茯苓药渣酶解工艺及其产物初步分析

Optimization of Enzymatic Preparation Process from the Residue of Poria cocos(Schw.) Wolf and its Products Analysis

目的优化茯苓药渣酶解工艺,制备水溶性酶解产物,并初步分析酶解产物的结构。方法通过Box-Behnken设计-响应面法优化制备工艺,酶解产物经Sevage法脱蛋白,Sephadex G-75、DEAE Sepharose F F以及Sephadex G-50纯化,采用PMP柱前衍生-高效液相色谱法(HPLC)和高效凝胶渗透色谱-多角度激光光散射-示差折光联用(HPGPC-MALLS-RI)测定其单糖组成和分子量,刚果红实验检测三股螺旋结构的有无。结果使用蜗牛酶,反应温度49℃,pH 6.2,反应时间34 h,酶用量2.6%,酶解得率达到36.97%左右,纯化后产物主要是由葡萄糖组成的低聚糖,由3个组分构成,重均分子量分别为1.739×10^(5)、811.9、435.4,其中重均分子量为811.9的低聚糖为中性糖,占92.1%,是主要组成部分,纯化酶解产物不具有三股螺旋结构。结论蜗牛酶适用于降解茯苓药渣,工艺稳定可行,所得产物得率较高,水溶性好,为进一步开发利用茯苓资源奠定了基础。

Objective To optimize the preparation process of water-soluble polysaccharides or oligosaccharide from the residue of Poria cocos(Schw.)Wolf and to preliminarily analyze the structure of enzymolysis products.Methods Box-Behnken design-response surface method was used to study the optimum preparation conditions.The protein was deproteinized by Sevage method,and the enzymolysis products was purified by Sephadex G-75,DEAE Sepharose F F and Sephadex G-50 column chromatography.The monosaccharide composition and molecular weight was determined by HPLC with PMP precolumn derivatization and HPGPC-MALLS-RI.The Congo red assay was used to detect the existence of three-stranded helical structures.Results Using snailase,with the optimum hydrolysis condition for Poria cocos enzymatic hydrolysis of 49℃,pH 6.2,34 h,the enzyme dosage was 2.6%,the extraction rate of pachyman was about 36.97%.After purified,polysaccharide were main composed of glucose was made up of three components,which Mw was 1.739×10^(5),811.9,and 435.4,and the oligosaccharide of the main component having Mw of 811.9 accounted for 92.1%.There was no triple helix structure existed in this fraction.Conclusion The processing technology of selecting Snailase and hydrolyzing polysaccharides optimized by Box-Behnken design-response surface method has the advantages of stable and a new processing technology for water solubility enzymolysis products.which provides some ideas for the further development and utilization of poria cocos resources.