Electromigration Behavior of a Mixture of Chitosan Oligomers at Different Concentrations MOHAMMED AIDER, † JOSEPH ARUL, † MIRCEA-ALEXANDRU MATEESCU, ‡ SERGE BRUNET, § AND LAURENT BAZINET* ,† Institute of Nutraceuticals and Functional Foods (I NAF) and Department of Food Science and Nutrition, Laval University, Que ´bec (Que ´bec), Canada G1K 7P4, Department of Chemistry and Biochemistry, Universite ´ du Que ´bec a ` Montre ´al, CP 8888, Succursale A, Montre ´al (Que ´bec), Canada H3C 3P8, and ISM Biopolymer Inc., 220, Denison E. Granby (Que ´bec), Canada J2H 2R6 In this study, the effect of the concentration of a chitosan oligomer mixture on its electrophoretic behavior was studied as a function of pH and ionic strength added. It was shown that the concentration has a significant effect on the average electrophoretic mobility of the chitosan oligomer mixture and on isoelectric point. At a concentration of 3%, the ionic strength added did not show any effect on the electromigration behavior of the chitosan oligomer mixture. By decreasing the concentration of the chitosan oligomer mixture, ionic strength showed a significant effect on the average electrophoretic mobility but not on the isoelectric point. The highest shift of the isoelectric point was recorded in water at 0.003% concentration of the oligomer mixture. Under these conditions, the isoelectric point was at pH 5 whereas it was at pH 8 at 3% concentration of chitosan oligomer mixture. Electrophoretic measurements were also taken in water/ethanol aqueous medium. By adding ethanol to the medium, the average electrophoretic mobility decreased. This would have been caused by the increase in viscosity of the medium. Increasing ethanol ratio in the running medium, the isoelectric point moved from pH 5 in water up to pH 6-8 dependently on chitosan oligomer mixture concentration and ethanol content of the medium. KEYWORDS: Chitosan oligomer mixture; concentration; isoelectric point; ethanol; dielectric constant; electrophoretic mobility INTRODUCTION Chitosan is the N-deacetylated product of chitin found in the shells of crabs and shrimps and as cell wall components of most fungi, yeasts, and molds. Chitosan has applications in several fields such as biomedical, personal care products, biotechnology, pharmaceutical, nutraceutical, and food. Chitosan oligomers with a degree of polymerization of about 6 are potentially useful as medicinal agents and as food ingredients because of their biological and therapeutic values (1-4). To produce these bioactive oligosaccharides, biocatalytical (enzymatic) and chemi- cal procedures are used (5-7). The yield of the biocatalytical reaction is not high (8). By combining physical methods (shear- force treatment) and acid hydrolysis, the molecular weight of the chitosan oligomer can be decreased (2, 9, 10). Although execution of these physical methods is not difficult, fast degradation rates and random reactions result in product variability (8). In acid hydrolysis, 10% acetic acid is generally used as a solvent, with 5% NaNO 3 added for the deacetylation reaction. This method can decompose chitosan into units of one to six N-acetylglucosamines, and such products are soluble at pH 7 (11). All these methods can serve to produce a mixture of molecules with various molecular weights. To separate these chitosan oligomer mixtures to obtain pure or enriched oligomer, it is essential to find a suitable technique. The exploitation of the electric properties of these molecules could offer a solution (12). The chitosan oligomer has one or more amine functional groups depending on the degree of polymerization (13). Under specific pH conditions, these groups are charged because of the protonation of the amine group (NH 3 + ). Therefore, they will migrate under the effect of an external electric field (13). The migration speed of each molecule should be different from others because of differences of their molecular weight, electric charge, and concentration. That would make possible the separation of various chitosan oligomers by exploiting the electrophoretic properties of each molecule. To carry out this objective, it is important to study and understand the electrophoretic behaviors of the chitosan oligomers under various conditions of the medium and different concentrations of the mixture. In a previous study (14), it was shown that electromobility of chitosan D-glucosamine (mono- mer) and individual chitosan oligomer (dimer, trimer, tetramer, * Author to whom correspondence should be addressed. Phone: 418- 656-2131 ext. 7445; fax: 418-656-3353; e-mail: laurent.bazinet@aln.ulaval.ca. † Laval University. ‡ Universite ´ du Que ´bec a ` Montre ´al. § ISM Biopolymer Inc. 10170 J. Agric. Food Chem. 2006, 54, 10170-10176 10.1021/jf061653n CCC: $33.50 © 2006 American Chemical Society Published on Web 12/01/2006