Improved Pharmacokinetics and Stability Properties of Catalase by Chemical Glycosidation with End-Group Activated Dextran Reynaldo Villalonga, 1 Aymara Valdivia, 1 Yunel Pe ´rez, 1 Bertha Chongo 2 1 Center for Enzyme Technology, University of Matanzas, Matanzas, CP 44740, Cuba 2 Institute of Animal Sciences, San Jose ´ de las Lajas, La Habana 32 700, Cuba Received 22 May 2006; accepted 13 June 2006 DOI 10.1002/app.25019 Published online in Wiley InterScience (www.interscience.wiley.com). ABSTRACT: Catalase was chemically modified with a monoactivated dextran derivative having a carboxylate group at its reducing end residue. The modified enzyme retained 77% of its initial specific activity and was 3-fold more resistant to tryptic degradation. The plasma half-life time was increased to 7.3-fold after glycosidation. Ó 2006 Wiley Periodicals, Inc. J Appl Polym Sci 102: 4573–4576, 2006 Key words: catalase; dextran; drug delivery systems; enzymes; polysaccharides INTRODUCTION Several proteins are currently used as therapeutic drugs for numerous diseases, but their major appli- cation is limited by the rapid clearance of these bio- molecules after in vivo administration. This clearance is mediated by different mechanisms such as the rec- ognition by the immune system, proteolytic degrada- tion, and glomerular filtration through the kidney. 1 That is why the development of new delivery sys- tems for these drugs is required. Modification of proteins with biocompatible poly- mers constitutes a successful approach for improving their pharmacokinetics and stability properties. 2 In special, poly(ethylene glycol) (PEG) has been widely used for site-directed modification of protein drugs, and there are many PEG-based protein conjugates in the market with increased circulatory half-life. 3 However, PEG is a non biodegradable macromole- cule which is slowly cleared from the human body, and the occurrence of anaphylactic reactions and a severe bronchospasm have been reported for PEG- modified therapeutic enzymes. 4 Chemical crosslinking of enzymes with polyacti- vated macromolecules have been extensively used as a method for stabilizing such kind of catalysts. 5–8 Through this approach the catalytic and functional activity of several enzymes have been improved. 5–8 However, the adducts prepared showed undefined chemical structure and in some case lower catalytic behavior. 9 For this reason, the development of alter- native modification methods for preparing pharma- cologically active polymer–protein conjugates with more defined composition receives considerable at- tention in pharmacological sciences. In the present article, we describe the synthesis of an end-group monocarboxylated dextran derivative and its attachment to the surface of catalase (EC 1.11.1.6). Dextran is a biocompatible and biodegrad- able polymer widely employed in biomedical fields. 10 Among these applications, crosslinking of pharmaco- logical active proteins is included. 11 As target enzyme we used catalase, a tetrameric antioxidant enzyme with potential application in the therapy of several diseases mediated by reactive oxygen species. 12 MATERIALS AND METHODS Materials Bovine liver catalase (14.6 U/g) was from Roche Mo- lecular Biochemistry (Mannheim, Germany). Dextran 5000 was from Serva Electrophoresis (Heidelberg, Germany). Bovine pancreatic trypsin and Fractogel EMD BioSEC (S) were from Merck (Darmstadt, Germany). All other chemicals were analytical grade. Synthesis of end-group carboxylated dextran Dextran (2 g), dissolved in 10 mL distilled water was treated with 1 g e-aminocaproic acid and stirred for Correspondence to: R. Villalonga (reynaldo.villalonga@ umcc.cu). Contract grant sponsor: International Foundation for Science, Stockholm, Sweden. Contract grant sponsor: Organisation for the Prohibition of Chemical Weapons, The Hague, The Netherlands; contract grant number: F/3004-2. Journal of Applied Polymer Science, Vol. 102, 4573–4576 (2006) V V C 2006 Wiley Periodicals, Inc.