目的 制备交联纳米SiO₂固定化蜗牛酶，并对其理化性质、酶解淫羊藿苷为宝藿苷I的最适酶解条件及酶学性能进行考察。方法 采用共价偶联法将蜗牛酶固定在戊二醛交联纳米SiO₂上，以相对酶活力为指标，对固定化条件进行优化；采用透射电镜（TEM）、傅里叶红外光谱（FTIR）和元素分析法对固定化蜗牛酶的理化性质进行表征；并以淫羊藿苷为底物，以游离酶为对照，考察固定化蜗牛酶的最适酶解条件及酶解动力学参数、重复利用性、热稳定性等酶学性能。结果 固定化蜗牛酶制备的最佳酶载比为1：3，固定化时间为6 h；其转化淫羊藿苷的最适酶解条件为pH 5.0，转化温度60℃，酶与底物质量比1：2，转化时间12 h。固定化蜗牛酶的酶解动力学参数最大反应速率（V_max）为0.43 μg/min，米氏常数（K_m）为0.78 mmol/L，重复使用5次后的相对酶活仍能保持在70%以上。结论 制备的交联纳米SiO₂固定化蜗牛酶机械强度高、稳定性好，不但可重复使用，还可定向水解淫羊藿苷为活性更强的宝藿苷I，工艺简单易行，转化效率高，适宜于工业化生产。

Objective To prepare the immobilized snailase on cross-linked nano-SiO₂, characterize its physicochemical properties, and investigate the optimal conditions of hydrolyzing icariin to baohuoside Ⅰ and enzymatic properties. Methods The snailase was immobilized on the glutaraldehyde cross-linked nano-SiO₂ by covalent coupling method. The immobilization conditions were optimized using the relative enzyme activity as the test index. Transmission electron microscopy (TEM), fourier transform infrared spectroscopy (FTIR), and elemental analysis methods were used to characterize the physicochemical properties of immobilized snailase. Using icariin as the substrate and free enzyme as the control, the optimal enzyme conditions, enzymatic kinetic parameters, recyclability, and thermal stability of the immobilized snailase were also investigated. Results The optimal mass ratio of enzyme to carrier was 1:3 and the optimal immobilization time was 6 h for the preparation of the immobilized snailase. The optimum hydrolysis conditions of the immobilized snailase were as follows:pH 5.0, conversion temperature 60℃, mass ratio of enzyme to substrate 1:2, and transformation time 12 h. The V_max and K_m of the immobilized snailase was 0.43 μg/min and 0.78 mmol/L, respectively. After repeated use for 5 times, the residual relative enzyme activity of the immobilized snailase maintained above 70%. Conclusion The immobilized snailase on cross-linked nano-SiO₂ has high mechanical strength, strong stability, and good reusability.It can hydrolyze the icariin to the baohuoside Ⅰ with a better activity. The hydrolysis process was simple, easy, and suitable for industrial production.

国家自然科学基金青年基金项目（81703714）；江苏省科教强卫医学重点人才项目（ZDRCA2016036）