**1. Introduction**

Chitosan is a semi-synthetic linear copolymer composed of a variable number of *β*-(1-4) linked units of 2-acetamide-2-deoxy-*β*-d-glucopyranose (GlcNAc) and 2-amino-2-deoxy-*β*-dglycopyranose (GlcN) [1]. The two monomers differ with respect to the C2-substituent in the sugar ring, which is either an amino or acetamide group (Figure 1). Chitosan is obtained via the alkaline deacetylation of chitin. However, the deacylation reaction hardly proceeds completely in a normal heterogeneous reaction, leading to a random distribution of GlcNAc and GlcN residues in the chitosan polymer [2, 3]. The degree of acetylation of a polymer is a measure of the average number of GlcNAc per 100 chitosan monomers in percentile unit. The degree of acetylation governs important physical-chemical properties of the chitosan polymer such as solubility and conformation, being critical for the effectiveness of various technological applications [4-6]. The threshold for the conversion of chitin into chitosan depends on the solubility of the oligosaccharide in a slightly acid solution (0,1 mol/L of acetic acid). Conventionally, chitin polymers with a level of acetylation below 50% is considered as chitosan. The level of protonated amino groups in the glucosamine monomers dictates the solubility of chitosan, conferring the cationic nature to the polymer [7].

Chitin is the most abundant amino polysaccharide, being produced in the amount of one hundred billion tons per year in nature [8]. Its main source is the exoskeleton of crabs and shrimps, whose availability in nature makes chitin a renewable source of chitosan. In the last decades chitosan has emerged as a biomaterial with unique properties for advanced applications in green chemistry, biomedical, pharmaceutical, food [9] and agriculture [10].

© 2012 Cunha et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2012 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

The variety of applications of chitosan is determined by its chemical structure, which varies with respect to size (average molecular weight; MW), degree of acetylation (DA) and numerous chemical modifications [6, 11-15]. An increasing number of chitosan chemical modifications have been described in the literature [14, 15]. Chitosan is also a highly absorptive material used as heavy-metal chelators in water [16, 17]. The chelation of metals occurs via electrostatic interactions with chitosan reactive groups (hydroxyl, acetamide and primary amino groups). These functional groups are also responsible for properties such as high hydrophilicity, reactivity, and structural flexibility that make chitosan soluble in nearneutral acid solution [17]. The soluble adsorptive properties of chitosan enable its use for removal of pesticide and dyes from water, for adsorption of proteins, as flocculant agent, and even as a catalyst support for biodiesel production [18-24]. Besides being an abundant renewable resource, chitosan stands out due to some unique properties such as exceptional biocompatibility, biodegradability, non-toxicity and the easiness of production of chemically modified forms [14, 25, 26]. Furthermore, chitosan exhibits antimicrobial and antifungal activity [27-30]. These singular properties makes chitosan welll-suited for a wide range of biomedical applications such as drug delivery, platform for neural stem cell growth, tissue engineering (bone, cartilage, nerve, skin), immunoprophylaxis, gene therapy, wound healing and treatment of infections [31-44].

**Figure 1.** Schematic representation of a disaccharide formed by 2-amino-2-deoxy-*β*-d-glycopyranose (GlcN) bound to 2-acetamide-2-deoxy-*β*-d-glucopyranose (GlcNac). and dihedral angles are shown in the scheme in red and here are represented by atoms: O5-C1-O1-C4' and C1-O1-C4'-C5', respectively. GlcN and GlcNac are the two units forming chitosan. Solubility is typically achieved when GlcN > 50%.

This short review will cover the structural dynamics of chitosan from a microscopic perspective, focusing on the interplay between its conformational variability and macroscopic properties such as solubility and aggregation. It is not the goal of the review to provide a detailed summary of the extensive literature on carbohydrate structure and its characterization. Excellent reviews on the subject can be found in the literature [45-47]. The text is organized in four main sections. First, we present an overview of chemical interactions between chitosan and biological materials, in particular lipid bilayers. In what follows, we describe the advantages and limitations of experimental and computational techniques used for the structural characterization of oligosaccharides, emphasizing the necessity of combining different approaches in order to obtain high-resolution structural data on chitosan. In this section we also introduce the theoretical principles underlying molecular dynamics (MD) simulations, which has been widely used to study the structural dynamics of carbohydrates in solution. The third section, we review the types of secondary structure observed for chitin and chitosan in the crystalline state. In the final section, we offer a detailed account of the structure and conformational dynamics of chitosan in solution as unveiled by computational simulations carried out in our group.

## **2. The molecular interactions underlying chitosan bioactivity**

230 The Complex World of Polysaccharides

healing and treatment of infections [31-44].

The variety of applications of chitosan is determined by its chemical structure, which varies with respect to size (average molecular weight; MW), degree of acetylation (DA) and numerous chemical modifications [6, 11-15]. An increasing number of chitosan chemical modifications have been described in the literature [14, 15]. Chitosan is also a highly absorptive material used as heavy-metal chelators in water [16, 17]. The chelation of metals occurs via electrostatic interactions with chitosan reactive groups (hydroxyl, acetamide and primary amino groups). These functional groups are also responsible for properties such as high hydrophilicity, reactivity, and structural flexibility that make chitosan soluble in nearneutral acid solution [17]. The soluble adsorptive properties of chitosan enable its use for removal of pesticide and dyes from water, for adsorption of proteins, as flocculant agent, and even as a catalyst support for biodiesel production [18-24]. Besides being an abundant renewable resource, chitosan stands out due to some unique properties such as exceptional biocompatibility, biodegradability, non-toxicity and the easiness of production of chemically modified forms [14, 25, 26]. Furthermore, chitosan exhibits antimicrobial and antifungal activity [27-30]. These singular properties makes chitosan welll-suited for a wide range of biomedical applications such as drug delivery, platform for neural stem cell growth, tissue engineering (bone, cartilage, nerve, skin), immunoprophylaxis, gene therapy, wound

**Figure 1.** Schematic representation of a disaccharide formed by 2-amino-2-deoxy-*β*-d-glycopyranose (GlcN) bound to 2-acetamide-2-deoxy-*β*-d-glucopyranose (GlcNac). and dihedral angles are shown in the scheme in red and here are represented by atoms: O5-C1-O1-C4' and C1-O1-C4'-C5', respectively. GlcN and GlcNac are the two units forming chitosan. Solubility is typically achieved when GlcN > 50%.

This short review will cover the structural dynamics of chitosan from a microscopic perspective, focusing on the interplay between its conformational variability and macroscopic properties such as solubility and aggregation. It is not the goal of the review to provide a detailed summary of the extensive literature on carbohydrate structure and its characterization. Excellent reviews on the subject can be found in the literature [45-47]. The text is organized in four main sections. First, we present an overview of chemical interactions between chitosan and biological materials, in particular lipid bilayers. In what follows, we describe the advantages and limitations of experimental and computational techniques used for the structural characterization of oligosaccharides, emphasizing the necessity of combining different approaches in order to obtain high-resolution structural data on chitosan. In this section we also introduce the theoretical principles underlying molecular dynamics (MD) simulations, which has been widely used to study the structural Chitosan is a very promising material with wide range of biomedical applications. This oligosaccharide incorporates highly sought properties for biomedical applications (biocompatibility, biodegradability and bioresorbability) with the easy processing into gels, membranes, nanofibers, beads, microparticles, nanoparticles, scaffolds and sponges forms [48-52]. Yet, chitosan has a flexible, hydrophilic helical structure with reactive amine groups, which offers a multitude of possible inter- and intra-molecular interactions. A detailed understanding of the effects of different materials and environmental conditions on such interactions can enable the design of novel chitosan-based technologies.

Chitosan amino groups are the major players in metal chelating processes. However, it has been previously shown that electrostatic interactions involving the protonated amino groups *per se* in low pH are not sufficient to explain the biological behavior of chitosan in presence of biological membranes [53]. A comparative study between chitosan and a fully cationic polymer has shown that hydrophobic interactions play an important role in the polysaccharide action. A study on the effect of the pH and the molecular weight of chitosan in multilamellar vesicles of dipalmitoylphosphatidylcholine (DPPC) has shown that increasing the biopolymer molecular weight (213 kDa) and decreasing of pH can lead to disruption of the membrane [54]. In contrast, another study has shown that the interactions between chitosan and DPPC lipids in liposomes led to an increase in thermodynamic stability of the composite. It has been proposed that this stabilization results from a shielding mechanism based on the electrostatic interactions between the chitosan chains and phospholipids polar heads [55]. In contrast, anti-fungal and bactericidal activity of chitosan has been attributed to the ability of chitosan to disrupt the inner and outer membranes of cells [56, 57]. The contribution of electrostatic, hydrophobic and hydrogen bond interactions between chitosan and three different lipids have been evaluated using Langmuir films to mimic the interaction of chitosan and bacterial membranes [29]. It has been shown that chitosan had a negligible effect on DPPC monolayers but it distinctly affected dipalmitoylphosphatidylglycerol (DPPG) and cholesterol monolayers [29]. The effect on DPPG was found to decrease with increasing pH, ascribed to the charge-mediating action of chitosan, whereas the pH did not affect the cholesterol monolayers where interactions occurs mainly via hydrogen bonding. A recent study has suggested that sensitivity of fungi to chitosan depends of the membrane fluidity and dynamics [58]. The same group has also suggested in a previous work that chitosan kills fungal cells by an unknown mechanism that does not involve endocytosis [59]. Although chitosan has been used in a variety of biologically relevant applications involving interactions with lipids, proteins, inorganic and organic compounds, a microscopic picture of these interactions remains lacking. The conformational flexibility of chitosan has hampered the acquisition of high-resolution

structural data through of X-ray crystallography and NMR spectroscopy [46, 47]. However, current molecular modeling techniques can be used to bridge the gap of experimental resolution, thus providing complementary information to measurements.
