The influence of surface and thermal denaturation of adsorbed β-lactoglobulin (β-Lg) on the flocculation
of hydrocarbon oil droplets was measured at pH 3 and compared with that at pH 7. Oil-in-water emulsions
(5 wt % n-hexadecane, 0.5 wt % β-Lg, pH 3.0) were prepared that contained different levels of salt (0−150
mM NaCl) added immediately after homogenization. The mean particle diameter (d
43) and particle size
distribution of diluted emulsions were measured by laser diffraction when they were either (i) stored at
30 °C for 48 h or (ii) subjected to different thermal treatments (30−95 °C for 20 min). In the absence of
salt, little droplet flocculation was observed at pH 3 or 7 because of the strong electrostatic repulsion
between the droplets. In the presence of 150 mM NaCl, a progressive increase in mean particle size with
time was observed in pH 7 emulsions during storage at 30 °C, but no significant change in mean particle
diameter with time (d
43 ∼ 1.4 ± 0.2 μm) was observed in the pH 3 emulsions. Droplet aggregation became
more extensive in pH 7 emulsions containing salt (added before thermal processing) when they were
heated above 70 °C, which was attributed to thermal denaturation of adsorbed β-Lg leading to interdroplet
disulfide bond formation. In contrast, the mean particle size decreased and the creaming stability improved when pH 3 emulsions were heated above 70 °C. These results suggest that the droplets in the pH
3 emulsions were weakly flocculated at temperatures below the thermal denaturation temperature of β-Lg
(T < 70 °C) but that flocs did not form so readily above this temperature, which was attributed to a
reduction in droplet surface hydrophobicity due to protein conformational changes. The most likely
explanation for the difference in behavior of the emulsions is that disulfide bond formation occurs much
more readily at pH 7 than at pH 3.