DEVELOPMENT OF COMPOSITE OF CHITOSAN AND DEXTRAN FOR SEPARATION OF PROTEINS AND ITS APPLICATION FOR INDUSTRIAL LARGE SCALE PROTEIN SEPARATION

壳聚糖与右旋糖酐复合蛋白质分离材料的研制及其在工业化大规模蛋白质分离中的应用

基本信息

批准号：
07650926
负责人：
YOSHIDA Hiroyuki
金额：
$ 1.41万
依托单位：
Osaka Prefecture University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

Ion exchange chromatography is useful for large industrial-scale separation of proteins. A number of ion exchangers have been developed. Dextran-DEAE such as DEAE Sephadex A-50 (Pharmacia LKB Biotechnology) is a very good protein separator. However since it is very soft, it is compressed in a column even in very slow flow rate and therefore it can not be used in industrial-scale protein separation.In the present work, we have developed a composite of chitosan and dextran-DEAE to make the dextran-DEAE hard. The supporter of the dextran-DEAE was a chitosan bead, Chitopearl 2501 (Ch-2501, Fuji Spinning Co.), which was a hard gel and an adsorber of protein in itself. We let dextran penetrate into the macropores of Ch-2501 by contacting with dextran solution. Thereafter, the dextran in the macropore was cross-linked and then DEAE was introduced on the dextran network. The finished product (hereafter called Ch-Dx-DEAE-2) was hard and was not compressed in a column at all. The equilibrium iso … More therm for adsorption of BSA was affected by pH considerably. This means that Ch-Dx-DEAE-2 is feasible for separation of proteins. The isotherms were correlated well by the Langmuir equation. The saturation capacity for adsorption of BSA on Ch-Dx-DEAE-2 was 4.5,2.5, and 1.3 times larger than those of DEAE Sepharose Fast Flow (Pharmacia LKB Biotechnology), Ch-2501, and DEAE Sephadex A-50, respectively. The experimental breakthrough curve agreed well with the theoretical breakthrough curve obtained by considering the intraparticle diffusion, liquid film diffusion, and axial dispersion in the bed. The intraparticle effective diffusivities of BSA were determined by matching the experimental breakthrough curves with the theoretical ones. The diffusivity in Ch-Dx-DEAE-2 was the largest in the above ion exchangers. It was ten times larger than that in Ch-2501. BSA adsorbed on Ch-Dx-DEAE-2 could be desorbed using NaCl aqueous solution easily. The elution curve of BSA from the bed was very sharp.The value of the peak was 870 times larger than that of feed concentration of BSA in adsorption process. This is 4 times larger than that obtained using the best commercial ion exchanger which is used commonly in the world. Less

离子交换色谱可用于大规模工业规模的蛋白质分离，已开发出许多离子交换剂，例如 DEAE Sephadex A-50（法玛西亚 LKB 生物技术公司），但由于其性能非常差。它是软的，即使在非常慢的流速下也会被压缩在柱中，因此它不能用于工业规模的蛋白质分离。在目前的工作中，我们开发了一种壳聚糖和葡聚糖-DEAE使葡聚糖-DEAE变硬葡聚糖-DEAE的支持体是壳聚糖珠Chitopearl 2501(Ch-2501，Fuji Spinning Co.)，其本身是硬凝胶并且是蛋白质的吸附剂。通过与葡聚糖溶液接触，让葡聚糖渗透到Ch-2501的大孔中，之后，大孔中的葡聚糖发生交联。然后将 DEAE 引入到葡聚糖网络上，最终产物（以下称为 Ch-Dx-DEAE-2）很硬，根本没有在柱中压缩。BSA 吸附的平衡热受 pH 影响。这意味着 Ch-Dx-DEAE-2 可以通过 Langmuir 方程很好地分离蛋白质。 Ch-Dx-DEAE-2 上 BSA 的吸附饱和能力为。分别比 DEAE Sepharose Fast Flow (Pharmacia LKB Biotechnology)、Ch-2501 和 DEAE Sephadex A-50 大 4.5、2.5 和 1.3 倍。实验突破曲线与考虑颗粒内获得的理论突破曲线非常吻合。通过将实验突破曲线与理论曲线相匹配，确定了 BSA 的扩散、液膜扩散和轴向扩散。 Ch-Dx-DEAE-2 中的扩散系数是上述离子交换剂中最大的，比 Ch-2501 中的扩散系数大 10 倍，Ch-Dx-DEAE-2 上吸附的 BSA 很容易用 NaCl 水溶液解吸。 BSA从床层的洗脱曲线非常尖锐。峰值比吸附过程中BSA的进料浓度大870倍，比使用时获得的值大4倍。世界上最常用的最好的商业离子交换器。