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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 黃良得(Lean-Teik Ng) | |
| dc.contributor.author | Chun-Han Su | en |
| dc.contributor.author | 蘇俊翰 | zh_TW |
| dc.date.accessioned | 2021-06-17T00:10:50Z | - |
| dc.date.available | 2015-07-20 | |
| dc.date.copyright | 2012-07-20 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-07-14 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/65740 | - |
| dc.description.abstract | 舞菇 (Grifola frondosa；GF) 為傳統上用來促進免疫功能及治療糖尿病之藥用真菌。第二型糖尿病為最普遍的慢性病之一，而抑制其相關消化酶活性之抑制劑可降低餐後高血糖，具有預防或治療第二型糖尿病之功效。因此，本研究目的為：(1) 評估舞菇不同萃取物對第二型糖尿病相關消化酶 (α-澱粉酶及α-葡萄糖苷酶) 之抑制效果及鑑定其活性成分；(2) 探討舞菇活性成分對α-葡萄糖苷酶之抑制特性；及 (3) 利用模擬小腸環境模式分析舞菇活性成分對澱粉水解的影響。舞菇子實體不同萃取物及多醣體對α-澱粉酶及α-葡萄糖苷酶的抑制能力均以比色法進行測定。結果顯示，所有舞菇成分對α-澱粉酶之抑制效果皆不佳，但非極性成分 (GF-H 及GF-oil) 對α-葡萄糖苷酶則具有極佳之抑制效果。以氣相層析質譜儀分析得知，此非極性成分中之主要化合物為油酸 (IC50 = 0.0779 mM) 和亞麻油酸 (IC50 = 0.112 mM)，兩者對α-葡萄糖苷酶具極佳之抑制活性，且抑制效果皆優於標準品acarbose (IC50 = 2.91 mM)。構效關係亦指出在不同脂肪酸中，油酸及亞麻油酸確實對α-葡萄糖苷酶具有較強之抑制能力。抑制動力學分析結果指出油酸和亞麻油酸皆屬於競爭型抑制劑，而螢光淬滅之實驗結果顯示其能和α-葡萄糖苷酶結合形成複合體，但圓二色光譜分析結果則指出油酸及亞麻油酸對α-葡萄糖苷酶之二級結構影響不大。另外，油酸和亞麻油酸在模擬小腸環境的透析模式中，可顯著降低澱粉水解率，代表其可延緩澱粉在小腸中的消化速率，進而降低餐後高血糖。本研究首次指出，屬於不飽和脂肪酸的油酸及亞麻油酸對α-葡萄糖苷酶活性具有極佳之抑制效果，故具開發為副作用較少之α-葡萄糖苷酶抑制劑的潛力，以用於預防或治療第二型糖尿病。 | zh_TW |
| dc.description.abstract | Grifola frondosa (GF), also known as Wu-gu (舞菇), is a medicinal fungus traditionally used to enhance immune functions and for treating diabetes. Type 2 diabetes is one of the most common chronic diseases, and inhibitors for type 2 diabetes-related digestive enzymes which reduce postprandial hyperglycemia play a key role in preventing and treating type 2 diabetes. Therefore, the aims of this study were: (i) to evaluate the inhibitory effects of different GF components on type 2 diabetes-related digestive enzymes (α-amylase and α-glucosidase) and to identify their bioactive components; (ii) to investigate the α-glucosidase inhibitory characteristics of GF
bioactive components; and (iii) to examine the effects of GF bioactive components on starch hydrolysis in the simulated small intestine conditions. The inhibitory effects of different extracts, fractions, and polysaccharides of GF fruiting bodies on α-amylase and α-glucosidase were evaluated by in vitro colorimetric methods. Results showed that all GF components exhibited weak inhibitory effects on α-amylase activity, whereas the non-polar components possessed good anti-α-glucosidase activity, with the strongest potency on this activity was noted on GF-H and GF-oil. GC-MS analysis showed that the bioactive components contained mainly of oleic acid (IC50 = 0.0779 mM) and linoleic acid (IC50 = 0.112 mM), which were found to possess potent anti-α-glucosidase activity, and were noted to be stronger than acarbose (IC50 = 2.91 mM). Structure-activity relationship study further confirmed that among the different fatty acids, oleic acid and linoleic acid possessed the most potent inhibitory activity against α-glucosidase. Inhibition kinetics showed that oleic acid and linoleic acid were competitive inhibitors, and fluorescence quenching study showed that they would bind to α-glucosidase to form a complex, while the results of circular dichroism spectroscopy indicated that these inhibitors had little effects on the secondary structure of α-glucosidase. In addition, oleic acid and linoleic acid significantly decreased the hydrolysis rate of starch in a dialysis model which mimics the small intestine conditions, indicating that they could retard the digestion of starch in the small intestine and hence decrease the postprandial blood glucose level. This is the first report to show that unsaturated fatty acids oleic acid and linoleic acid possessed strong inhibitory effects on α-glucosidase activity, suggesting that they have the potential to be developed as new α-glucosidase inhibitors with less side effects for the treatment of type 2 diabetes. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T00:10:50Z (GMT). No. of bitstreams: 1 ntu-101-R99623001-1.pdf: 7054260 bytes, checksum: e200f6da1f619e7cee4c49be9a2c43ad (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | Content
口試委員會審定書 ........................................................................................................... I 誌謝 .................................................................................................................................. II 摘要 ................................................................................................................................ III Abstract ........................................................................................................................... IV Content ........................................................................................................................... VI Figure content ................................................................................................................... X Table content .................................................................................................................. XII Chapter 1 Introduction ...................................................................................................... 1 1.1. Medicinal mushrooms and G. frondosa ............................................................. 3 1.2. Bioactive polysaccharides of G. frondosa ......................................................... 6 1.3. Pharmacological activities of G. frondosa ......................................................... 8 1.3.1. Immunomodulation ................................................................................ 8 1.3.2. Anti-cancer ............................................................................................. 8 1.3.3. Anti-hyperglycemia .............................................................................. 10 1.3.4. Anti-hyperlipidemia and anti-hypertension .......................................... 10 1.3.5. Anti-inflammation ................................................................................. 11 1.3.6. Anti-oxidation ........................................................................................ 11 1.3.7. Anti-virus, hepatoprotection, and modulation of sebaceous lipogenesis ............................................................................................ 12 1.4. Diabetes mellitus ............................................................................................. 13 1.4.1. Classification ........................................................................................ 13 1.4.2. Treatments of type 2 diabetes ............................................................... 14 Chapter 2 Inhibitory effects of G. frondosa components on type 2 diabetes-related digestive enzymes and identification of their bioactive components ............ 18 2.1. Introduction ..................................................................................................... 18 2.2. Materials and methods ..................................................................................... 21 2.2.1. Materials ............................................................................................... 21 2.2.2. G. frondosa extract preparation ............................................................ 21 2.2.3. Crude polysaccharide preparation ........................................................ 22 2.2.4. G. frondosa fraction preparation ........................................................... 22 2.2.5. Fractionation of n-hexane extract ......................................................... 23 2.2.6. α-Amylase inhibition assay .................................................................. 27 2.2.7. α-Glucosidase inhibition assay ............................................................. 27 2.2.8. Identification and quantification of the bioactive components ............ 28 2.2.8.1. Preparation of standard solutions and sample for gas chromatography-mass spectrometry (GC-MS) analysis ......... 28 2.2.8.2. GC-MS analysis ....................................................................... 29 2.3. Results and discussion ..................................................................................... 30 2.3.1. Yields of different G. frondosa components ........................................ 30 2.3.2. α-Amylase and α-glucosidase inhibitory activities .............................. 30 2.3.3. Identification and quantification of the bioactive components of G. frondosa ........................................................................................... 35 2.3.4. Structure-activity relationship .............................................................. 47 2.4. Conclusions ..................................................................................................... 51 Chapter 3 Physical and chemical characteristics of the interaction between the bioactive components of G. frondosa and α-glucosidase .............................. 52 3.1. Introduction ..................................................................................................... 52 3.2. Materials and methods ..................................................................................... 56 3.2.1. Materials ............................................................................................... 56 3.2.2. Sample preparation ............................................................................... 56 3.2.3. Kinetic assay of α-glucosidase inhibition ............................................. 56 3.2.4. Fluorescence spectra measurements ..................................................... 57 3.2.5. Circular dichroism measurements ........................................................ 57 3.3. Results and discussion ..................................................................................... 58 3.3.1. Kinetic assay of α-glucosidase inhibition ............................................. 58 3.3.2. Fluorescence spectra ............................................................................. 62 3.3.3. Mechanism of fluorescence quenching ................................................ 62 3.3.4. Circular dichroism spectra .................................................................... 67 3.4. Conclusions ..................................................................................................... 70 Chapter 4 Study on the effects of G. frondosa bioactive components on starch hydrolysis ...................................................................................................... 71 4.1. Introduction ..................................................................................................... 71 4.2. Materials and methods ..................................................................................... 72 4.2.1. Materials ............................................................................................... 72 4.2.2. Sample preparation ............................................................................... 72 4.2.3. Determination of starch hydrolysis rate................................................ 72 4.2.4. Statistical analysis ................................................................................ 73 4.3. Results and discussion ..................................................................................... 74 4.3.1. Establishment of a dialysis model which mimics the small intestine conditions ............................................................................................. 74 4.3.2. Determination of the starch hydrolysis rate .......................................... 74 4.4. Conclusions ..................................................................................................... 79 Chapter 5 Overall conclusions ........................................................................................ 80 References ...................................................................................................................... 82 Publications .................................................................................................................... 96 Figure content Figure 1. Dried fruiting bodies of G. frondosa ................................................................. 5 Figure 2. Structure of the β-glucan in D-fraction ............................................................. 7 Figure 3. Mechanism of action of different types of oral anti-diabetic agents ............... 17 Figure 4. α-Glucosidase inhibitors inhibit the activity of α-glucosidase enzymes in the brush border of small intestinal enterocytes ......................................... 20 Figure 5. An overview of the G. frondosa sample preparation ...................................... 25 Figure 6. Structures of ergosterol and ergosterol peroxide ............................................. 26 Figure 7. An overview on the applications of GC/MS and LC/MS techniques for selected compound classes ............................................................................. 38 Figure 8. Structures of derivatization reagents and the trimethylsilylation reaction of sample and reagents ................................................................................... 39 Figure 9. Total ion chromatograms ................................................................................. 40 Figure 10. Structures of identified fatty acids in GF-H and GF-oil ............................... 41 Figure 11. Mass spectra of identified peaks of GF-H ..................................................... 42 Figure 12. Structures of cis-unsaturated fatty acids with different carbon chain lengths and different numbers of double bonds ............................................ 50 Figure 13. Different types of reversible inhibition and their Lineweaver-Burk plots .... 54 Figure 14. Linear regression curves of reaction rates versus different volumes of α-glucosidase with or without samples ......................................................... 59 Figure 15. Reaction curves of α-glucosidase with or without samples .......................... 60 Figure 16. Lineweaver-Burk plot of the reactions of α-glucosidase with or without samples .......................................................................................................... 61 Figure 17. Fluorescence spectra of α-glucosidase interacted with different concentrations of samples ............................................................................. 64 Figure 18. The Stern-Volmer curves of α-glucosidase interacted with different concentrations of oleic acid and linoleic acid ............................................... 65 Figure 19. The Perrin curves of α-glucosidase interacted with different concentrations of oleic acid and linoleic acid ............................................... 65 Figure 20. Circular dichroism spectra of α-glucosidase interacted with different concentrations of samples ............................................................................. 68 Figure 21. Starch hydrolysis curves of samples at different concentrations .................. 76 Figure 22. Starch hydrolysis curves of different samples at the concentration of 1.2 mg/mL ................................................................................................. 77 Table content Table 1. Yields of G. frondosa components .................................................................... 33 Table 2. Inhibitory concentrations of G. frondosa components on α-amylase and α-glucosidase activities ..................................................................................... 34 Table 3. Peak identification of trimethylsilyl (TMS) derivatives of fatty acids ............. 43 Table 4. Calibration curves of fatty acids ....................................................................... 44 Table 5. Contents of fatty acids in GF-H and GF-oil ..................................................... 45 Table 6. Inhibitory concentrations of fatty acids on α-amylase and α-glucosidase activities ............................................................................................................ 46 Table 7. Inhibitory concentrations of different saturated fatty acids and unsaturated fatty acids on α-amylase and α-glucosidase activities ...................................... 49 Table 8. Effects of reversible inhibitors on apparent Km and apparent Vmax ................... 55 Table 9. Ki values of oleic acid and linoleic acid on α-glucosidase ............................... 61 Table 10. KSV and Kq values of the interaction between α-glucosidase and samples ..... 66 Table 11. Kp values of the interaction between α-glucosidase and samples ................... 66 Table 12. Effects of oleic acid, linoleic acid and GF-H on the secondary structure contents of α-glucosidase ............................................................................... 69 Table 13. Starch hydrolysis rates of control, oleic acid, linoleic acid, GF-H and acarbose at different time periods ................................................................... 78 | |
| dc.language.iso | en | |
| dc.subject | 第二型糖尿病 | zh_TW |
| dc.subject | 舞菇 | zh_TW |
| dc.subject | α-澱粉酶 | zh_TW |
| dc.subject | α-葡萄糖苷 | zh_TW |
| dc.subject | 脂肪酸 | zh_TW |
| dc.subject | fatty acids | en |
| dc.subject | α-amylase | en |
| dc.subject | α-glucosidase | en |
| dc.subject | type 2 diabetes | en |
| dc.subject | Grifola frondosa | en |
| dc.title | 舞菇抗第二型糖尿病相關消化酶之研究 | zh_TW |
| dc.title | Study on the inhibitory effects of Grifola frondosa on type 2 diabetes-related digestive enzymes | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 林松洲(Song-Chow Lin),徐駿森(Chun-Hua Hsu) | |
| dc.subject.keyword | 舞菇,α-澱粉酶,α-葡萄糖苷,酶,第二型糖尿病,脂肪酸, | zh_TW |
| dc.subject.keyword | Grifola frondosa,α-amylase,α-glucosidase,type 2 diabetes,fatty acids, | en |
| dc.relation.page | 97 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-07-16 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 農業化學研究所 | zh_TW |
| 顯示於系所單位： | 農業化學系 | |
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