异淀粉酶在α-环糊精制备中的应用及条件优化

Application and Reaction Condition Optimization of Isoamylase in α-cyclodextrin Production

环糊精具有外侧亲水内部空腔疏水的桶形结构,可与众多的各类大小、形状合适的疏水性客体分子形成包合物,改变其理化性质。其中,α-环糊精(α-CD)具有较小的空腔、溶解性好及耐酸解,因此在很多领域具有特殊的应用。目前,单独采用环糊精葡萄糖基转移酶(α-CGTase)生产α-CD的底物利用率低,而已开发的双酶法不仅底物成本高,而且环糊精总转化率及α-CD比例也有待提高。因此,为了提高α-CD的得率,降低生产成本,本研究将α-CGTase与异淀粉酶复配,以木薯淀粉为底物,对酶转化条件进行了优化。结果表明,当底物浓度15%,液化时间10 min,温度40℃,反应p H 5.5,异淀粉酶加量144 U/g底物,α-CGTase加量15 U/g底物,正癸醇浓度5%(v/v),转化周期18 h,环糊精总转化率为90.5%,α-CD所占比例为96.3%,α-CD的转化率为国内外报道的最高水平。

Cyclodextrins(CD) have a cylindrical shape with a hydrophobic inside and a hydrophilic outside,which can form inclusion bodies with many hydrophobic molecules,changing their physical and chemical properties.α-cyclodextrin(α-CD) has smaller internal cavity,highest resistance to enzymatic hydrolysis and is more resistant to hydrolysis in acid solutions,so it has special applications in industry. At present,substrate utilization was low when produced α-CD by cyclodextrin glycosyltransferase(α-CGTase) alone. However,the developed double enzyme method not only has high cost on substrate,but also has not so good total yield of CD as well as α-CD ratio which are still remained to improve. Thus,in order to enhance the yield of α-CD and reduce production costs,the condition for α-CD preparation by conjunction use of isoamylase with α-CGTase,using the cassava starch as substrate,was optimized. Heat-treatment of 15% cassava starch at 70℃ for 10 min in the presence of 10 U α-CGTase per gram substrate allowed the starch liquefaction. At p H 5.5,40℃,5%(v/v) 1-decanol,144 U isoamylase per gram substrate,15 U α-CGTase per gram substrate incubated for 18 hours,90.5% of the starch was transformed into CD,the proportion of α-CD was 96.3%. This is the highest level of α-CD conversion by enzyme method.