Thermal Density Fluctuations and Polymorphic Phase Transitions of Ethane (C2D6) in the Gas/Liquid and Supercritical States.

The phase behavior of the liquid C2D6 below and above the critical point was investigated using small-angle neutron scattering (SANS) in temperature and pressure ranges from 10 to 45 °C and 20 to 126 bar, respectively. The scattering of thermal fluctuations of the molecular density was determined and thus the gas-liquid and Widom lines. At the same time, we observed additional scattering of droplets of more densely packed C2D6 molecules above the gas-liquid line and in the supercritical fluid regime from just below the critical point for all temperatures at about ΔP = 10 bar above the Widom line. This line is interpreted as the Frenkel line. These results are consistent with our previous studies on CO2 and thus indicate a universal phase behavior for monomolecular liquids below and above the critical point. The interpretation of the Frenkel line as the lower limit of a polymorphic phase transition is in contrast to the usual interpretation as the limit of a dynamic process. The correlation lengths (ξ) of the thermal density fluctuations at the critical point and at the Widom line are determined between 20 and 35 Å and thus in the range of the droplet radius between 60 and 80 Å. These long-range fluctuations appear to suppress the formation of droplets, which can only form at about 10 bar above the critical point and the Widom line when ξ becomes smaller than 10 Å.

Small-Molecular-Weight Additives Modulate Calcification by Interacting with Prenucleation Clusters on the Molecular Level.

Small-molecular-weight (MW) additives can strongly impact amorphous calcium carbonate (ACC), playing an elusive role in biogenic, geologic, and industrial calcification. Here, we present molecular mechanisms by which these additives regulate stability and composition of both CaCO3 solutions and solid ACC. Potent antiscalants inhibit ACC precipitation by interacting with prenucleation clusters (PNCs); they specifically trigger and integrate into PNCs or feed PNC growth actively. Only PNC-interacting additives are traceable in ACC, considerably stabilizing it against crystallization. The selective incorporation of potent additives in PNCs is a reliable chemical label that provides conclusive chemical evidence that ACC is a molecular PNC-derived precipitate. Our results reveal additive-cluster interactions beyond established mechanistic conceptions. They reassess the role of small-MW molecules in crystallization and biomineralization while breaking grounds for new sustainable antiscalants.

The nano- and meso-scale structure of amorphous calcium carbonate.

Understanding the underlying processes of biomineralization is crucial to a range of disciplines allowing us to quantify the effects of climate change on marine organisms, decipher the details of paleoclimate records and advance the development of biomimetic materials. Many biological minerals form via intermediate amorphous phases, which are hard to characterize due to their transient nature and a lack of long-range order. Here, using Monte Carlo simulations constrained by X-ray and neutron scattering data together with model building, we demonstrate a method for determining the structure of these intermediates with a study of amorphous calcium carbonate (ACC) which is a precursor in the bio-formation of crystalline calcium carbonates. We find that ACC consists of highly ordered anhydrous nano-domains of approx. 2 nm that can be described as nanocrystalline. These nano-domains are held together by an interstitial net-like matrix of water molecules which generate, on the mesoscale, a heterogeneous and gel-like structure of ACC. We probed the structural stability and dynamics of our model on the nanosecond timescale by molecular dynamics simulations. These simulations revealed a gel-like and glassy nature of ACC due to the water molecules and carbonate ions in the interstitial matrix featuring pronounced orientational and translational flexibility. This allows for viscous mobility with diffusion constants four to five orders of magnitude lower than those observed in solutions. Small and ultra-small angle neutron scattering indicates a hierarchically-ordered organization of ACC across length scales that allow us, based on our nano-domain model, to build a comprehensive picture of ACC formation by cluster assembly from solution. This contribution provides a new atomic-scale understanding of ACC and provides a framework for the general exploration of biomineralization and biomimetic processes.

Multiple scattering and resolution effects in small-angle neutron scattering experiments calculated and corrected by the software package MuScatt.

This article deals with multiple scattering effects that are important for the method of small-angle neutron scattering (SANS). It considers three channels for the coherent elastic, the incoherent elastic and the incoherent inelastic scattering processes. The first channel contains the desired information on the experiment. Its multiple scattering effects can be desmeared, as shown in the later sections of the article. The other two channels display a nearly constant background as a function of the scattering angle. The incoherent elastic scattering is treated by the theory of Chandrasekhar, allowing for multiple scattering even at large scattering angles. The transfer to a single representative thermalized wavelength by the inelastic scattering - as a simplification - is assumed to happen by a single scattering event. Once the transition to this altered wavelength has happened, further incoherent multiple scattering is considered. The first part of the paper deals with the multiple scattering effects of light water. In the later part of the article, deconvolution algorithms for multiple scattering and instrumental resolution of the elastic coherent signal as implemented in the program MuScatt are described. All of these considerations are interesting for both reactor-based instruments with velocity selectors and time-of-flight SANS instruments and may improve the reliability of the data treatment.

Silica Fouling in Reverse Osmosis Systems-Operando Small-Angle Neutron Scattering Studies.

We present operando small-angle neutron scattering (SANS) experiments on silica fouling at two reverse osmose (RO) membranes under almost realistic conditions of practiced RO desalination technique. To its realization, two cells were designed for pressure fields and tangential feed cross-flows up to 50 bar and 36 L/h, one cell equipped with the membrane and the other one as an empty cell to measure the feed solution in parallel far from the membrane. We studied several aqueous silica dispersions combining the parameters of colloidal radius, volume fraction, and ionic strength. A relevant result is the observation of Bragg diffraction as part of the SANS scattering pattern, representing a crystalline cake layer of simple cubic lattice structure. Other relevant parameters are silica colloidal size and volume fraction far from and above the membrane, as well as the lattice parameter of the silica cake layer, its volume fraction, thickness, and porosity in comparison with the corresponding permeate flux. The experiments show that the formation of cake layer depends to a large extent on colloidal size, ionic strength and cross-flow. Cake layer formation proved to be a reversible process, which could be dissolved at larger cross-flow. Only in one case we observed an irreversible cake layer formation showing the characteristics of an unstable phase transition. We likewise observed enhanced silica concentration and/or cake formation above the membrane, giving indication of a first order liquid-solid phase transformation.

MATSAS: a small-angle scattering computing tool for porous systems.

MATSAS is a script-based MATLAB program for analysis of X-ray and neutron small-angle scattering (SAS) data obtained from various facilities. The program has primarily been developed for sedimentary rock samples but is equally applicable to other porous media. MATSAS imports raw SAS data from .xls(x) or .csv files, combines small-angle and very small angle scattering data, subtracts the sample background, and displays the processed scattering curves in log-log plots. MATSAS uses the polydisperse spherical (PDSP) model to obtain structural information on the scatterers (scattering objects); for a porous system, the results include specific surface area (SSA), porosity (Φ), and differential and logarithmic differential pore area/volume distributions. In addition, pore and surface fractal dimensions (D p and D s, respectively) are obtained from the scattering profiles. The program package allows simultaneous and rapid analysis of a batch of samples, and the results are then exported to .xlsx and .csv files with separate spreadsheets for individual samples. MATSAS is the first SAS program that delivers a full suite of pore characterizations for sedimentary rocks. MATSAS is an open-source package and is freely available at GitHub (https://github.com/matsas-software/MATSAS).

Beyond structural models for the mode of action: How natural antimicrobial peptides affect lipid transport.

HYPOTHESIS: Most textbook models for antimicrobial peptides (AMP) mode of action are focused on structural effects and pore formation in lipid membranes, while these deformations have been shown to require high concentrations of peptide bound to the membrane. Even insertion of low amounts of peptides in the membrane is hypothesized to affect the transmembrane transport of lipids, which may play a key role in the peptide effect on membranes. EXPERIMENTS: Here we combine state-of-the-art small angle X-ray/neutron scattering (SAXS/SANS) techniques to systematically study the effect of a broad selection of natural AMPs on lipid membranes. Our approach enables us to relate the structural interactions, effects on lipid exchange processes, and thermodynamic parameters, directly in the same model system. FINDINGS: The studied peptides, indolicidin, aurein 1.2, magainin II, cecropin A and LL-37 all cause a general acceleration of essential lipid transport processes, without necessarily altering the overall structure of the lipid membranes or creating organized pore-like structures. We observe rapid scrambling of the lipid composition associated with enhanced lipid transport which may trigger lethal signaling processes and enhance ion transport. The reported membrane effects provide a plausible canonical mechanism of AMP-membrane interaction and can reconcile many of the previously observed effects of AMPs on bacterial membranes.

Polymorphic phase transition in liquid and supercritical carbon dioxide.

We present experiments on molecular density fluctuations in liquid and supercritical (SC) CO2 using small-angle neutron scattering. Thermal density fluctuations in SC-CO2 determine susceptibility and correlation length identifying the Widom line at their maxima. Droplet formation occurs at the gas-liquid line and between 20 and 60 bar above the Widom line, the corresponding borderline identified as the Frenkel line. The droplets start to form spheres of constant radius of ≈ 45 Å and transform into rods and globules at higher pressure. Droplet formation represents a liquid-liquid (polymorphic) phase transition of the same composition but different density, whose difference defines its order parameter. Polymorphism in CO2 is a new observation stimulating interesting discussions on the topics of gas-like to liquid-like transition in SC fluids and polymorphism since CO2 represents a "simple" van der Waals liquid in contrast to water, which is the most widely studied liquid showing polymorphism in its supercooled state.

Size Measurement of Extracellular Vesicles and Synthetic Liposomes: The Impact of the Hydration Shell and the Protein Corona.

Size characterization of extracellular vesicles (EVs) and drug delivery liposomes is of great importance in their applications in diagnosis and therapy of diseases. There are many different size characterization techniques used in the field, which often report different size values. Besides technological biases, these differences originate from the fact that various methods measure different physical quantities to determine particle size. In this study, the size of synthetic liposomes with nominal diameters of 50nm and 100nm, and red blood cell-derived EVs (REVs) were measured with established optical methods, such as dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA), and with emerging non-optical methods such as microfluidic resistive pulse sensing (MRPS) and very small-angle neutron scattering (VSANS). The comparison of the hydrodynamic sizes obtained by DLS and NTA with the sizes corresponding to the excluded volume of the particles by MRPS enabled the estimation of the thickness of the hydration shell of the particles. The comparison of diameter values corresponding to the boundary of the phospholipid bilayer obtained from VSANS measurements with MRPS size values revealed the thickness of the polyethylene glycol-layer in case of synthetic liposomes, and the thickness of the protein corona in case of REVs.

Tale of Two Tails: Molecular Exchange Kinetics of Telechelic Polymer Micelles.

Telechelic polymers contain two chain ends that are able to promote self-assembly into "flowerlike" or interconnected micellar structures. Here, we investigate the molecular exchange kinetics of such micelles using time-resolved small-angle neutron scattering. We show that the activation energies of monofunctional and telechelic chain exchange are identical. This demonstrates that the two chain ends are not simultaneously released in a single event. Instead, the results show that, contrary to regular micelles, the kinetics occurs in a multistep process involving a collision-induced single-molecule exchange mechanism where the exchange rate is directly proportional to the polymer concentration. We show that this novel mechanism can be quantitatively explained by a simple kinetic model.

Morphology of Thin Film Composite Membranes Explored by Small-Angle Neutron Scattering and Positron-Annihilation Lifetime Spectroscopy.

The morphology of thin film composite (TFC) membranes used in reverse osmosis (RO) and nanofiltration (NF) water treatment was explored with small-angle neutron scattering (SANS) and positron-annihilation lifetime spectroscopy (PALS). The combination of both methods allowed the characterization of the bulk porous structure from a few Å to µm in radius. PALS shows pores of 4.5 Å average radius in a surface layer of about 4 m thickness, which become 40% smaller at the free surface of the membranes. This observation may correlate with the glass state of the involved polymer. Pores of similar size appear in SANS as closely packed pores of 6 Å radius distributed with an average distance of 30 Å. The main effort of SANS was the characterization of the morphology of the porous polysulfone support layer as well as the fibers of the nonwoven fabric layer. Contrast variation using the media H2O/D2O and supercritical CO2 and CD4 identified the polymers of the support layers as well as internal heterogeneities.

Inner structure and dynamics of microgels with low and medium crosslinker content prepared via surfactant-free precipitation polymerization and continuous monomer feeding approach.

The preparation of poly(N-isopropylacrylamide) microgels via classical precipitation polymerization (batch method) and a continuous monomer feeding approach (feeding method) leads to different internal crosslinker distributions, i.e., from core-shell-like to a more homogeneous one. The internal structure and dynamics of these microgels with low and medium crosslinker concentrations are studied with dynamic light scattering and small-angle neutron scattering in a wide q-range below and above the volume phase transition temperature. The influence of the preparation method, and crosslinker and initiator concentration on the internal structure of the microgels is investigated. In contrast to the classical conception where polymer microgels possess a core-shell structure with the averaged internal polymer density distribution within the core part, a detailed view of the internal inhomogeneities of the PNIPAM microgels and the presence of internal domains even above the volume phase transition temperature, when polymer microgels are in the deswollen state, are presented. The correlation between initiator concentration and the size of internal domains that appear inside the microgel with temperature increase is demonstrated. Moreover, the influence of internal inhomogeneities on the dynamics of the batch- and feeding-microgels studied with neutron spin-echo spectroscopy is reported.

Cooperativity during Melting and Molecular Exchange in Micelles with Crystalline Cores.

Molecular exchange processes are important equilibration and transport mechanisms in both synthetic and biological self-assembled systems such as micelles, vesicles, and membranes. Still, these processes are not entirely understood, in particular the effect of crystallinity and the interplay between cooperative melting processes and chain exchange. Here we focus on a set of simple polymer micelles formed by binary mixtures of poly(ethylene oxide)-mono-n-alkyl-ethers (C_{n}-PEO5) which allows the melting point to be tuned over a wide range. We show that the melting transition is cooperative in the confined 4-5 nm micellar core, whereas the exchange process is widely decoupled and unimeric in nature. As confirmed by differential scanning calorimetry, the total activation energy for ejecting a molecule out of the micellar core below the melting point is the sum of the enthalpy of fusion and the corresponding activation energy in the melt state. This suggests that a "local, single-chain melting process" preludes the molecular diffusion out of the micelle during chain exchange.

A comparison of the network structure and inner dynamics of homogeneously and heterogeneously crosslinked PNIPAM microgels with high crosslinker content.

Poly(N-isopropylacrylamide) microgel particles were prepared via a "classical" surfactant-free precipitation polymerization and a continuous monomer feeding approach. It is anticipated that this yields microgel particles with different internal structures, namely a dense core with a fluffy shell for the classical approach and a more even crosslink distribution in the case of the continuous monomer feeding approach. A thorough structural investigation of the resulting microgels with dynamic light scattering, atomic force microscopy and small angle neutron scattering was conducted and related to neutron spin echo spectroscopy data. In this way a link between structural and dynamic features of the internal polymer network was made.

Design of Interactions Between Nanomaterials and Proteins: A Highly Affine Peptide Tag to Bare Iron Oxide Nanoparticles for Magnetic Protein Separation.

Superparamagnetic nanoparticles have recently gained much attention due to their broad range of applicability including medical in vivo technologies, sensors, and as supports for catalysts. As magnetic affinity materials, they can be utilized for the development of new purification strategies for pharmaceuticals and other target molecules from crude lysates. Here, a short peptide tag based on a glutamate sequence is introduced and the adsorption of pure protein as well as protein from crude cell lysate at different conditions is demonstrated. Fused to a model protein this tag can be used to recognize and purify this protein from a fermentation broth by bare iron oxide nanoparticles (BIONs). Binding of up to 0.2 g protein per g nanoparticles can be achieved and recovered easily by switching to a citrate buffered system. For a deeper understanding of the separation process, the aggregation and agglomeration of the nanoparticle protein systems were monitored for binding and elution steps. Furthermore, an upscaling of the process to the liter scale and the separation of a green fluorescent protein (GFP) containing the affinity tag to purities of 70% from Escherichia coli fermentation broth was possible in a one step process by means of high gradient magnetic separation (HGMS).

Ionic Dependence of Gelatin Hydrogel Architecture Explored Using Small and Very Small Angle Neutron Scattering Technique.

The hierarchical structure of gelatin hydrogels mimics a natural extracellular matrix and provides an optimized microenvironment for the growth of 3D structured tissue analogs. In the presence of metal ions, gelatin hydrogels exhibit various mechanical properties that are correlated with the molecular interactions and the hierarchical structure. The structure and structural response of gelatin hydrogels to variation of gelatin concentration, pH, or addition of metal ions are explored by small and very small angle neutron scattering over broad length scales. The measurements of the hydrogels reveal the existence of a two-level structure of colloid-like large clusters and a 3D cage-like gel network. In the presence of Fe3+ ions the hydrogels show a highly dense and stiff network, while Ca2+ ions have an opposite effect. The results provide important structural insight for improvement of the design of gelatin based hydrogels and are therefore suitable for various applications.

Structural control of caspase-generated glutamyl-tRNA synthetase by appended noncatalytic WHEP domains.

Aminoacyl-tRNA synthetases are ubiquitous, evolutionarily conserved enzymes catalyzing the conjugation of amino acids onto cognate tRNAs. During eukaryotic evolution, tRNA synthetases have been the targets of persistent structural modifications. These modifications can be additive, as in the evolutionary acquisition of noncatalytic domains, or subtractive, as in the generation of truncated variants through regulated mechanisms such as proteolytic processing, alternative splicing, or coding region polyadenylation. A unique variant is the human glutamyl-prolyl-tRNA synthetase (EPRS) consisting of two fused synthetases joined by a linker containing three copies of the WHEP domain (termed by its presence in tryptophanyl-, histidyl-, and glutamyl-prolyl-tRNA synthetases). Here, we identify site-selective proteolysis as a mechanism that severs the linkage between the EPRS synthetases in vitro and in vivo Caspase action targeted Asp-929 in the third WHEP domain, thereby separating the two synthetases. Using a neoepitope antibody directed against the newly exposed C terminus, we demonstrate EPRS cleavage at Asp-929 in vitro and in vivo Biochemical and biophysical characterizations of the N-terminally generated EPRS proteoform containing the glutamyl-tRNA synthetase and most of the linker, including two WHEP domains, combined with structural analysis by small-angle neutron scattering, revealed a role for the WHEP domains in modulating conformations of the catalytic core and GSH-S-transferase-C-terminal-like (GST-C) domain. WHEP-driven conformational rearrangement altered GST-C domain interactions and conferred distinct oligomeric states in solution. Collectively, our results reveal long-range conformational changes imposed by the WHEP domains and illustrate how noncatalytic domains can modulate the global structure of tRNA synthetases in complex eukaryotic systems.

Densification of Supercritical Carbon Dioxide Accompanied by Droplet Formation When Passing the Widom Line.

Thermal density fluctuations of supercritical CO_{2} were explored using small-angle neutron scattering (SANS), whose amplitude (susceptibility) and correlation length show the expected maximum at the Widom line. At low pressure, the susceptibility is in excellent agreement with the evaluated values on the basis of mass density measurements. At about 20 bar beyond the Widom line, SANS shows the formation of droplets accompanied by an enhanced number density of the supercritical fluid. The corresponding borderline is interpreted as a Frenkel line separating gas- and liquidlike regimes.

Luminescence of Eu3+ ions in hybrid polymer-inorganic composites based on poly(methyl methacrylate) and zirconia nanoparticles.

Spherical nanoparticles of ZrO2 with 2 and 10 mol% EuO1.5 up to 20 nm size were prepared by the method of hydrothermal synthesis for luminescent functionalization of the polymer-inorganic nanocomposites based on poly(methyl methacrylate). Surface modification of oxide nanoparticles was carried out by 3-(trimethoxysilyl)propyl methacrylate, dimethoxymethylvinyl silane and 2-hydroxyethyl methacrylate to provide uniform distribution and to prevent agglomeration of nanosized filler in the polymer matrix. Polymer-inorganic composites were synthesized by in situ free radical polymerization in bulk. Structuring of ZrO2 -EuO1.5 nanoparticles in the poly(methyl methacrylate) was studied by very-small-angle neutron scattering. According to the results, the dependence of photoluminescent properties of ZrO2 -EuO1.5 nanoparticles on the content of lanthanide, the symmetry of the crystal field, surface treatment and the polymer matrix were established. A correlation was shown between Stark splitting in luminescence spectra of ZrO2 -EuO1.5 nanoparticles and their phase composition. Using MMT-assay it was shown that composites based on poly(methyl methacrylate) and ZrO2 -EuO1.5 nanoparticles do not have cytotoxic properties, which makes it possible to use them as prosthesis materials with contrasted and luminescent imaging properties.

Pressure-Dependence of Poly(N-isopropylacrylamide) Mesoglobule Formation in Aqueous Solution.

Above their cloud point, aqueous solutions of the thermoresponsive polymer poly(N-isopropylacrylamide) (PNIPAM) form large mesoglobules. We have carried out very small-angle neutron scattering (VSANS with q = 0.21-2.3 × 10-3 Å-1) and Raman spectroscopy experiments on a 3 wt % PNIPAM solution in D2O at atmospheric and elevated pressures (up to 113 MPa). Raman spectroscopy reveals that, at high pressure, the polymer is less dehydrated upon crossing the cloud point. VSANS shows that the mesoglobules are significantly larger and contain more D2O than at atmospheric pressure. We conclude that the size of the mesoglobules and thus their growth process are closely related to the hydration state of PNIPAM.