A Study of the Optimal Conditions for Starch Hydrolysis Through Thermostable Α - Amylase

2137 Words Jul 10th, 2013 9 Pages
T. Kolusheva, A. Marinova Journal of the University of Chemical Technology and Metallurgy, 42, 1, 2007, 93-96

T. Kolusheva, A. Marinova

University of Chemical Technology and Metallurgy 8 Kl. Ohridski, 1756 Sofia, Bulgaria E-mail: e-mail: manahova@abv.bg.

Received 10 July 2006 Accepted 12 November 2006

ABSTRACT The present work determines the optimal conditions for starch hydrolysis by thermostable α -amylase (EC produced by Bac.subtilis strain XÊ-86. The hydrolysis reaction has the greatest rate at pH = 7.0, starch substrate concentration 250 g.l-1, enzyme concentration – 12 enzyme units per ml suspension and 90o C temperature. We
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Phosphate buffer containing 1/15 mol l-1 solutions of KH2PO4 and Na2HPO4, pH = 7.0 is used as medium for carrying out the enzyme hydrolysis of starch. The experiments are performed in a glass cell with 150 ml volume and constant stirring of the suspension. The suspensions of the substrate are prepared with phosphate buffer solution to which the starch is added gradually while constantly stirring. The obtained suspension is heated to the desired temperature and after its attainment, the enzyme is progressively added. At various intervals, samples of 1.00 ml volume are taken from the obtained hydrolysate and placed in a volumetruk flask of 100 ml. Immediately 0.5 ml 2 mol/l-1 HCl is added to the sample to stop the hydrolysis. The reducing sugars obtained during the starch hydrolysis are determined by the complexometric method we have developed earlier [8]. According to this method, 25 ml distilled water and 25 ml of both Fehling I and II solutions are added to the sample hydrolysate. The sample is heated with a boiling water bath for 5 min. The flask is cooled down and filled up to the mark with distilled water, after which the sample is homogenized. The red residue of Cu2O thus obtained is filtered through a pa-

per filter for small crystalline residues. A sample with 25 ml volume is taken from the filtrate and transferred to an Erlenmeyer flask of 300 ml volume. 8 ml acetate buffer solution (2 mol.l-1, pH = 5 - 5.1) and 0.10

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