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Study on effect of culture condition on polysaccharide production of Phyllanthus urinaria L. cells
|Authors: ||Yi-Cheng Cheng|
|Contributors: ||NTOU:Department of Food Science|
|Issue Date: ||2011-06-22T08:40:05Z
|Abstract: ||葉下珠 (Phyllanthus urinaria L.) 是本草綱目拾遺中所列嶺南重要的生藥之一，具有保肝、制菌及抗發炎等作用。研究報告顯示一般藥用植物的多醣體具有抗發炎及抗腫瘤效果。 利用植物細胞培養技術，可作為生產二次代謝產物或多醣體之工具，為了利用細胞培養法大量生產多醣體及細胞，本研究以葉下珠細胞進行培養探討不同培養條件對其生產多醣體的影響。採用B5配方的基本培養基，誘導癒合組織，進行懸浮細胞培養。接種葉下珠的種細胞到125 mL 三角培養瓶，於 25℃，無光照條件下培養 42 天，實驗結果顯示培養第14 天時細胞產量達最高為22 g/L，多醣體則在 21 天才達最高點，多醣體產生隨細胞量增加而慢慢增加。在培養液中添加生長調節素NAA (naphthalene acetic acid)、2,4-D (2,4 – dichloropheno -xyacetic acid) 對葉下珠細胞生長及多醣體生產皆有顯著增加，兩者對多醣體之產量增加差異不大。不同碳源對於細胞和生長和多醣體生產之影響，大致相同。碳源濃度以12％蔗糖濃度組之胞外多醣體產量3.82 g/L最佳，顯示碳源對葉下珠細胞成長及多醣體產出很重要。不同有機氮源培養，對多醣體產量以 Casein 培養者最佳，可達 1.9 g/L。葉下珠細胞培養產生的胞外粗多醣體，其分子量經分析得知分佈在150,000∼780,000 Da之間。 另外以葉下珠細胞培養生產多醣體與葉下珠藥材萃取者做比較，顯示細胞培養多醣體產率高達80 g/kg/3週，優於野生或田間栽培藥材的12 g/kg/年。所以利用中草藥植物細胞培養生產多醣體之方法，是一項具有很高競爭力之生物技術。|
Phyllanthus urinaria L. has long been used as an important Chinese medicinal herb with functions of preventing liver dysfunction, healing diabetes and jaundice. Recent reports showed that polysaccharides obtained from plants have activities of anti-inflammation and anti-tumor. Plant cell culture technology is an alternative to produce bioactive secondary metabolites and polysaccharides. To investigate the effects of culture conditions on polysaccharide production from cell cultures of Phyllanthus urinaria L. Callus induction, acclimation of suspension cultures, and optimization of cultivation conditions for polysaccharide production were conducted based on B5 medium. The maximum level of cell growth was 2.2 g/L after subculturing 14 days, while that of polysaccharides was reached after 21 days. Addition of growth regulator to the culture medium could increase the cell growth and polysaccharides production. No obvious effect was observed using different growth regulators NAA or 2,4-D and different carbon sources, while 3.82 g/L polysaccharides was obtained when 12% sucrose were added in the medium. The effects of different organic nitrogen concentration on production rate of polysaccharides were investigated, in which casein showed higher polysaccharide production (1.9 g/L). The molecular weight of crude cell culture-derived polysaccharides ranged approximately from 150,000 to 780,000. The yield of crude polysaccharides extracted from herbal materials harvested from the field was 12 g/kg/year, while that from cell cultures was 80 g/kg/21 days, demonstrating the potential availability and applicability of plant cell cultures in polysaccharide production.
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