目的 制备微球固定化蜗牛酶,优化微球固定化蜗牛酶的制备方法及其生物转化淫羊藿苷的工艺条件。方法 利用微乳化-凝胶法将蜗牛酶包埋固定于海藻酸钡中,单因素优化蜗牛酶的固定条件。考察微球固定化蜗牛酶生物转化淫羊藿苷的最适反应温度、pH值、底物比、底物质量浓度,并与游离蜗牛酶比较,同时考察微球固定化蜗牛酶的酶学性质。结果 利用1.0%海藻酸钠、1.0% BaCl₂及适量乳化剂(质量分数为2.5%司盘-20和1.5%聚山梨酯80),通过微乳化凝胶的方法制备微球固定化蜗牛酶。与游离蜗牛酶相比,微球固定化蜗牛酶的热稳定性、pH值稳定性均增强。微球固定化蜗牛酶转化淫羊藿苷的最适条件:转化温度50 ℃,pH值5,微球固定化蜗牛酶与淫羊藿苷质量比3:1,淫羊藿苷质量浓度为0.4 mg/mL,转化时间为2.0 h,经3次转化,转化率仍可保持(53.84±3.41)%。结论 微球固定化蜗牛酶转化淫羊藿苷工艺简单,酶学性质稳定,可重复使用,为淫羊藿苷生物转化的工业生产奠定基础。

Objective To prepare microspheres immobilized snailase, then optimize the method for making microspheres immobilized snailase and the process conditions for transformation of icariin. Methods The snailase was embedded in barium-alginate by the method of microemulsion-coacervation. The preparation process was optimized by orthogonal test. Moreover, to optimize and compare the conditions for transforming icariin, such as temperature, pH values, ratio of substrate, and concentration, and to investigate the properties of snailase. Results Sodium alginates (1.0%), BaCl₂ (1.0%), and emulgator were used for making microspheres immobilized snailase by emulsified and immobilized method. Compared with the free snailase, the stability of temperature and pH value of microspheres immobilized snailase was improved. The optimized conditions of microspheres immobilized snailase were 50 ℃, pH 5, microspheres immobilized snailase mass ratio of 3:1, icariin concentration of 0.4 mg/mL, conversion 2 h, conversion 3 times, transformational ratio up to (53.84 ±3.41)%. Conclusion Microspheres immobilized snailase is stable and reusable. Besides, the process of transform icariin is moderate, so it is important and fundamental for industrialization.

国家自然科学基金资助项目(81173557);中国中医科学院中央级公益性科研院所基本科研业务费专项资金资助(ZZ08080016)