Biomaterials 25 (2004) 1937–1945 Enzyme immobilization in novel alginate–chitosan core-shell microcapsules Ehab Taqieddin, Mansoor Amiji* Department of Pharmaceutical Sciences, School of Pharmacy, Northeastern University, 110 Mugar Life Sciences Building, Boston, MA 02115, USA Received 31 March 2003; accepted 11 August 2003 Abstract Alginate–chitosan core-shell microcapsules were prepared in order to develop a biocompatible matrix for enzyme immobilization, where the protein is retained either in a liquid or solid core and the shell allows permeability control over substrates and products. The permeability coefficients of different molecular weight compounds (vitamin B2, vitamin B12, and myoglobin) were determined through sodium tripolyphosphate (Na-TPP)-crosslinked chitosan membrane. The microcapsule core was formed by crosslinking sodium alginate with either calcium or barium ions. The crosslinked alginate core was uniformly coated with a chitosan layer and crosslinked with Na-TPP. In the case of calcium alginate, the phosphate ions of Na-TPP were able to extract the calcium ions from alginate and liquefy the core. A model enzyme, b-galactosidase, was immobilized in the alginate core and the catalytic activity was measured with o-nitrophenyl-b-d-galactopyranoside (ONPG). Change in the activity of free and immobilized enzyme was determined at three different temperatures. Na-TPP crosslinked chitosan membranes were found to be permeable to solutes of up to 17,000Da molecular weight. The enzyme loading efficiency was higher in the barium alginate core (100%) as compared to the calcium alginate core (60%). The rate of ONPG conversion to o-nitrophenol was faster in the case of calcium alginate–chitosan microcapsules as compared to barium alginate–chitosan microcapsules. Barium alginate–chitosan microcapsules, however, did improve the stability of the enzyme at 37  C relative to calcium alginate–chitosan microcapsules or free enzyme. This study illustrates a new method of enzyme immobilization for biotechnology applications using liquid or solid core and shell microcapsule technology. r 2003 Elsevier Ltd. All rights reserved. Keywords: Alginate; Chitosan; Core-shell microcapsules; Controlled permeability; Enzyme immobilization; b-galactosidase 1. Introduction Enzyme immobilization has attracted a wide range of interest from fundamental academic research to many different industrial applications [1]. The basic idea behind enzyme immobilization is to entrap the protein in a semi-permeable support material, which prevents the enzyme from leaving while allowing substrates, products, and co-factors to pass through [2]. Although the exact requirements for the immobilizing matrix are dictated by the type of enzyme and the intended application, it is clear that the material should at least be non-degradable and compatible with the enzymes. The process for immobilization should also be mild enough so as not to denature the enzyme during preparation. When an immobilized enzyme is used in vivo, the support material should also prevent immune recognition, especially if the enzyme is of non- human origin [3]. Alginate is by far the most widely used polymer for immobilization and microencapsulation technologies [4,5]. Alginate is a seaweed extract composed of chains of alternating a-l-guluronic acid and b-d-mannuronic acid residues [6]. Alginate supports are usually made by crosslinking the carboxyl group of the a-l-guluronic acid with a solution of a cationic crosslinker such as calcium chloride, barium chloride, or poly(l-lysine) [7,8]. Alginate matrices crosslinked with Ca 2+ ions, however, are unstable in the physiological environment or in common buffer solutions with high concentration of phosphate and citrate ions that can extract Ca 2+ from the alginate and liquefy the system. ARTICLE IN PRESS *Corresponding author. Tel.: +1-617-373-3137; fax: +1-617-373- 8886. E-mail address: m.amiji@neu.edu (M. Amiji). 0142-9612/$-see front matter r 2003 Elsevier Ltd. All rights reserved. doi:10.1016/j.biomaterials.2003.08.034