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題 名 | 臺灣黃蘗細胞懸浮培養生產小蘗鹼之研究=Berberine Production in Cell Suspension Cultures of Phellodendron amurense Rupr. var. wilsonii (Hayata & Kanehira) |
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作 者 | 劉季函; 賴瑞聲; 李宏謨; 吳明哲; | 書刊名 | 臺灣農業研究 |
卷 期 | 61:3 2012.09[民101.09] |
頁 次 | 頁196-208 |
分類號 | 414.34 |
關鍵詞 | 臺灣黃蘗; 癒合組織; 懸浮細胞; 小蘗鹼; 高效液相層析; Phellodendron amurense var. wilsonii; Callus; Cell suspension; Berberine; High performance liquid chromatograph; HPLC; |
語 文 | 中文(Chinese) |
中文摘要 | 黃蘗藥材來源為芸香科(Rutaceae)黃蘗屬(Phellodendron spp.)植物之樹皮,具有清熱解毒、消炎止痢、滋陰通便的功效,黃蘗樹皮含有小蘗鹼(berberine),為西藥製劑的重要原料,廣泛被使用於治療腸胃炎、細菌性痢疾等。台灣黃蘗 [Phellodendron amurense Rupr. var. wilsonii(Hayata & Kanehira)] 為台灣特有種,小蘗鹼含量高,是黃蘗藥材優異來源,但須生長6-8年才可利用,且原生地已過度採集,因此,建立台灣黃蘗懸浮細胞培養系統,是生產小蘗鹼的另一選擇。本研究以台灣黃蘗幼株為材料,進行癒合組織誘導,並探討建立台灣黃蘗細胞懸浮培養之條件,並以小蘗鹼含量為評估指標。結果顯示癒合組織誘導以葉柄為最佳培植體,經暗處理培養於0.5mg/L2,4-D、1.0mg/LBA之MS基礎鹽類培養基時,可於28天後誘導出癒合組織(callus),其型態為淡黃色、質地鬆軟、較鬆散,適合作為懸浮細胞培養來源。懸浮細胞培養以WPM培養基添加0.5mg/L2,4-D及1.0mg/LBA為基礎培養基,於黑暗中以100rpm轉速震盪培養,生長20天時細胞生長量可達9倍,但癒合組織及前述懸浮細胞培養經75%乙醇萃取及HPLC檢測分析皆無小蘗鹼含量。本研究於懸浮細胞培養第18天添加2mg/LBA及6mg/LNAA可順利誘導小蘗鹼生合成,於第28天時懸浮細胞之小蘗鹼含量為188.68 μg/gdw,本研究可作為台灣黃蘗懸浮細胞培養量產小蘗鹼之基礎,懸浮細胞如以不同生產調節劑處理,也可能誘導出棕櫚鹼及黃蘗鹼等其他藥用成分,具有製藥應用潛力。 |
英文摘要 | Barks of Huang-Po (Phellodendron spp.) contain berberine and it is used as an important herbal medicine for the treatment of fever, inflammation, stomach ache and intestinal illness. Taiwan Huang-Po [Phellodendron amurense Rupr. var. wilsonii (Hayata& Kanehira)] is a native species in Taiwan and the bark of this plant is used as an important medicine because of its high berberine content. However, it would take more than 6-8 years to grow Taiwan Huang-Po trees for harvesting barks. In addition, the supply of barks of Taiwan Huang-Po from natural habitat is dwindling due to excessive harvesting of this plant. The objective of this study was to establish a cell suspension culture method for production of berberine from Taiwan Huang-Po, as an alternative method for production of berberine from barks of plants grown in natural habitat. Two months old seedlings were used as explants to induce callus in this study. Leaf petiole was proven to be the best tissue for callus induction. Leaf petiles were placed on Murashige and Skoog (MS) basal medium containing 0.5 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.0 mg/L 6-benzylaminopurine (BA) and incubated in dark for 28 days to form callus tissues which were used as inoculum for cell suspension cultures. Callus tissues were inoculated on a medium containing WPM basal salt amended with 0.5mg/L 2,4-D and 1.0 mg/L BA and the cell suspension was cultured on a shaker at 100 rpm for 20 days and then used for testing amount of cells and berberine in the cultures. Results showed that the total amount of cells in the cell suspension cultures increased by 9 folds after incubation for 20 days but no berberine was detected when the cells were extracted by 75% ethanol and analyzed by high performance liquid chromatography (HPLC). In contrast, suspension cells grown in the medium containing 2 mg/L BA and 6 mg/L α-naphthaleneacetic acid (NAA) for 18 days, resulted in production of berberine. Berberine accumulation reached 188.68 μg/g dry weight in the 28-day-old, cell-suspension culture. Thus, this method of employing growth regulators in cell suspension culture may be important in eastablishing protocol for industrial production of berberine from cell suspension cultures of the endemic species P. amurens. |
本系統中英文摘要資訊取自各篇刊載內容。