Production Of High-Purity FOS By Immobilized Mixed-Enzyme System And Nanofiltration Technology

Fructooligosaccharides(FOS), a functional sweetener used widely in healthfood, have lots of favorable biological and functional properties such as lowcalorie and noncariogenic sweeteners, consecutive improvement of the intestinalmicrobial flora, relief of constipation, decrease of total cholesterol and lipid inthe serum. The G-type FOS production converted by immobilizedfuctosyltransferase(FTS)or zymogen is not suit for the diabetics because inwhich the content of glucose and sucrose that have no biological properties istoo high and is about 35% and 15%, respectively.In the course of FOS producing, immobilized glucose oxidase(GOD) andimmobilized catalase(CAT) were used for removing glucose which iscompetitive inhibitor to immobilized FTS. As the result, sucrose was convertedfurthest and high-content FOS (more than 80%, FOS/total saccharide) was get.And then, used the high-content FOS as stuff, P-type FOS (FOS/total saccharide≥90%) was obtained by nanofiltration for glucose-removing.The optimal immobilization condition for GOD and CAT were investigatedand their enzymatic properties were studied on. The optimal immobilizationcondition for GOD was as follows: carrier—D380 positive ion exchange resin,carries/GOD—1g/2mg, gluctarldehyde concentration—0.2%, temperature—30℃,cross-link time--8h, pH7.0. It was found that free and immobilized GOD havethe pH optimum of 5.5 and 6.0, and were stable over a pH range of 5.0-7.0. Theoptimal temperature and thermostability of the immobilized GOD was improvedover that of the free enzyme, resulting in a temperature range for activity of35-45℃and a retention of 56% of its optimal activity when subject to 55℃for60 min, compared to the free enzyme which exhibited a 30-40℃activity and aretention of 28% of its optimal activity. The Km of the immobilized GOD(59.8g/ml) was about 2.1 fold that of the free enzyme (28.6g/ml).The optimal immobilization condition for CAT was as follows: PF resin,carries/CAT—1 g/3mg, gluctarldehyde concentration—0.15%, temperature—30℃,cross-link time—10h, pH7.0. It was found that the immobilized CAT have the pHoptimum of 6.0 and was stable over a pH range of 5.0-7.0. The immobilizedCAT have the temperature optimum of 40℃and was stable over athermostability range30-45℃. The optimal proportion to immobilized GOD andimmobilized CAT for catalyzing glucose is 1/2(g/g). Used 100 ml 30% sucrosesolution as substrate, added immobilized FTS 1.5 g, immobilized GOD 1.5g andimmobilized CAT 3.0g as catalyzer, intermittently added CaCO₃ to maintain pH, for reacting 20 h at 30℃, high content FOS(about 80%) was get. And then, usednanofiltration for glucose-removing, P-type FOS(FOS/total saccharide≥90%)was obtained.