固定化黑曲霉β-葡萄糖苷酶制备染料木素活性苷元的研究

Study on Isoflavone Active Aglycone Preparation by Immobilized β-glucosidase from Aspergillus niger

摘要:

比较了以海藻酸钠为载体，用胶囊法、包埋-交联法、交联-包埋法三种不同方法固定化黑曲霉β-葡萄糖苷酶的效果，并研究了最佳固定化方法的固定化条件和固定化酶的部分性质。结果表明，交联-包埋法即β-葡萄糖苷酶与0.20%戊二醛交联后再用2.0%海藻酸钠包埋的固定化方法中酶结合效率和酶活力回收率最高。海藻酸钠浓度和戊二醛浓度对酶结合效率影响较大，戊二醛浓度和包埋颗粒直径大小对酶活力回收率影响显著。与游离酶相比，制备的固定化酶最适温度、最适pH值和K_m值分别由50℃、4.5和2.57 μg/mL下降到40℃、4.0和2.02 μg/mL。固定化酶具有更强的耐酸性和稳定性。该固定化酶用于大豆异黄酮活性苷元染料木素的合成，重复使用6次后，固定化酶的活力仍保持84.94%，染料木苷转化率为56.04%。

Abstract:

With sodium alginate as a carrier and glutaraldehyde as the crosslinking agent, an improved immobilization method of β-glucosidase for production of soybean genistein was developed. As compared with entrapment or entrapment-crosslinkage, crosslinkage-entrapment that β-glucosidase was treated with glutaraldehyde and then entrapped in sodium alginate remained high loading efficiency and activity recovery, Effects of bead sizes, concentrations of alginate and glutaraldehyde as well, on the loading efficiency and activity recovery were assessed. When compared with the free enzyme, the optimum temperature, pH value and K_m of the immobilized β-glucosidase were respectively shifted from 50℃ to 40℃, 4.5 to 4.0 and 2.57 μg/mL to 2.02 μg/mL. The stabilities of the immobilized β-glucosidase were considerably better than that of the native enzyme. The immobilized β-glucosidase was employed to genistein production, 84.94% of the activity and 56.04% of conversion were kept after consecutive use of 6 times.