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Separation and Purification of Biochemicals by Affinity Combined with Batch-wise Adsorption and Column-wise Elution

亲和结合分批吸附和柱式洗脱分离纯化生化物质

基本信息

批准号：
62470107
负责人：
GOTO Shigeo
金额：
$ 2.18万
依托单位：
Engineering Faculty, Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1987
资助国家：
日本
起止时间：
1987 至 1988
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-62470107/
关键词：
Biochemicals Separation Purification Concentration Affinity Chromatography 吸着

项目摘要

A new process in affinity separation of biochemicals was proposed. The adsorption stage was subsituted from the conventional column-wise operation to batch-wise operation. In a batch-wise mode, the solute can be adsorbed quickly from a large quantity of very dilute soution. Then, the combined process with batch-wise adsorption and column-wise elution brings a effcient purification in a short time. Two (small and large) sizes of experiemntal apparatus were constructed. It consisted of a bubble slurry tank for adsorption and a fixed bed column for elution. The air bubble is used for stirring in the adsorber to protect mechanically weak adsorbent gel. After the solute is adsorbed in the bubble slurry tank, adsorbent is transferred to the lower column to from a fixed bed. The adsorbent is washed with buffer solution and then the adsorbed component is eluted by the eluent. After the adsorbent is washed with buffer solution, the adsorbent is moved to the upper tank for the next adsorption stage. The column-wise adsorption was carried out by using the lower column to compare with the batch-wise adsorption. Purifications of albumin by Blue-Sepharose CL-6B and trypsin by Soybean Trypsin Inhibitor and desalting by ion-exchange resin were conducted for the combined process. The adsorption isotherms for albumin and trypsin were Langmuir type and Freundlich type, respectively. It was found from experimental results of trypsin that the operating time in the batch-wise adsorption was shorter by 0.14 times than that in the column-wise adsorption. The ratio of the elution coucentration to the feed concentration was 14 times for albumin and it was 35 times for trypsin. Therefore, it was clarified from experimental and theoretical results that the combined process was effective to shorten the total operating time per one cycle.

提出了一种生物化学物质亲和分离的新工艺。吸附阶段由传统的柱式操作改为间歇操作。在分批模式下，溶质可以从大量的极稀溶液中快速吸附。然后，分批吸附和柱式洗脱相结合的工艺在短时间内实现了有效的纯化。建立了小型和大型两种尺寸的体验仪。它由用于吸附的气泡浆液槽和用于洗脱的固定床柱组成。气泡用于在吸附器内搅拌，以保护机械弱吸附凝胶。溶质在气泡浆液槽中吸附后，将吸附剂从固定床转移到下塔。用缓冲液洗涤吸附剂，然后用洗脱液洗脱被吸附的组分。吸附剂用缓冲溶液洗涤后，移至上槽进行下一步的吸附。采用下柱进行柱状吸附，并与间歇吸附进行比较。采用Blue-SepharoseCL-6B纯化白蛋白，大豆胰酶抑制剂纯化胰酶，离子交换树脂脱盐。白蛋白和胰酶的吸附等温线分别为Langmuir型和Freundlich型。对胰酶的实验结果表明，间歇吸附的操作时间比柱式吸附的操作时间短0.14倍。洗脱浓度与料液浓度之比，白蛋白为14倍，胰酶为35倍。因此，从实验和理论结果可以看出，该组合工艺有效地缩短了每一周期的总运行时间。