Saccharification and Isomerization by GA and GI 193 Applied Biochemistry and Biotechnology Vol. 102–103, 2002 193 *Author to whom all correspondence and reprint requests should be addressed. Copyright © 2002 by Humana Press Inc. All rights of any nature whatsoever reserved. 0273-2289/02/102–103/0193/$11.75 Effect of pH on Simultaneous Saccharification and Isomerization by Glucoamylase and Glucose Isomerase ABHA MISHRA* AND MEERA DEBNATH (DAS) School of Biochemical Engineering, Institute of Technology, Banaras Hindu University, Varanasi 221005, India, E-mail: misrabha91@rediffmail.com Abstract pH and temperature play critical roles in multistep enzymatic conver- sions. In such conversions, the optimal pH for individual steps differs greatly. In this article, we describe the production of glucoamylase (from Aspergillus oryzae MTCC152 in solid-state fermentation) and glucose isomerase (from Streptomyces griseus NCIM2020 in submerged fermentation), used in indus- tries for producing high-fructose syrup. Optimum pH for glucoamylase was found to be 5.0. For glucose isomerase, the optimum pH ranged between 7.0 and 8.5, depending on the type of buffer used. Optimum temperature for glucoamylase and glucose isomerase was 50 and 60°C, respectively. When both the enzymatic conversions were performed simultaneously at a com- promised pH of 6.5, both the enzymes showed lowered activity. We also studied the kinetics at different pHs, which allows the two-step reaction to take place simultaneously. This was done by separating two steps by a thin layer of urease. Ammonia generated by the hydrolysis of urea consumed the hydrogen ions, thereby allowing optimal activity of glucose isomerase at an acidic pH of 5.0. Index Entries: Glucoamylase; glucose isomerase; Aspergillus oryzae; Strep- tomyces griseus. Introduction In multienzyme systems, the product of one enzymatic reaction becomes the substrate for the other. One such example is the production of high-fructose syrup from low-cost starchy materials. High-fructose syrup, an equilibrium mixture of glucose and fructose, is approximately one to three times sweeter than sucrose. It is preferred in the food industry because it does not pose a problem of crystallization, as happens with sucrose.