Cristina Stefaniu

Langmuir monolayers as models to study processes at membrane surfaces

The use of new sophisticated and highly surface sensitive techniques as synchrotron based X-ray scattering techniques and in-house infrared reflection absorption spectroscopy (IRRAS) has revolutionized the monolayer research. Not only the determination of monolayer structures but also interactions between amphiphilic monolayers at the soft air/liquid interface and molecules dissolved in the subphase are important for many areas in material and life sciences. Monolayers are convenient quasi-two-dimensional model systems. This review focuses on interactions between amphiphilic molecules in binary and ternary mixtures as well as on interfacial interactions with interesting biomolecules dissolved in the subphase. The phase state of monolayers can be easily triggered at constant temperature by increasing the packing density of the lipids by compression. Simultaneously the monolayer structure changes are followed in situ by grazing incidence X-ray diffraction or IRRAS. The interactions can be indirectly determined by the observed structure changes. Additionally, the yield of enzymatic reaction can be quantitatively determined, secondary structures of peptides and proteins can be measured and compared with those observed in bulk. In this way, the influence of a confinement on the structural properties of biomolecules can be determined. The adsorption of DNA can be quantified as well as the competing adsorption of ions at charged interfaces. The influence of modified nanoparticles on model membranes can be clearly determined. In this review, the relevance and utility of Langmuir monolayers as suitable models to study physical and chemical interactions at membrane surfaces are clearly demonstrated.

Nanotubes Protruding from Poly(allylamine hydrochloride)-Graft-Pyrene Microcapsules

Protrusion of one-dimensional nanotubes or nanorods from the poly(allylamine hydrochloride)-graft-pyrene (PAH-Py) microcapsules was discovered when the microcapsules were incubated in pH 0 and pH 2 solutions, respectively. Micelles assembled from deliberately synthesized PAH-Py polymers were also able to transform into one-dimensional structures, demonstrating the chemistry driven nature of the phenomenon. The one-dimensional nanotubes consisted of only 1-pyrenecarboxaldehyde with ordered π-π stacking, and showed a helical structure and anisotropic property. The hydrolysis of Schiff base and its rate at different pH values (10 times slower at pH 0 than at pH 2) played a key role in determining the final nanostructures, and the linear PAH directed the regular building up process especially for the nanotubes. Hollow capsules budded with nanotubes or nanorods mimicking the cellular protrusion of filopodia were successfully prepared by tuning the incubation pH and time. These results and the proposed mechanism open new opportunities for design of novel micronanostructures and materials for nanoscience, and biological and other advanced technologies.

Functional carbon nanosheets prepared from hexayne amphiphile monolayers at room temperature

Carbon nanostructures that feature two-dimensional extended nanosheets are important components for technological applications such as high-performance composites, lithium-ion storage, photovoltaics and nanoelectronics. Chemical functionalization would render such structures better processable and more suited for tailored applications, but typically this is precluded by the high temperatures needed to prepare the nanosheets. Here, we report direct access to functional carbon nanosheets of uniform thickness at room temperature. We used amphiphiles that contain hexayne segments as metastable carbon precursors and self-assembled these into ordered monolayers at the air/water interface. Subsequent carbonization by ultraviolet irradiation in ambient conditions resulted in the quantitative carbonization of the hexayne sublayer. Carbon nanosheets prepared in this way retained their surface functionalization and featured an sp(2)-rich amorphous carbon structure comparable to that usually obtained on annealing above 800 °C. Moreover, they exhibited a molecularly defined thickness of 1.9 nm, were mechanically self-supporting over several micrometres and had macroscopic lateral dimensions on the order of centimetres.

Subgel Phase Structure in Monolayers of Glycosylphosphatidylinositol Glycolipids

Glycosylphosphatidylinositols (GPIs) are complex glycolipids that are commonly found in eukaryotic cells as a posttranslational modification of proteins or as free GPIs displayed on the cell surface. Although their main function is to anchor the attached protein (AP) to the cell membrane, the conserved nature of the complex pseudopentasaccharide core of GPIs (Figure 1A) suggests biological roles beyond simple physical anchoring. The GPI-APs and free GPIs show non-Brownian density fluctuations on cell surfaces, and associate with membrane microdomains, known as lipid rafts. While protein–protein interactions may contribute to the observed clustering of GPI-APs on the cell membranes, such interactions cannot be responsible for clustering of free GPIs and their association with lipid rafts. It is, therefore, conceivable that the interactions between GPI molecules and their interactions with other membrane associated species play a role in the heterogeneous distribution of GPIs in cell membranes. Some proteins that do not form clusters in the cytosol do so on the cell surface when expressed as a GPI-AP construct. Clustering of the free GPIs has also been observed on the cell surface of some parasitic protozoa. GPIs and GPI-APs are involved in a variety of biological processes, such as signal transduction, protein sorting and transport, intermembrane transfers, parasitic infections, and pathophysiology of prion diseases. Insights into the behavior of GPIs and GPI-APs in cell membranes could contribute to the understanding of the roles GPIs play in these processes. Monolayers formed with GPI-APs have been investigated, but the limited scope did not provide fundamental insights into the behavior of GPIs in cellular membranes. We aim to elucidate the structural characteristics and conformational behavior of GPIs in well-defined membrane models. Herein, we report the unprecedented ordering in two-dimensional monolayers of GlcNa1!6myoIno-1-phosphodistearylglycerol fragment of GPIs (1; Figure 1 A) observed by grazing-incidence X-ray diffraction (GIXD). Synthetic GPI mimic 1 represents a minimal fragment that might adequately emulate GPI behavior. While 1 lacks the trimannose portion present in all known GPIs, the glucosaminephosphoinositol moiety features both the amino and phosphate groups largely seen as major determinants of the behavior of the charged head groups. Compound 1 was studied in 2D films confined at the air/ liquid interface that are easy-to-handle model systems of one membrane leaflet (Figure 1A). The reduced dimensionality of the system is advantageous to better understand the role of different interactions for structure formation. To gain first insights into the molecular interactions and the possible phase transitions of compound 1 in monolayers, surface pressure/ molecular area isotherms were recorded on different subphases (Figure 1B). The water subphase was used in the presence and absence of Ca ions to test for differences that Figure 1. A) Conserved GPI structure and the structure of GlcNa1! 6myoIno-1-phosphodistearoylglycerol fragment 1. B) Surface pressure/ molecular area (p/A) isotherms on the surface of water (c), PBS (10 mm, pH 7.4, 150 mm NaCl; a), and pH 2 solution (0.01m HCl; b). C) GIXD patterns of monolayers of 1 on PBS at 20 8C (2 mN m 1 c, 30 mN m 1 a).

Biocompatible Magnetite Nanoparticles Trapped at the Air/Water Interface

In the last decade, iron oxide nanoparticles (NPs)—known as super-paramagnetic iron oxide nanoparticles (SPION)—have received significant attention due to their applicability in a variety of domains. In the biomedical field, iron oxide nanoparticles are very promising as drug-delivery systems, magneticresonance-imaging contrast enhancers (clinical diagnosis), inflammation-responsive or anti-cancer agents, or for labeling and cell separation. In addition to their potential medical applications, magnetic nanoparticles are of great interest in materials science, for the development of magnetic data-recording media and nanocomposite permanent magnets, as well as in catalysis and colloid chemistry, where ferrofluids represent a topic of high interest. This study is focused on Fe3O4 NPs grafted with the thermosensitive and biocompatible copolymer MEO2MA/OEGMA, [10] whose interfacial behavior is yet unknown. Many papers have been dedicated to the formation, organization, and stability of Langmuir films of iron oxide (Fe2O3 or Fe3O4) NPs and their Langmuir–Blodgett transfer. The system studied herein is completely different from the iron oxide NPs already studied at the air/water interface due to the unique properties of the polymer used. A monodisperse population of uncharged Fe3O4 cores with a diameter of 6.4 nm, which are grafted with catechol-terminated copolymers of 2-(2-methoxyethoxy) ethyl methacrylate (MEO2 MA) and oligo(ethylene glycol) methacrylate (OEGMA), has been investigated (Figure 1). Due to the presence of oligo(ethylene glycol) side groups on the surfaces, the Fe3O4@MEO2MA90-co-OEGMA10 NPs (90 and 10 represent the molar fractions of MEO2MA and OEGMA, respectively) can be dispersed in water, exhibiting a high colloidal stability against salt and, at the same time, they can be well dispersed in organic solvents such as chloroform, ethanol or toluene. This specific copolymer in water (no salt) is hydrophilic and becomes hydrophobic at temperatures above 40 8C. The experimental details are presented in the Supporting Information. Since the polymer and the polymer-dressed NPs behave exactly in the same way, showing that the polymer dictates the surface activity, we will describe here mainly the results obtained with the polymer-dressed NPs because this system can be investigated with a larger number of methods giving complementary information. The surface activity of the Fe3O4@MEO2MA90-co-OEGMA10 NPs is based on the amphiphilic character of the copolymer shell. This (oligo ethylene glycol) methyl ether methacrylate polymer has a graft structure (Figure 1) composed of an apolar carbon–carbon backbone which leads to a competitive hydrophobic effect and multiple oligo(ethylene glycol) side chains of which ether oxygen atoms form stabilizing hydrogen bonds with water. Moreover, the ethylene oxide motif can adopt a configuration with the oxygen atoms on one side of the molecule and with the two methylene groups on the other, thus giving the molecule both a hydrophilic and a hydrophobic surface. The hydrophobicity can be tuned by changing the molar fraction of the two monomers. The polymer can be dispersed both in water and in chloroform; therefore, the corresponding films have been prepared either by adsorption from aqueous bulk solution or by spreading from a chloroform solution at the interface. Using, for example, a NP bulk concentration of 1.5 10 3 mg mL , a constant surface pressure value of approximately 23 mN m 1 is reached after 20 h (Figure 2 A). Compression isotherms of Langmuir layers formed by spreading certain amounts of NPs are shown in Figure 2 B. By compression, the surface pressure increases continuously to 25 mN m 1 (critical pressure pc of the polymer as well as the NP film). During further compression, a plateau region appears at which the surface pressure increases only slightly up to a maximum value of 27 mN m . It is very important to highlight that no hysteresis of the compression/ expansion isotherms is observed when the interfacial film is compressed to surface pressures below the critical pressure of the Langmuir layer (Figure 2 C), suggesting that no loss of material from the interface occurs. Based on this observation, we can calculate the interfacial concentration of NPs corresponding to the critical pressure. The critical concentration amounts to (7.7 0.6) 10 4 mg cm . This value is in good agreement with the NP interfacial concentration of 8.2 10 4 mg cm , which can be calFigure 1. Schematic representation of the Fe3O4 @ MEO2MA90-co-OEGMA10 NPs

Langmuir and Gibbs Magnetite NP Layers at the Air/Water Interface

The interfacial properties of Fe(3)O(4)@MEO(2)MA(90)-co-OEGMA(10) NPs, recently developed and described as promising nanotools for biomedical applications, have been investigated at the air/water interface. These Fe(3)O(4) NPs, capped with catechol-terminated random copolymer brushes of 2-(2-methoxyethoxy) ethyl methacrylate (MEO(2)MA) and oligo(ethylene glycol) methacrylate (OEGMA), with molar fractions of 90% and 10%, respectively, proved to be surface active. Surface tension measurements of aqueous dispersions of the NPs showed that the adsorption of the NPs at the air/water interface is time- and concentration-dependent. These NPs do not behave as classical amphiphiles. Once adsorbed at the air/water interface, they do not exchange with NPs in bulk, but they are trapped at the interface. This means that all NPs from the bulk adsorb to the interface until reaching maximum coverage of the interface, which corresponds to values between 6 × 10(-4) and 8 × 10(-4) mg/cm(2) and a critical equilibrium surface tension of ∼47 mN/m. Moreover, Langmuir layers of Fe(3)O(4)@MEO(2)MA(90)-co-OEGMA(10) NPs have been investigated by measuring surface pressure-area compression-expansion isotherms and in situ X-ray fluorescence spectra. The compression-expansion isotherms showed a plateau region above a critical surface pressure of ∼25 mN/m and a pronounced hysteresis. By using a special one-barrier Langmuir trough equipped with two surface pressure microbalances, we have shown that the NPs are squeezed out from the interface into the aqueous subphase, and they readsorb on the other side of the barrier. The results have been supported by TEM as well as AFM experiments of transferred Langmuir-Schaefer films on solid supports. This study shows the ability of Fe(3)O(4)@MEO(2)MA(90)-co-OEGMA(10) NPs to transfer from hydrophilic media (an aqueous solution) to the hydrophobic/hydrophilic interface (air/water interface) and back to the hydrophilic media. This behavior is very promising, opening studies of their ability to cross biological membranes.

Grazing incidence X-ray diffraction studies of condensed double-chain phospholipid monolayers formed at the soft air/water interface.

The use of highly brilliant synchrotron light sources in the middle of the 1980s for X-ray diffraction has revolutionized the research of condensed monolayers. Since then, monolayers gained popularity as convenient quasi two-dimensional model systems widely used in biophysics and material science. This review focuses on structures observed in one-component phospholipid monolayers used as simplified two-dimensional models of biological membranes. In a monolayer system the phase transitions can be easily triggered at constant temperature by increasing the packing density of the lipids by compression. Simultaneously the monolayer structure changes are followed in situ by grazing incidence X-ray diffraction. Competing interactions between the different parts of the molecule are responsible for the different monolayer structures. These forces can be modified by chemical variations of the hydrophobic chain region, of the hydrophilic head group region or of the interfacial region between chains and head groups. Modifications of monolayer structures triggered by changes of the chemical structure of double-chain phospholipids are highlighted in this paper.

Monolayer characteristics of a long-chain N,O-diacyl substituted ethanolamine at the air/water interface.

The N- and/or O-acylation of amphiphilic ethanolamine attracts particular attention because of its interesting biological, pharmaceutical, and medicinal properties. Tetradecanoic acid-2-[(1-oxotetradecyl)amino]ethyl ester (TAOAE) as the selected N,O-diacyl derivative of ethanolamine has been synthesized in order to obtain first information about its main interfacial characteristics, such as the surface pressure-area (π-A) isotherms, the morphology of the condensed phase domains, the lattice structure of the condensed phase, and information about the existence of interfacial hydrogen bonds (-NH···O═C-). The π-A isotherms of TAOAE, similar to those of the most usual monolayers of amphiphiles, show a sharp break point (A(c)) indicating the first-order phase transition from the fluid (liquid-expanded (LE), gaseous (G)) to the condensed (liquid-condensed (LC)) phase. On the mesoscopic scale, the dendritic domains homogeneously reflecting suggest an orientation of the alkyl chains perpendicular to the aqueous surface. The grazing incidence X-ray diffraction (GIXD) studies reveal hexagonal packing of the TAOAE molecules oriented perpendicular to the surface in an LS phase. The existence of a hydrogen-bonding network in the monolayer is supported by infrared reflection absorption spectroscopy (IRRAS) experiments.

Bilayer Properties of 1,3-Diamidophospholipids

A series of 1,3-diamido phosphocholines was synthesized, and their potential to form stable bilayers was investigated. Large and giant unilamellar vesicles produced from these new lipids form a wide variety of faceted liposomes. Factors such as cooling rates and the careful choice of the liposome preparation method influence the formation of facets. Interdigitation was hypothesized as a main factor for the stabilization of facets and effectively monitored by small-angle X-ray scattering measurements.

Rigid Urea and Self-Healing Thiourea Ethanolamine Monolayers

A series of long-tail alkyl ethanolamine analogs containing amide-, urea-, and thiourea moieties was synthesized and the behavior of the corresponding monolayers was assessed on the Langmuir-Pockels trough combined with grazing incidence X-ray diffraction experiments and complemented by computer simulations. All compounds form stable monolayers at the soft air/water interface. The phase behavior is dominated by strong intermolecular headgroup hydrogen bond networks. While the amide analog forms well-defined monolayer structures, the stronger hydrogen bonds in the urea analogs lead to the formation of small three-dimensional crystallites already during spreading due to concentration fluctuations. The hydrogen bonds in the thiourea case form a two-dimensional network, which ruptures temporarily during compression and is recovered in a self-healing process, while in the urea clusters the hydrogen bonds form a more planar framework with gliding planes keeping the structure intact during compression. Because the thiourea analogs are able to self-heal after rupture, such compounds could have interesting properties as tight, ordered, and self-healing monolayers.