Screening Of High Temperature Resistant Amylase Producing Strains And Studies On Enzyme Production Conditions And Enzymatic Properties

Using starch hydrolysis circle as the screening model, the strain E10 with high production capacity of thermostable amylase was achieved in this paper by screening the soil collected from the paddy field near the school. In order to improve the enzyme production capability of the E10, single factor experiments were done for the kinds and mass fractions of carbon source, nitrogen source and inorganic salt; as well as for the pH value, seed age, inoculate volume, temperature and time. The L9(34)orthogonal optimization test for the fermentation medium and conditions was done by selecting the major factors influencing the enzyme production of fermentation medium(the mass fractions of wheat brain and potassium nitrate, calcium chloride, pH value) and fermentation conditions (the seed age, inoculate volume, temperature and time). The thermostable amylase was purified by the methods of salting out, dialysis and column chromatography. The enzyme's formula weight was determined by SDS gel electrophoresis, and its enzymological properties were also studied. The results showed that the optimal fermentation medium of the enzyme production was wheat bran 2.0%, KNO30.1%, CaC120.25%, with a starting pH value of 6.0. The optimal fermentation conditions of the enzyme production were:24 hours of seed age,7% of inoculate volume,36? of fermentation temperature,48 hours of time. Under the optimized conditions, the enzyme production of E10 could reach 180.94 IU/mL. The amylase after column chromatography was electrophoretically pure, with the optimum temperature at 70? and the optimum pH value at 7.0. The results of the enzyme stability test showed that the enzyme was stable under 70? at the pH value range of 6.0-7.0. The K+ ion promotes the activity of amylase, while the Mn2+, Cu2+, Fe2+, Ca2+ and Mg2+ ion restrain the activity of amylase.