LLG-5為Higuchi的研究團隊從海星Linckia laevigata身上純化分離所得到的神經節苷脂，並鑑定其結構為8-OMeNeuGcα2→11NeuGcα2→11NeuGcα2→3Galβ1→4Glcβ1→1Cer，同時亦證明在神經生長因子的輔助下，LLG-5具有神經增生的活性，顯示此化合物具有成為治療帕金森氏症等神經系統疾病的藥物潛力。
LLG-5前趨物之合成方法如下：為了可以快速建立植物鞘胺醇（phytosphingosinee）上三個立體中心的結構，我們以來蘇糖（D-lyxose） 做為起始物，經由Wittig反應引入所需的碳鏈長度，得到植物鞘胺醇的類似物，接著再進行Mitsunobu反應將C-2位置的羥基轉為疊氮官能基。接著植物鞘胺醇受體和乳糖衍生物予體進行醣基化反應得到植物神經鞘乳糖脂質（lactosyl phytosphingosine）。具有Neu5Cbz之GM3衍生物合成則是利用建構好的植物神經鞘乳糖脂質為受體與CMP-Neu5Cbz予體，在唾液酸轉移酶的催化下，進行唾液酸醣基化反應，即可生成單一α-位向唾液酸苷鍵結之唾液酸基化醣脂質。總結上述，以乳糖為起始物，經過5個步驟成功合成出LLG-5前趨物，總產率12%。

A new ganglioside, LLG-5 was purified from starfish Linckia laevigata by Higuchi group in 2005 and the structure was determined as 8-OMe-NeuGcα2→11NeuGcα2→11NeuGcα2→3Galβ1→4Glcβ1→1Cer. Moreover, LLG-5 displays neuritogenic activity toward rat pheochromocytoma PC-12 cells in the presence of nerve growth factor. The activity is greater than that of the mammalian ganglioside GM1. Therefore, LLG-5 is regarded as a potential drug to cure nervous system disease such as Parkinson's disease.
In our synthetic approach toward LLG-5, D-lyxose was chosen as the starting material to efficiently construct three stereogenic centers of phytosphingosine, and the lipid chain was introduced via Wittig olefination. After coupling of D-lyxose with lipid Wittig reagent, the hydroxyl group on C2 of the resulting hydroxyl lipid was converted to azido group by Mitsunobu reaction. With the phytosphingosine acceptor in hand, different lactose donors were prepared to investigate their glycosylation with phytosphingosine. The GM3 derivative with Neu5NCbz was synthesized by enzymatic sialylation using CMP-Neu5NCbz and the above lactosyl lipid. By using enzymatic synthesis, we can obtain the exclusively α sialylated glycolipid. In conclusion, we have successfully synthesized the LLG-5 precursor in 12% yield with 5 steps (from lactose).

總目錄
摘要 I
Abstract II
謝誌 III
總目錄 VI
圖目錄 IX
流程目錄 XI
表目錄 XII
縮寫表 XIII
第一章　緒論 1
1-1　前言 1
1-2　神經節苷脂 3
1-3　神經節苷脂LLG-5的發現及其相關研究 4
1-4　神經節苷脂LLG-3合成文獻回顧 6
1-4-1　Sato教授的方法 7
1-4-2　Kiso教授的方法 8
1-4-3　Withers教授的方法 10
1-5　植物鞘胺醇的合成方法 12
1-6　β-位向醣苷鍵的建立 15
1-7　以酵素合成方法建構α-2,3唾液酸苷 17
1-7-1　唾液酸醛縮酶 18
1-7-2　胞苷單磷酸唾液酸合成酶 20
1-7-3　α-2,3-唾液酸轉移酶 21
1-8　研究動機與目的 24
第二章　結果與討論 26
2-1　研究構想與逆合成路徑分析 26
2-2　醣受體-植物鞘胺醇之建構 29
2-2-1　Wittig試劑34之製備 29
2-2-2　合成植物鞘胺醇受體44 31
2-3　植物神經鞘乳糖脂質之合成 36
2-3-1　醣予體全乙醯化之乳糖衍生物45及47之合成 36
2-3-2　全三甲基乙醯化之乳糖衍生物55作為醣予體前驅物 41
2-3-3　植物神經鞘乳糖脂質衍生物56進行全去保護反應 42
2-4　以酵素進行α-2,3唾液酸醣基化 45
2-4-1　合成Neu5Ac-α-(2,3)-Lac-β-phytosphingosine 59 45
2-4-2　合成Neu5Cbz-α-(2,3)-Lac-β-phytosphingosine 26 50
第三章　結論 54
第四章　實驗部分 56
4-1　Materials and Methods 56
4-2　Synthetic Procedures and Characterization 57
第五章　參考文獻 76

1. 郭哲睿，中原大學 碩士論文，藉由化學選擇性醣苷化反應策略以研究具神經再生活性醣苷神經鞘胺脂質SJG-1的全合成，民國101年
2. Schneider, J. S.; Gollomp, S. M.; Sendek, S.; Colcher, A.; Cambi, F.; Du, W. A Randomized, Controlled, Delayed Start Trial of GM1 Ganglioside in Treated Parkinson's Disease Patients. J. Neurol. Sci. 2013, 324, 140-148.
3. Svennerholm, L. Gangliosides -- A new therapeutic agent against stroke and Alzheimer's disease. Life Sci. 1994, 55, 2125-2134.
4. Schengrund, C. L. Gangliosides: glycosphingolipids essential for normal neural development and function. Trends in Biochem. Sci. 2015, 40, 397-406.
5. Molander-Melin, M.; Blennow, K.; Bogdanovic, N.; Dellheden, B.; Månsson, J.-E.; Fredman, P. Structural Membrane Alterations in Alzheimer Brains Found to be Associated with Regional Disease Development; Increased Density of Gangliosides GM1 and GM2 and Loss of Cholesterol in Detergent-Resistant Membrane Domains. J. Neurochem. 2005, 92, 171-182.
6. Kracun, I.; Kalanj, S.; Talan-Hranilovic, J.; Cosovic, C. Cortical distribution of Gangliosides in Alzheimer's Disease. Neurochem. Int. 1992, 20, 433-438.
7. Cuello, A. C. Glycosphingolipids that can Regulate Nerve Growth and Repair. Adv. Pharmacol. 1990, 21, 1-50.
8. Higuchi, R.; Inagaki, M.; Yamada, K.; Miyamoto, T. Biologically active gangliosides from echinoderms. J. Nat. Med. 2007, 61, 367-370.
9. Kaneko, M.; Yamada, K.; Miyamoto, T.; Inagaki, M.; Higuchi, R. Neuritogenic Activity of Gangliosides from Echinoderms and their Structure-Activity Relationship. Chem. Pharm. Bull. 2007, 55, 462-463.
10. Inagaki, M.; Miyamoto, T.; Isobe, R.; Higuchi, R. Biologically Active Glycosides from Asteroidea, 43. Isolation and Structure of a New Neuritogenic-Active Ganglioside Molecular Species from the Starfish Linckia laevigata. Chem. Pharm. Bull. 2005, 53, 1551-1554.
11. Inagaki, M.; Isobe, R.; Higuchi, R. Isolation and Structure of a New Ganglioside Molecular Species. Eur. J. Org. Chem. 1999, 1999, 771-774.
12. Hanashima, S.; Ishikawa, D.; Akai, S.; Sato, K.-i. Synthesis of the Starfish Ganglioside LLG-3 Tetrasaccharide. Carbohydr. Res. 2009, 344, 747-752.
13. Tamai, H.; Ando, H.; Tanaka, H.-N.; Hosoda-Yabe, R.; Yabe, T.; Ishida, H.; Kiso, M. The Total Synthesis of the Neurogenic Ganglioside LLG-3 Isolated from the Starfish Linckia laevigata. Angew. Chem. Int. Ed. 2011, 50, 2330-2333.
14. Rich, J. R.; Withers, S. G. A Chemoenzymatic Total Synthesis of the Neurogenic Starfish Ganglioside LLG-3 Using an Engineered and Evolved Synthase. Angew. Chem. Int. Ed. 2012, 51, 8640-8643.
15. Ndonye, R. M.; Izmirian, D. P.; Dunn, M. F.; Yu, K. O. A.; Porcelli, S. A.; Khurana, A.; Kronenberg, M.; Richardson, S. K.; Howell, A. R. Synthesis and Evaluation of Sphinganine Analogues of KRN7000 and OCH. J. Org. Chem. 2005, 70, 10260-10270.
16. Goff, R. D.; Gao, Y.; Mattner, J.; Zhou, D.; Yin, N.; Cantu, C.; Teyton, L.; Bendelac, A.; Savage, P. B. Effects of Lipid Chain Lengths in α-Galactosylceramides on Cytokine Release by Natural Killer T Cells. J. Am. Chem. Soc. 2004, 126, 13602-13603.
17. Azuma, H.; Tamagaki, S.; Ogino, K. Stereospecific Total Syntheses of Sphingosine and its Analogues from L-Serine. J. Org. Chem. 2000, 65, 3538-3541.
18. Plettenburg, O.; Bodmer-Narkevitch, V.; Wong, C.-H. Synthesis of α-Galactosyl Ceramide, a Potent Immunostimulatory Agent. J. Org. Chem. 2002, 67, 4559-4564.
19. Luo, S.-Y.; Thopate, S. R.; Hsu, C.-Y.; Hung, S.-C. Synthesis of D-ribo-C18-Phytosphingosine from D-Glucosamine via the D-Allosamine Derivatives as Key Intermediates. Tetrahedron Lett. 2002, 43, 4889-4892.
20. Figueroa-Pérez, S.; Schmidt, R. R. Total Synthesis of α-Galactosyl Cerebroside. Carbohydr. Res. 2000, 328, 95-102.
21. Graziani, A.; Passacantilli, P.; Piancatelli, G.; Tani, S. 2-Deoxy- Disaccharide Approach to Natural and Unnatural Glycosphingolipids Synthesis. Tetrahedron: Asymmetry 2000, 11, 3921-3937.
22. Nakamura, T.; Shiozaki, M. Stereoselective Synthesis of D-erythro- sphingosine and L-lyxo-phytosphingosine. Tetrahedron 2001, 57, 9087-9092.
23. Martin, C.; Prünck, W.; Bortolussi, M.; Bloch, R. Enantioselective Synthesis of lyxo-(2R,3R,4R)-C18-Phytosphingosine Using Double Stereodifferentiation1. Tetrahedron: Asymmetry 2000, 11, 1585-1592.
24. He, L.; Byun, H.-S.; Bittman, R. A Stereocontrolled, Efficient Synthetic Route to Bioactive Sphingolipids:  Synthesis of Phytosphingosine and Phytoceramides from Unsaturated Ester Precursors via Cyclic Sulfate Intermediates. J. Org. Chem. 2000, 65, 7618-7626.
25. Lin, C. C.; Fan, G. T.; Fang, J. M. A concise route to phytosphingosine from lyxose. Tetrahedron Lett. 2003, 44, 5281-5283.
26. Demchenko, A. V. Handbook of Chemical Glycosylation: Advanced in Stereoselectivity and Therapeutic Relevance. 4-6.
27. Guo, J., Ye, X.-S. Protecting Groups in Carbohydrate Chemistry: Influence on Stereoselectivity of Glycosylations. Molecules 2010, 15, 7235-7265.
28. Daniels, A. D.; Campeotto, I.; van der Kamp, M. W.; Bolt, A. H.; Trinh, C. H.; Phillips, S. E. V.; Pearson, A. R.; Nelson, A.; Mulholland, A. J.; Berry, A. Reaction Mechanism of N-Acetylneuraminic Acid Lyase Revealed by a Combination of Crystallography, QM/MM Simulation, and Mutagenesis. ACS Chem. Biol. 2014, 9, 1025-1032.
29. Lau, K.; Yu, H.; Thon, V.; Khedri, Z.; Leon, M. E.; Tran, B. K.; Chen, X. Sequential Two-Step Multienzyme Synthesis of Tumor-Associated Aialyl T-Antigens and Derivatives. Org. Biomol. Chem. 2011, 9, 2784-2789.
30. Yu, H.; Chokhawala, H.; Karpel, R.; Yu, H.; Wu, B.; Zhang, J.; Zhang, Y.; Jia, Q.; Chen, X. A Multifunctional Pasteurella multocida Sialyltransferase:  A Powerful Tool for the Synthesis of Sialoside Libraries. J. Am. Chem. Soc. 2005, 127, 17618-17619.
31. Wu, W.-Y.; Jin, B.; Kong, D. C. M.; von Itzstein, M. A Facile Synthesis of a Useful 5-N-Substituted-3,5-Dideoxy-D-Glycero-D-Galacto-Nonulo- sonic Acid from 2-Acet-amido-2-Deoxy-D-Glucose. Carbohydr. Res. 1997, 300, 171-174.
32. Knorst, M.; Fessner, W.-D. CMP-Sialate Synthetase from Neisseria meningitidis − Overexpression and Application to the Synthesis of Oligosaccharides Containing Modified Sialic Acids. Adv. Synth. Catal. 2001, 343, 698-710.
33. Morley, T. J.; Withers, S. G. Chemoenzymatic Synthesis and Enzymatic Analysis of 8-Modified Cytidine Monophosphate-Sialic Acid and Sialyl Lactose Derivatives. J. Am. Chem. Soc. 2010, 132, 9430-9437.
34. Yu, C.-C.; Withers, S. G. Recent Developments in Enzymatic Synthesis of Modified Sialic Acid Derivatives. Adv. Synth. Catal. 2015, 357, 1633-1654.
35. Gilbert, M.; Brisson, J.-R.; Karwaski, M.-F.; Michniewicz, J.; Cunningham, A.-M.; Wu, Y.; Young, N. M.; Wakarchuk, W. W. Biosynthesis of Ganglioside Mimics in Campylobacter jejuni OH4384: Identificatin of the Glycosyltransferase genes, Enzymatic Synthesis of Model Compounds, and Characterization of Nanomole Amounts by 600-MHz 1H and 13C NMR Analysis. J. Biol. Chem. 2000, 275, 3896-3906.
36. Izumi, M.; Shen, G.-J.; Wacowich-Sgarbi, S.; Nakatani, T.; Plettenburg, O.; Wong, C.-H. Microbial Glycosyltransferases for Carbohydrate Synthesis:  α-2,3-Sialyltransferase from Neisseria gonorrheae. J. Am. Chem. Soc. 2001, 123, 10909-10918.
37. Fan, G. T.; Pan, Y. S.; Lu, K. C.; Cheng, Y. P.; Lin, W. C.; Lin, S.; Lin, C. H.; Wong, C. H.; Fang, J. M.; Lin, C. C. Synthesis of α-Galactosyl Ceramide and the Related Glycolipids for Evaluation of their Activities on Mouse Splenocytes. Tetrahedron 2005, 61, 1855-1862.
38. 陳彥男，國立清華大學 博士論文，藉由氟化學的應用及Julia-Kociensky烯烴化反應建構alpha-GalCer分子庫，民國100年
39. Sawant, R. C.; Hung, J.-T.; Chuang, H.-L.; Lin, H.-S.; Chen, W.-S.; Yu, A. L.; Luo, S.-Y. Synthesis of Hydroxylated Analogues of α-Galactosyl Ceramide (KRN7000) with Varying Stereochemistry. Eur. J. Org. Chem. 2013, 2013, 7611-7623.
40. Schmidt, R. R.; Maier, T. Synthesis of D-ribo and L-lyxo-phytosphingosine: Transformation into the corresponding lactosyl- ceramides. Carbohydr. Res. 1988, 174, 169-179.
41. Zhang, Y.; Toyokuni, T; Ruan, F.; Hakomori, S. A One Pot Synthesis of Mono- and Di-Lactosyl Sphingosines. Glycocongugate J. 2001, 18, 557-563.
42. Thompson, M. J.; Hutchinson, E. J.; Stratford, T. H.; Bowler, W. B.; Blackburn, G. M. Sugar Conjugates of Fulvestrant (ICI 182,780): Efficient General Procedures for Glycosylation of the Fulvestrant Core. Tetrahedron Lett. 2004, 45, 1207-1210.
43. Xia, C.; Schümann, J.; Emmanuel, R.; Zhang, Y.; Chen, W.; Zhang, W.; De Libero, G.; Wang, P. G. Modification of the Ceramide Moiety of Isoglobotrihexosylceramide on Its Agonist Activity in Stimulation of Invariant Natural Killer T Cells. J. Med. Chem. 2007, 50, 3489-3496.
44. Wei, G.; Zhang, L.; Cai, C.; Cheng, S.; Du, Y. Selective Cleavage of Sugar Anomeric O-Acyl Groups Using FeCl3·6H2O. Tetrahedron Lett. 2008, 49, 5488-5491.
45. Yao, Q.; Song, J.; Xia, C.; Zhang, W.; Wang, P. G. Chemoenzymatic Syntheses of iGb3 and Gb3. Org. Lett. 2006, 8, 911-914.
46. 官亭君，國立清華大學 博士論文，合成硫鍵結之α-2,8唾液酸寡醣體與GD3抗原，民國103年
47. Greene, T. W.; Wuts, P. G. M. Protective Groups in Organic Synthesis 3rd Edition. 170-172.
48. Nicolaou, K. C.; Caulfield, T. J.; Kataoka, H.; Stylianides, N. A. Total Synthesis of Tumor-Associated Lex Family of Glycosphingolipid. J. Am. Chem. Soc. 1990, 112, 3693-3695.
49. Ogilvie, K. K.; Iwacha, D. J. Use of tert-Butyldimethylsilyl Group of Protecting the Hydroxyl Functions of Nucleosides. Tetrahedron Lett. 1973, 14, 317-319.
50. Gassman, P. G.; Schenk, W. N. A General Procedure for the Base-Promoted Hydrolysis of Hindered Esters at Ambient Temperatures. J. Org. Chem. 1977, 42, 918-920.
51. van Boeckel, C. A. A.; van Boom, J. H. Synthesis of Glucosylphophatidylglycerol via a Phosphotriester Intermediate. Tetrahedron Lett. 1979, 20, 3561-3564.
52. Pukin, A. V.; Weijers, C. A. G. M.; van Lagen, B.; Wechselberger, R.; Sun, B.; Gilbert, M.; Karwaski, M.-F.; Florack, D. E. A.; Jacobs, B. C.; Tio-Gillen, A. P.; van Belkum, A.; Endtz, H. P.; Visser, G. M.; Zuilhof, H. GM3, GM2 and GM1 Mimics Designed for Biosensing: Chemoenzymatic Synthesis, Target Affinities and 900 MHz NMR Analysis. Carbohydr. Res. 2008, 343, 636-650.
53. Sparks, M. A.; Williams, K. W.; Lukacs, C.; Schrell, A.; Priebe, G.; Spaltenstein, A.; Whitesides, G. M. Synthesis of Potential Inhibitors of Hemagglutination by Influenza Virus: Chemoenzymatic Preparation of N-5 Analogs of N-Acetylneuraminic Acid. Tetrahedron 1993, 49, 1-12.
54. Morita, M.; Sawa; E; Yamaji, K.; Sakai, T.; Natori, T; Koezuka, Y; Fukushima, H.; Akimoto, K. Practical Total Synthesis of (2S,3S,4R)-1-O-(α-D-Galactopyranosyl)-N-hexa-cosanoyl-2-amino-1,3,4- octadecanetriol, the Antitumorial and Immunostimulatory α-Galactosylcer- amide, KRN7000. Biosci. Biotechnol. Biochem. 1996, 60, 288–292.
55. Nanda, S.; Yadav, J. S. Asymmetric synthesis of unnatural (Z,Z,E)-ocatadecatrienoid and eicosatrienoid by lipoxygenase-catalyzed oxygenation. Tetrahedron: Asymmetry 2003, 14, 1799-1806.
56. Martinková, M.; Pomikalová, K.; Gonda, J.; Vilková, M. A Common Approach to the Total Synthesis of L-arabino-, L-ribo-C18-Phytos- phingosines, ent-2-epi-Jaspine B and 3-epi-Jaspine B from D-Manose. Tetrahedron 2013, 69, 8228-8244.
57. Chiu, H. Y.; Tzou, D. L. M.; Patkar, L. N.; Lin, C. C. A Facile Synthesis of Phytosphingosine from Diisopropylidene-D-mannofuranose. J. Org. Chem. 2003, 68, 5788-5791.
58. Costantino, V.; Fattorusso, E.; Imperatore, C.; Mangoni, A. Immunomodulating Glycosphingolipids: an Efficient Synthesis of a 2'-Deoxy-α-Galactosyl-GSL. Tetrahedron 2002, 58, 369-375.