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Nanofibrils of β-lactoglobulin can be assembled into bundles by site-specific noncovalent cross-linking with high-methoxyl pectin (Hettiarachchi et al. Soft Matter 2016, 12, 756). Here we characterized the nanomechanical properties of bundles using atomic force microscopy and force spectroscopy. Bundles had Gaussian cross sections and a mean height of 17.4 ± 1.4 nm. Persistence lengths were calculated using image analysis with the mean-squared end-to-end model. The relationship between the persistence length and the thickness had exponents of 1.69-2.30, which is consistent with previous reports for other fibril types. In force spectroscopy experiments, the bundles stretched in a qualitatively different manner to fibrils, and some of the force curves were consistent with peeling fibrils away from bundles. The flexibility of pectin-linked nanofibril bundles is likely to be tunable by modulating the stiffness and length of fibrils and the ratio of pectin to fibrils, giving rise to a wide range of structures and functionalities.

The displacement of spread protein films from the air/water interface by surfactant was followed using Brewster angle microscopy (BAM) and interfacial rheology. The displacement of beta-lactoglobulin and beta-casein by a nonionic surfactant was monitored as a function of both surface pressure and time. In both cases, protein displacement occurred over the same surface pressure range that had been observed previously by atomic force microscopy (AFM). In the case of the beta-lactoglobulin, surfactant domains grew large enough in the protein film to be visible in the BAM images. The shapes of the domains were very similar to those seen previously by AFM in the late stages of displacement. The results from both proteins confirm the results published previously while highlighting some implications for the application of the "orogenic" model of displacement for large protein films. The surface rheological data showed that the beta-lactoglobulin/surfactant mixed film retained much of its elasticity until the latter stages of displacement. This indicates that at least in the early stages of displacement, the mixed film was dominated by the behavior of the protein in the film.

The displacement of the proteins (beta-lactoglobulin and beta-casein) from an air-water interface by the nonionic (Tween 20 and Tween 60) and ionic (sodium dodecyl sulfate, cetyltrimethylammonium bromide, and lyso-phosphatidylcholine-lauroyl) surfactants has been visualized by atomic force microscopy (AFM). The surface structure has been sampled by the use of Langmuir-Blodgett deposition onto mica substrates to allow imaging in the AFM. In all cases, the displacement process was found to occur through the recently proposed orogenic mechanism (Mackie et al. J. Colloid Interface Sci. 1999, 210, 157-166). In the case of the nonionic surfactants, the displacement involved nucleation and growth of surfactant domains leading to failure of the protein network and subsequent loss of protein into the bulk phase. The surface pressure dependence of the growth of surfactant domains and the failure of the network were found to be the same for both Tween 20 and Tween 60, demonstrating that the breakdown of the protein film was dominated by the mechanical properties of the network. The displacement of protein by ionic surfactants was found to be characterized by nucleation of surfactant domains with little domain growth prior to failure of the network. The size of the domains formed by ionic surfactants was found to be limited by the strong intersurfactant repulsive forces between the charged headgroups. Screening of these charges led to an increase in the size of the domains. The surface pressure at which the network continuity was lost was found to be dependent on the type of surfactant and, in all cases, to occur at higher surface pressures than that required for nonionic surfactants. This has been attributed to surfactant-protein binding that initially strengthens the protein network at low surfactant concentrations. Evidence obtained from surface shear rheology supports this assertion.