Immobilization of starch phosphorylase from cabbage leaves: Production of glucose-1-phosphate

Starch phosphorylase has been isolated from cabbage (Elephantotus scabar) leaves and partially purified using ammonium sulfate fractionation. The partially purified enzyme was desalted using Sephadex-G-25 chromatography. In the direction of polysaccharide synthesis, the enzyme showed optimum activity at pH 6.0 with two half pH optima at pH 5.3 and pH 7.1 whereas in the direction of glucose-1-phosphate formation, it showed optimum pH at pH 7.0 with half pH-optima at pH 6.4 and 7.6. The optimum temperature for the enzyme activity has been found to be 37 degrees C with two half temperature optima at 34 degrees C and 40 degrees C. The partially purified enzyme has been immobilized using egg shell as solid Support. The percentage retention of the enzyme on egg shell was nearly 56%. After immobilization, specific activity of the enzyme increased from 0. 0225 to 0.0452. Upon immobilization, there was a slight alkaline shift in the optimum pH when assayed in both the directions. The immobilized enzyme also displayed increased optimum temperature and thermo-stability and could be reused number of times. The increase in thermo-stability and reusability of the immobilized enzyme has been exploited for the production of glucose-1-phosphate a cytostatic compoound used in card io-th erapy. The glucose-1-phosphate produced has been purified with nearly 95% purity after adsorption chromatography on norite and ion exchange chromatography on DEAE cellulose.