Optimal Operating Conditions for Cultivation of Plant Cells in an External Loop Airlift Bubble Colum

外循环气升式气泡塔中植物细胞培养的最佳操作条件

• 批准号: 08455386
• 负责人: NAKAO Katsumi
• 金额: $ 4.8万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08455386/
• 关键词: Carrot Callus; beta-Carotene; External Loop Airlift; Kinetic Model; Optimal Operating Condition; Immobilized Carrot Callus

The present research employed the cultivation of carrot calluses for production of beta-carotene as a model system of plant cell culture and the external loop airlift bubble column as a bioreactor suitable to large scale culture under lower and more uniform hydodynamic shear stress. The purpose of this work is (a) to examine the effects of the various operating conditions on cell growth and beta-carotene production, (b) to determine the optimum of each operating variable taking the content, yield and productivity of beta-carotene as the objective functions, and (c) to establish kinetic model to predict the time courses and to search for an optimal operating condition. The results obtained for each research subject are summarized as follows.1. Effects of bioreactor geometry and operating conditions on cell growth and beta-carotene production of suspended carrot cellsAlthough the optimum values of each operating variable were almost the same for both airlift bioreactor and shaking flask, … More the beta-carotene content and productivity in the former was grater than those in the latter. The production under the combined optimal operating conditions was several times higher than that under the control conditions. The higher temperature favored both growth and production in the earlier stage of cultivation, while the lower ne favored those in the later stage. This suggested a two-phase cultivation shifting the temperature from higher to lower one. The characteristics of airlift bioreactor was also investigated.2. Effects of bioreactor geometry and operating conditions on beta-carotene production of immobilized carrot cellsThe carrot cells immobilized in the three different calcium alginate gel beads gave the lower growth and beta-carotene content than the corresponding suspended cells by factors of 1/2 to 1/3 and 1/5 to 1/10, respectively. The higher molecular weight of the alginates used reduced the growth and enhanced the content to a greater extent. This verified the negative correlation of beta-carotene content with cell growth. A further study on excretion of beta-caratene will be continued by using a novel glycolipid.3. Comparison of various culture methods and search for optimal operating conditionsThe kinetic model assuming an inhibited cell growth and a nonlinear dependence of beta-carotene production on growth well reproduced the observed time courses. The kinetic parameters in the model were correlated with the operating conditions in the dimensionless form based on the respective variables and parameters for the control cultures. The model with the correlations fairly predicted the time courses and the optimal operating conditions. In the solid cultivation on the gellan gum medium, the four different cell lines gave the content of no greater than 1/10 of that for the corresponding suspended cells. This revealed that no great decrease in cell growth due to solid cultivation resulted in a remarkable increase in the content although the negative correlation of content with growth was observed even in the solid cultivation. Less

本研究采用胡萝卜愈伤组织的培养来生产β-胡萝卜素作为植物细胞培养的模型系统，并采用外循环气升式泡罩塔作为适合在较低且更均匀的流体力学剪切应力下大规模培养的生物反应器。这项工作的目的是（a）检查各种操作条件对细胞生长和β-胡萝卜素生产的影响，（b）以β-胡萝卜素的含量、产量和生产率为目标函数确定每个操作变量的最佳值，以及（c）建立动力学模型来预测时间进程并寻找最佳操作条件。每个研究课题获得的结果总结如下： 1．生物反应器几何形状和操作条件对悬浮胡萝卜细胞生长和β-胡萝卜素产生的影响虽然气升式生物反应器和摇瓶的每个操作变量的最佳值几乎相同，但前者的β-胡萝卜素含量和生产率高于后者。组合最佳操作条件下的产量比对照条件下高出数倍。较高的温度有利于栽培前期的生长和产量，较低的温度有利于栽培后期的生长和产量。这表明两阶段培养将温度从较高阶段转变为较低阶段。并对气升式生物反应器的特点进行了研究。 2．生物反应器几何形状和操作条件对固定化胡萝卜细胞产生β-胡萝卜素的影响固定在三种不同海藻酸钙凝胶珠中的胡萝卜细胞的生长和β-胡萝卜素含量分别比相应的悬浮细胞低1/2至1/3和1/5至1/10。所用海藻酸盐的分子量较高，在更大程度上减少了生长并提高了含量。这验证了β-胡萝卜素含量与细胞生长的负相关性。将通过使用新型糖脂继续对β-胡萝卜素的排泄进行进一步的研究。3．比较各种培养方法并寻找最佳操作条件假设细胞生长受到抑制和β-胡萝卜素产量对生长的非线性依赖性，动力学模型很好地再现了观察到的时间进程。基于对照培养物的相应变量和参数，模型中的动力学参数与无量纲形式的操作条件相关。具有相关性的模型相当准确地预测了时间进程和最佳操作条件。在结冷胶培养基上的固体培养中，四种不同的细胞系的含量不大于相应悬浮细胞的1/10。这表明，虽然即使在固体培养中也观察到含量与生长呈负相关，但固体培养并未导致细胞生长的大幅下降导致含量显着增加。较少的

项目成果

中尾勝實: "ニンジン細胞培養によるβ-カロチン生産の速度論的解析" 化学工学会姫路大会講演要旨集. 308-309 (1996)

Katsumi Nakao：“胡萝卜细胞培养产生β-胡萝卜素的动力学分析”化学工程师学会姬路会议记录308-309（1996）。

K.Nakao et al.: "Mass Transfer Properties of Bubble Columns Suspending Immobilized Glucose Oxidase Gel Beads for Gluconic Acid Production" Preprints of 4th Japanese / German Symposium Bubble. 124-131 (1997)

K.Nakao 等人：“用于葡萄糖酸生产的悬浮固定化葡萄糖氧化酶凝胶珠的气泡柱的传质特性”第四届日本/德国气泡研讨会预印本。

中尾勝實: "固定化グルコースオキシダーゼゲル粒子を用いるエアリフト型気泡塔の物質移動係数の測定" 化学工学会第29回秋季大会研究発表講演要旨集. 第1巻. 140-140 (1996)

Katsumi Nakao：“使用固定化葡萄糖氧化酶凝胶颗粒测量气升式气泡塔的传质系数”，化学工程师学会第 29 届秋季会议摘要，第 1 卷 140-140（1996 年）。

K.Nakao, K.Fukunaga, H.Funahashi et al.: "Influence of Culture Conditions on the Production of β-and α-Carotenes by Carrot Cells" Chemical Engineering Symposium Series. No.61. 149-153 (1998)

K.Nakao、K.Fukunaga、H.Funahashi 等人：“培养条件对胡萝卜细胞生产 β-和 α-胡萝卜素的影响”化学工程研讨会系列第 61 期（1998 年）。

K.Nakao, K.Fukunaga, H.Funahashi et al.: "Optimal Operating Conditions for β-Carotene Production Using Carrot Cells Cultivated in an External Loop Airlift Bubble Column" Proceedings of 7th International Conference on Computer Application in Biotechnology.

K. Nakao、K. Fukunaga、H. Funahashi 等人：“使用外环气升式气泡塔中培养的胡萝卜细胞生产 β-胡萝卜素的最佳操作条件”第七届生物技术计算机应用国际会议论文集。