Previous research on the binding and gelation of calcium/alginate in aqueous solution were mostly conducted in the (semi-)concentrated regime, and it did not provide details of the binding process and the formation of egg-box junctions. In the present investigation, the binding of calcium to alginate, of low and high molecular weight and different guluronate/mannuronate ratios, was investigated in dilute solutions using isothermal titration calorimetry (ITC), Ca2+-selective potentiometry, and viscometry techniques. The results reveal three distinct and successive steps in the binding of calcium to alginate with increased concentration of Ca ions. They were assigned to (i) interaction of Ca2+ with a single guluronate unit forming monocomplexes; (ii) propagation and formation of egg-box dimers via pairing of these monocomplexes; and (iii) lateral association of the egg-box dimers, generating multimers. The third step has different association modes depending on the molecular weight of alginate. The boundaries between these steps are reasonably critical, and they closely correlate with the Ca/guluronate stoichiometry expected for egg-box dimers and multimers with 2/1 helical chains. The formation of egg-box dimers and their subsequent association are thermodynamically equivalent processes and can be fitted by a model of independent binding sites. The binding of Ca to alginates of different guluronate contents is controlled by a balance between enthalpy and entropy.

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The structural characteristics of the gum exudate of Acacia senegal (gum arabic) have been investigated by monitoring the composition and physicochemical properties before and after treatment with proteolytic enzyme and various alkaline systems. Molecular mass ( M w) and radius of gyration ( R g) measurements were performed using gel permeation chromatography (GPC) coupled to refractive index, UV absorbance, and multiangle light scattering detectors and indicated that the macromolecules present have a compact structure. It was found that treatment with proteolytic enzyme caused the arabinogalactan-protein component (AGP) with average molecular mass approximately 2 x 10 (6) Da to degrade, yielding material of molecular mass approximately 4 x 10 (5) Da, whereas the bulk of the material corresponding to the protein-deficient arabinogalactan component (AG) with molecular mass 4 x 10 (5) remained unaffected. Barium hydroxide was found to hydrolyze the polysaccharide component (AG) itself in addition to the proteinaceous component as demonstrated in control experiments using dextran. However, sodium borohydride/sodium hydroxide treatments were unable to hydrolyze dextran and were assumed to hydrolyze only the proteinaceous component of gum arabic. The AGP component was completely degraded, yielding material of molecular mass approximately 4.5 x 10 (4) Da. It has been concluded, therefore, that the enzyme did not fully hydrolyze all of the protein present and that the AGP component of gum arabic consists of carbohydrate blocks of approximately 4.5 x 10 (4) Da linked to a polypeptide chain consistent with the wattle blossom structure. Because the AGP was degraded to differing extents using a mild and more severe sodium borohydride/sodium hydroxide treatment, it was concluded that the polysaccharide moieties were linked through both O-serine and O-hydroxyproline residues. The gum arabic sample was deglycosylated by treatment with anhydrous hydrogen fluoride and revealed the presence of two putative core proteins of approximately 3 x 10 (4) and approximately 5 x 10 (3) Da, respectively, which correspond to proteins of approximately 250 and 45 amino acids in length. A new model for the structure of the AGP component has been proposed.

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The complexation between bovine serum albumin (BSA) and sugar beet pectin (SBP) was studied in situ by coupling glucono-δ-lactone (GDL) induced acidification with dynamic light scattering and turbidity measurements. Individual measurements at specific pHs and mixing ratios were also carried out using zeta potentiometry, gel permeation chromatography-multiangle laser light scattering (GPC-MALLS), and isothermal titration calorimetry (ITC). These investigations together enabled the establishment of a phase diagram of BSA/SBP and the identification of the molecular events during protein/polysaccharide complexation in relation to the phase diagram, which showed five regions: (I) a stable region of mixed individual soluble polymers, (II) a stable region of intramolecular soluble complexes, (III) a quasi-stable region of intermolecular soluble complexes, (IV) an unstable region of intermolecular insoluble complexes, and (V) a second stable region of mixed individual soluble polymers, on lowering pH. We found for the first time that the complexation could take place well above the critical pH(c), the value that most previous studies had regarded as the onset occurrence of complexation. A model of structural transitions between the regions was proposed. The borderline between region II and region III represents the BSA/SBP stoichiometry for intramolecular soluble complex at a specific pH, while that between region III and region IV identifies the composition of the intermolecular insoluble complex. Also studied was the effect of NaCl and CaCl(2) on the phase diagram and structural transitions.