Hydration of Lysozyme Studied by Raman Spectroscopy

Hydration plays a fundamental role in maintaining the three-dimensional structure and function of proteins. In this study, Raman spectroscopy was used to probe the hydration induced structural changes at various sites of lysozyme under isothermal conditions in the range of water contents from 0 to 44 wt %. Raman hydration curves were constructed from detailed analysis of marker bands. Transition inflection points (<i>w</i>_m) and onsets determined from the hydration curves have shown that structural changes start at 7–10 and end at about 35 wt % water. The onset of structural changes coincides with the onset of the broad glass transition earlier observed in this system. The increase of α-helix content starts at very low concentrations of water with <i>w</i>_m = 12 wt %. Monitoring the development of importance for enzymatic action hydrophobic clusters has revealed <i>w</i>_m = 15 wt % and completion of the process at 25 wt %. The parameters of 621 cm^–1 (Phe) and 1448 cm^–1 (CH₂ bending) modes were found to be sensitive to hydration, suggesting changes in organization of water molecules near the protein surface. The native structure of lysozyme was achieved at 35 wt % water where its content is high enough for filling the space between lysozyme molecules

Adsorption of Lipid Liquid Crystalline Nanoparticles on Cationic, Hydrophilic, and Hydrophobic Surfaces

Investigation of nonlamellar nanoparticles formed by dispersion of self-assembled lipid liquid crystalline phases is stimulated by their many potential applications in science and technology; resulting from their unique solubilizing, encapsulating, and space-dividing nature. Understanding the interfacial behavior of lipid liquid crystalline nanoparticles (LCNPs) at surfaces can facilitate the exploitation of such systems for a number of potentially interesting uses, including preparation of functional surface coatings and uses as carriers of biologically active substances. We have studied the adsorption of LCNP, based on phosphatidylcholine/glycerol dioleate and Polysorbate 80 as stabilizers, at different model surfaces by use of in situ ellipsometry. The technique allows time-resolved monitoring of the layer thickness and the amount adsorbed, thereby providing insights into the restructuring of the lipid nanoparticle upon adsorption. The effects of solvent condition, electrolyte concentration, particle size, and surface chemistry on adsorbed layer properties were investigated. Furthermore, the internal structures of the particles were investigated by cryo-transmission electron microscopy and small angle X-ray diffraction on the corresponding liquid crystalline phases in excess water. LCNPs are shown to form well-defined layers at the solid–liquid interface with a structure and coverage that are determined by the interplay between the self-assembly properties of the lipids and lipid surface interactions, respectively. At the hydrophobic surface, hydrophobic interaction results in a structural transition from the original LCNP morphology to a monolayer structure at the interface. In contrast, at cationic and hydrophilic surfaces, relaxation is a relatively slow process, resulting in much thicker adsorbed layers, with thickness and adsorption behavior that to a greater extent reflect the original bulk LCNP properties

Adsorption of Lipid Liquid Crystalline Nanoparticles: Effects of Particle Composition, Internal Structure, and Phase Behavior

Controlling the interfacial behavior and properties of lipid liquid crystalline nanoparticles (LCNPs) at surfaces is essential for their application for preparing functional surface coatings as well as understanding some aspects of their properties as drug delivery vehicles. Here we have studied a LCNP system formed by mixing soy phosphatidylcholine (SPC), forming liquid crystalline lamellar structures in excess water, and glycerol dioleate (GDO), forming reversed structures, dispersed into nanoparticle with the surfactant polysorbate 80 (P80) as stabilizer. LCNP particle properties were controlled by using different ratios of the lipid building blocks as well as different concentrations of the surfactant P80. The LCNP size, internal structure, morphology, and charge were characterized by dynamic light scattering (DLS), synchrotron small-ange X-ray scattering (SAXS), cryo-transmission electron microscopy (cryo-TEM), and zeta potential measurements, respectively. With increasing SPC to GDO ratio in the interval from 35:65 to 60:40, the bulk lipid phase structure goes from reversed cubic micellar phase with <i>Fd</i>3<i>m</i> space group to reversed hexagonal phase. Adding P80 results in a successive shift toward more disorganized lamellar type of structures. This is also seen from cryo-TEM images for the LCNPs, where higher P80 ratios results in more extended lamellar layers surrounding the inner, more dense, lipid-rich particle core with nonlamellar structure. When put in contact with a solid silica surface, the LCNPs adsorb to form multilayer structures with a surface excess and thickness values that increase strongly with the content of P80 and decreases with increasing SPC:GDO ratio. This is reflected in both the adsorption rate and steady-state values, indicating that the driving force for adsorption is largely governed by attractive interactions between poly­(ethylene oxide) (PEO) units of the P80 stabilizer and the silica surface. On cationic surface, i.e., silica modified with 3-aminopropltriethoxysilane (APTES), the slightly negatively charged LCNPs give rise to a very significant adsorption, which is relatively independent of LCNP composition. Finally, the dynamic thickness measurements indicate that direct adsorption of intact particles occurred on the cationic surface, while a slow buildup of the layer thickness with time is seen for the weakly interacting systems

Complementary X-ray, ellipsometry and neutron data from Non-lamellar lipid assembly at interfaces: controlling layer structure by responsive nanogel particles

Figure S1. Effect of temperature on the lattice parameter, a, of a cubic phase composed of GMO-50/DGMO (40/60 weight ratio) alone (open markers) or containing 10 wt% nanogel (black solid markers). Figure S2. SAXS data for the formulations of nanogels dispersed in lipids (85 wt % lipids composed of GMO-50/DGMO at a 40/60 weight ratio) with 15 wt % ethanol at 25 °C and 40 °C. Figure S3. Spectroscopic ellipsometry parameters, Δ (pink circles) and ψ (black triangles), as a function of wavelength for a film of GMO-50:DGMO (40:60 wt%) on silica. Figure S4. (A) Schematic representation of magnetic contrast surfaces used in this study. Figure S5. Neutron reflectivity data for a spin coated film of GMO-50:DGMO (60:40 wt% ratio) containing nanogel (10wt%) at (A) 25°C and (B) 40°C. Figure S6. Diffraction pattern extracted from the off-specular neutron reflectivity patterns in Figure 6 of lipid-only and lipid-nanogel layers at 25 °C and 40 °C