Studies on immobilization of alfa- amylase to cyanogen bromide activated sephadex g 200
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Date
1977
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University of Sri Lanka (Colombo Campus): UC(MED).
Abstract
Studies were made to characterise soluble Alfa-Amylase (Bacterial). The kinetics of Alfa-amylase on starch is zero order at room temperature (28 0 c) for the first 5 minutes. Hence the activity of Alfa-amylase was measured in terms of mg maltose released during first five minutes. Alfa-amylase showed optimal activity at pH 6.0 Cyanogen bromide was prepared and was used to activate sephadex G200 at pH 11.5. The volume of the cyanogen bromide activated gel, at this pH decreased by about 50 percent, compared to that of the unactivated Sephadex G200. Alfa-amylase was coupled to cyanogen bromide activated Sephadex G200 at pH8.3 and 7.0. Coupling of the enzyme led to further decrease in volumes of about 15 percent and 6 percent at pH 8.3 and 7.0 respectively. The amounts of protein in the immobilized Alfa-amylase prepared at pH 8.3 and 7.0 were estimated by, Kjeldhal method, by the tryptophan content and from the difference in the amount of protein present in the original solution and that in the washings. It was observed that 27 percent of the added protein got immobilized at pH 8.3, Where as at pH 7.0 it was only 11 percent. The activities of the Alfa-amylases immobilized at pH 8.3 and 7.0 were, 0.75 percent and 0.52 percent of the total soluble Alfa-amylase respectively. The loss of activity at pH 8.3 and 7.0 were 97.4 percent and 96 percent respectively, which is significant at both pHs. The immobilized Alfa-amylase (prepared at pH 8.3) was stable on storage for a period of 3 weeks at 4 0 C in 0.4M phosphate buffer (pH 6.9) containing 0.02 percent sodium azide. The drop in activity during this period was only 8 percent. A sample of immobilized Alfa-amylase (prepared at pH 8.3) was tested thrice in 3 weeks for stability on usage and it retained 85 percent of its original activity.
Description
Dissertation: M.Sc., University of Sri Lanka (Colombo Campus): UC(MED), 1977.
Keywords
Amylases
Citation
ATTYGALLE, AB, Studies on immobilization of alfa- amylase to cyanogen bromide activated sephadex g 200, University of Sri Lanka (Colombo Campus) UC(MED), 1977: 50p.