Solvation and expansion of neutral and charged chains of a carbohydrate polyelectrolyte: Galacturonan in water. A critical revisiting.

Experimental and theoretical data have been revisited to shed light onto the aspects of hydration and chain expansion of pectic acid (galacturonan) upon charging. The prediction of the variation of the number of solvation water molecules between the two limit ionization states from theoretical calculations was confirmed to a very high accuracy by the corresponding number evaluated form dilatometric measurements. The relevance of hydration to the mechanism of bonding of calcium ions by sodium pectate is discussed. Characterization of polymer expansion has been obtained by calculating the values of the characteristic ratio and/or the persistence length on the respective populations and comparing the theoretical predictions with experimental data. The results show that a charged chain in typical conditions of ionic strength is more expanded than its neutral counterpart, whereas the ideal limit (31 and 21) helical conformations in the uncharged and totally charged conditions, respectively, share the same value of the linear advance of the helical repeat, when the ionic strength tends to infinite. Total divergence between theoretical predictions and experimental evidence rules out the possibility that carboxylate charge reduction by protonation and by methyl esterification are equivalent in determining the solution behavior of galacturonan.

On the Molecular Mechanism of the Calcium-Induced Gelation of Pectate. Different Steps in the Binding of Calcium Ions by Pectate.

Pectic acid/sodium pectate is one of the most widespread hydrocolloid used in the food industry. It is able to form strong ionotropic gels by the addition of ions, in particular, calcium ions. The initial steps of binding Ca2+ ions to a sample of sodium pectate with a composition close to 90% of ideal Na+-poly(galacturonate) were investigated by means of circular dichroism (CD), microcalorimetry, dilatometry, viscosity, and membrane osmometry, as a function of increasing Rj, Rj being the ratio of the molar concentrations of Ca2+ and pectate repeating units. Data were collected in aqueous NaClO4 at 25 °C. The key instrument of interpretation has been the counterion condensation theory (CCT) of linear polyelectrolytes, modified to include the presence of both specific affinity of the divalent counterion for the polysaccharide ("territorial binding"), and, very importantly, strong chemical bonding (not a covalent bonding, though) of Ca2+ on conformationally well-defined sites on the polymer, with local charge annihilation. Intrinsic viscosity and number-average molar mass data as a function of Rj showed that calcium bonding brings about chain association right from the beginning of addition to pectate. The analysis of the microcalorimetric curve using the modified CCT revealed two types of bonding. In the order of development as a function of Rj, the first mode (type 1) could be reconciled with the "tilted egg-box" type, recently proposed for Ca2+ binding to alginate and the second mode (type 2) with the "shifted egg-box" proposed for calcium pectate on the basis of conformational analysis investigation. Likewise, the two types of bonding turned out to be superimposable with similar bonding categories proposed for alginate and low-methoxyl pectin (LMP), on the one side, and for the association of semiflexible polyelectrolytes, on the other. The analysis allowed us to obtain standard Gibbs free energy, enthalpy, entropy, and volume molar values both for the affinity and the chemical bonding processes. Interestingly, the analysis of the dependence of the gelation temperatures, Tg, of LMP upon increasing additions of calcium ions provided the values of Tg and standard Gibbs free-energy of calcium-to-pectate association coinciding with those obtained from calorimetry for the type-2 bonding process. This finding corroborated previously reported evidence on the enthalpic nature of the elasticity of Ca2+-pectate gels. Finally, comparative analysis of different techniques, but of CD in particular, enabled proposing a "loose-21-helix" as the starting conformation of sodium pectate in aqueous solution.

Butyrate-Loaded Chitosan/Hyaluronan Nanoparticles: A Suitable Tool for Sustained Inhibition of ROS Release by Activated Neutrophils.

Tissue damage caused by excessive amounts of neutrophil-derived reactive oxygen species (ROS) occurs in many inflammatory diseases. Butyrate is a short-chain fatty acid (SCFA) with known anti-inflammatory properties, able to modulate several neutrophil functions. Evidence is provided here that butyrate inhibits neutrophil ROS release in a dose and time-dependent fashion. Given the short half-life of butyrate, chitosan/hyaluronan nanoparticles are next designed and developed as controlled release carriers able to provide cells with a long-lasting supply of this SCFA. Notably, while the inhibition of neutrophil ROS production by free butyrate declines over time, that of butyrate-loaded chitosan/hyaluronan nanoparticles (B-NPs) is sustained. Additional valuable features of these nanoparticles are inherent ROS scavenger activity, resistance to cell internalization, and mucoadhesiveness. B-NPs appear as promising tools to limit ROS-dependent tissue injury during inflammation. Particularly, by virtue of their mucoadhesiveness, B-NPs administered by enema can be effective in the treatment of inflammatory bowel diseases.

Polyelectrolyte study of the calcium-induced chain association of pectate.

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).