能提供分析物較高內能，促使醣碎裂。與其他二級質譜比較，此種方法較容易造成醣的跨環碎裂並且產生獨特的碎片。透過控制分析物碎裂的程度，這個方法可以用來鑑定蔗醣和乳醣等雙醣異構物，LewisA和LewisX三醣異構物，以及LewisB和LewisY四醣異構物。另外，在本論文中也對奈米粒子的組成材料在游離及碎裂機制中所扮演的角色加以探討，推測出下列的結論：在DHB@MNP中，氧化鐵核會吸收吸收雷射能量傳遞給分析物造成游離及碎裂。包覆在其外的二氧化矽層保護氧化鐵核以大幅減低其碎裂產生的背景訊號，並且緩衝氧化鐵核傳遞的能量使低濃度奈米粒子可用於完整分子量分析。而共價修飾在奈米粒子表面的基質亦可以吸收雷射，提高醣分析物以及其碎片的游離效率。

Before advances in tandem MALDI MS had been developed for unambiguous structure determination, MALDI-TOF MS has traditionally been used only for the molecular weight determination of biomolecules. However, analysis of low molecular weight analytes in MALDI-TOF MS is limited due to the severe interference by matrix signals. Recent progresses on the functionalized nanomaterials have impacted the background-free analysis of small molecule. By using matrix functionalized nanoparticles for background-free measurement, we reported a tunable ionization/fragmentation strategy to generate analyte product ions in the MS mode by tuning different concentration of surface modified nanoparticles for rapid structural characterization.
Among these biomolecules, the analysis of carbohydrates is relatively challenging due to their inherent low ionization efficiency. Using the isomeric saccharides as model system, soft and background-free measurement can be achieved with enhanced signals under low nanoparticle concentration. On the other hand, high concentration of nanoparticle imparted high internal energy to the analytes and promoted fragmentation. Compared with other MS/MS strategies, DHB@MNP is likely to induce oligosaccharide corss-ring cleavage and generate unique fragments. Based on the controllable extent of the soft/harsh ionization, this strategy distinguished isomeric sucrose and lactose, LewisA and LewisX trisaccharides, and LewisB and LewisY tetrasaccharides. Also, the investigation of key components of the composition of nanoparticle in the ionization and fragmentation mechanism suggested three important features of DHB@MNP: (1) the energy pooling by the Fe3O4 core may account for ionization and fragmentation of the oligosaccharides at high concentration of the MNP, (2) the coating of SiO2 reduces the background signals and mitigates the internal energy imparted by the core, and (3) the DHB conjugated on the surface enhance the signals of the analyte and its fragments.

謝誌 i
中文摘要 ii
ABSTRACT iii
CONTENTS v
LIST OF FIGURES vii
LIST OF TABLE ix
ABBREVIATIONS x
1 Introduction 1
1.1 MALDI MS analysis for small molecule 1
1.2 Nanoparticle-based MALDI-TOF MS 1
1.3 Ionization Mechanism-Soft ionization and Harsh ionization 3
1.3.1 Electron Ionization (EI) 4
1.3.2 Fast Atom Bombardment (FAB) 4
1.3.3 Laser Desorption Ionization (LDI) and Matrix-assisted Laser Desorption/Ionization (MALDI) 5
1.3.4 Electrospray Ionization (ESI) 6
1.4 Oligosaccharide Analysis by MALDI-TOF MS 6
1.5 Thesis Objective 8
2 Materials and Methods 11
2.1 Materials 11
2.2 Fabrication of Nanoparticles 11
2.2.1 Fabrication of Fe3O4 MNP 12
2.2.2 Fabrication of SiO2@MNP 12
2.2.3 Fabrication of DHB@MNP 13
2.3 Sample Deposition Method 13
2.4 MALDI-TOF MS Analysis 14
2.5 The Survival Yield Calculation 15
2.6 FAB and ESI MS Analysis 15
3 Results and Discussion 16
3.1 Nanoparticle-assisted MALDI-TOF MS 16
3.2 Comparison with Different Nanoparticles 17
3.3 DHB@MNP Concentration-Dependent Fragmentation 19
3.4 Validation for Oligosaccharides Fragments by MALDI-TOF MS/MS 22
3.5 Comparison with other MS/MS methods 24
3.6 Laser intensity effect 27
3.7 The Role of Each Component on DHB@MNP in the Energy-Transfer Mechanism 29
3.8 Differentiation between Isomeric Oligosaccharides 31
4 Conclusion 38
Reference 40

1.Karas, M.; Bachmann, D.; Hillenkamp, F. Anal. Chem. 1985, 57, 2935-2939.
2.Tanaka, K.; Waki, H.; Ido, Y.; Akita, S.; Yoshida, Y.; Yoshida, T.; Matsuo, T. Rapid Commun Mass Spectrom 1988, 2, 151-153.
3.Carre, V.; Aubriet, F.; Scheepers, P. T.; Krier1, G.; Muller, J.-F. Rapid Commun Mass Spectrom 2005, 19, 871-880.
4.Sleno, L.; Volmer, D. A. Rapid Commun Mass Spectrom 2005, 19, 1928-1936.
5.Guo, Z.; Zhang, Q.; Zou, H.; Guo, B.; Ni, J. Anal. Chem. 2002, 74, 1637-1641.
6.Cohen, L. H.; Gusev, A. I. Anal Bioanal Chem 2002, 373, 571-586.
7.Cohen, L.; Go, E. P.; Siuzdak, G. Small-molecule desorption/ionization mass analysis, In MALDI MS: A Practical Guide to Instrumentation, Methods and Applications; Hillenkamp, F., Peter-Katalinić, J., Eds.; Wiley-VCH Verlag GmbH & Co. KGaA: Weinheim, 2007
8.Wei, J.; Buriak, J. M.; Siuzdak, G. Nature 1999, 399, 243-246.
9.Trauger, S. A.; Go, E. P.; Shen, Z.; Apon, J. V.; Compton, B. J.; Bouvier, E. S. P.; Finn, M. G.; Siuzdak, G. Anal. Chem. 2004, 76, 4484-4489.
10.Go, E. P.; Apon, J. V.; Luo, G.; Saghatelian, A.; Daniels, R. H.; Sahi, V.; Dubrow, R.; Cravatt, B. F.; Vertes, A.; Siuzdak, G. Anal. Chem. 2005, 77, 1641-1646.
11.Xu, S.; Li, Y.; Zou, H.; Qiu, J.; Guo, Z.; Guo, B. Anal. Chem. 2003, 75, 6191-6195.
12.Pan, C.; Xu, S.; Hu, L.; Su, X.; Ou, J.; Zou, H.; Guo, Z.; Zhang, Y.; Guo, B. J. Am. Soc. Mass Spectrom. 2005, 16, 883-892.
13.Ren, S.-f.; Zhang, L.; Cheng, Z.-h.; Guoa, Y.-l. J. Am. Soc. Mass Spectrom. 2005, 16, 333-339.
14.Huang, Y.-F.; Chang, H.-T. Anal. Chem. 2006, 78, 1485-1493.
15.Shrivas, K.; Wu, H.-F. Rapid Commun Mass Spectrom 2008, 22, 2863-2872.
16.Yuan, M.; Shan, Z.; Tian, B.; Tu, B.; Yang, P.; Zhao, D. Micropor. Mesopor. Mater. 2005, 78, 37-41.
17.Shana, Z.; Hana, L.; Yuana, M.; Denga, C.; Zhaoa, D.; Tu, B.; Yang, P. Anal. Chim. Acta 2007, 593, 13-19.
18.Chiang, C.-K.; Chiang, N.-C.; Lin, Z.-H.; Lan, G.-Y.; Lin, Y.-W.; Chang, H.-T. J. Am. Soc. Mass Spectrom. 2010, 21, 1204-1207.
19.Chen, W.-T.; Chiang, C.-K.; Lin, Y.-W.; Chang, H.-T. J. Am. Soc. Mass Spectrom. 2010, 21, 864-867.
20.Chiang, C.-K.; Yang, Z.; Lin, Y.-W.; Chen, W.-T.; Lin, H.-J.; Chang, H.-T. Anal. Chem. 2010, 82, 4543-4550.
21.Lin, P.-C.; Tseng, M.-C.; Su, A.-K.; Chen, Y.-J.; Lin, C.-C. Anal. Chem. 2007, 79, 3401-3408.
22.Tseng, M.-C.; Obena, R.; Lu, Y.-W.; Lin, P.-C.; Lin, P.-Y.; Yen, Y.-S.; Lin, J.-T.; Huang, L.-D.; Lu, K.-L.; Lai, L.-L.; Lin, C.-C.; Chen, Y.-J. J. Am. Soc. Mass Spectrom. 2010, 21, 1930-1939.
23.Pokrovskii, V. A.; Mosin, V. V. Theoret. Exp. Chem. 1987, 23, 58-73.
24.Varki, A. Glycobiology 1993, 3, 97-130.
25.Dwek, R. A.; Butters, T. D. Chem. Rev. 2002, 102, 283-284.
26.Zaia, J. Mass Spectrom. Rev. 2004, 23, 161-227.
27.Viseux, N.; Costello, C. E.; Domon, B. J. Mass Spectrom. 1999, 34, 364-376.
28.Chernushevich, I. V.; Thomson, B. A. Anal. Chem. 2004, 76, 1754-1760.
29.Opsal, R. B.; Owens, K. G.; Reilly, J. P. Anal. Chem. 1985, 57, 1884-1889.
30.Burlingame, A. L. Collision-induced dissociation (CID) of peptides and proteins, In Biological mass spectrometry; Burlingame, A. L., Ed.; Gulf Professional Publishing: 2005
31.Tseng, K.; Lindsay, L.-L.; Penn, S.; Hedrick, J.-L.; Lebrilla, C.-B. Anal. Biochem. 1997, 250, 18-28.
32.Cancilla, M. T.; Wong, A. W.; Voss, L. R.; Lebrilla, C. B. Anal. Chem. 1999, 71, 3206-3218.
33.Domon, B.; Costello, C. E. Glycoconjugate J. 1988, 5, 397-409.
34.Yamagaki, T.; Nakanishi, H. J. Mass Spectrom. 2000, 35, 1300-1307.
35.Ferreira, J. A.; Domingues, M. R. M.; Reis, A.; Monteiro, M. A.; Coimbra, M. A. Anal. Biochem. 2010, 397.
36.Obena, R. P., University of the Philippines, Diliman, 2011.
37.Wang, M.-T.; Liu, M.-H.; Wang, C. R. C.; Chang, S. Y. J. Am. Soc. Mass Spectrom. 2009, 20, 1925-1932.
38.Sandeep, C. S. S.; Samal, A. K.; Pradeep, T.; Philip, R. Chem. Phys. Lett. 2010, 486, 326-330.
39.Willis, A. L.; Turro, N. J.; O''Brien, S. Chem. Mater. 2005, 17, 5970-5975.
40.Li, Y.-S.; Church, J. S.; Woodhead, A. L.; Mouss, F. Spectrochim. Acta. Part A. 2010, 76, 484-489.
41.Gorecka-Drzazga, A.; Bargiel, S.; Walczak, R.; Dziuban, J. A.; Kraj, A.; Dylag, T.; Silberring, J. Sensor Actuator B-Chem. 2004, 103, 206-212.
42.Zhigilei, L. V.; Kodali, P. B. S.; Garrison, B. J. J. Phys. Chem. B 1997, 101, 2028-2037.
43.Zhigilei, L. V.; Garrison, B. J. Appl. Phys. Lett. 1999, 74, 1341 - 1343.
44.Knochenmuss, R. The Analyst. 2006, 131, 966-986.
45.Orlando, R.; Bush, C. A.; Fenselau, C. Biol. Mass Spectrom. 1990, 19, 747-754.