Lipase entrapment in PVA/Chitosan biodegradable film for reactor coatings Karla A. Batista a , Flavio Marques Lopes a, b , Fabio Yamashita c , Kátia Flávia Fernandes a, ⁎ a Departamento de Bioquímica e Biologia Molecular, Laboratório de Química de Proteínas, Universidade Federal de Goiás, Cx. Postal 131, 74001-970, Goiânia, GO, Brazil b Unidade Universitária de Ciências Exatas e Tecnológicas, Universidade Estadual de Goiás, Anápolis, GO, Brazil c Departamento de Tecnologia de Alimentos e Medicamentos, Laboratório de Tecnologia, Universidade Estadual de Londrina, Cx. Postal 6001, CEP 86051-990, Londrina, PR, Brazil abstract article info Article history: Received 2 February 2012 Received in revised form 3 December 2012 Accepted 24 December 2012 Available online 31 December 2012 Keywords: Immobilization Olive oil hydrolysis Stability Repeated use Bioreactor This study reports the development and characterization of novel biodegradable film, based on chitosan and polyvinyl alcohol containing lipase entrapped. The films showed a thickness of 70.4 and 79 μm to PVA/Chitosan and PVA/Chitosan/Lipase, respectively. The entrapment of lipase in PVA/Chitosan film resulted in increasing of 69.4% tensile strength (TS), and 52.4% of elongation. SEM images showed the formation of a continuous film, without pores or cracks. The lipase entrapment efficiency was estimated in 92% and the films were repeatedly used for 25 hydrolytic cycles, maintaining 62% of initial activity. The PVA/Chitosan/Lipase film was used for olive oil hydrolysis of high performance. These results indicate that PVA/Chitosan/Lipase is a promising material for biotechnology applications such as triacylglycerol hydrolysis and biodiesel production. © 2012 Elsevier B.V. All rights reserved. 1. Introduction Lipases are enzymes that catalyze the hydrolysis of acylglycerides and other esters at the interface between water and insoluble sub- strates. In organic medium, lipases also catalyze the reverse reaction of synthesis of esters or transesterification [1]. The microbial lipases have attracted considerable attention due to their potential, such as high production, good stability, and many stereo-specific properties. Most of the reports on lipase-catalyzed ester hydrolysis or synthesis in- volves lipases from Candida rugosa, Rhizomucor miehei, Rhizopus oryzae, Humicola lanuginosa, Thermomyces lanuginosus and others [2,3]. Lipolase® is a commercial lipase preparation from the fungus T. lanuginosus, which is produced on industrial scale using Aspergillus oryzae as a host organism [4]. In industrial processes, the feasibility of the use of enzymes is mainly determined by the cost of the biocatalyst, easy recovery, and improvement of its stability [3,5]. Thus, the immobi- lization of enzymes in supports that allow their recovery, and that may also help to increase the operational stability is very attractive. Immobi- lization also contributes to stabilize protein structure; thus, reducing biocatalyst inactivation, either thermal or due to interactions with sol- vents [6–8]. Regarding to lipases, the chemical nature of the material used in immobilization represents an important component in the reac- tion, once lipases actuate at water/oil interface and the support must have a chemical characteristic that allows the reaction to take place. In this scenario, blends of chitosan and polyvinyl alcohol (PVA) are promising biodegradable materials due to their highly controllable chem- ical and physical properties. Chitosan is a deacetylated polysaccharide obtained from chitin, the main exoskeleton component in crustaceans [9]. Chitosan has promising applications in biotechnological fields due to its good biodegradability, nontoxicity, biocompatibility, and ability to form membranes and films, beads, fibers, gels and actuate as gas and aroma barrier [10–13]. Polyvinyl alcohol is a biodegradable, water-soluble synthetic poly- mer that presents good biocompatible properties and excellent film forming capacity. Owing characteristics such as easy preparation, chemi- cal resistance, and mechanical properties, the polyvinyl alcohol has been used in many biomaterial applications [9,11]. Considering the good biological properties of chitosan and polyvinyl alcohol, a combination of these polymers may have beneficial effects on the biological characteristics of the blended films [14]. Furthermore, the chemical cross-linking in the blend formed between PVA/Chitosan may improve mechanical strength, thermal stability, keeping the intrinsic properties of transparency and swelling ability [15]. Thus, the purpose of this work was to immobilize lipase in PVA-chitosan films for reactor coating, and test this new material as a slow enzyme delivery system for batching reactions. 2. Methodology 2.1. The preparation of PVA/Chitosan and PVA/Chitosan/Lipase film In order to select the most appropriate combination of lipase and mannitol in the film composition a central composite rotatable design (CCRD) was performed. The ultimate goal was to investigate the effect of lipase and mannitol concentration on the tensile strength and % elon- gation. For the experimental design, the two variables were confined in two levels: 0.05% (low level) to 0.2% (high level) for mannitol and 25 μL Materials Science and Engineering C 33 (2013) 1696–1701 ⁎ Corresponding author. Tel.: +55 62 3521 1492; fax: +55 62 3521 1190. E-mail address: katia@icb.ufg.br (K.F. Fernandes). 0928-4931/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.msec.2012.12.082 Contents lists available at SciVerse ScienceDirect Materials Science and Engineering C journal homepage: www.elsevier.com/locate/msec