African Journal of Biotechnology Vol. 6 (23), pp. 2715-2720, 3 December 2007   

ISSN 1684–5315 © 2007 Academic Journals        

Full Length Research Paper

Immobilization of starch phosphorylase from seeds of Indian millet (Pennisetum typhoides) variety KB 560

Sarika Garg and Anil Kumar*

School of Biotechnology, Devi Ahilya University, Khandwa Road, INDORE-452001, India.

*Corresponding author. E-mail: ak_sbt@yahoo.com. Tel: 91-731-2470372, 2470373. Fax: 91-731-2470372.

Accepted 6 November, 2007

Starch phosphorylase has been isolated from the seeds of millet (Pennisetum typhoides) variety KB560 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 5.4 and pH 7.0 whereas in the direction of glucose-1-phosphate formation, it showed optimum pH at pH 7.6 with half pH optima at pH 6.7 and 7.9. The optimum temperature for the enzyme activity has been found to be 37^oC with two half temperature optima at 34 and 40^oC. The partially purified enzyme has been immobilized using brick dust as solid support. The percentage retention of the enzyme on brick dust was nearly 80%. After immobilization, specific activity of the enzyme increased from 0.816 to 2.89. 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 compound used in cardio-therapy. The glucose-1-phosphate produced has been purified with nearly 95% purity after adsorption chromatography on norite and ion exchange chromatography on DEAE cellulose.  

Key words: Starch phosphorylase, immobilization, glucose-1-phosphate, brick dust, millet.