In several experimental techniques D2O rather then H2O is often used as a solvent for proteins. Concerning the influence of the solvent on the stability of the proteins, contradicting results have been reported in literature. In this paper the influence of H2O-D2O solvent substitution on the stability of globular protein structure is determined in a systematic way. The differential scanning calorimetry technique is applied to allow for a thermodynamic analysis of two types of globular proteins: hen's egg lysozyme (LSZ) with relatively strong internal cohesion ("hard" globular protein) and bovine serum albumin (BSA), which is known for its conformational adaptability ("soft" globular protein). Both proteins tend to be more stable in D2O compared to H2O. We explain the increase of protein stability in D2O by the observation that D2O is a poorer solvent for nonpolar amino acids than H2O, implying that the hydrophobic effect is larger in D2O. In case of BSA the transitions between different isomeric forms, at low pH values the Nm and F forms, and at higher pH values Nm and B, were observed by the presence of a supplementary peak in the DSC thermogram. It appears that the pH-range for which the Nm form is the preferred one is wider in D2O than in H2O.

The Lifshitz-van der Waals, acid-base, and total surface free energies of various wood species were calculated from contact angle measurements. For spruce (Picea abies) and meranti (Shorea spp.) the following three methods were compared: capillary rise in wood powder columns (based on the Washburn equation), dynamic contact angle measurements (according to the Wilhelmy-plate principle), and sessile drop measurements along and across the grain of the wood. The capillary rise method was limited to nonswelling solvents, which means that only the Lifshitz-van der Waals component could be measured. With the dynamic contact angle measurement, the wettability during the first immersion was decreased compared to that of the sessile drop. This was probably due to reduced capillary penetration, but with the second immersion the presence of an adsorbed solvent layer increased the wettability and hence affected the surface energy data. The sessile drop measurements were highly dependent on the direction of measurement. Increasing the wood moisture content decreased the Lifshitz-van der Waals component and increased the basic surface energy parameter of the wood. All of the wood species tested were characterized as having low-energy surfaces with a dominant Lifshitz-van der Waals component. Measurement of acid and base parameters of wood surfaces seemed not to be very reliable because of its strong dependence on the measuring conditions. With respect to this, it should be noted that thermodynamic equilibrium conditions assumed by Young's equation are generally not fulfilled with wood surfaces because of chemical heterogeneity, surface roughness, and the adsorption of the test solvent.

The cross-linking behavior of mussel adhesive protein Mefp-1 was studied by measuring the rate of aggregation of the protein by photon correlation spectroscopy. To be able to calculate the aggregation numbers, the hydrodynamic radius of monomer Mefp-1 (10 nm) was determined under reducing conditions. The aggregation is controlled by the redox potential of the solution, and the aggregation number varied, independent of pH, over a factor 2 within the experimentally accessible redox potential window. A kinetic model for cross-linking, based on the intricate interplay of the oxidation and auto-oxidation of the hydroquinones of Mefp-1, is proposed. The oxidation rate strongly depends on redox potential. The cross-linking rate is taken to be proportional to the rate of auto-oxidation. The model correctly predicts the experimentally observed phenomena. When the oxidation rate is slower than the auto-oxidation rate, cross-linking is efficient and controlled by the oxidation rate. When the rate of auto-oxidation rate is slower than the oxidation rate, the cross-linking is inefficient due to the quick exhaustion of the hydroquinones. The experimentally determined rate constant for cross-linking is found to be much smaller than those found for auto-oxidation of hydroquinones because of the excluded volume interactions imposed by the protein backbone. Tuning the interplay between oxidation and auto-oxidation presents the potential of controlling cross-linking density independent of the density of reactive groups.

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