梳状亲水性环氧基载体固定化Pseudomonas stutzeri LC2-8脂肪酶

Immobilization of Pseudomonas stutzeri LC2-8 Lipase by a Comblike and Hydrophilic Support with Epoxy Groups

以氯乙酰化聚苯乙烯微球(PS-acyl-Cl)为引发剂,亲水性丙烯酰胺(AM)和甲基丙烯酸缩水甘油酯(GMA)为单体,CuCl及联吡啶(Bipy)为催化体系,通过原子转移自由基聚合反应(ATRP)制备得到梳状亲水性环氧基柔性载体(PS-acyl-P(AM-co-GMA)),通过改变AM与GMA配比,使载体环氧基含量和亲水性得到控制。用该柔性载体固定Pseudomonas stutzeri LC2-8脂肪酶,优化了固定化条件,并对柔性固定化酶性质进行考察。结果显示,当n(AM):n(GMA)=20:60,固定化时间为24 h,固定化温度为30℃,固定化pH为7.0时,固定化酶活力达到最高,为24.1U g 1。固定化酶的最适pH为8.0,最适温度为30℃,其热稳定性比游离酶高,重复使用8次,剩余酶活力80%左右。以上表明,以ATRP法合成的载体PS-acyl-P(AM-co-GMA)可成功用于脂肪酶的固定化,有效提高脂肪酶的稳定性和实用性。

A comblike and hydrophilic support with epoxy groups （PS-acyI-P（AM-co-GMA）） was synthesized by transfer radical polymerization （ATRP） method with hydrophilic acrylamide （AM） and glycidyl methacrylate as monomers, CuCI and 2-2＇-bipyridine （Bipy） complex as catalyst. The oxirane content and hydrophilicity of the prepared support could be controlled via changing the monomer＇s mass ratio of GMA to AM. The Pseudomonas stutzeri LC2-8 lipase was immobilized by the support and the main factors influencing the immobilization process and the properties of the prepared immobilized lipase were also studied. The results show that, with n（AM）：n（GMA）=20：60, immobilizing time of 24 h, coupling temperature of 30℃ and pH of 7.0, the activity of the prepared immobilized lipase can achieve its maximum value of 24.1 U.g-1. It was found that the optimal pH and temperature for the preparation of immobilized lipase are 8.0 and 30℃, respectively. The experimental results show that the thermal stability of the immobilized lipase was improved in comparison with the free lipase. The immobilized lipase could retain above 80% of its activity after 8 consecutive operations The results indicate that the support PS-acyI-P（AM-co-GMA） synthesized by ATRP method can be successfully used for the immobilization of the lipase and improve the stability and usefulness of the lipase effectively.