Thomas Arnebrant

Protein adsorption to solid surfaces

The phenomenon of protein adsorption to solid surfaces affects the performance of many materials and processes, in areas ranging from medicine to biochemical engineering. Controlling protein adsorption, from solutions of single proteins as well as from more complex mixtures, requires an understanding of the mechanism(s) by which it occurs. This, in turn, entails detailed characterization of both the protein and the solid surface and identification of those factors controlling the adsorption process.

Protein interactions at solid surfaces

Abstract In this review article we discuss a large number of the studies of interactions between protein-coated surfaces that has been presented in the literature. We also demonstrate how to relate surface force data to results from other techniques in order to provide a more full picture of protein behaviour at interfaces. One aim of the article is to discuss the experimental procedure and the difficulties with surface force measurements in protein systems. It is particularly important to point out how the sensitivity of this technique differs from that of other techniques, e.g. in determining structural changes in adsorbed proteins and in detecting proteins adsorbed on top of an inner firmly bound layer. It is also important to realize which surface force data cannot easily be compared with findings from other techniques (one example is the kinetics of adsorption and desorption). We have tried to group proteins into different classes depending on their size and structure, and to try to find results that are common within these classes. It was found that some observations for unordered proteins with amphiphilic character, and for the small compact proteins, appear consistently within the respective class. Hence, for these types of protein common features/principles of the interfacial behaviour are identified. The very large and flexible glycoproteins behave in a similar way to synthetic polymers, but we found it hard to draw any firm conclusions based on the surface force studies presented so far. Perhaps, the most complicated surface behaviour is observed for soft globular proteins that undergo large-scale conformational changes upon adsorption and when the layers are held under a high compressive force.

Adsorption of β-lactoglobulin onto silica, methylated silica, and polysulfone.

Milk and whey are widely processed by membrane filtration, often using polysulphone membranes. Adsorption of β-lactoglobulin onto polysulphone was studied at protein concn. of 0.1 and 1.0%, as well as 12% to represent concn. encountered during ultrafiltration. Adsorption onto silica and methylated silica surfaces (representing strongly hydrophilic and strongly hydrophobic surfaces resp.) was also studied. Protein was dissolved in 0.01 smallcap˜M phosphate buffer pH 7.0 containing 0.15 smallcap˜M NaC1 and adsorption/desorption was monitored in situ using a Rudolph Thin Film ellipsometer. Polysulphone surfaces adsorbed the greatest amount of β-lactoglobulin and silica the least; methylated silica was intermediate. Differences between methylated silica and polysulphone may reflect differences in surface roughness. Adsorption to polysulphone and methylated silica was not reversed on dilution, whereas adsorption to silica was partially reversible. Pretreatment of polysulphone and methylated silica surfaces with 0.1% β-lactoglobulin markedly reduced subsequent adsorption from 12% β-lactoglobulin (equivalent to adsorption from 0.1% solution alone); preadsorption to silica surfaces had much less effect on subsequent adsorption. Methylated silica was concluded to be a representative model for a polysulphone surface. (Less)

Adsorption of α-lactalbumin and β-lactoglobulin on metal surfaces versus temperature

Abstract The adsorption of α-lactalbumin and β-lactoglobulin on a hydrophilic chromium surface was followed in situ, using ellipsometry. The experiments were performed at temperatures up to and exceeding (in the case of α-lactalbumin) the thermal denaturation temperatures of the proteins. The denaturation temperatures and the reversibility of the denaturation of the proteins were estimated by differential scanning calorimetry (DSC). The residual transition enthalpy for the thermal denaturation of β-lactoglobulin, as a function of preheat time at temperatures dose to the denaturation temperature, was also recorded. In agreement with earlier reports, the denaturation of β-lactoglobulin was found to be irreversible, whereas the denaturation of α-lactalbumin was highly reversible. When approaching its denaturation temperature, the adsorption behavior of β-lactoglobulin indicates that an aggregation occurs at the surface starting after a time lag of several minutes. No such behavior was found for α-lactalbumin, where both the initial and the plateau values of the adsorbed amount increased gradually as a function of temperature. Preheated solutions of β-lactoglobulin had nearly the same adsorption behavior at 25°C as the native protein, while the thermally denatured form of α-lactalbumin seemed to be more surface active than the native.

Sequential and competitive adsorption of β-lactoglobulin and κ-casein on metal surfaces☆

The adsorbed amount of K-casein and β-lactoglobulin on metal surfaces was measured in situ by ellipsometry as a function of time. The measurements were made in phosphate-buffered solutions. Glass slides with vacuum-deposited chromium, treated in different ways to obtain hydrophilic as well as hydrophobic surfaces, were used. The effect of a monolayer of one protein on the subsequent adsorption of the other protein was studied, and competitive adsorption of the two proteins was followed using ellipsometry and 14C-labeled β-lactoglobulin. The results of the sequential adsorption indicate that κ-casein adsorbs after the plateau value of adsorption of β-lactoglobulin has been reached, whereas no adsorption of β-lactoglobulin takes place after the plateau value of κ-casein has been reached. When the two proteins were added simultaneously the surface energy of the substrate influences both the total adsorbed amount and the composition of the adsorbed layer.

Interaction of cetyltrimethylammonium bromide and sodium dodecyl sulfate with β-lactoglobulin and lysozyme at solid surfaces

The interaction of two ionic surfactants, CTAB and SDS, with β-lactoglobulin and lysozyme at surfaces was monitored by in situ ellipsometry. The effects of the surfactants on proteins adsorbed at a surface as well as the adsorption from protein/surfactant mixtures were studied. The behavior at four different surfaces, silicon oxide, chromium oxide, nickel oxide, and methylated silica, was investigated. The adsorbed amounts of protein were in all cases below or in the range of what would be expected for monolayer adsorption. The elutability of protein seemed to be most complete at the methylated silica surface while for the oxide surfaces the degree of elution was decreasing in the order silica, chromium oxide, and nickel oxide. The effects of surfactants on adsorbed protein films with no protein present in the solution were described according to four adsorption/displacement models: (1) Removal of the protein upon addition of surfactant. (2) Replacement of the protein by the surfactant. (3) Reversible adsorption of the surfactant to the surface with adsorbed protein. (4) Partial removal of protein according to model 1 or 2. When surfactants were premixed with protein prior to adsorption, the interaction between protein and surfactant in solution had to be taken into account. This was reflected in differences in the amounts adsorbed obtained after adsorption as well as after rinse between these experiments and those where surfactant was added after preadsorption of protein. Under certain conditions the presence of surfactant completely prevented protein adsorption.

An ellipsometry study of ionic surfactant adsorption on chromium surfaces

Abstract The adsorption of dodecyl sulfate and cetyltrimethylammonium surfactant on chromium was studied by in situ ellipsometry. Two types of chromium surfaces were used: clean hydrophilic surfaces and hydrophobized surfaces. The amounts adsorbed are constant at concentrations exceeding the critical micelle concentration (CMC). In general, less surfactant is adsorbed on the hydrophilic surface than on the hydrophobized surface. No adsorption of surfactant molecules can be observed on the clean hydrophilic surface, unless the surfactant carries an opposite charge in relation to the surface. The amounts of surfactant adsorbed increase as the initial charge of the oppositely charged surface increases. The adsorption on the hydrophobic surface was found to be less pH dependent and surfactant molecules are adsorbed even if the surface and the surfactant have the same charge. An increase in the ionic strength generally leads to larger amounts adsorbed. The adsorption of cetyltrimethylammonium surfactant was found to be dependent on the type of counterion present. As opposed to bromide counterions, the hydroxide and chloride counterions lower the amount of surfactant adsorbed.

Adsorption of insulin on metal surfaces in relation to association behavior

Abstract The adsorption of insulin on metal surfaces from aqueous solutions was monitored by in situ ellipsometry. Clean (hydrophilic) chromium and titanium surfaces as well as chromium surfaces treated to be hydrophobic were used. The adsorbed amount was found to be higher on the hydrophilic than on the hydrophobic surfaces. The presence of the divalent metal ions zinc and calcium increased the adsorbed amount and also the fraction desorbable upon rinsing. This was related to the behavior in the bulk solution, where zinc and calcium increase the association. The increase in adsorbed amount was observed also when zinc was added to an adsorbed layer of Zn-free insulin in contact with insulin solution. Addition of citrate to an adsorbed layer of insulin from a solution containing four zinc ions per hexamer had an effect similar to that of rinsing, indicating a removal of zinc in both cases. The effect of the pH was investigated in the range from pH 7 to pH 8, showing an increase in adsorbed amounts at lower pH, especially pronounced for the samples with a high zinc content, four zinc ions per insulin hexamer. The calcium-containing samples did not show such a strong pH dependence.

The effect of delmopinol on salivary pellicles, the wettability of tooth surfaces in vivo and bacterial cell surfaces in vitro

Delmopinol is a low molecular weight surface active compound, which has been shown to be effective against dental plaque both in vitro and in vivo, and against gingivitis in vivo. The purpose of the present study was to investigate the influence of delmopinol on salivary pellicles and on the adhesiveness of tooth surfaces, bacterial cell surfaces and pellicle‐coated bacterial cell surfaces in order to elucidate its mode of action. The interaction of delmopinol with salivary pellicles showed that the effect was strongly dependent on the surface properties of the material on which the pellicle was formed, ranging from reversible binding of delmopinol to removal of pellicle material. The binding of delmopinol appeared to be reversible upon rinsing in all cases. The results from the contact angle measurements showed that the accessibility of polar units increased after adsorption, suggesting that delmopinol exposes the polar part of the molecule outwards after adsorption, but this effect was short‐lived. It i...

Adsorption of globular model proteins to silica and methylated silica surfaces and their elutability by dodecyltrimethylammonium bromide

The interaction between a cationic surfactant (dodecyltrimethylammonium bromide) and six model proteins adsorbed on to methylated silica and silica surfaces was investigated. The proteins were bovine serum albumin, cytochrome c, β-lactoglobulin, α-lactalbumin, lysozyme and ovalbumin. The adsorption of the proteins at pH 7 and their subsequent removal by surfactant were studied by in situ ellipsometry. The degree of desorption upon dilution and the degree of elutability were compared and no relationship between these parameters could be found, which indicates that the mechanisms behind the two ways of protein removal are quite different. Further, the degree of elutability by surfactant was related to the physicochemical properties of the proteins. It was found that the size, charge, temperature of denaturation and adiabatic compressibility influenced the degree of elutability at the hydrophilic negatively charged silica surfaces for those of the model proteins that were still adsorbed after buffer rinsing. Negatively charged proteins with high denaturation temperatures, indicating high structural stability, did not adsorb on to this surface (ovalbumin) or adsorbed to a very low degree and were desorbed upon rinsing with buffer (β-lactoglobulin). All proteins adsorbed on to the hydrophobic methylated silica and the parameters that seemed to influence the degree of elutability were size and shell hydrophobicity of the proteins.