John R. P. Webster

Recent advances in the study of chemical surfaces and interfaces by specular neutron reflection

The use of specular neutron reflection to study a wide variety of problems in surface and interfacial chemistry is introduced and discussed. Recent developments in neutron reflectometry instrumentation, and their implementation in the design of the SURF reflectometer at the ISIS pulsed neutron source, are described. The design of the SURF reflectometer has been optimised for the surface chemistry of soft matter and new experimental results that exploit the novel features of this second generation neutron reflectometer are presented and discussed in the context of the opportunities for future studies that the technique and the new instrumentation presents. The examples from the broad programme in surface chemistry include surfactant and polymer adsorption at the air/liquid and liquid/solid interfaces, adsorption at the liquid/liquid interface; Langmuir–Blodgett films and liquid crystalline alignment layers, thin solid polymer films and interfaces; liquid mixtures and in situ electrochemistry.

Structural conformation of lysozyme layers at the air/water interface studied by neutron reflection

The adsorption of chicken egg white lysozyme at the air/water interface has been studied by specular neutron reflection. The variation of the total thickness of the lysozyme layer at the surface of water under varying solution conditions has been determined. The use of mixed H2O and D2O allowed the determination of the extent of immersion of the layer in water at all concentrations. The measured layer thickness combined with the globular dimensions of lysozyme suggests that the adsorbed lysozyme molecules retain their globular structure with no significant denaturation. Measurements were made over a lysozyme concentration range of 9×10-4 g dm-3 to 4 g dm-3 at pH 7 and at an ionic strength of 0.02 M. The thickness of the layer was determined by measuring neutron reflectivities in null reflecting water (NRW) where the signal is only from the adsorbed protein layer. Below 0.1 g dm-3 the surface coverage increases with bulk concentration but the thickness of the layer is constant at 30±3 A, suggesting that lysozyme is adsorbed sideways-on. As the bulk concentration increases, the layer thickness gradually increases to a value of 47±3 A2 at a bulk concentration of 1 g dm-3, suggesting that the molecules switch from sideways-on to longways-on orientations. The area per molecule at 1 g dm-3 was found to be 950±50 A2 which is close to the limit of 30×30 A2 for a saturated layer of longways-on molecules. The extent of mixing of the layer with water was determined directly by measuring reflectivity profiles in mixed H2O and D2O. A two layer model was found to be appropriate with an upper layer in air and a lower layer fully immersed in water. The thickness of the layer in air was found to vary from 15±5 Aat the lowest bulk concentration to 9±3 Aat the highest concentration studied. The results show that as the total layer thickness increases with bulk concentration the fraction of the layer immersed in water increases from 50 to 85%. At the highest concentration of 4 g dm-3 the adsorbed layer is better described by a two layer model consisting of a close packed top layer of thickness 47±3 Aand a loosely packed sublayer of 30±3 A.

Low resolution structure and dynamics of a Colicin-Receptor complex determined by neutron scattering

Background: In order to kill E. coli, colicins need to cross the bacterial outer membrane. Results: Neutron scattering data show colicin N at the protein-lipid interface of its receptor OmpF. Conclusion: Colicins can unfold and penetrate membranes via the outside wall of their receptors. Significance: The protein-lipid interface may be the route that colicins take into the cell. Proteins that translocate across cell membranes need to overcome a significant hydrophobic barrier. This is usually accomplished via specialized protein complexes, which provide a polar transmembrane pore. Exceptions to this include bacterial toxins, which insert into and cross the lipid bilayer itself. We are studying the mechanism by which large antibacterial proteins enter Escherichia coli via specific outer membrane proteins. Here we describe the use of neutron scattering to investigate the interaction of colicin N with its outer membrane receptor protein OmpF. The positions of lipids, colicin N, and OmpF were separately resolved within complex structures by the use of selective deuteration. Neutron reflectivity showed, in real time, that OmpF mediates the insertion of colicin N into lipid monolayers. This data were complemented by Brewster Angle Microscopy images, which showed a lateral association of OmpF in the presence of colicin N. Small angle neutron scattering experiments then defined the three-dimensional structure of the colicin N-OmpF complex. This revealed that colicin N unfolds and binds to the OmpF-lipid interface. The implications of this unfolding step for colicin translocation across membranes are discussed.

The effects of solvency on the structure of an adsorbed polymer layer and dispersion stability

Abstract Small-angle neutron scattering (SANS) and photon correlation spectroscopy (PCS) have been used to determine the volume fraction profiles, adsorbed amounts and hydrodynamic thickness of poly (vinyl alcohol-co-acetate) and poly (ethylene oxide) adsorbed on polystyrene latex from water. The effects of solvency on the structure of the adsorbed layer and the stability of the dispersions have been investigated by varying temperature in the presence of electrolyte. The results show that the thickness of the layer contracts on approach to flocculation and that the volume fraction of segments near the interface increases together with a concomitant increase in the adsorbed amount. The profiles are used to calculate the steric barrier due to interpenetration and the results are discussed in terms of dispersion stability.

Neutron Reflectivity Study of the Structure of pH-Responsive Polymer Brushes Grown from a Macroinitiator at the Sapphire-Water Interface

Poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) brushes have been grown by surface-initiated atom transfer radical polymerization (SI-ATRP) from a polyanionic macroinitiator adsorbed at the sapphire-water interface, and neutron reflectivity has been used to characterize the structures and pH response of the brushes. The polymer brushes are well-described by Gaussian density profiles with an additional thin, dense layer close to the solid-liquid interface for the thicker brushes at pH 7 and 9, which produces a spike in the density profile. The spike in the distribution accounts for less than 5% of the polymer and disappears as the brushes swell at pH 3. The observed swelling behavior has been used in combination with the predictions of scaling theory and previous experimental measurements to determine the grafted density of PDMAEMA chains.

An Accurate In Vitro Model of the E. coli Envelope.

Gram-negative bacteria are an increasingly serious source of antibiotic-resistant infections, partly owing to their characteristic protective envelope. This complex, 20 nm thick barrier includes a highly impermeable, asymmetric bilayer outer membrane (OM), which plays a pivotal role in resisting antibacterial chemotherapy. Nevertheless, the OM molecular structure and its dynamics are poorly understood because the structure is difficult to recreate or study in vitro. The successful formation and characterization of a fully asymmetric model envelope using Langmuir–Blodgett and Langmuir–Schaefer methods is now reported. Neutron reflectivity and isotopic labeling confirmed the expected structure and asymmetry and showed that experiments with antibacterial proteins reproduced published in vivo behavior. By closely recreating natural OM behavior, this model provides a much needed robust system for antibiotic development.

Competitive adsorption of lysozyme and C12E5 at the air/liquid interface

We have studied the adsorption of lysozyme and pentaethylene glycol monododecyl ether (C12E5) at the air/water interface using neutron reflection and surface tension measurements. The effect of C12E5 concentration was examined at three fixed lysozyme concentrations of 0.01, 1 and 4 g dm−3. The surface tension showed little variation with the addition of C12E5 over the low surfactant concentration region, but with the increase of C12E5 concentration, the surface tension gradually became identical to that corresponding to pure C12E5. These results suggest a progressive replacement of lysozyme by C12E5 and that the observed surface event is dominated by competitive adsorption. The parallel neutron measurements showed that, at low surfactant concentration, the surface was predominantly occupied by lysozyme. At intermediate C12E5 concentrations, the surface layer consisted of both lysozyme and C12E5, with the C12E5 eventually completely replacing the adsorbed lysozyme as the surfactant concentration was further increased. While the neutron results confirm the inference from surface tension measurement, structural analysis clearly showed the partial breakdown of the globular structure of lysozyme induced by the nonionic surfactant. Furthermore, neutron data showed that the adsorbed C12E5 molecules are present at the top surface layer only, suggesting no preferential association or binding between the surfactant and any immersed protein fragments at the interface.

Interfacial dynamic adsorption and structure of molecular layers of peptide surfactants.

Short peptide surfactants have recently emerged as a new class of amphiphiles, with tremendous potential in improving surface biocompatibility and mediating interfacial DNA immobilization. To establish their basic interfacial adsorption properties, cationic peptide surfactants V(m)K(n) have been studied by combining the measurements of spectroscopic ellipsometry (SE), neutron reflection (NR) and atomic force microscopy (AFM). Our results showed that changes in peptide structure, concentration, solution pH and ionic strength all affected their interfacial assembly. Increases in m and decreases in n reduced the critical aggregation concentration (CAC), but increased the amount of adsorption, showing the strong influence of the amphiphilic balance between hydrophilic and hydrophobic moieties. While the surface adsorbed amount increased with time and peptide concentration, an increase in ionic strength decreased peptide adsorption due to surface charge neutralization. Changes in solution pH did not affect the equilibrium surface adsorbed amount on the weakly negative SiO(2) surface, but did alter the adsorption dynamics. Neutron reflection revealed that V(6)K readily formed a bilayer structure of 35 A thickness at the interface, with the main part of the V(6) fragments being packed back-to-back to form a 15 A hydrophobic core and the two outer K regions being incorporated with a minor amount of V fragments forming the headgroup layers of 9 A each. AFM imaging revealed a sheet-like membrane structure incorporating defects of holes but the thicknesses probed by AFM were consistent with neutron reflection. It was demonstrated that the V(6)K peptide bilayer was effective for immobilization of DNA. The amount of DNA immobilized followed approximate 1:1 charge neutralization between the outer leaf peptide sublayer and the negatively charged DNA.

Interfacial assembly of cationic peptide surfactants

Short surfactant-like peptides bearing the molecular architecture of hydrophilic and hydrophobic moieties are attractive for a wide range of biological and technological applications. However, to realize the benefits offered by them we need to understand the basic physiochemical properties arising from their interfacial self-assembly and solution aggregation. In this paper, we use a combined approach of spectroscopic ellipsometry (SE), atomic force microscopy (AFM) and neutron reflection (NR) to characterize the interfacial assembly of cationic peptide V6K2 at the hydrophilic silica–water interface. The SE measurement revealed that the peptide dynamic adsorption was characterised by a fast initial process within the first 3–5 min, followed by a slow molecular reorientation within the next 30–40 min. AFM imaging revealed the formation of a dense peptide layer incorporating defects and some large vesicles. This interfacial structural feature from AFM offered a useful starting point for fitting the neutron reflectivity profiles measured. At 0.05 wt% V6K2, the reflectivity profiles were well fitted using a two-layer model with a dense 40 A inner layer containing about 50% peptide close to the oxide surface and a loose 40 A outer layer containing some 8% peptide on the solution side. With the help of partial deuteration to the peptide (hV6hK2 and dV6hK2 and the solvent isotopic contrasts (D2O, H2O), we found that the densely packed peptide region was comprised of a sandwiched peptide bilayer with their hydrophobic tails (V6) attracted to each other and the cationic head groups (K2) projected towards the oxide surface and the bulk water. This peptide bilayer structure is similar to that formed by conventional cationic surfactants when adsorbed at the same anionic SiO2–water interface, indicating the dominant effect of hydrophobic interaction. This study has demonstrated that the combined measurements provide a useful account of structure and dynamics of interfacial peptide self-assembly.

Surface modification of polyethylene with multi-end-functional polyethylene additives.

We have prepared and characterized a series of multifluorocarbon end-functional polyethylene additives, which when blended with polyethylene matrices increase surface hydrophobicity and lipophobicity. Water contact angles of >112° were observed on spin-cast blended film surfaces containing less than 1% fluorocarbon in the bulk, compared to ~98° in the absence of any additive. Crystallinity in these films gives rise to surface roughness that is an order of magnitude greater than is typical for amorphous spin-cast films but is too little to give rise to superhydrophobicity. X-ray photoelectron spectroscopy (XPS) confirms the enrichment of the multifluorocarbon additives at the air surface by up to 80 times the bulk concentration. Ion beam analysis was used to quantify the surface excess of the additives as a function of composition, functionality, and molecular weight of either blend component. In some cases, an excess of the additives was also found at the substrate interface, indicating phase separation into self-stratified layers. The combination of neutron reflectometry and ion beam analysis allowed the surface excess to be quantified above and below the melting point of the blended films. In these films, where the melting temperatures of the additive and matrix components are relatively similar (within 15 °C), the surface excess is almost independent of whether the blended film is semicrystalline or molten, suggesting that the additive undergoes cocrystallization with the matrix when the blended films are allowed to cool below the melting point.