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The present paper explores the effect of boric acid on Chitlac, a lactose-modified chitosan which had previously shown interesting biological and physical-chemical features. The herewith-reported experimental evidences demonstrated that boric acid binds to Chitlac, producing conformational and association effects on the chitosan derivative. The thermodynamics of boric acid binding to Chitlac was explored by means of 11B NMR, circular dichroism (CD), and UV–vis spectroscopy, while macromolecular effects were investigated by means of viscometry and dynamic light scattering (DLS). The experimental results revealed a chain–chain association when limited amounts of boric acid were added to Chitlac. However, upon exceeding a critical boric acid limit dependent on the polysaccharide concentration, the soluble aggregates disentangle. The rheological behavior of Chitlac upon treatment with boric acid was explored showing a dilatant behavior in conditions of steady flow. An uncommonly high dependence in the scaling law between the zero-shear viscosity and the concentration of Chitlac was found, i.e., η0 ∝ CCTL5.8, pointing to interesting potential implications of the present system in biomaterials development.

Polysaccharide networks, in the form of hydrogels and dried membranes based on chitosan and on the cross-linker tripolyphosphate (TPP), were developed using a novel approach. TPP was incorporated into chitosan by slow diffusion to favor a controlled gelation. By varying chitosan, TPP, and NaCl concentration, transition from inhomogeneous to homogeneous systems was achieved. Rheology and uniaxial compression tests enabled to identify the best performing hydrogel composition with respect to mechanical properties. FTIR, (31)P NMR, and spectrophotometric methods were used to investigate the interaction chitosan-TPP, the kinetics of phosphates diffusion during the dialysis and the amount of TPP in the hydrogel. A freeze-drying procedure enabled the preparation of soft pliable membranes. The lactate dehydrogenase assay demonstrated the biocompatibility of the membranes toward fibroblasts. Overall, we devised a novel approach to prepare homogeneous macroscopic chitosan/TPP-based biomaterials with tunable mechanical properties and good biocompatibility that show good potential as novel polysaccharide derivatives.

The present manuscript deals with the setup and evaluation of two analytical methods, namely Circular Dichroism (CD) and 23Na longitudinal relaxation, to determine the composition of binary mixtures of the sodium salts of alginate and hyaluronan in solution. The quantitative results obtained were confirmed by 1H NMR, and the developed methods were applied to the determination of the composition and polymer release of mixed hydrogels. Although focused on the specific case of binary mixtures of alginate and hyaluronan, the approach drawn in the present manuscript can find applications in different binary systems composed of natural or synthetic non-interacting polymers.

The initial binding of calcium ions by alginate chains was investigated in dilute solution. The use of viscometry, light scattering, circular dichroism, and fluorescence quenching performed on both Ca(2+) and Mg(2+) alginate systems allowed collecting new experimental data in addition to those already reported in the literature. This led us to propose an ion multicomplex binding modality and to disprove the Ca(2+)-alginate monocomplex formation. The first mode of bonding was proposed to be formed by four G residues from facing chains in a conformation which, albeit ordered, is different from the well-known "egg box" one. This was indicated as a locally tilted conformation (tilted egg-box). The addition of further bonding ions caused the well-known cooperative egg-box conformation (strong bonding) with notable variation in the physical-chemical properties of the dilute solution.

Alginate/hydroxyapatite composite scaffolds were developed using a novel production design. Hydroxyapatite (HAp) was incorporated into an alginate solution and internal gelling was induced by addition of slowly acid hydrolyzing d-gluconic acid delta-lactone (GDL) for the direct release of calcium ions from HAp. Hydrogels were then freeze-casted to produce a three-dimensional isotropic porous network. Scanning electron microscopy (SEM) observations, confocal laser scanning microscopy (CLSM) and microcomputed tomography (micro-CT) analysis of the scaffolds showed an optimal interconnected porous structure with pore sizes ranging between 100 and 300 microm and over 88% porosity. Proliferation assay and SEM observations demonstrated that human osteosarcoma cell lines were able to proliferate, maintain osteoblast-like phenotype and massively colonize the scaffold structure. Overall, these combined results indicate that the novel alginate based composites efficiently support the adhesion and proliferation of cells showing at the same time adequate structural and physical-chemical properties for being used as scaffolds in bone tissue engineering strategies.

The theoretical model devised in the previous paper (Donati, I.; Benegas, J. C.; Cesàro, A.; Paoletti, S. Biomacromolecules 2006, 7 (5), 1587-1596) for the description of ion-induced chain aggregation is here applied to the case of chain dimerization of poly(galacturonate) in the presence of calcium ions. Particular attention has been directed toward the initial stage of dimer formation [i.e., in the low regime of calcium-to-polymer ratio (Rj)]. Circular dichroism (CD) data allowed evaluation of the fraction, theta, of calcium ions bound within chain dimers according to the "egg-box"-model. The theoretical model was able to reproduce satisfactorily the total molar enthalpy variation experimentally determined; the contributions of affinity (specificity in territorial condensation) and chemical bonding of calcium counterions to the thermodynamic properties of the system (i.e., enthalpy and entropy) were calculated. The intrinsic molar enthalpy of bonding, DeltaH(bond,0), displayed a peculiar sigmoid dependence on Rj. In particular, its decrease toward more negative values was interpreted as stemming from a (cooperative) calcium-induced conformational change that accompanies pectate chain pairing upon junction formation. The calculated pKin of instability of the Ca-(GalA-)4 complex was 10.80, in very good agreement with the corresponding value reported for the Ca-EDTA complex (i.e., 10.96). Significant contributions to the complex stability were the enthalpy of ion pairing (DeltaH(ionpairing,bond) = -5.1 kcal (mol calcium)-1, in good agreement with the value reported for calcium-EDTA: approximately -5.4 kcal (mol calcium)-1), and the entropy of desolvation (DeltaS(desolv,bond) = 43.7 cal mol-1 K-1, well within the range of values reported for calcium-EDTA: 42-57 cal mol-1 K-1).

Polyuronates such as pectate and alginate are very well-known examples of biological polyelectrolytes undergoing, upon addition of divalent cations, an interchain association that acts as the junction of an eventually formed stable hydrogel. In the present paper, a thermodynamic model based on the counterion condensation theory has been developed to account for this cation-induced chain pairing of negatively charged polyelectrolytes. The strong interactions between cross-linking ions and uronate moieties in the specific binding site have been described in terms of chemical bonding, with complete charge annihilation between the two species. The chain-pairing process is depicted as progressively increasing with the concentration of cross-linking counterions and is thermodynamically defined by the fraction of each species. On these bases, the total Gibbs energy of the system has been expressed as the sum of the contributions of the Gibbs energy of the (single) chain stretches and of the (associated) dimers, weighted by their respective fractions 1 - theta and theta. In addition, the model assumes that the condensed divalent counterions exhibit an affinity free-energy for the chain, G(C)(aff,0), and the junction, G(D)(aff,0), respectively. Moreover, a specific Gibbs energy of chemical bonding, G(bond,0), has been introduced as the driving force for the formation of dimers. The model provides the mathematical formalism for calculating the fraction, theta, of chain dimers formed and the amount of ions condensed and bound onto the polyelectrolyte when two different types of counterions (of equal or different valence) are present. The effect of the parameter G(bond,0) has been investigated and, in particular, its difference from G(C,D)(aff,0) was found to be crucial in determining the distribution of the ions into territorial condensation and chemical bonding, respectively. Finally, the effect of the variation of the molar ratio between cross-linking ions and uronic groups in the specific binding sites, sigma0, was evaluated. In particular, a remarkable decrease in the amount of condensed counterions has been pointed out in the case of sigma0 = 1/3, with respect to the value of sigma0 = 1/4, characterizing the traditional "egg-box" structure, as a result of the drop of the charge density of the polyelectrolyte induced by complete charge annihilation.

Light scattering (LS) investigations have additionally confirmed the intramolecular nature of the fundamental ordered conformation (single helix) of κ-carrageenan in aqueous 0.1 M NaI. The partial replacement of sodium with cesium ions induces a (thermoreversible) intermolecular association of the polysaccharide in the ordered conformation. The association-inducing effect can be observed already at the lowest investigated concentration of the cation. We found no significant change of the thermodynamic properties related to an alleged composition threshold value in the range of the cesium equivalent fraction, X Cs , from 0.12 to 0.40. The extent and the rate of the association process strongly increase with the concentration of cesium. Equilibrium values for the association constant could be obtained except for the case of X Cs = 0.40, where the process is too massive to allow long-time LS investigation. In the latter condition, some additional stiffening of the chain was noticed.

The temperature-induced helix-to-coil conformational transition accompanying the thermoreversible gelation of syndiotactic poly(methyl methacrylate) in toluene was studied by rheology and FTIR and 1H NMR spectroscopy. Both rheological and FTIR measurements, carried out dynamically as a function of temperature, revealed a marked hysteresis between the heating and the cooling cycles. It was not possible to assess a quantitative difference between the values of the temperature of onset of the physical variations between the rheological and the spectroscopic experiments. It is then possible to conclude that a conformational change of the backbone (from the evolution of the band at 860 cm-1) is the molecular basis of the macroscopic formation of the gel. Spin−spin relaxation time, or T2, measurements were used in the 1H NMR study. The data obtained from the relaxation behavior of the α-methyl group, indicated a fraction of 0.28 for the chain repeating units incorporated in highly cooperative helical stretches,...

The conformational transition of ι-carrageenan in different salt solutions under nongelling conditions has been studied by light scattering (WALLS). The focus is on the thermodynamic interactions and the concentration dependence of the reduced scattering intensity at zero angle. An upward curvature of (KC p /R 0 ) in conditions of conformational ordering has been interpreted as due to a concentration-dependent reversible chain association. The open association model of Elias has been applied to analyze the data. In conditions of conformational ordering a decrease of polymer concentration, Cp, is accompanied by a decrease of (M w ) assoc . These results have been confirmed by using low-angle light scattering (LALLS). In the lowest range of Cp, the decreasing values of (M w ) assoc are already very close to that of the single-stranded disordered conformation. In the same range, the values of the specific optical activity remain constant. This is a confirmatory evidence that the fundamental ordered conformation of ι-carrageenan is a single helix.

A large number of light-scattering data indicate that the sulfated polysaccharide K-carrageenan undergoes an intramolecular conformational transition under suitable conditions from a semiflexible, disordered conformation to a rather rigid, helical one. By varying the parameters controlling the conformational phase diagram (i.e., by increasing the concentration of polymer or that of the supporting electrolyte, or both, or by decreasing temperature), a tertiary structure is manifested, through a progressive increase of the weight-average molar mass (M w ) as determined by light scattering. This can be very well explained in terms of a reversible association of stretches of such intramolecular helical structures and quantitatively described by a model of open association of two or more chains. The association of κ-carrageenan was demonstrated to take place at around room temperature also in the presence of iodide ions, as long as their concentration is ≥0.20 M. In this way the apparent inconsistency of a number of published M w results can be reconciled. Additional light-scattering results indicated that the demonstrated thermodynamic tendency to reversible interchain association may lead to an irreversible aggregation of polymer chains if an improper solution preparation procedure is used.