Biochemical Engineering Journal 73 (2013) 72–79 Contents lists available at SciVerse ScienceDirect Biochemical Engineering Journal journa l h o me pa ge: www.elsevier.com/locate/bej Regular article A surfactant-coated lipase immobilized in magnetic nanoparticles for multicycle ethyl isovalerate enzymatic production Iram Mahmood a,c,∗ , Ishfaq Ahmad b , Guo Chen c , Liu Huizhou c a School of Chemical and Materials Engineering, National University of Sciences and Technology, Islamabad, Pakistan b Department of Mathematics and Statistics, International Islamic University, Islamabad, Pakistan c National Key Laboratory of Green Process and Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China a r t i c l e i n f o Article history: Received 1 June 2012 Received in revised form 3 January 2013 Accepted 24 January 2013 Available online 31 January 2013 Keywords: Magnetic nanoparticles Gum arabic Lipase Esterification a b s t r a c t Gum arabic coated magnetic Fe 3 O 4 nanoparticles (GAMNP) were prepared by chemical co-precipitation method and their surface morphology, particle size and presence of polymer-coating was confirmed by various measurements, including transmission electron microscopy (TEM), X-ray diffraction (XRD), thermo gravimetric analysis (TGA), and Fourier transform infra red (FTIR) analysis. Magnetic particles were employed for their potential application as a support material for lipase immobilization. Glutaralde- hyde was used as a coupling agent for efficient binding of lipase onto the magnetic carrier. For this purpose, the surface of a Candida rugosa lipase was initially coated with various surfactants, to stabilize enzyme in its open form, and then immobilized on to the support. This immobilized system was used as a biocatalyst for ethyl isovalerate, a flavor ester, production. The influence of various factors such as type of surfactant, optimum temperature and pH requirement, organic solvent used, amount of surfactant in coating lipase and effect of enzyme loadings on the esterification reaction were systematically studied. Different surfac- tants were used amongst which non-ionic surfactant performed better, showing about 80% esterification yield in 48 h as compared to cationic/anionic surfactants. Enhanced activity due to interfacial activation was observed for immobilized non-ionic surfactant–lipase complex. The immobilized surfactant coated lipase activity was retained after reusing seven times. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Esters of short chain fatty acids and alcohols are important com- ponents of natural aromas and flavors [1]. Hence they are in high demand and frequently used in the food, beverages, pharmaceu- tical and cosmetic industries. For instance, ethyl isovalerate is a derivative of valeric acid, mainly found in fruits (one of the prin- cipal component of blueberry) and widely used in perfumery and fragrance. However, plant extracted natural flavor esters are often either too scarce or expensive for commercial use. The use of enzymes to catalyze esterification has become a more promising method than acid- or base-catalyzed reactions for ester production [2,3]. In this context, the concept of a natural ester made by enzymatic synthesis with lipase and natural substrate is an attractive alternative to those routes [4,5]. However, stability and reusability of the enzymes have been of major concern in non- aqueous enzymatic synthesis [6,7]. Hence, utility of such enzymes ∗ Corresponding author at: School of Chemical and Materials Engineering, National University of Sciences and Technology, Sector H-12, Islamabad 44000, Pakistan. Tel.: +92 51 9085 5112; fax: +92 51 9085 5002. E-mail address: dr.iram@scme.nust.edu.pk (I. Mahmood). in industrial processes is limited by their tendency to being dena- ture and become inactivated when exposed to organic solvents. Many researchers have recently overcome the problem and use of enzymes in organic media has attracted much interest for effec- tive biocatalysis [8,9]. Some approaches to overcome this problem have been successfully developed [10,11] such as modifying the enzyme with high molecular weight synthetic polymers [12,13], or coating them with surfactants. The most significant advantages of the surfactant coated enzymes are their simple preparation proce- dure and good solubility in a wide range of organic solvents [13]. The physical modification of enzymes with surfactants is more suitable for surface hydrophobic enzymes as it can be applied in lipase assay to increase the lipid–water interfacial area by main- taining surface charge and hydrophobicity which, in turn, enhance the observed rates of lipase-catalyzed reactions [14], a process called bio-imprinting. In this regard, Mingarro et al. [15] reported a novel concept for the activation of lipolytic enzymes, interfa- cial activation-based molecular bioimprinting (IAMI), for use in nonaqueous media. Since the preparation procedure for surfactant coated enzymes is conceptually identical to the IAMI method, the conformation of lipase in the resultant complex could be considered to be an open active form. However, to fully exploit the technical and economical advantages of lipases, it is recommended to use 1369-703X/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.bej.2013.01.017