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研究生:陳世傑
研究生(外文):Chen, Shih-Chieh
論文名稱:葛組織培養之研究
論文名稱(外文):Studies on tissue culture of Pueraria lobata (Willdenow) Ohwi
指導教授:蔡新聲, 陳忠川
指導教授(外文):Tsay Shin-Shen, Chen Chung-Shong
學位類別:碩士
校院名稱:中國醫藥學院
系所名稱:中國藥學研究所
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:1997
畢業學年度:85
語文別:中文
論文頁數:132
中文關鍵詞:葛根組織培養
外文關鍵詞:Pueraria lobataTissue culture
相關次數:
  • 被引用被引用:9
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葛根為豆科葛屬植物的根。首載於神農本草經中品,歷代藥用植物學及生
藥學文獻均有記載。自古即為有名的解熱、發汗、鎮痛、消炎、生津止瀉
藥材,臨床上用於熱性病之口渴、嘔氣、惡寒、頭痛、心絞痛,特別用於
感冒之頸部及肩背強急之治療,扁桃腺炎、蕁麻疹、蓄膿症等之初期症狀
,「葛根湯」尤為使用頻繁之著名方劑。葛根主成分為黃酮類的葛根素 (
puerarin )、大豆黃酮( daidzein ) 及大豆黃酮( daidzin ) 等,具
有解熱、鎮痙、降血糖、雌激素樣作用,並能增加腦及冠狀血管血流量而
降低血管阻力,是當今文明病的對症良藥。台灣市場之葛根大多以
Pueraria lobata ( WILLDENOW ) OHWI 為主,本研究主要目的在藉由組
織培養技術,建立P. lobata大量繁殖的方法,及尋找利用癒合組織生產
daidzein和puerarin之可行性,結果摘要如下:1. 不同培殖體誘導不定
芽之能力,以莖節和頂芽較佳。其中以莖節為培殖體培養於含有 2 mg/l
BA 及 0.2 mg/l NAA之MS基本鹽類培養基中,可獲得最多之不定芽。2.
自莖節誘導之組織培養苗,再培養於含有2 mg/l BA, 0.2 mg/l NAA, 3 %
sucrose及 2 g/l peptone之MS基本鹽類培養基中,小苗生長較好且健康
,也可得到較多之次生不定芽。3. NAA, MS無機鹽和sucrose都能影響不
定芽之發根。其中以含有1/2 MS無機鹽, 2 mg/l NAA, 0.2 mg/l BA和 3%
sucrose之培養基對發根之效果最好。4. 將組織培養苗移植於混合不同
比例之介質中, 5 週後調查存活率,結果發現以soil:vermiculite:
peat moss=1:1:1 之介質存活率最高可達90 %。5. 將根、莖和葉培
養於含有1 mg/l 2,4-D, 0.5 mg/l BA之MS基本鹽類培養基中可誘導癒合
組織形成。葉片誘導得之癒合組織極易褐化、死亡。6. 將根、莖和葉三
種來源的癒合組織培養於2 mg/l BA, 0.2 mg/l NAA, 3%sucrose, 20%
coconut milk及0.4% gelrite之MS基本鹽類培養基中,無法使其產生器官
的分化。7. 20%椰子汁有防止癒合組織褐化之效果,且癒合組織鮮重最
重。8. 含有1.0 mg/l 2,4-D, 0.5 mg/l BA, 500 mg/l casein
hydrolysate, 1 g/l peptone, 20% coconut milk, 3% sucrose, 0.4%
gelrite 及 pH 5.7之MS基本鹽類培養基最適合癒合組織之生長,約經20
至25天培養之癒合組織鮮重最重。9. 利用高效液相層析儀,可測得培養
苗和癒合組織中daidzein和puerarin的含量。
Puerariae Radix ( Ke-Ken ) is the dried root of some Pueraria
species of Leguminosae. It was first recorded in Shen-Nung-Pen-
Ts''ao-Ching under the middle category and it was also recorded
in the successive Pen-Ts''ao of the descending dynasties. Ke-Ken
has been used as diaphoretic, antipyretic and relaxant for
thirsty, nausea and headache. It is also very effective for
stiff neck and back in common cold. Ke-Ken-Tang is also a famous
prescription commonly used. The improvely discovery of recent
times, it contains two therapeutic ingredients : daidzein and
puerarin, which possess antihyertensive and antidiabetic
properties. Most of the Puerariae Radix sold in Taiwan market
are from the origin of Pueraria lobata, but some other Ke-Ken
imported is still unknown. The purpose of this study was tried
to establish a method of mass propagation by tissue culture
techniques on P. lobata and to seek a possibility of daidzein
and puerarin production from the callus culture. The
experimental results are summarized as follows : 1. The stem-
node and shoot tip possessed the highest ability for
adventitious bud formation when explants were cultured on a
medium containing MS salts with 2 mg/l BA and 0.2 mg/l NAA. The
stem-node was the best source organ for adventitious buds
formation.2. The plantlet produced from a stem-node cultured on
MS medium containing 2 mg/l BA, 0.2 mg/l NAA, 3 % sucrose and 2
g/l peptone showed healthy, better growth, and more adventitious
buds. Stem-node was the best among three source organs tested.3.
The rooting from adventitious buds was affected by NAA, MS salts
and sucrose concentrations in the medium. The best medium for
rooting was 1/2 MS salts with 2 mg/l NAA, 0.2 mg/l BA and 3%
sucrose.4. The plantlet of P. lobata produced from callus
culture could be transplanted into a mixture of soil,
vermiculite and peat moss ( v/v 1:1:1), and growth well under
outdoor conditions. The survival rate was 90%.5. The callus
could be induced from root, stem and leaf cultured on a medium
containing 1 mg/l 2,4-D, 0.5 mg/l BA. The leaf-derived callus
was found the callus easy to browning and die.6. The callus of
P. lobata induced had no differentiation ability, when it was
cultured on a medium containing MS salts with 2 mg/l BA, 0.2 mg/
l NAA, 3%sucrose and 0.4% gelrite.7. Addition of coconut milk
(CM) could protect the browning of the callus, and addition of
20%CM increase the largest fresh weight of callus.8. The callus
of P. lobata adapted to culture on a modified medium with MS
salts supplemented 1.0 mg/l 2,4-D, 0.5 mg/l BA, 500 mg/l casein
hydrolysate, 1 g/l peptone, 20% coconut milk, 3% sucrose, pH
value of 5.7 and 0.4% gelrite. And the cultured duration between
20 to 25 days could obtain the largest quantities of callus.9.
Diadzein and puerarin were detected from plantlet and callus of
P. lobata by using high performance liquid chromatograph (HPLC)
technique.
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