青钱柳悬浮细胞的培养及其基质消耗的规律

Establishment of the Cell Suspension Culture System of Cyclocarya paliurus and Matrix Consumption Laws

摘要:

青钱柳为我国特有的胡桃科青钱柳属单种属珍稀植物，也是国家重点保护的濒危植物之一，青钱柳叶含三萜、黄酮、多糖及微量元素等活性成分，具有降糖、调脂、增强免疫等多种生理活性和药理功能，是很好的天然保健食品资源。本文建立青钱柳细胞悬浮培养体系，测定悬浮培养细胞的生长曲线，并研究了培养代数、接种量、装液量、pH值和激素组合对青钱柳悬浮体系建立的影响，同时研究了悬浮培养青钱柳细胞对培养液中主要基质的消耗规律。结果表明：适合青钱柳悬浮培养的培养基为：MS+0.1 mg/L 2,4-D+0.5 mg/L NAA+1 mg/L KT+30 g/L蔗糖，接种量为8%，摇瓶装液量的体积分数为40%，初始pH 5。青钱柳细胞生长与碳源、氮源和磷的消耗存在着密切关系，在培养8 d，糖和PO43-基本消耗完毕；在氮源的吸收上，NH4+优于NO3-，NH4+的吸收基本上是呈直线下降。

Abstract:

Cyclocarya paliurus is a unique plant resource in China. As one of the threatened plants, it has been received increasing attention. Cyclocarya paliurus is rich in flavonoids, polysaccharides, triterpenoids and other bioactive substances. However, the its application is restricted for the problems of its sexual and asexual reproduction. Nowadays, employing cell culture technology to get more bioactive compounds is a vital way to make full use of Cyclocarya paliurus. The suspension cell system influenced by the factors of inoculum density, the liquid medium volume per flask, pH, sucrose concentration, phytohormone combinations and the consumption rules for culture medium in the process of cells proliferation were studied. The dynamic curve of cells proliferation and total flavonoids, polysaccharides and triterpenoids synthesized in the suspension cell system were measured. The result revealed that the preferable conditions for suspension cells proliferation were as follows: MS medium supplemented with 1.0 mg/L KT, 0.1 mg/L 2,4-D, 0.5 mg/L NAA and sucrose 30 g/L. The optimum culture situations were seed volume 8%, liquid medium volume per flask 40% and initial pH 5. Moreover, the proliferation of Cyclocarya paliurus cells correlated closely with the consumption of sugar, nitrate and phosphate. After 8 days, sugar and phosphate in the medium were absorbed completely, while NH4+ was utilized faster than NO3-, and the concentration of NH4+ showed a linear decreasing trend.