食用蘑菇富有營養價值。近年來，食用蘑菇多醣的生物活性引發了越來越多的關注。因此，爲了理解結構與活性（SAR），就必須瞭解多醣體的化學結構。在本篇研究中，選用了常被食用的的姬松茸進行研究。通過冷水萃取姬松茸中多醣，接著通過冷丙酮將多醣沉澱，分子篩層析法和離子交換樹脂法進行多醣的分離與純化。生物活性的部分則針對多醣對免疫刺激活性以及α-葡萄糖苷酶的抑制能力進行研究。純化後的多醣體則通過氣相層析質譜儀（GC-MS）和核磁共振（NMR）光譜進行解析。
實驗結果表明ASP-1具有α-葡萄糖苷酶抑制活性，在濃度為2.5 mg/mL時，抑制能力達58%。通過氣相層析質（GC-MS）和核磁共振（NMR）光譜分析，證明姬松茸多醣（ASP-1）主要由66%的半乳糖，以及15%的岩藻糖、10%的葡萄糖和9%的甘露糖所組成。藉由醣鍵結分析姬松茸多醣（ASP-1）的骨架是由1,4-連接的葡萄糖基、1,2-連接的半乳糖基、1,6-連接的半乳糖基、1,2,6-連接的半乳糖基以及具末端鍵結的岩藻糖、葡萄糖和甘露糖組成。
本研究表明姬松茸多醣（ASP-1）具有抑制α-葡萄糖苷酶的活性，希望在不久的將來姬松茸多醣（ASP-1）可以用作為食品添加劑或是食品補充劑用來治療第二型糖尿病。

Edible mushrooms are a good source of valuable nutritious ingredients. Recently, mushroom polysaccharides attract increasing attentions on their biological activities. Therefore, elucidation of their chemical structures is imperative for understanding structure-activity relationship (SAR). In the present study, a popular mushroom species, Agaricus blazei Murrill (姬松茸), was selected for studying. The polysaccharide of Agaricus blazei Murrill was extracted by cold water first and precipitated by cold-acetone, and sequentially purified by size exclusion and ion exchange chromatographies. Biological activities of polysaccharides were investigated to study immunological activity and α-d-glucosidase inhibitory activity. Chemical structure of purified polysaccharide was analyzed by gas chromatography-mass spectroscopy (GC-MS) and nuclear magnetic resonance (NMR) spectroscopy.
Experimental results revealed that water-soluble polysaccharide ASP-1 had some inhibitory activity on α-glucosidase in a dose-dependent manner, with the inhibitory rate of 58% when the concentration reached 2.5 mg/mL. The chemical structure of ASP-1 was analyzed by gas chromatography-mass spectroscopy and NMR spectra. The results demonstrated that the polysaccharide of Agaricus blazei Murrill (ASP-1; MW~70 kDa) was composed of galactose (66%), fucose (15%), glucose (10%) and mannose (9%). According to the combination of chemical and instrumental analysis, the results indicated that ASP-1 had a backbone consisting of (1 → 4)-linked-α-D-glucopyranosyl, (1 → 2)-linked-α-D- galactopyranosyl, (1 → 6)-linked-α-D-galactopyranosyl and (1 → 2, 6)-linked-α-D-glucopyranosyl linkage, and terminal mannosyl, fucosyl and galactosyl linkage, respectively.
The present study indicated that ASP-1 has inhibitory activity of α-glucosidase, which may have potential as a food additive or as a food supplement for an enzyme-targeted treatment of Type 2 diabetes in the near future.

中文摘要 i
Abstract ii
Contents iv
List of Figure vii
List of Table ix
1. Introduction 1
1.1. Mushroom 1
1.2. Fungal Polysaccharides 2
1.3. Agaricus blazei Murrill 4
1.3.1. Morphology of Agaricus blazei Murrill 5
1.3.2. Taxonomy of Agaricus blazei Murrill 6
1.3.3. Chemical composition of Agaricus blazei Murrill 6
1.3.4. Ethnomedical significance of Agaricus blazei Murrill 7
1.3.5. Biological activities of polysaccharides of Agaricus blazei Murrill 8
1.4. Structural analysis of polysaccharides 10
1.5. Nuclear Magnetic Resonance spectroscopy in polysaccharide analysis 11
2. Materials and methods 13
2.1. Materials 13
2.2. Extraction of polysaccharides 14
2.3. Purification of polysaccharides by size exclusion chromatography 16
2.4. Limulus amebocyte lysate ( LAL) assay 17
2.5. Cell cultures 18
2.6. Cytokine production assay 18
2.7. Purification of the polysaccharides by ion exchange chromatography 19
2.8. Determination of α-glycosidase inhibitory activity 20
2.9. Deacetylation of polysaccharides 21
2.10. Monosaccharide composition 21
2.11. Linkage analysis 24
2.12. Nuclear magnetic resonance (NMR) spectroscopy 26
3. Results 27
3.1. Extraction and purification of water-soluble polysaccharides 27
3.2. LAL assay 28
3.3. Cytokine measurement 28
3.4. Purification of the polysaccharides by ion exchange chromatography 30
3.5. α-Glucosidase inhibitory activity of ASP-1 and deacetylated ASP-1 33
3.6. Monosaccharide composition analysis of ASP-1 by GC-MS 35
3.7. Linkage analysis of ASP-1 36
3.8. Determination of the sugar residues and their sequence by NMR 38
3.8.1. 1D 1H NMR Spectrum 38
3.8.2. 1D 13C NMR spectrum 40
3.8.3. 1D 13C DEPT-135 spectrum 41
3.8.4. 2D 1H-1H COSY 42
3.8.5. 2D 1H-13C HSQC 45
3.8.6. 2D 1H-13C HMBC 47
3.8.7. Summary of the NMR spectrum analysis of ASP-1 49
4. Discussion 51
5. References 54
Appendix 67

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