Research Article Received: 4 April 2016 Revised: 20 June 2016 Accepted article published: 18 July 2016 Published online in Wiley Online Library: (wileyonlinelibrary.com) DOI 10.1002/jctb.5060 Continuous cane sugar inversion process using immobilized invertase Aditya C Koli and Vilas G Gaikar * ABSTRACT BACKGROUND: Sugarcane juice is mostly used by industry for manufacturing sucrose and invert sugars. However, during enzymatic processing of raw sugarcane juice for an extended period of time, maintaining enzyme stability is a major bottleneck in food and agricultural industries. The present work reports use of invertase immobilized on glutaraldehyde crosslinked chitosan beads, for continuous inversion of sugarcane juice for an extended period of time. RESULTS: Complete inversion of clarified sugarcane juice was obtained in a continuous process for 4 days at an optimized flow rate of 0.3 cm 3 min -1 and at a room temperature of 303 K. Only 4% decrease was seen in inversion efficiency up to the 7th day. Adsorptive behaviour of invert sugars on chitosan in the column led to a delay in product equilibrium. The immobilized invertase exhibited the maximum rate of reaction at a sucrose concentration of 0.5 mol dm -3 showing good stability in the pH range 3.0 to 6.0 and temperature range 30 ∘ C to 60 ∘ C. CONCLUSIONS: The covalent immobilization of invertase on chitosan beads enhanced enzyme stability and as a result, the enzyme became more resistant to denaturation during storage and reuse compared with the free enzyme. Thus, better conversion was obtained in the continuous inversion of sugarcane juice for extended periods of time. © 2016 Society of Chemical Industry Keywords: immobilization; chitosan; crosslinking; sugarcane juice; filtration; continuous inversion; process; adsorption NOTATION C Molar equilibrium concentration (mol dm -3 ) k Freundlich adsorption constant n Adsorption intensity q amount of solute adsorbed per kg of adsorbant (mol kg -1 ) INTRODUCTION Sugarcane juice is a natural source of sucrose which is mostly used in industries for manufacturing sucrose or sugar. Sucrose can be hydrolyzed to invert sugar, i.e. a mixture of fructose and glucose. Invert sugar is mainly used in food, beverages, confec- tionary and pharmaceutical industries because of increased sweet- ness over sucrose and improved shelf life of the product, as it does not crystallize easily. 1,2 The conventional acid hydrolysis pro- cess for sugar inversion produces poor quality invert syrup as it becomes contaminated by undesired coloured compounds such as furfural, and has low conversion efficiency even at higher tem- peratures of 75–80 ∘ C where it is normally operated. 3 Many food industries would, therefore, prefer greener enzymatic processes. The operational stability of invertase as the enzyme catalyst for extended periods of operation remains, however, the major con- cern apart from loss of the enzyme, if used in a homogeneous state. Immobilization is an option to produce the invert sugar with concomitant reutilization of the catalyst. In addition, use of contin- uous processing in an enzymatic reactor gives additional benefits of forming low amounts of by-products over the acid hydrolysis process. 4 – 9 Monsan and Combes 8 have reported 30% higher effi- ciency for continuous hydrolysis of sucrose using a packed bed of invertase immobilized on corn grits, compared with a contin- uous stirred tank reactor using the same catalyst. Boudrant and Cheftel 7 too have reported continuous hydrolysis of sucrose using invertase immobilized on Amberlite ion exchange resin, showing constant conversion of 76% over 190 h. However, the reaction time in the reactor probably was not enough to take the conversion to completion. Polymeric adsorbents based on hydrocarbons are available in bulk quantity with reproducible characteristics, but are known to denature enzymes over time with prominent loss of activity. Mansfeld et al. 9 carried out continuous hydrolysis of concentrated sucrose solutions using polystyrene bound inver- tase, showing better stability of the immobilized invertase over one year. For a carrier to be suitable for enzyme immobilization, it must have a large surface area with a large number of reactive groups per gram of adsorbent, good mechanical and chemical stabilities and good flow properties. The most frequently used supports in biotechnology for covalent enzyme immobiliza- tion are natural polymers such as chitin, cellulose and synthetic hydrophilic polymers such as acrylate, methacrylate, acryl amides and polyvinylalcohol. 1,4 – 16 Chitosan is a nontoxic, biocompatible ∗ Correspondence to: VG Gaikar, Department of Chemical Engineering, Institute of Chemical Technology, Matunga, Mumbai – 400 019, India. E-mail: vg.gaikar@ictmumbai.edu.in Department of Chemical Engineering, Institute of Chemical Technology, Matunga, Mumbai – 400 019, India J Chem Technol Biotechnol (2016) www.soci.org © 2016 Society of Chemical Industry