黑曲霉所产α-L-鼠李糖苷酶酶学性质及其在异槲皮素制备上应用

Study on the enzymatic properties of alpha-L-rhamnosidase by Aspergillus niger and its application in the preparation of isoquercitrin

本文前期筛选得到一株产α-L-鼠李糖苷酶的黑曲霉,并利用该酶转化芦丁制备异槲皮素。该黑曲霉在以芦丁作为诱导剂的培养基中生长,对发酵液用70%硫酸铵沉淀,透析得到α-L-鼠李糖苷酶的粗酶液,酶活为798 U/m L,用此酶液对芦丁进行水解制得异槲皮素。采用SDS-PAGE电泳测定粗酶液的酶蛋白分子量为55 k Da,经优化该酶的最适温度为55℃,最适p H为5.5,在此条件芦丁转化45 min后的转化率高达到95%以上。用柱层析处理能得到纯度99%以上的异槲皮素。该酶同样具有良好的热稳定性和高催化活性。本研究可为α-L-鼠李糖苷酶用于天然产物的脱糖基化这一类生物转化提供有效借鉴。

In this paper we studied one strain Aspergillus niger to produce alpha- L-rhamnosidase by fermentation, and isoquercitrin was prepared by curve enzyme with rutin as raw material.The Aspergillus niger was grown in the medium which containing rutin as an agent, and secretion alpha-L-rhamnosidase.The enzyme fermentation broth was precipitated by 70% ammonium sulfate,and the crude enzyme solution of alpha-L-rhamnosidase was obtained by dialysis.The enzyme activity was detected as 798 U/mL.The crude enzyme was then used to hydrolyze rutin into isoquercitrin.The molecular weight of the enzyme protein was 55 kDa by SDS- PAGE electrophoresis. The optimal temperature and pH of the enzyme were 55℃ and 5.5, respectively.The conversion rate of rutin reached 95% through in the optimum conditions described above. Using column chromatography method, isoquercetin purity could reach more than 99% .The enzyme also showed good thermal stability, and it can be used to convert rutin into quercetin, which has the advantages of high conversion rate,less by-products and high purity of the product.This study can provide an effective reference for the biotransformation of natural products.