Adsorption behaviour and surfactant elution of cationic salivary proteins at solid/liquid interfaces, studied by in situ ellipsometry

Abstract

Adsorption of the cationic salivary proteins lactoferrin, lactoperoxidase, lysozyme and histatin 5 to pure (hydrophilic) and methylated (hydrophobized) silica surfaces was investigated by in situ ellipsometry. Effects of concentration (≤10 g ml −1 , for lysozyme ≤200 g ml −1 ) and dependence of surface wettability, as well as adsorption kinetics and elutability of adsorbed films by buffer and sodium dodecyl sulphate (SDS) solutions were investigated. Results showed that the amounts adsorbed decreased in the order lactoferrin ≥ lactoperoxidase > lysozyme ≥ histatin 5. On hydrophilic silica, the adsorption was most likely driven by electrostatic interactions, which resulted in adsorbed amounts of lactoferrin that indicated the formation of a monolayer with both side-on and end-on adsorbed molecules. For lactoperoxidase the adsorbed amounts were somewhat higher than an end-on monolayer, lysozyme adsorption showed amounts corresponding to a side-on monolayer, and histatin 5 displayed adsorbed amounts in the range of a side-on monolayer. On hydrophobized substrata, the adsorption was also mediated by hydrophobic interactions, which resulted in lower adsorbed amounts of lactoferrin and lactoperoxidase; closer to side-on monolayer coverage. For both lysozyme and histatin 5 the adsorbed amounts were the same as on the hydrophilic silica. The investigated proteins exhibited fast adsorption kinetics, and the initial kinetics indicated mass transport controlled behaviour at low concentrations on both types of substrates. Buffer rinsing and SDS elution indicated that the proteins in general were more tightly bound to the hydrophobized surface compared to hydrophilic silica. Overall, the surface activity of the investigated proteins implicates their importance in the salivary film formation.

Lactoperoxidase and Histatin 5 – their Adsorption Behaviour on Silica and Hydrophobized Silica Surfaces, and Implications on their Role in the Initial Salivary Film Formation

“…As can be seen, the surface area per molecule of lactoperoxidase was larger on hydrophobized silica substrates compared to hydrophilic substrates, revealing that a larger number of molecules were adsorbed to the pure silica substrate. These results are in agreement with those in previous studies [12,27]. The main driving force for adsorption on the hydrophilic substrate is most likely electrostatic attraction between the cationic protein and the an- Variations between measurements were < 10%.…”

“…Further, the overall character of the pellicle has been reported to be anionic [11], and the presence of these cationic components may increase the cohesiveness and thickness of the film by complex formations with anionic pellicle proteins. This study is a complement to a preceding, more detailed study performed on the adsorption behaviour of lactoferrin, lactoperoxidase, lysozyme and histatin 5, that investigated the adsorption in terms of concentration dependence, influence of surface wettability, kinetics and elutability by buffer and surfactant solutions [12]. The present investigation focused on the adsorption of lactoperoxidase and histatin 5.…”

The salivary mucin MUC5B and lactoperoxidase can be used for layer-by-layer film formation

“…Most of the data in this table is taken from the literature [34] except that of MUC5B and α-amylase, respectively, which were measured by ellipsometry at similar conditions as in Svendsen et al [34]. All the individual proteins adsorb on hydrophilic silica although the extension of adsorption is considerably larger for lactoferrin and lactoperoxidase.…”

Activity of lactoperoxidase when adsorbed on protein layers

“…Higher adsorbed masses, up to 3.2 mg/m 2 of LPO depending on enzyme concentration in solution, were obtained on hydrophilic surfaces [15]. One problem with silica surfaces is that it is difficult to measure the activity of the adsorbed enzyme, which is of obvious importance for the development of antimicrobial surfaces.…”