Journal of Molecular Catalysis B: Enzymatic 96 (2013) 6–13 Contents lists available at SciVerse ScienceDirect Journal of Molecular Catalysis B: Enzymatic jo u r n al homep age: www.elsevier.com/locate/molcatb Immobilization of -amylase on gum acacia stabilized magnetite nanoparticles, an easily recoverable and reusable support Venkatanarasimhan Swarnalatha, Rani Aluri Esther, Raghavachari Dhamodharan ∗ Department of Chemistry, Indian Institute of Technology Madras, Tamil Nadu 600036, India a r t i c l e i n f o Article history: Received 6 March 2013 Received in revised form 25 May 2013 Accepted 25 May 2013 Available online 3 June 2013 Keywords: -Amylase Immobilization of enzyme Gum acacia Magnetite nanoparticles a b s t r a c t In this work, -amylase is immobilized, using glutaraldehyde, onto magnetite nanoparticles prepared using gum acacia as the steric stabilizer (GA-MN), for the first time. The immobilization of amylase to GA- MN is very fast and the synthesis of GA-MN is very simple. The use of GA enables higher immobilization of -amylase (60%), in contrast to the unmodified magnetite nanoparticles (∼20%). The optimum pH and temperature for maximum enzyme activity for the immobilized amylase are identified to be 7.0 and 40 ◦ C, respectively, for the hydrolysis of starch. The kinetic studies confirm the Michaelis–Menten behavior and suggests overall enhancement in the performance of the immobilized enzyme with reference to the free enzyme. Similarly the thermal stability of the enzyme is found to increase after the immobilization. The GA-MN bound amylase has also been demonstrated to be capable of being reused for at least six cycles while retaining ∼70% of the initial activity. By using a magnetically active support, quick separation of amylase from reaction mixture is enabled. The catalytic rate of amylase is actually found to enhance by twofold after the immobilization, which is extremely advantageous in industry. At higher temperature, the immobilized enzyme exhibits higher enzyme activity than that of the free enzyme. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Enzymes are biological macromolecules that are used as bio- catalysts. Despite having many advantages viz. mild reaction conditions and high stereo-, regio- and enantiospecificity, enzymes suffer few demerits [1]. In contrast to conventional catalysts, enzymes tend to degrade on long-standing. The storage and processing conditions also affect their stability. It is always tricky to recover and reuse the enzymes after the catalytic transformations. So to accomplish the recovery and to boost the thermal stability of enzymes, enzyme immobilization is implemented. In enzymol- ogy, enzyme immobilization is a process where the enzyme is immobilized onto a stationary phase prior to the reaction. This immobilization protocol simplifies the separation of the enzymes from the products, once the reaction is done. Immobilization also facilitates the simultaneous multi-enzyme catalysis reactions and the enzymes can be very easily recovered for recycling, unlike free enzyme catalysis. The immobilized enzymes also possess greater thermal stability and extended life-time relative to the un- immobilized analogs. Amylases are hydrolase enzymes, which hydrolyze starch or its hydrolytic products to reducing sugars. They are one among ∗ Corresponding author. Tel.: +91 4422574204. E-mail address: damo@iitm.ac.in (R. Dhamodharan). the most largely consumed enzymes and their sale is estimated to be approximately US $ 225 million throughout the world [2]. Amylases, which are produced by a wide range of organisms, con- stitute an approximate 25% of the entire global enzyme market [3]. Sources of amylases play a crucial role during the commer- cial applications as they alter the operational parameters such as pH, temperature etc., markedly. -Amylase (1,4--d-glucan- glucanhydrolase, EC.3.2.1.1), which is one of the several amylase enzymes, hydrolyzes starch to maltose by randomly cleaving the internal -1,4-linkages (endoamylase). The major industrial sec- tors, which expansively spend -amylase are for the production of high fructose corn syrup, ethanol and paper recycling. Various types of carriers have been previously investigated for the effective immobilization of -amylase such as functionalized glass beads [4], mesoporous silica [5], amberlite MB 150, chitosan beads [6], gelatin [7], alginate [8], poly(hydroxyethyl methacrylate), copolymers of styrene and hydroxyethyl methacrylate [9], poly(acrylamide) [10], modified poly(N-isopropylacrylamide) [11], polyaniline [12], porous nitrocellulose [13], silver nanoparticles doped gum acacia- gelatin-silica nanohybrid [14] and tamarind gum-silica nanohybrid [15] etc. In the past few decades, materials of nano dimension have been dramatically emerging and have eventually reached an inevitable position in all the disciplines of science and tech- nology. Among these nanomaterials, magnetic nanoparticles are considered quite unique since they possess excellent and unusual 1381-1177/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.molcatb.2013.05.022