Extraction, Purification And Application Of β-amylase From Malt

The advantages ofβ-amylase from plant have high activity, good thermo-stability, width pH range and so on. The paper is not only research the condition of extraction,purification and characterization, but also the application.The activities ofβ-amylase in different time and extraction conditions were compared in the experiment. The extraction process ofβ-amylase from germinated barley was researched on the basis of one-factor tests and the Box-Behnken center-united experimental design principle. The method of response surface analysis with 5 factors and 3 levels was adopted. A second order quadratic equation for extraction ofβ-amylase was built. Based on the analysis of the response surface plots,effects of extraction conditions were explored. The results showed that the extraction activity ofβ-amylase from germinated barley in third day was the highest; By using this new method, the optimum extraction condition was obtained as follows: liquid to material ratio 1:17, extraction temperature 44℃, buffer-solution pH value 6.4, extraction time 2.3 h, NaHSO3 concentration 1.64 g/L. Under the optimized condition, the average extraction activity ofβ-amylase from 1 gram was 1230.22 U.The crude enzyme was purified to ammonium sulphate precipitation,and then used two successive steps including DEAE-650M and HW-55F. The final recovery rate of the total enzyme activity was 41.1%, the activity increase of 21.8 folds, the specific activity was 1462.7 U/mg. The molecular weight of the enzyme was estimated to be 60 kDa by SDS-PAGE.The optimal temperature and pH of theβ-amylase were 50℃and 5.5, respectively. It was relatively stable in 40-60℃and pH 4.5-6.5. When the enzyme was kept 70℃for one hour, the remained activity was 6%. Mg2+,Ca2+ and Zn2+ had activation effect on the enzyme activity seriously. Mn2+,Cu2+ and Fe2+ had inhibitory effect on the enzyme activity seriously. Na+,K+ and NH4+ had no effect on the enzyme activity . The advantages ofβ-amylase from plant have high better thermo-stability and width pH range than that from microorganism. The preparation of high maltose syrups using potato starch mixed with enzyme was studied.The liquefying and mashing process of potato were engaged using heat-stableα-amylase ,β-amylase made by ammonium sulphate precipitation and pullulanase respectively, the best liquefying technique and mashing technology were obtained. The optimum liquefying condition was: 40% starchpulp, pH6.3, heat-stableα-amylase: 10.2 U/g starch , temperature 98℃, 10min,and the optimum mashing condition was: pH5.5,β-amylase 455.67 U/g starch , pullulanase 0.864 PUN/g starch, 48h, 50℃. The content of maltose in this condition was reached to 68.37%, which could meet standard requirement. It suggests that the maltβ-amylase made by ammonium sulphate precipitation could be applicated in industry.In the same conditions, the content of maltose made by maltβ-amylase is higher than made by microorganismβ-amylase.