An Overview of the Biopharmaceutical Applications of Chitosan and Alginate

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BioPharm International, BioPharm International-11-01-2003, Volume 16, Issue 11

Chitosan and alginate hydrogels are high-molecular-weight carbohydrate polymers used in an extensive array of pharmaceutical applications. While many applications depend upon the ionized state of the polymer, other factors play an important role in the pharmacological activity of complexes that include these agents.

Chitosan and alginate hydrogels are high-molecular-weight carbohydrate polymers used in an extensive array of pharmaceutical applications. While many applications depend upon the ionized state of the polymer, other factors play an important role in the pharmacological activity of complexes that include these agents. This review focuses upon ionic versus nonionic interactions, and illustrates where modifications of these ionic sites dramatically alter their pharmacological activity and pharmaceutical applications.

Both of these hydrogels are natural polysaccharides, and when hydrated, they dramatically increase the viscosity of aqueous solutions. (A hydrogel is a colloid in which the dispersion medium is water.) In addition to their swelling and dispersal properties, the ionic characteristics of these polymers are important. Additionally, the sequence of the monosaccharide units in these polymers affects their physical and pharmacological properties. Applications include encapsulating enzymes, formulating macromolecular complexes, and gene transfection.

Figure 1. A partial structure of chitosan.

General Characteristics

Chitosan is a polysaccharide hydrogel derived from acid extraction of chitin. The most common source of chitin is the shells of crabs and other crustaceans.

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Chitosan can be isolated in complexes with a molecular weight exceeding 10

6

daltons.

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Upon acid extraction of chitin, the N-acetylglucosamine subunits undergo partial deacetylation to yield a chitosan polymer composed of poly D-glucosamine. A partial structure of chitosan is shown in Figure 1. The presence of aminosugars makes chitosan soluble in dilute solutions of organic acids. Chitosan has many applications based upon its ability to form complexes with anions and composite films with other polysaccharides.

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Hydration and dissolution readily occur at a pH below 5.5.

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Alginate or alginic acid is a polysaccharide hydrogel derived from brown seaweed.5 Alginate polymers can be isolated with a molecular weight exceeding 100,000 daltons.6 The repeating units are composed of mannuronic and guluronic acid monosaccharide subunits as shown in Figure 2. These monosaccharides are arranged to facilitate the complexation of divalent cations between the carboxyl groups on adjacent alginate chains.7 A rigid chelate structure forms when an alginate mix is added to a divalent cation solution. This property is used to encapsulate enzymes, macromolecular complexes, and cells.8,9 Alginate also forms films that can be readily hydrated.

Ionic and Nonionic Interactions

In their ionized forms, chitosan is positively charged and alginate is negatively charged. Drug release and gene transfection appear to depend primarily upon the charge of the polymer.

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Furthermore, the carbohydrate backbone confers some selective characteristics to the polymers. For instance, the degree of deacetylation affects chitosan's binding characteristics.13 The sequence of mannuronic and guluronic acid residues play an important role in the application and activity of alginate polymers.14 These residues determine the orientation of the carboxyl group for complexation with cations.

Figure 2. Monosaccharide subunits of alginate.

While studies have highlighted the importance of charge density and orientation, the presence of a polysaccharide chain is also very important. Chitosan has been reported to be an effective gene transfection agent,12 and has been reported to be selective for HeLa cells and related cell lines.11 Binding of chitosan to DNA results in condensation and interaction with sulfate groups in the extracellular matrix, internalization of the gene, and transcription.15 This mechanism does not account for the selectivity of chitosan-mediated transfection in certain cell types, since sulfate groups appear to be ubiquitous in the extracellular matrix of most cell types. The ionic binding of sulfate with the amino group of chitosan is the basis of this interaction.

Studies at other labs indicate that cellular binding to chitosan could be disrupted with methyl α-D-mannopyranoside, a non-ionized, non-reducing monosaccharide.16 These findings suggest that cell surface interactions with chitosan are due in part to the interaction of the carbohydrate chains of chitosan (the backbone) and the extracellular matrix. This may explain, in part, the selectivity of transfection in certain cell types independent of the cell's ability to transcribe the DNA introduced in the transfer process.

Studies of alginate-mediated transfection reveal the importance of the carbohydrate backbone. Unmodified alginate is a poor transfection agent in most systems, presumably due to the lack of positively charged groups necessary to complex with DNA. Modification of the carboxyl groups of alginate with aromatic and heterocyclic amines containing two or more amino groups did not consistently yield an effective transfection agent.17 Among these derivatives, the alginate and basic fuchsin conjugate was the only effective transfection agent. Modification of alginate with basic fuchsin results in the incorporation of two widely-separated amino groups into the carbohydrate backbone, facilitating transfection. The results from these studies strongly suggest that the carbohydrate backbone of these hydrogels must affect the orientation of the amino groups and their interaction with certain cell types.

Future Directions

Chitosan and alginate are both excellent film-formers with opposite ionic charges. These characteristics provide applications for composites of these polymers in the same system.

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Their excellent film forming abilities are currently under investigation in the design of a bilayer complex for selective blotting and removal of nucleic acids and other macromolecules. In this bilayer system, an external chitosan layer can be introduced onto cellulose fibers or other substrate. DNA will readily bind to the chitosan film. A calcium alginate layer can be formed over the chitosan, resulting in a bilayer film (Figure 3). The calcium alginate layer - as well as any inclusion complexes in the alginate - can be readily removed with any dilute citrate without significantly affecting the position of the DNA-chitosan blot. Potential applications include molecular watermarking in forensics as well as forming a substrate and a matrix for gene transfection. Other derivatives of these hydrogels are currently under development. These natural polymers and their broad applications will be a fertile area for innovation.

Figure 3. Alginate and chitosan bilayer film.

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