Biomaterials 26 (2005) 819–826 Synthesis and in vitro evaluation of a novel thiolated chitosan Krum Kafedjiiski a , Alexander H. Krauland b , Martin H. Hoffer c , Andreas Bernkop-Schn . urch a, * a Department of Pharmaceutical Technology, Institute of Pharmacy, Leopold-Franzens-University Innsbruck, Innrain 52, Josef M . oller Haus, A-6020 Innsbruck, Austria b Center of Pharmacy, Institute of Pharmaceutical Technology and Biopharmaceutics, University of Vienna, Althanstr. 14, A-1090 Vienna, Austria c MucoBiomer GmbH, Industriezeile 6, A-2100 Leobendorf, Austria Received 4 November 2003; accepted 13 March 2004 Abstract In order to achieve the same properties as chitosan–4-thio-butyl-amidine and to overcome at the same time its insufficient stability, the aim of this study was to evaluate the imidoester reaction of isopropyl-S-acetylthioacetimidate for the chemical modification of chitosan and to study the properties of the resulting chitosan–thioethylamidine (TEA) derivative. The thioalkylamidine substitute was introduced without the formation of N-substituted non-thiol products. The resulting conjugates exhibited 1.0570.17% or 139.68717.13 mmol immobilized free thiol groups per gram polymer and a total amount of reduced and oxidized thiol groups of 1.8170.65% or 179.46767.95 mmol/g polymer. By the immobilization of thiol groups mucoadhesion was strongly improved due to the formation of disulfide bonds with mucus glycoproteins. Chitosan–TEA was investigated regarding to its mucoadhesive properties via tensile studies and the rotating cylinder method. In tensile studies the total work of adhesion of chitosan–TEA was increased 3.3-fold in comparison to unmodified chitosan. Results from the rotating cylinder method showed an improvement ratio of 8.9 for chitosan–TEA compared with unmodified chitosan. In spite of the immobilization of thiol groups onto chitosan its swelling behavior in aqueous solutions was not significantly altered. Cumulative release studies out of matrix tablets comprising the chitosan–TEA and the model compound fluorescence labeled dextrane (FD 4 ) demonstrated a controlled release over 3h with a trend toward a pseudo-zero-order kinetic. Because of these features the new chitosan thioamidine conjugate might represent a promising new polymeric excipient for various drug delivery systems. r 2004 Elsevier Ltd. All rights reserved. Keywords: Thiolated chitosan; Chitosan–thioethylamidine conjugates; Isopropyl-S-acetylthioacetimidate.HCl; Mucoadhesion 1. Introduction Chitosan is a natural, cationic aminopolysaccharide copolymer of glucosamine and N-acetylglucosamine. It is obtained by the alkaline, partial deacetylation of chitin, which originates from shells of crustaceans such as crabs and prawns [1]. Chitosan is a biodegradable, biocompatible, less toxic and mucoadhesive biopolymer [2]. Its use as a pharmaceutical excipient is meanwhile well-established [3]. It has been reported to enhance drug permeation across the intestinal, nasal and buccal mucosa [4–6]. In order to further enhance the mucoadhesive proper- ties of chitosan, different chitosan derivatives have been developed based on various theories explaining the mechanism of mucoadhesion [7]. By applying the known concept of the immobilization of thiol groups to the primary amino groups of chitosan, the above-mentioned properties of the polymer were strongly improved [8– 10]. In such a way chitosan–cysteine conjugates, chitosan–thioglycolic acid conjugates and chitosan– 4thio-butyl-amidine conjugates (chitosan–TBA) have been obtained [11–13]. Among them chitosan–TBA turned out to exhibit the most promising features. The modifying reagent for chitosan–TBA conjugates is 2- iminothiolane (a cyclic thioimidate), which reacts with amino groups and introduces a sulfhydryl residue via a positively charged amidine substructure. Thus, when 2-iminothiolane is applied for the modification of ARTICLE IN PRESS *Corresponding author. Tel.: +43-512-507-53-83; fax: +43-512- 507-29-33. Email-address: andreas.bernkop@uibk.ac.at (A. Bernkop-Schn . urch). 0142-9612/$-see front matter r 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.biomaterials.2004.03.011