Comparative investigation of the binding characteristics of poly-l-lysine and chitosan on alginate hydrogel

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Abstract

The binding properties of poly-l-lysine and chitosan to alginate have been evaluated quantitatively and compared. Poly-l-lysine bound to alginate hydrogel more rapidly than chitosan as poly-l-lysine has a smaller molar hydrodynamic volume. In addition, poly-l-lysine showed a much higher binding capacity (6.14:1) for alginate hydrogel beads than chitosan (2.71:1), and a little higher binding stoichiometry (0.58) to sodium alginate molecules in solution than chitosan (0.49). An exothermic heat of alginate-poly-l-lysine complexes formation of 2.02 kJ/mol was detected. For alginate–chitosan complexes, the binding enthalpy has been seen to be −3.49 kJ/mol. The stability of the polyelectrolyte complexes was related to their binding enthalpy. The alginate-poly-l-lysine complexes could be disintegrated and rebuilt. By contrast, chitosan was bound with alginate in a steady state. These results provide fundamental insights regarding the structure and property relationships of macromolecules, and will be helpful in designing and selecting appropriate polymers.

Introduction

Microcapsules with a hydrogel core and a polyanion–polycation membrane have been proved to possess various applications in biomedical and pharmaceutical areas [1], [2], [3]. Although many polymers have been attempted, alginate-poly-l-lysine and alginate–chitosan microcapsules are still the commonly used microcapsule systems [4], [5]. Alginate is composed of linear chains of α-l-guluronic acid and β-d-mannuronic acid residues and bound through 1, 4-glycosidic linkages [6], [7]. Poly-l-lysine is a peptide having excellent structural precision in terms of molecular weight and secondary structure elements [8]. Chitosan is a linear copolymer polysaccharide, which is made up of various amounts of β (1-4)-linked 2-amino-2-deoxy-d-glucose (d-glucosamine) and 2-acetamido-2-deoxy-d-glucose (N-acetyl-d-glucosamine) units [3], [9]. The carboxylic acid groups on alginate make it can crosslink with multiple divalent cations and thus form hydrogels [10]. Amino groups on polycations can interact with alginate hydrogel to form microcapsule membrane on the surface of the hydrogel.

The microcapsule membrane has been demonstrated to be of great importance not only for transport properties (semipermeable membrane) but as well for stability. Previous studies indicated that microcapsules made of poly-l-lysine and chitosan showed different swelling behavior and mechanical properties [11], [12]. The stability of alginate-poly-l-lysine microcapsules in physiological conditions was reduced due to the swelling behavior. Alginate-poly-l-lysine microcapsules were ruptured in shaking flask. This limited stability of microcapsules hampered their application [13]. As the polyanion–polycation membrane is formed by the reaction between carboxylate moieties on alginate and protonated amines on chitosan or poly-l-lysine, it is believed that the physicochemical properties of polyanion–polycation membrane could be tailored by controlling the degree of association between the functional groups [14], [15], [16]. Also, as the membrane is formed by binding of polycations on calcium alginate hydrogel beads, control of the molecules at this level requires a comprehensive understanding of the binding properties of this system.

The differences in molecular structure will affect their binding with alginate. It was reported that there were different interaction mechanisms between chitosan–alginate and poly-l-lysine–alginate during the assembly process [17]. Poly-l-lysine had the ability to diffuse “in” and “out” of the alginate-based polyelectrolyte films while chitosan was typically confined to the films [18], [19], which suggested that the binding affinity of alginate was much stronger with chitosan than poly-l-lysine. However, this appeared to contradict the known facts that poly-l-lysine has a larger charge density than chitosan, and a resulting stronger electrostatic interaction with alginate [4], [16]. Studying the thermodynamics of interactions between oppositely charged polymers is helpful in designing novel structures of fundamental and applicative interests [20]. Though De and Robinson tried to investigate the interactions of alginate-poly-l-lysine and alginate–chitosan, the complexity of the polymeric interaction made it difficult to determine and compare the key thermodynamic parameters, such as enthalpy and stoichiometry [16]. In addition, the structure of the polycation will not only affect its interaction with alginate, but also affect the stability of polyelectrolyte complexes. The relationship between the binding enthalpy and microcapsule stability has not been established yet.

In this work, the binding properties of poly-l-lysine and chitosan on calcium alginate hydrogel beads have been evaluated quantitatively and compared, including their binding kinetics, binding capacity, binding enthalpy, and stoichiometry. The molecular state and conformation of these polycations were investigated to understand the origin of the differences in binding capacity. In addition, the properties of alginate-poly-l-lysine and alginate–chitosan complexes, including stability and mechanical strength, were measured and compared. The correlation between the stability of polyelectrolyte complexes and their thermodynamic properties was established.

Section snippets

Materials components

Sodium alginate (Qingdao Crystal Salt Bioscience and Technology Corporation, Qingdao, China) with a viscosity of 100 cP at a concentration of 1% (w/v) aqueous solution at 25 °C was purchased. The compositions of the alginate molecules were characterized by 1H NMR with G/M ratio of 34/66, with the molecular weight (Mw) of 430 kDa. Poly-l-lysine hydrobromide samples with Mw of 4000–15,000, 15,000–30,000, 30,000–70,000 by viscosity were from Sigma Aldrich Chemical Co (USA). The poly-l-lysine with the

Formation kinetics of alginate-polycation microcapsules

The binding kinetics of poly-l-lysine and chitosan with alginate hydrogel beads was measured. Polycation molecules diffused into hydrogels while alginate migrated outwards to form microcapsules, Fig. 1(a). In this work, poly-l-lysine and chitosan molecules with the same Mw of 10 kDa were both bound with alginate beads quickly. If not specifically notes, we used poly-l-lysine and chitosan with the same Mw of 10 kDa in the following sections. In Fig. 1(a), poly-l-lysine was able to bind with

Conclusions

Insights into the mechanisms of interaction between alginate and the two polycations, poly-l-lysine and chitosan, were provided in this work. The comparison study was performed on several fundamental aspects to gain a better understanding of the interactions, including thermodynamic and kinetic properties, binding capacities and conformational changes of polycations. Chitosan had a stronger binding enthalpy than poly-l-lysine during the interaction with alginate. Meanwhile, the stoichiometry

Acknowledgements

This study was supported by “Strategic Priority Research Program” of the Chinese Academy of Sciences, Stem Cell and Regenerative Medicine Research (No. XDA01030303), National Natural Science Foundation of China (No. 51103157) and Ocean Public Welfare Scientific Research Project of China (No. 201305016-4, No. 201405015-3). This work was also supported by Open Funding Project of the State Key Laboratory of Bioreactor Engineering.

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