Ca-alginate gels were studied by small-angle X-ray scattering and rheology to determine relations between chemical composition and concentrations of the alginate and the elasticity and structure of the gels. The gels were prepared by in situ release of Ca 2+ from either Ca-EGTA or CaCO3 with total Ca 2+ concentration in the range 5-30 mM. Alginates with low (39%), intermediate (50%), and high (68%) fractions of R-L-GulA originating from the brown algae Ascophyllum nodosum, Laminaria hyperborea leaf, and Laminaria hyperborea stipe, respectively, were employed. Two to three different degrees of polymerization for each chemical composition were used in the experiments. The excess small-angle X-ray scattering for the alginates in solution yielded linear cross-sectional Guinier plots, and the cross-sectional radius of gyration, R g,c, was determined to be 3.1-4.6 Å. The SAXS profiles of the alginate gels depended on the alginate concentration, Ca 2+ concentration, and the alginate composition. The SAXS data suggested that dimerization of chain segments was the principal association mode at low fractional Ca 2+ saturation of guluronic acid of the alginate. Increasing the fractional Ca 2+ saturation of guluronic acid, either by the concentrations or selection of alginate source, yielded coexisting lateral association modes, as manifested in a curvature in the cross-sectional plots. The coexisting junction zone multiplicities occur because of a delicate balance between the block length distribution of the R-L-GulA residues, polymer concentration, and Ca 2+ . These results are quantitative extensions of the "egg-box" model used to describe ionotropic gelation of alginate and hence enhance the understanding of the structure-function relationship of alginate gels.

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The enzymes mannuronan C-5 epimerases catalyze conversion of beta-D-mannuronic acid to alpha-L-guluronic acid in alginates at the polymer level and thereby introduce sequences that have functional properties relevant to gelation. The enzymatic conversion by recombinant mannuronan C-5 epimerases AlgE4 and AlgE2 on alginate type substrates with different degree of polymerization and initial low fraction of alpha-L-guluronic acid was investigated. Essentially no enzymatic activity was found for fractionated mannuronan oligomer substrates with an average degree of polymerization, DP(n), less than or equal 6, whereas increasing the DP(n) yielded increased epimerization activity. This indicates that these enzymes have an active site consisting of binding domains for consecutive residues that requires interaction with 7 or more consecutive residues to show enzymatic activity. The experimentally determined kinetics of the reaction, and the residue sequence arrangement introduced by the epimerization, were modeled using Monte Carlo simulation accounting for the various competing intrachain substrates and assuming either a processive mode of action or preferred attack. The comparison between experimental data and simulation results suggests that epimerization by AlgE4 is best described by a processive mode of action, whereas the mode of action of AlgE2 appears to be more difficult to determine.