CNBr-Papain-Sepharose 4B亲和填料的制备及其性质研究

Preparation and Properties of CNBr-Papain-Sepharose 4B Affinity Media

为探求从鱼类废弃组织及漂洗水中回收Cystatin的高效方法,首先自制了溴化氢-木瓜蛋白酶-琼脂糖凝胶(CNBr-Papain-Speharose 4B)亲和填料,确定了适宜的配基Papain偶联密度,之后测定了该亲和填料的热稳定性、酸碱稳定性、贮藏性能及使用的重复性,最后以该制备的填料对鱼糜漂洗水中的CPIs活性进行了初步回收。以对重组鲢鱼Cystatin吸附率为评价指标,确定适宜Papain偶联密度,即CNBr-Speharose 4B和Papain溶液最佳料液比为1∶7(g∶mL)。CNBr-Papain-Speharose 4B的适宜使用温度为4~40℃,且在pH 3.0~12.0之间稳定。CNBr-Papain-Speharose 4B在4℃下,6个月的贮藏期内,表现较好的稳定性。将填料装柱后进行验证实验,证明该填料的使用重复性能稳定,且最终成功高效地回收了鲢鱼鱼糜漂洗水中的CPIs。

To explore an efficient method of recycling Cystatin from fish waste tissue and rinsing water,the self-made affinity media CNBr-Papain-Speharose 4B was firstly prepared.The optimal coupling density of Papain was further determined.Then the thermal stability,acid-base stability,storage performance and repeatability were respectively detected.At last,the active CPIs in silver carp surimi rinse water was preliminary recycled by the self-made affinity media.The optimal coupling density of Papain was evaluated by the adsorption rate of recombinant carp cystatin.The best liquid-solid ratio of CNBr-Speharose 4B toPapain was indicated as 1∶7(g∶mL).The optimal temperature of CNBr-Papain-Speharose 4B was 4~40℃during usage,and it was stable between pH 3.0~12.0.The storage detection suggested that CNBr-Papain-Speharose 4B was stableat 4℃within 6 months.The affinity media's repeatability was verified to be stable.Finally,CNBr-Papain-Speharose 4B affinity media successfully recycled CPIs from silver carpsurimi rinse water with high efficiency.