Research On Efficiently Extract And Purify Of β-amylase From The Soybean Whey Wastewater

Large-scale manufacturing of soybean protein isolate results in large amount of soybean whey "wastewater", which cannot be discharged into the environment, which typically contains ca.10% remaining soybean proteins and oligosaccharides, and a substantial amount of the whey protein is β-amylase (α-1,4-glucan maltohydrolase; EC.3.2.1.2).β-amylase from the soybean possesses good properties of thermo stability and a wide range of working pH, which is widely used in relative industries.A new, pragmatic process has been developed for preparing food-grade β-amylase out of such effluent and is centered by 2 ultrafiltration (UF) steps using 3 kDa and 30 kDa molecular weight cut-offs polyethersulfone membranes respectively, following with gel filtration chromatography. Before the first UF with volume concentration ratio (VCR) ca.6-8, the crude effluent is adjusted from pH 4.5 to 7 and a centrifugal supernatant is subsequently obtained. The first UF retentate is added by 1.2 M (NH4)2SO4 and followed by adjusting to pH 4. The resulted centrifugal supernatant is re-adjusted to pH 7 before being fed into the second UF, ended at VCR 10. The second UF retentate has an enzyme activity of 674 U/ml, specific activity of 14 U/mg, which is more than the request of industrialization, and is associated with 74% combined enzyme activity recovery. After gel filtration,β-amylase was purified 24 fold, giving a final specific activity of 68.6 U/mg.SDS-PAGE of the purified enzyme showed a single band and a subunit weight of about 55 kDa. Moreover, the amylase was stable within pH scope of 5-7.5, and was stable in the temperature range (55～70℃). K+、Ca2+、NH4+ and Zn2+,these four metal ions had an active effect on the β-amylase activity, while Cu2+ and Fe2+ did a negative effect on the β-amylase activity, Na+ and Mg2+ had no obvious effect on the β-amylase activity.