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研究生:易宗瑛
研究生(外文):Chung-Ying I
論文名稱:牛樟芝深層醱酵菌絲體對第一型糖尿病大白鼠耐糖能力及醣類代謝的影響
論文名稱(外文):Effects of the mycelium from submerged cultured Antrodia camphorata on glucose tolerance and carbohydrate metabolism in type 1 diabetic rat
指導教授:沈賜川吳明昌
指導教授(外文):Szu-Chuan ShenMing-Chang Wu
學位類別:碩士
校院名稱:國立屏東科技大學
系所名稱:食品科學系所
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2008
畢業學年度:96
語文別:中文
論文頁數:100
中文關鍵詞:菌絲體牛樟芝糖尿病鼠碳水化合物代謝安氏試驗
外文關鍵詞:MyceliumAntrodia camphoratadiabetic ratscarbohydrate metabolismAmes test
相關次數:
  • 被引用被引用:3
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論文摘要內容:
本研究目的在探討樟芝深層醱酵菌絲體,對於以Streptozotocin (STZ)注射Wistar大鼠所誘發第一型糖尿病耐糖不良動物模式之降血糖效果及其對肝臟碳水化合物代謝的影響,並以安氏試驗分析樟芝深層醱酵菌絲體之抗致突變活性。
結果顯示,在急性試驗中,樟芝深層醱酵菌絲體水溶液有明顯延緩糖尿病大鼠血糖上升的效果,且高劑量 (400 mg/Kg BW) 的效果似乎比低劑量 (200 mg/Kg BW) 佳;而在進行六週的長效性試驗,灌食樟芝深層醱酵菌絲體水溶液,在本試驗濃度範圍內 (400 mg/kg BW),並無明顯改善第一型糖尿病大鼠葡萄糖耐受性之效果。
試驗動物於第六週犧牲後分析其肝臟中參與碳水化合物代謝之酵素活性變化情形,結果發現樟芝深層醱酵菌絲體水溶液能活化肝臟 hexokinase(HKase)、phosphofructokinase (PFKase)、glucose-6-phosphate dehydrogenas(G6PDHase)活性並降低 fructose-1,6-bisphosphatase(F1, 6 BPase)、Glucose -6-phosphatase(G6Pase)活性,顯示其能促進葡萄糖進入 glycolysis pathway 及 pentose monophosphate shunt 並減少肝臟葡萄糖的釋放。灌食樟芝深層醱酵菌絲體之糖尿病鼠實驗組,其肝臟酵素包括HKase 、PFKase 、G6PDHase之活性皆高於糖尿病鼠控制組;且其F1,6BPase、G6Pase活性也較糖尿病控制組低並有顯著性差異。
安氏試驗 (Ames test) 係以Salmonella typhimurium TA100為測試菌
株,探討樟芝深層醱酵菌絲體水萃物之致突變性,實驗結果顯示,在0.06-1.00 mg/plate的測試濃度範圍內均不具毒性及致突變性,且對直致突變劑(4-NQO) 及間接致突變劑B【a】P均具有抑制效果。
綜合上述實驗結果,樟芝深層醱酵菌絲體可改善第一型糖尿病鼠肝臟碳水化合物代謝酵素的活性,但在本實驗劑量範圍內 (400 mg/kg BW),對改善第一型糖尿病鼠耐糖能力及血糖方面並無預期之效果。
The contents of abstract in this thesis:
The purpose of this study is to investigate the effect of the mycelium of submerged cultured Antrodia camphorata in lowing blood sugar and hepatic carbohydrate metabolism. Wistar rats were treated with streptozotocin (STZ) to become induced diabetic rats and applied to the study. Ames test was used to analyze the anti-mutagenic effect for the mycelium of submerged cultured Antrodia camphorata.
The acute oral glucose tolerance test (OGTT) results showed that the mycelium of submerged cultured Antrodia camphorata can alleviate the rising of blood sugar in STZ-induced diabetic rats. Especially, the effects in high dose of OGTT (fed with 400 mg/kg of sugar) were more obvious than in low dose of OGTT (200 mg/kg). After 6-week studies, the diabetic rats fed with mycelium of submerged cultured Antrodia camphorata doesn’t show significantly in improvement on glucose tolerance.
The rats were sacrificed after 6 weeks. The activities of liver enzymes
participating in carbohydrate metabolism were analyzed. The results showed that mycelium of submerged cultured Antrodia camphorata could increase the activities of hexokinase (HKase), phosphofructokinase (PFKase),
glucose-6-phosphate dehydrogenas (G6PDHase), but decreased the
activities of fructose-1,6-bisphosphatase (F1, 6 BPase) and Glucose-6-phosphatase (G6Pase). It suggests that the mycelium of submerged cultured Antrodia camphorata can promote glucose to go in glycolysis pathway and pentose monophosphate shunt, and reduce the release of glucose from liver. Liver enzyme activities, included HKase, PFKase, G6PDHase, are higher in diabetic rats fed with mycelium of submerged cultured Antrodia camphorata than STZ-induced diabetic rats. On the other hand, the activities of F1, 6BPase, G6Pase of diabetic rats fed with mycelium of submerged cultured Antrodia camphorata are significantly lower than STZ-induced diabetic controls.
Salmonella typhimurium TA100 was used for Ames test to analyze the mutagenesis capability of aqueous extract from mycelium of submerged cultured Antrodia camphorata. The results showed that the aqueous extract
have wither toxicity nor mutagenesis characteristic in the test concentration of 0.06-1.0 mg/plate. Anti-mutagenetic activity of aqueous extract from mycelium of submerged cultured Antrodia camphorata show inhibitory effect efficiently treated with 4-NQO than treated with B[a]P.
In conclusions, the mycelium of submerged cultured Antrodia camphorata can improve the hepatic carbohydrate metabolic enzyme activities in STZ-induced diabetic rats, nevertheless, this effect in OGTT and blood sugar reduction was not significant in our study.
壹 前言............................................................................................. 1
貳 文獻回顧..................................................................................... 2
一 樟芝的介紹......................................................................... 2
(一)簡介................................................................................. 2
(二)生理活性成分................................................................. 4
(三)生理功能研究................................................................. 5
二 糖尿病........................................................................................ 7
(一)糖尿病的介紹................................................................ 7
(二)糖尿病的定義................................................................. 7
(三)糖尿病的常見症狀........................................................ 9
(四)糖尿病診斷標準............................................................. 11
(五)糖尿病的分類................................................................. 13
(六)糖尿病併發症................................................................ 16
(七)糖尿病的流行病學......................................................... 17
三 致突變性試驗............................................................................. 22
(一)安姆試驗法簡介............................................................. 22
(二)安姆試驗檢測原理......................................................... 22
(三)試驗菌株......................................................................... 22
(四)轉化酵素......................................................................... 22
四 抗致突變性試驗......................................................................... 25
(一)食品中的抗致突變物..................................................... 25
(二)抗致突變物之作用機制................................................. 25
五 糖尿病研究之動物模式............................................................. 29

(一)研究糖尿病的動物模式................................................. 29
(二)STZ化學誘導糖尿病動物模式..................................... 30
參 材料與方法................................................................................. 33
一 實驗材料來源......................................................................... 33
二 儀器設備................................................................................. 33
三 試劑藥品................................................................................. 34
四 實驗架構................................................................................. 36
五 實驗方法................................................................................. 39
(一) 一般成分分析................................................................. 39
1. 水份測定...................................................................... 39
  2. 灰份測定...................................................................... 39
  3. 粗脂肪測定................................................................. 40
4. 粗蛋白測定….............................................................. 40
5. 總醣測定….................................................................. 41
(二)動物實驗........................................................................... 42
1.實驗動物購入及飼養.................................................... 42
2.誘發劑的配製................................................................ 42
3.實驗動物誘導及分組.................................................... 42
4.急性試驗…………………........................................... 43
5.長效性試驗.................................................................... 43
6.動物犧牲與檢體之收集............................................... 44
7.葡萄糖 (glucose) 濃度測定......................................... 44
8.肝臟分析........................................................................ 44
(1)粗酵素液的製備......................................................... 44
(2)肝臟酵素測定............................................................. 45
a. Glucose-6-phosphate dehydrogenase活性的測定. 45
b. Glucose-6-phosphatase活性的測定....................... 45
c. Fructose-1,6-biphosphatase活性的測定................ 45
d. Phosphofructokinase活性的測定........................... 45
e. Hexokinase活性的測定.......................................... 46
f. 蛋白質濃度之測定................................................. 46
(三)牛樟芝深層醱酵菌絲體抗致突變性之研究................ 46
1.萃取物之製備............................................ ........... 46
2.試驗菌株................................................................ 47
3.溶液的配備……………………............ ............... 47
4.試驗菌株活化……................................................ 47
5.毒性試驗.................................................................................. 47
6.致突變性試驗......................................................................... 48
7.抗致突變性試驗...................................................................... 48
(四)統計分析........................................................................ 53
肆 結果與討論................................................................................. 54
一 一般成分分析..................................................................... 54
(一)水份................................................................................. 54
(二)灰份................................................................................. 55
(三)粗脂肪............................................................................. 55
(四)粗蛋白............................................................................. 55
(五)碳水化合物..................................................................... 55
二 動物試驗............................................................................. 58
(一)急性試驗-探討急性灌食牛樟芝深層醱酵菌絲體以STZ誘發第一型糖尿病大白鼠葡萄糖耐受性的影響................................................................................. 58
(二)長效試驗-探討連續灌食牛樟芝深層醱酵菌絲體六週期間對以對STZ誘發第一型糖尿病大白鼠模式的影響................................................................................. 61
1. 灌食牛樟芝醱酵菌絲體對糖尿病大白鼠耐糖的影響............................................................................... 61
2. 灌食牛樟芝菌絲體溶液對糖尿病大白鼠於試驗期間生長之影響........................................................... 62
(1)飲水量................................................................... 62
(2)攝食量................................................................... 62
(3)重量及相對臟器的影響....................................... 62
3. 灌食牛樟芝菌絲體對第一型糖尿病大白鼠醣類代謝的影響................................................................... 74
(1) Glucose -6-phosphatase.......................................... 74
(2) Fructose-1,6-biphosphatase..................................... 74
(3) Hexokinase............................................................... 75
(4) Phosphofructokinase................................................ 75
(5) Glucose-6-phosphate dehydrogenase....................... 75
三 牛樟芝深層醱酵菌絲體抗致突變性之研究....................... 82
(一)毒性試驗............................................................................ 82
  (二)致突變性試驗..................................................................... 84
  (三)抗致突變性試驗................................................................. 86
伍 結論............................................................................................. 91
參考文獻............................................................................................. 92
作者簡介………………………………………………………….... 100


圖表目錄

圖1、 糖尿病代謝混亂結果.......................................................... 8
圖2、 糖尿病分類.......................................................................... 15
圖3、 主要抗致突變食品成分之作用模式.................................. 28
圖4、 STZ之化學結構.................................................................. 31
圖 5、 STZ造成大鼠胰臟β-細胞死亡機制.................................. 32
圖6、 毒性試驗流程圖.................................................................. 50
圖7、 致突變試驗流程圖.............................................................. 51
圖8、 抗致突變性試驗流程圖...................................................... 52
圖9、 急性牛樟芝菌絲體溶液200 mg/kg BW及400 mg/kg BW 並進行口服葡萄糖耐受性試驗之大白鼠血漿葡萄糖濃度化................................................................................ .... 60
圖10、 長期灌食牛樟芝菌絲體溶液第0週之口服葡萄糖耐受性 試驗大白鼠血漿葡萄糖濃度的變化.............................. ..
64
圖11、 長期灌食牛樟芝菌絲體溶液第4週之口服葡萄糖耐受性試驗大白鼠血漿葡萄糖濃度的變化.............................. ..
66
圖12、 長期灌食牛樟芝菌絲體溶液第6週之口服葡萄糖耐受性試驗大白鼠血漿葡萄糖濃度的變化.............................. ..
68
圖13、 大白鼠灌食牛樟芝菌絲體溶液六週飲水量之變化........ . 70
圖14、 大白鼠灌食牛樟芝菌絲體溶液六週攝食量之變化......... 71
圖15、 大白鼠餵食牛樟芝菌絲體溶液六週體重之變化............ . 72
圖16、 灌食牛樟芝菌絲體溶液六週後肝臟中Glucose-6-phosphatase的酵素活性.................................... 77
圖17、 灌食牛樟芝菌絲體溶液六週後肝臟中 fructose-1,6-biphosphatase的酵素活性.......................... ... 78
圖18、 灌食牛樟芝菌絲體溶液六週後肝臟中hexokinase的酵素活性..................................................................................... 79
圖19、 灌食牛樟芝菌絲體溶液六週後肝臟中phosphofructokinase的酵素活性........................................ 80
圖20、 灌食牛樟芝菌絲體溶液六週後肝臟中glucose-6-phosphate dehydrogenase 的酵素活性………..
81
圖21、 灌食牛樟芝深層醱酵菌絲體水萃物於TA100對 4NQO之抗致突變活性.................................................................
89
圖22、 灌食牛樟芝深層醱酵菌絲體水萃物於TA100對 B【a】P之抗致突變活性.................................................................
90
表1、 糖尿病診斷標準.................................................................. 12
表2、 糖尿病及耐糖障礙 (IGT) 人口數及盛行率.................... 20
表3、 六十五歲及以上國人糖尿病(以空腹血糖 ≧126 mg/dL或有服降血糖藥物者為定義糖尿病的標準)盛行率 - 依性別、年齡層別之比較................................................. 21
表4、 致突變性檢測方法............................................................. 24
表5、 食品中的抗致突變/抗癌物................................................. 27
表6、 牛樟芝深層醱酵菌絲體一般成分分析............................... 57
表7、 急性灌食牛樟芝菌絲體溶液200 mg/kg BW及400 mg/kg BW 並進行口服葡萄糖耐受性試驗之大白鼠血漿葡萄糖濃度變化......................................................................... 59
表8、 長期灌食牛樟芝菌絲體溶液第0週之口服葡萄糖耐受性 試驗大白鼠血漿葡萄糖濃度的變化.................................
65
表9、 長期灌食牛樟芝菌絲體溶液第4週之口服葡萄糖耐受性 試驗大白鼠血漿葡萄糖濃度的變化.................................
67
表10、 長期灌食牛樟芝菌絲體溶液第6週之口服葡萄糖耐受性試驗大白鼠血漿葡萄糖濃度的變化.................................
69
表11、 食牛樟芝菌絲體溶液六週後大白鼠臟器重量及相對重量之變化.................................................................................
73
表12、 牛樟芝深層醱酵菌絲體水萃物之毒性試驗...................... 83
表13、 牛樟芝深層醱酵菌絲體水萃物之致突變性試驗. ….…… 85
表14、牛樟芝深層醱酵菌絲體水萃物對4-NQO 抗致突變驗.........................................................................................
87
表15、 牛樟芝深層醱酵菌絲體水萃物對B【a】P 抗致突變試驗.........................................................................................
88
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