褐藻胶裂解酶AlgL17酶解工艺优化及酶解产物分析

Optimization of Enzymatic Hydrolysis Process and Analysis of Hydrolysates by Alginate Lyase AlgL17

为探索重组褐藻胶裂解酶AlgL17酶解工艺的优化条件,采用3,5-二硝基水杨酸法(3,5-dinitrosalicylic acid,DNS)和苯酚-硫酸法分别测定反应生成的还原糖及总糖含量,计算平均聚合度,从反应温度、反应pH值、底物质量分数、加酶量及振荡速率5方面探讨各因素对酶解反应的影响,确定褐藻胶裂解酶酶解工艺的最优条件。结果表明,AlgL17酶解海藻酸钠的最优工艺条件:反应温度为35℃,pH=8. 0,初始底物质量分数为1. 1%,加酶量为1. 16 U,不振荡反应180 min。在此条件下,产生的还原糖质量浓度为7. 09 g/L,酶解产物的平均聚合度为2。液相色谱质谱联用(liquid chromatography-mass spectrometry,LC-MS)分析海藻酸钠酶解终产物为单糖、二糖、三糖和四糖。

In order to explore the optimal conditions for enzymatic hydrolysis process of the recombinant alginate lyase AlgL17,the contents of reducing sugar and total sugar in sodium alginate hydrolysates were measured by DNS method and phenol-sulfuric acid method,respectively.The average degree of polymerization was calculated.The effects of various factors,such as reaction temperature,pH,substrate concentration,amount of enzyme added and oscillating rate,on enzymatic hydrolysis were investigated.The results showed that the optimum conditions for the hydrolysis of sodium alginate by AlgL17were as follows:temperature35℃,pH=8.0,initial substrate mass concentration1.1%,enzyme1.16U,reaction time180min with non-oscillation.At these conditions,the mass concentration of the produced reducing sugar was7.09g/L and the average poly-merization degree was2.LC-MS analysis showed that the final enzymatic hydrolysates of sodium alginate were monosaccharide,disaccharide,trisaccharide and tetrasccharide.