Soft matter probed by nonlinear scattering: self-assembly, interfaces, hydration, and long-range order

All organisms are made to a large extent of soft matter - macromolecules such as proteins and polysaccharides, or assemblies of small molecules such as lipids embedded in an aqueous environment. Understanding the role of order in soft matter presents a challenge for experimentalists because such systems are neither completely disordered nor perfectly crystalline. This is especially the case for the aqueous environment itself whose order fluctuates on femto- and pico-second time scales. In this thesis, we attempt to elucidate the presence, extent, changes, and implications of orientational order in several soft matter systems: from polyelectrolyte solutions to lipid membrane interfaces and supramolecular structures. To perform this task, we employ nonlinear optical techniques: second-harmonic and sum-frequency scattering. In the first two chapters, we explore the structure of water in aqueous polyelectrolyte solutions. Polyelectrolytes of high molecular mass are shown to induce orientational order in water that spans hundreds of nanometers. Using an adapted model of second-harmonic scattering from charged particles, we can explain the observed ordering in terms of weakly screened long-range electrostatic interactions that perturb the structure of water via charge-dipole interactions. Upon exchanging ordinary (light) water for heavy water, we observe a significant nuclear quantum effect in the induced order, indicating that bulk water-water correlations are enhanced as well by polyelectrolytes. In the following chapter, we exploit the same nuclear quantum effect in polyelectrolyte solutions to show that the polyelectrolyte-induced orientational order correlates with an anomalous increase in the reduced viscosity. We propose that the viscosity anomaly arises from enhanced order in a solvent which hinders the viscous flow. This observation represents a rare direct link between a microscopic property and a macroscopic observable. In the last two chapters, we change focus from homogeneous solutions to dispersions of liposomes which serve as models of biological membranes. In Chapter 5 we explore the binding of the protein alpha-synuclein to anionic vesicles from the perspective of interfacial water. We show that the adsorption of the protein to a vesicle decreases the order in the interfacial water by 30% but does not affect the surface nor the zeta potential. We propose that the changes in the structure of interfacial water can serve as a label-free probe of protein binding. In the last chapter, we investigate the supramolecular chemistry of inclusion complexes of methyl-Î²-cyclodextrin with various lipids. By combining multiple nonlinear scattering techniques, we show that the complexes self-assemble into micrometer-long directed fibers. Based on measurements of different lipids, we propose that the self-assembly process is driven by hydrogen bonding between the lipid headgroup and cyclodextrin. Lastly, we show that the long-range order of the self-assembled structure is transmitted into the structure of the hydrating water.

Molecular ordering effects in bulk and at lipid/water nanoscopic interfaces: A story of water, light, and order

Nathan Dupertuis

Water is ubiquitous on Earth, playing a critical role for a plethora of structures, processes and chemical reactions in nature. Its unique physicochemical properties originate mainly from its hydrogen bond network. Despite its recognized importance, many aspects of the structure of water as a liquid remain elusive. It is especially difficult to understand the complex nature of fluctuations and ordering in pure water, or to get a complete picture of aqueous interfaces at the molecular level. In this thesis, our aim is to improve our understanding of the underlying interactions between water and ions, macromolecules, and interfaces, especially the long range interactions. To this aim, we use two nonlinear optical techniques, second harmonic scattering (SHS) and vibrational sum frequency scattering (SFS), on liquid water as well as on model lipid membranes mimicking the cell membranes.We investigate with SHS how the orientational ordering of water molecules is affected by temperature changes and nuclear quantum effects. Scattering measurements of pure water reveal that the intermolecular correlations undergo a symmetry transformation with increasing temperature. On the other hand, with increasing temperature, aqueous electrolyte solutions exhibit a diminished influence of the combined electrostatic field on the water- water correlations. This trend is predicted qualitatively by a Debye-Hückel model, but not quantitatively due to its non-inclusion of hydrogen bonding and nuclear quantum effects. These insights facilitate the elaboration of future nuclear quantum mechanical models of water.The same interactions are further studied by a comparison of pure liquid water and pure liquid carbon tetrachloride. SHS measurements and simulations reveal that transient nanoscale voids exist in water and carbon tetrachloride. The coherent SHS emissions observed in water are generated by charge density fluctuations around the cavities. These measurements strongly hint towards non-uniformity in the structure of water, on femtosecond timescale and nanometric range.Next, we turn towards the structure of water around model lipid membranes. We explore first the three-dimensional confinement of liquid water inside liposomes of various size. By analyzing SHS patterns of zwitterionic and anionic liposomes, we observe that the effect of confinement is visible on a range of a hundred nanometers, larger compared to the previously reported ranges on the order of a nanometer or about ten nanometers. The water orientational ordering at the interface as well as the surface potential of the liposomes are retrieved with a nonlinear light scattering modeling, showing that the confinement effect is mainly attributable to the hydrogen bond network. The effect is in particular 8 times larger for light water than for heavy water.Finally, we investigate how the water structure is modified around lipid monolayers and bilayers. The hydration of lipid membranes is asymmetric with respect to charge. SHS and vibrational SFS reveal that the charge hydration asymmetry is drastically different depending on the nature of the lipid layer. It depends on a delicate balance between electrostatic and hydrogen bonding interactions. The charge hydration asymmetry is further modified by the presence of charges on the lipid headgroups on both sides of the bilayers.

EPFL2021

Lipid membrane characterization with second harmonic scattering

Cornelis Ulrich Lütgebaucks

Membranes, composed of a variety of lipids and other biomolecules, mediate signaling processes between cells and their aqueous environment. To fulfill this function, membranes can vary their composition leaflet-specific and thus alter their surface properties. To fully understand the impact of these processes on the molecular level, it is necessary to develop tools that can access the molecular properties of free-floating model membranes label-free. These tools are ideally surface-specific. In this thesis, we apply the nonlinear optical techniques second harmonic scattering (SHS) and vibrational sum-frequency scattering (SFS) together with electrokinetic measurements to label-free characterize the interfacial properties, hydration structure and surface potentials of liposomes in aqueous solutions. First, we generalize the nonlinear optical theory to describe the second-order surface response from interfaces with aqueous solutions independent of the ionic strength for reflection, transmission and scattering geometries. We demonstrate that interference effects from oriented water molecules in the bulk aqueous solution alter the probing depth and the expected second-order response at low ionic strengths. Then, we apply this theory to demonstrate that SHS patterns of liposomes and oil droplets contain all necessary information to extract the absolute surface potential of the respective particles without assuming a model for the interfacial structure. By analyzing scattering patterns that capture the orientational distribution of water around the particles, we find surface potentials of -38 mV for bare oil-droplets and -11 mV for zwitterionic liposomes in water. For anionic liposomes the surface potential varies between -150 mV and -23 mV in solutions containing different amounts of NaCl ranging from

\sim

0 mM to 10 mM. These values are remarkably different for solutions to the Gouy-Chapman model considering a fixed surface charge density. Next, we characterize the hydration and lipid asymmetries in binary mixed membranes using SHS and SFS. The liposomes exhibit hydration asymmetry between the inner and outer leaflet. The lipid number density between the inner and outer leaflet is the same, although geometrical packing arguments would suggest a different density. However, an asymmetric lipid distribution between the leaflets can be induced by fine tuning specific intermolecular interactions between the lipids. This is shown with dipalmitoylphosphoserine and dioleoylphosphocholine mixtures creating a membrane structure that allows intermolecular H-bonding between the phosphate and amine groups of the lipids. Finally, we quantify the surface properties of membranes composed of lipids containing phosphoserine and phosphocholine headgroups. Surprisingly, we find a very high degree of counterion condensation on anionic membranes in pure water: only 1 % of all lipids are ionized. This indicates a tightly packed layer of ions around the membrane that needs to be considered when modelling the interfacial structure around membranes.

EPFL2017

Label-free and charge-sensitive dynamic imaging of lipid membrane hydration on millisecond time scales

Sylvie Roke, Orly Bagunu Tarun

Biological membranes are highly dynamic and complex lipid bilayers, responsible for the fate of living cells. To achieve this function, the hydrating environment is crucial. However, membrane imaging typically neglects water, focusing on the insertion of probes, resonant responses of lipids, or the hydrophobic core. Owing to a recent improvement of second-harmonic (SH) imaging throughput by three orders of magnitude, we show here that we can use SH microscopy to follow membrane hydration of freestanding lipid bilayers on millisecond time scales. Instead of using the UV/VIS resonant response of specific membrane-inserted fluorophores to record static SH images over time scales of >1,000 s, we SH imaged symmetric and asymmetric lipid membranes, while varying the ionic strength and pH of the adjacent solutions. We show that the nonresonant SH response of water molecules aligned by charge−dipole interactions with charged lipids can be used as a label-free probe of membrane structure and dynamics. Lipid domain diffusion is imaged label-free by means of the hydration of charged domains. The orientational ordering of water is used to construct electrostatic membrane potential maps. The average membrane potential depends quadratically on an applied external bias, which is modeled by nonlinear optical theory. Spatiotemporal fluctuations on the order of 100-mV changes in the membrane potential are seen. These changes imply that membranes are very dynamic, not only in their structure but also in their membrane potential landscape. This may have important consequences for membrane function, mechanical stability, and protein/pore distributions.

2018