新型磁性亲和吸附剂的制备及其在尿激酶纯化中的应用研究

Preparation of Novel Magnetic Affinity Adsorbents and Application for Purification of Urokinase

合成了磁性聚乙烯醇和磁性聚甲基丙烯酸甲酯微球 ,并以它们为基质 ,用环氧氯丙烷、羰基二咪唑或溴化氰活化后 ,分别键合氨基乙酸、 6-氨基己酸、乙二胺或己二胺为间隔臂 ,用 1 -乙基 -3 -(3 -二甲基氨丙基 )碳二亚胺盐酸盐或羰基二咪唑为偶联试剂 ,分别偶联对氨基苯甲脒、 L-精氨酸甲酯、胍基乙酸或胍基己酸配体 ,合成了 1 7种磁性亲和吸附剂 ,并用于尿激酶粗品的纯化 .与前文制备的 8种磁性亲和吸附剂作对照 ,研究了基质、活化试剂、间隔臂分子、偶联试剂及配体等因素对尿激酶纯化效果的影响 .这些磁性亲和吸附剂在尿激酶的纯化中取得了较好的效果 ,大多数磁性亲和吸附剂的活性回收率在 40 %～ 70 %之间 ,纯化倍数为 1 5～ 40 ,吸附容量为 0 .0 8～ 0 .2 mg/m L.

Magnetic dextran microsphere, magnetic agarose microsphere, magnetic polyvinyl alcohol microsphere and magnetic polyhydroxyethyl methacrylate microsphere were activated by epichlorohydrin, N,N′ carbonyldiimidazole or cyanogen bromide on which 6 aminohexanoic acid, glycine, ethylene diamine or 1,6 diamino hexane were bonded as spacers. Then they were coupled with p aminobenzamide, L arginien methyl ester, guanidine hexanoic acid or guanidine acetic acid using N,N′ dicyclohexyl carbodiimide, N (3 dimethylaminopropyl) N′ ethylcarbodiimide hydrochloride or N,N′ carbonyl diimidazole as coupling reagents. Whereafter 25 magnetic affinity adsorbents were prepared. To test the resolving power of them, magnetic affinity adsorbents were applied to purifying crude urokinase in a batch mode. The effect of some factors such as activators, the length of spacer, coupling reagents and ligands on the rusults of purification was discussed. Good results were obtained. The bioactivity recovery is 40%-70%, the purification fold is between 15 to 40, and the adsorptive capacity varies from 0 08 to 0 2 mg per milliliter adsorbent.