Small-angle X-ray Scattering Analysis Research Articles

Identification of an intrinsically disordered region (IDR) in arginyltransferase 1 (ATE1).

Arginyltransferase 1 (ATE1) catalyzes arginylation, an important post-translational modification (PTM) in eukaryotes that plays a critical role in cellular homeostasis. The disruption of ATE1 function is implicated in mammalian neurodegenerative disorders and cardiovascular maldevelopment, while post-translational arginylation has also been linked to the activities of several important human viruses such as SARS-CoV-2 and HIV. Despite the known significance of ATE1 in mammalian cellular function, past biophysical studies of this enzyme have mainly focused on yeast ATE1, leaving the mechanism of arginylation in mammalian cells unclear. In this study, we sought to structurally and biophysically characterize mouse (Mus musculus) ATE1. Using size-exclusion chromatography (SEC), small angle X-ray scattering (SAXS), and hydrogen deuterium exchange mass spectrometry (HDX-MS), assisted by AlphaFold modeling, we found that mouse ATE1 is structurally more complex than yeast ATE1. Importantly, our data indicate the existence of an intrinsically disordered region (IDR) in all mouse ATE1 splice variants. However, comparative HDX-MS analyses show that yeast ATE1 does not have such an IDR, consistent with prior X-ray, cryo-EM, and SAXS analyses. Furthermore, bioinformatics approaches reveal that mammalian ATE1 sequences, as well as in a large majority of other eukaryotes, contain an IDR-like sequence positioned in proximity to the ATE1 GNAT active-site fold. Computational analysis suggests that the IDR likely facilitates the formation of the complex between ATE1 and tRNAArg, adding a new complexity to ATE1 structure and providing new insights for future studies of ATE1 functions.

Fabrication and stability of dual Pickering double emulsions stabilized with food-grade particles

The aim of this study was to create stable, dual Pickering-stabilized double emulsions (DEs) where both the internal and external droplets are stabilized by food grade particles. Water-in-oil (W/O) emulsions were stabilized by using cocoa butter-high oleic sunflower oil based oleogels (CBolg) and then W/O emulsion was used as the dispersed phase in water-in-oil-in-water (W/O/W) emulsions, latter stabilized by whey protein microgels (WPM). The capability of the CBolg (10 v/v% cocoa butter) to form W/O and subsequently a W/O/W emulsion was characterized using cross-polarized light microscopy, cryogenic scanning electron microscopy (cryo-SEM), confocal light microscopy (CLSM) and static light scattering. In addition, the characterization of the WPM's internal structure was evaluated by small angle X-ray scattering (SAXS). Results revealed dual Pickering stabilized W/O/W emulsions with a droplet size of 10 μm could be formed using whey protein microgel (WPM, 3–5 v/v%) as the Pickering stabilizer. Dynamic light scattering (DLS) analysis revealed WPM particles size averaging approximately 100 nm, with internal structures consisting of partially stretched protein strands with smaller compact subunits along the strand (partially folded proteins) was confirmed by SAXS analysis. The dual Pickering-stabilized DE droplets were stable over a month by formation a protective layer of WPM around the oil droplets, as observed in CLSM images, unlike the systems where the external phase was stabilized by non-microgelled whey protein, latter destabilized within hours. Such dual Pickering-stabilized DEs may offer new templates for designing healthy low-fat foods and for co-delivery of nutrients and active in multiple compartments.

Design and Application of a Gas Diffusion Electrode (GDE) Cell for Operando and In Situ Studies.

Presented here is an electrochemical three-electrode Gas Diffusion Electrode (GDE) cell tailored for operandoand in situ investigations of electrocatalytic processes, with a particular focus on X-ray scattering studies. The optimized cell is engineered to accommodate the minimal sample-detector distances requisite for comprehensive X-ray total scattering investigations. An in-depth understanding of catalytic processes requires their study under 'working' conditions. Configured as a flow-cell, the setup therefore enables the examination of electrocatalysts under high current densities and associated gas evolution phenomena, particularly pertinent for reactions like the oxygen evolution reaction (OER). Notably, its transparency simplifies cell alignment, troubleshooting, and facilitates scans through the catalyst layer, crucial for background corrections. Demonstrating its versatility, we showcase its utility through Small Angle X-ray Scattering (SAXS), X-ray Diffraction (XRD), and X-ray Pair Distribution Function (PDF) analyses of total scattering data.

Celecoxib-hydroxypropyl-β-cyclodextrin inclusion complex in a chitosan/PEO-PPO-PEO block copolymer matrix: Structural effect and drug release

HypothesisTriblock copolymers of poly(ethylene oxide) and poly(propylene oxide)-based matrices, such as Poloxamer 407 (P407) or Pluronic® F127, are extensively utilized in drug delivery and permeation systems due to their FDA approval and listing in the US and European Pharmacopoeias. The study hypothesizes that incorporating 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) and the celecoxib-HP-β-CD inclusion complex into a 16 wt% P407 and chitosan blend in an aqueous acetic acid solution will affect the system's rheological and structural properties. ExperimentsRheological, small-angle X-ray scattering (SAXS), and dynamic light scattering (DLS) experiments were conducted to assess the impact of acetic acid and chitosan on the 16 wt% P407 and chitosan blend. Additionally, in vitro drug release studies were performed to monitor the drug release profile over time. FindingsThe addition of HP-β-CD was found to inhibit gel formation in the 16 wt% P407 and chitosan blend. However, the presence of the celecoxib-HP-β-CD inclusion complex showed no significant structural effects compared to P407 blended with chitosan alone. Rheological and SAXS analyses demonstrated that acetic acid led to the formation of a lamellar phase due to the lower pH, facilitating injectability. The presence of chitosan in acetic acid resulted in the detection of a hexagonal phase, affecting the release of celecoxib.

Characterization of recombinant human lactoferrin expressed in Komagataella phaffii.

This work presents a thorough characterization of Helaina recombinant human lactoferrin (rhLF, Effera™) expressed in a yeast system at an industrial scale for the first time. Proteomic analysis confirmed that its amino acid sequence is identical to that of native human LF. N-linked glycans were detected at three known glycosylation sites, namely, Asparagines-156, -497, and -642 and they were predominantly oligomannose structures having five to nine mannoses. Helaina rhLF's protein secondary structure was nearly identical to that of human milk lactoferrin (hmLF), as revealed by microfluidic modulation spectroscopy. Results of small-angle X-ray scattering (SAXS) and analytical ultracentrifugation analyses confirmed that, like hmLF, Helaina rhLF displayed well-folded globular structures in solution. Reconstructed solvent envelopes of Helaina rhLF, obtained through the SAXS analysis, demonstrated a remarkable fit with the reported crystalline structure of iron-bound native hmLF. Differential scanning calorimetry investigations into the thermal stability of Helaina rhLF revealed two distinct denaturation temperatures at 68.7 ± 0.9 °C and 91.9 ± 0.5 °C, consistently mirroring denaturation temperatures observed for apo- and holo-hmLF. Overall, Helaina rhLF differed from hmLF in the N-glycans they possessed; nevertheless, the characterization results affirmed that Helaina rhLF was of high purity and exhibited globular structures closely akin to that of hmLF.

Insights into dynamic changes in ADC-7 and P99 cephalosporinases using small angle x-ray scattering (SAXS)

To counter the emergence of β-lactamase (BL) mediated resistance, design of new β-lactamase inhibitors (BLIs) is critical. Many high-resolution crystallographic structures of BL complexed with BLIs are available. However, their impact on BLI design is struggling to keep pace with novel and emerging variants. Small angle x-ray scattering (SAXS) in combination with molecular modeling is a useful tool to determine dynamic structures of macromolecules in solution. An important application of SAXS is to determine the conformational changes that occur when BLI bind to BL. To probe if conformational dynamics occur in class C cephalosporinases, we studied SAXS profiles of two clinically relevant class C β-lactamases, Acinetobacter baumannii ADC-7 and Enterobacter cloacae P99 in apo format complexed with BLIs. Importantly, SAXS data analysis demonstrated that in solution, these representative class C enzymes remain monomeric and did not show the associated assemblies that were seen in various crystal structures. SAXS data acquired for ADC-7 and P99, in apo and inhibitor bound states, clearly showed that these enzymes undergo detectable conformational changes, and these class C β-lactamases also close upon binding inhibitors as does BlaC. Further analysis revealed that addition of inhibitor led to the compacting of a range of residues around the active site, indicating that the conformational changes that both P99 and ADC-7 undergo are central to inhibitor recognition and efficacy. Our findings support the importance of exploring conformational changes using SAXS analysis in the design of future BLIs. Communicated by Ramaswamy H. Sarma

Confinement Effects on the Structure of Entropy-Induced Supercrystals.

Depletion-induced self-assembly is routinely used to separate plasmonic nanoparticles (NPs) of different shapes, but less often for its ability to create supercrystals (SCs) in suspension. Therefore, these plasmonic assemblies have not yet reached a high level of maturity and their in-depth characterization by a combination of in situ techniques is still very much needed. In this work, gold triangles (AuNTs) and silver nanorods (AgNRs) are assembled by depletion-induced self-assembly. Small Angle X-ray Scattering (SAXS) and scanning electron microscopy (SEM) analysis shows that the AuNTs and AgNRs form 3D and 2D hexagonal lattices in bulk, respectively. The colloidal crystals are also imaged by in situ Liquid-Cell Transmission Electron Microscopy. Under confinement, the affinity of the NPs for the liquid cell windows reduces their ability to stack perpendicularly to the membrane and lead to SCs with a lower dimensionality than their bulk counterparts. Moreover, extended beam irradiation leads to disassembly of the lattices, which is well described by a model accounting for the desorption kinetics highlighting the key role of the NP-membrane interaction in the structural properties of SCs in the liquid-cell. The results shed light on the reconfigurability of NP superlattices obtained by depletion-induced self-assembly, which can rearrange under confinement.

Interplay between the Pore Network Modifications and Dynamics of Confined Polyethyleneimine in Self-Assembled Polyethyleneimine–Silica Nanoparticle Microspheres

The CO2 adsorption performance of porous materials loaded with an amine-bearing functionality is determined by the pore network of porous adsorbents and the dynamics of the adsorbed polymeric chains bearing the amine functionality. Herein, we report the modifications in the pore network of evaporation-induced self-assembled microspheres of silica nanoparticles with varying loadings of polyethyleneimine (PEI) using positron annihilation lifetime spectroscopy (PALS) and small-angle X-ray scattering (SAXS). The dynamics of the loaded PEI in the pore network of microspheres have been investigated using temperature-dependent broadband dielectric spectroscopy. PALS investigation confirms microporosity at two different length scales available at the interparticle regions in pristine silica nanoparticle-based microspheres. With PEI loading through the self-assembly process, the pore size and pore volume accessed by PALS are observed to increase up to 20 wt % loading, followed by a decrease due to modification in the clustering behavior of nanoparticles and filling of pores with excess PEI. These variations are highly consistent with the nonmonotonic jamming of silica nanoparticles, as observed from SAXS analysis, arising due to complex interactions between silica nanoparticles and PEI. Both the segmental and localized relaxations of the adsorbed PEI are observed to slow down compared to bulk PEI due to the interactions with silica nanoparticles akin to polymer nanocomposites. The slowing down of the relaxations is consistent with the free-volume variations of the confined PEI determined using PALS. The segmental relaxation time of the loaded PEI in the microspheres varies nonmonotonically with the PEI content. The observations from complementary investigations unequivocally confirm that the packing behavior of silica nanoparticles in the evaporation-induced self-assembled microspheres is controlled by the PEI content. On the other hand, the dynamics of PEI confined within the pore network of the microsphere are determined by the clustering behavior of silica nanoparticles.

Extraction, Purification, and Small-Angle X-ray Scattering Analysis of the YsxC GTPase of Staphylococcus aureus

Extraction, Purification, and Small-Angle X-ray Scattering Analysis of the YsxC GTPase of Staphylococcus aureus

Structural and functional insights of the catalytic GH5 and Calx-β domains from the metagenome-derived endoglucanase CelE2

Cellulose is the most abundant natural polymer on Earth, representing an attractive feedstock for bioproducts and biofuel production. Cellulases promote the depolymerization of cellulose, generating short oligosaccharides and glucose, which are useful in biotechnological applications. Among the classical cellulases, those from glycoside hydrolase family 5 (GH5) are one of the most abundant in Nature, displaying several modular architectures with other accessory domains attached to its catalytic core, such as carbohydrate-binding modules (CBMs), Ig-like, FN3-like, and Calx-β domains, which can influence the enzyme activity. The metagenome-derived endoglucanase CelE2 has in its modular architecture an N-terminal domain belonging to the GH5 family and a C-terminal domain with a high identity to the Calx-β domain. In this study, the GH5 and the Calx-β domains were subcloned and heterologously expressed in E. coli, to evaluate the structural and functional properties of the individualized domains of CelE2. Thermostability analysis by circular dichroism (CD) revealed a decrease in the denaturation temperature values around 4.6 °C for the catalytic domain (CelE21–381) compared to CelE2 full-length. The CD analyses revealed that the Calx-β domain (CelE2382–477) was unfolded, suggesting that this domain requires to be attached to the catalytic core to become structurally stable. The three-dimensional structure of the catalytic domain CelE21–381 was determined at 2.1 Å resolution, showing a typical (α/β)8-barrel fold and a narrow active site compared to other cellulases from the same family. The biochemical characterization showed that the deletion of the Calx-β domain increased more than 3-fold the activity of the catalytic domain CelE21–381 towards the insoluble substrate Avicel. The main functional properties of CelE2, such as substrate specificity, optimal pH and temperature, thermal stability, and activation by CaCl2, were not altered after the deletion of the accessory domain. Furthermore, the Small Angle X-ray Scattering (SAXS) analyses showed that the addition of CaCl2 was beneficial CelE21–381 protein solvency. This work contributed to fundamental concepts about the structure and function of cellulases, which are useful in applications involving lignocellulosic materials degradation into food and feedstuffs and biofuel production.

Investigating the Role of Classical Ayurveda-Based Incineration Process on the Synthesis of Zinc Oxide Based Jasada Bhasma Nanoparticles and Zn2+ Bioavailability.

Jasada bhasma (JB) is a zinc oxide-based Indian traditional Ayurveda-based herbo-metallic nanoparticle used for the treatment of zinc (Zn) deficiency and autoimmune and inflammatory disorders. JB is made by following the Ayurveda-based guidelines using zinc oxide (ZnO) as a raw material and going through 17 cycles of the high-temperature incineration and trituration process known as "Ma̅raṇa" in the presence of herbal decoctions prepared from the leaves ofAzadirachta indica andAloe vera gel. These cycles improve the purity of the parent material and transform its physicochemical properties, converting it into nanoparticles. However, there still exists a knowledge gap regarding the role of incineration in the physicochemical transformation of the Zn raw material into JB nanoparticles and the biological interaction of the final product. In the present study, the JB samples obtained during different Ma̅raṇa cycles were carefully studied for their physicochemical transformation using analytical methods such as powdered X-ray diffraction (XRD), small-angle X-ray scattering (SAXS), field emission scanning electron microscopy (FESEM), energy-dispersive X-ray spectroscopy, Fourier transform infrared (FTIR) spectroscopy, Raman spectroscopy, and dynamic light scattering (DLS). According to the XRD results, the Zn and oxygen molecules in hexagonal ZnO wurtzite crystals gradually realigned as a result of repeated heat treatments that caused lattice tension and crystal size reduction from 53.14 to 42.40 nm. A morphological transition from 1.5 μm rod shape to 31 nm in the JB particles can be seen using FESEM and SAXS analyses. The existence of 10 nm-sized nanoparticles in the finished product was confirmed by HRTEM. The presence of ZnO was confirmed in all samples by FTIR and Raman spectroscopies. Cell viability analysis showed an inhibitory concentration 50% of >1000 μg/mL for JB nanoparticles, revealing no adverse effects in human colon Caco-2 cells. A dose-dependent uptake and intracellular accumulation of JB nanoparticles were observed in Caco-2 cells using inductively coupled plasma-based mass spectroscopy (ICP-MS). Bioavailability of Zn2+ ions (6% w/w) through JB dissolution in acidic pH 4.0 was observed, representing the stomach and intracellular lysosomal physiological conditions. Therefore, the study showed that the repeated incineration cycles produced biocompatible JB nanoparticles through the physicochemical transformation at molecular levels capable of delivering bioavailable Zn2+ ions under physiological conditions. In conclusion, the medicinal properties of JB nanoparticles described in Ayurveda were found to originate from their small size and dissolution properties, formed through the classical incineration-based synthesis process.

ATR-FTIR, SAXS and UV-Vis Studies of Silicone Hydrogel and Bio-Hydrogel Soft Contact Lenses

Aims: This study aimed to evaluate the effect of contact lens materials on the structural properties and to examine ultraviolet (UVA part) and visible (Vis) transmittance with and without UV filters of the commercially available silicone hydrogel (SiHy) and bio-hydrogel (bio-Hy) soft contact lenses (CLs) in vitro.
 Place and Duration of Study: Hacettepe University, Department of Physics, Ankara, Turkey, between May 2018 and May 2021.
 Methodology:Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) spectra of CLs were recorded (at removing from its package, after 10 min, 1 h and 1 day at room temperature) in the 4000-650 cm-1region to estimate water contents of CLs. Hierarchical Cluster Analysis (HCA) was performed to differentiate chemical structure of CLs based on the spectral differences. Ultraviolet (UVA) and visible light transmittance of (CLs) was measured in the 315 -800 nm region. Small Angle X-ray Scattering (SAXS) analyses were performed to obtain further structural information on nano-scale. 
 Results: One of the key observations in this study is the large influence of lens water content. The HCA analysis grouped all the CLs of same brand in same cluster based on their chemical similarity. The UVA transmittance results showed that CLs with UV blockers almost met ClassI and ClassII standards. The size (11.8-39.9 nm) and differences in morphologies of the nano globules were determined and correlated with equilibrium water content (EWC).
 Conclusion: This work was designed to explain important characteristics of commercial CLs and results will have implications for future experimental and clinical research regarding hydration/ dehydration experiments with CL polymers.

Estimation of the fiber orientation distribution of carbon fiber–reinforced plastics using small-angle X-ray scattering

• Carbon fiber orientation distributions in the CFRP were determined using their SAXS analysis. • Fast evaluation was achieved by introducing a calibration curve for integral width of the azimuthal direction. • Similar orientation distribution compared those obtained with X-ray CT could be obtained. The orientation of discontinuous carbon fibers (CFs) in CF-reinforced plastics (CFRPs) determines their properties, necessitating convenient evaluation methods. Herein, a fast and non-destructive method is proposed for the evaluation of the orientation distribution of CFs in CFRPs based on small-angle X-ray scattering (SAXS). The width of the equatorial streak in the SAXS pattern of (or azimuthal intensity distribution of scattering from) a CFRP depends on the orientation distribution of its CFs; a calibration curve for this relationship was derived by assuming the orientation distribution to be Gaussian. This calibration curve was used to determine the orientation distributions of CFRPs featuring CFs with different orientation distributions, which correspond well with the orientation distributions determined by X-ray computed tomography. Thus, the proposed method can be used to conveniently and quantitatively evaluate the orientation distribution of discontinuous CFs in CFRPs. Because this method is based on scattering from elongated nanovoids in CFs, it can be used to evaluate composite materials in which reinforcing fibers containing elongated scattering bodies, e.g., high-strength organic fibers containing fibrils, are preferentially oriented.

Structural and functional insights into the glycoside hydrolase family 30 xylanase of the rumen bacterium Ruminococcus flavefaciens

The family 30 glycoside hydrolase (RfGH30) from Ruminococcus flavefaciens expressing xylanase activity was homology modeled. The structure revealed a (β/α)8 TIM barrel topology along with an associated ‘side β-structure’. Secondary structure analysis by circular dichroism displayed 28.09% α-helices and 21.02% β-strands, which corroborated with the prediction results. Multiple Sequence Alignment and structure superposition of RfGH30 with its homologue indicate that Glu200 and Glu302 are the catalytic residues displaying a retaining-type of catalysis. Molecular Dynamics (MD) simulation of RfGH30 confirmed the structural compactness and steadiness of modeled structure. Molecular docking studies showed maximum binding affinity with xylobiose that was corroborated by the total binding Gibbs free energy of -160.5 kJ/mol calculated by g_mmpbsa tool. MD simulation of RfGH30-xylobiose confirmed the structural specificity of catalytic residues and its increased stability in presence of ligand. Small angle X-ray scattering (SAXS) analysis of RfGH30 confirmed its monodispersed, fully folded and elongated structure in solution form. Dummy atom model revealed monomeric form of RfGH30 at 2.8 mg/ml, while at 4 mg/ml as dimer. Dynamic light scattering analysis of RfGH30 showed monodispersity and increase in hydrodynamic radius (Rh) from 2.0 to 4.0 mg/mL, displaying possible dimerization of RfGH30 at 4.0 mg/mL as corroborated by SAXS analysis. The lowered zeta potential of RfGH30 at 4 mg/mL also indicated the formation of a dimer.

Biomolecules adsorption to trigger the self-assembly of nanospheres and nanorods

HypothesisThe self-assembly (SA) of nanostructures triggered by the adsorption of biomolecules (BSA) is investigated as a platform to develop diagnostics and functional applications. Mixing nanostructures of different morphologies; SiO2 nanospheres (NS) to a cellulose nanorod crystals (CNC) leads to the formation of short range ordered domains in the dried film. Adding biomolecules-treated SiO2 NS into the CNC develop the long range ordered assembly in the composite film. ExperimentsThe CNC-SiO2 composite films were prepared by mixing CNC suspension (3 wt%) with the SiO2 NS of different concentration (0.5–5 mg/mL) followed by drying at the ambient conditions. The CNC-SiO2 @BSA films are prepared by first mixing the BSA (1 mg/mL) with the SiO2 suspension of different concentrations. Later the SiO2 @BSA suspension was mixed with the CNC and left for drying at the ambient conditions. The self-assembly mechanisms of CNC, SiO2 and BSA are studied by combining SEM, DLS, UV-Vis, QCM-D and SAXS analysis. FindingsThe SA mechanism of aqueous suspensions of CNC blended with SiO2 NS pre-treated with BSA (by adsorption) is studied by a combination of small angle X-ray scattering (SAXS), scanning electron microscopy (SEM), UV-Vis spectroscopy (UV-Vis), dynamic light scattering (DLS) and quartz crystal microbalance with dissipation (QCM-D). Both CNC and SiO2 NS are negatively charged and have demonstrated self-assembly ability driven from a combination of long- and short- range order arrangement. Adding SiO2 NS to a CNC suspension disrupts the original SA of both CNC and SiO2, which leads to the formation of small, short- range ordered domains of large polydispersity. Surprisingly, adding BSA-treated SiO2 NS into the CNC network enhances the arrangement of CNC and NS towards a new system with long range ordered assembly. The SA mechanisms of cellulose-SiO2 composites result from the balance of electrostatic repulsion and van der Waal attraction in the structure factor, radial distance distribution and potential of mean force. Understanding the effect of biomolecule adsorption on the self-assembling mechanisms of CNC nanorods with SiO2 nanospheres enables the efficient engineering of catalysis, biosensors and diagnostics.

The plant nucleoplasmin AtFKBP43 needs its extended arms for histone interaction

The nucleoplasmin family of histone chaperones is a key player in governing the dynamic architecture of chromatin, thereby regulating various DNA-templated processes. The crystal structure of the N-terminal domain of Arabidopsis thaliana FKBP43 (AtFKBP43), an FK506-binding immunophilin protein, revealed a characteristic nucleoplasmin fold, thus confirming it to be a member of the FKBP nucleoplasmin class. Small-Angle X-ray Scattering (SAXS) analyses confirmed its pentameric nature in solution, and additional studies confirmed the nucleoplasmin fold to be highly stable. Unlike its homolog AtFKBP53, the AtFKBP43 nucleoplasmin core domain could not interact with histones and required the acidic arms, C-terminal to the core, for histone association. However, SAXS generated low-resolution envelope structure, ITC, and AUC results revealed that an AtFKBP43 pentamer with C-terminal extensions interacts with H2A/H2B dimer and H3/H4 tetramer in an equimolar ratio, like AtFKBP53. Put together, AtFKBP43 belongs to a hitherto unreported subclass of FKBP nucleoplasmins that requires the C-terminal acidic stretches emanating from the core domain for histone interaction.

Crystallographic and Small Angle X-ray Scattering Analyses Provide New Mechanistic Insight into the Protein Quality Control System by the ER-Resident Oxidoreductases and their Upstream Enzymes

Crystallographic and Small Angle X-ray Scattering Analyses Provide New Mechanistic Insight into the Protein Quality Control System by the ER-Resident Oxidoreductases and their Upstream Enzymes

Small-angle X-ray scattering (SAXS) and small-angle neutron scattering (SANS) study on the structure of sodium caseinate in dispersions and at the oil-water interface: Effect of calcium ions

The structure of sodium caseinate particles, as affected by the presence of calcium ions (Ca2+), in aqueous solution and in oil (toluene)-in-water emulsions, was investigated by small-angle X-ray and neutron scattering (SAXS and SANS). SAXS analyses indicated that the sodium caseinate dispersed in water as small particles with electrostatic interactions, which has a radius of gyration (Rg) of ~5 nm and an effective radius (Reff) of ~ 10 nm with an assuming spherical shape. In the presence of Ca2+, the caseinate particles aggregated as large particles with a hydrodynamic diameter > 100 nm as determined by dynamic light scattering. The networks within the large particles were self-assembled from the small Ca2+-cross-linked particles (Rg ~ 6.5–8.0 nm), as probed by SAXS. The fractal-like dimension increased from 2.5 to 3.4 with increasing protein and CaCl2 concentrations, suggesting a denser structure. The integrity of the caseinate particles at the oil-water interface was enhanced by Ca2+ cross-linking, as observed by transmission electron microscopy. The oilwater interface stabilised by Ca2+-cross-linked caseinate particles was ~ 30 nm thick, six times thicker than that stabilised by sodium caseinate (~ 5 nm) as analysed by SANS with contrast variation technique. Quantifying the structure of sodium caseinate in an aqueous solution and at the oil-water interface provides valuable insights for designing new casein-based functional materials.

Synthesis of cholesterol containing unsymmetrical dimers: a new series of liquid crystals

ABSTRACT We report an efficient and practical procedure for the synthesis of cholesterol containing unsymmetrical liquid crystals dimers. Liquid crystalline and thermal properties were studied by POM, TGA, DSC, and SAXS analysis. All final products (3a-d) presented liquid crystal properties. The presence of smectic (compounds 3a, 3b, and 3d) and chiral nematic (3 c) mesophases evidenced by polarised optical microscopy (POM) and further confirmed by small-angle X-ray scattering (SAXS) analyses. The compound 3 c remains supercooled until room temperature and presented enantiotropic mesomorphism at a low temperature. This characteristic of organisation and mesomorphism at room temperature makes compound 3 c an ideal candidate for technological applications. The SAXS analyses further suggest possibly the presence of smectic clusters within the nematic liquid crystalline phase. All compounds presented high thermal stability at around 300°C when tested for their thermal properties. The highest occupied molecular orbital (HOMO) energies were estimated from cyclic voltammetry (CV) and used together with the optical absorption measurements to determine the band gap and lowest unoccupied molecular orbital (LUMO) energies. The values obtained were: HOMO (–5.81 to – 6.03 eV), LUMO (–1.53 to – 2.12 eV), and band gap (3.88 to 4.28 eV).

Stabilization by Nano Spray Dryer of Pioglitazone Polymeric Nanosystems: Development, In Vivo, Ex Vivo and Synchrotron Analysis.

Pioglitazone-loaded PLGA-PEG nanoparticles (NPs) were stabilized by the spray drying technique as an alternative to the treatment of ocular inflammatory disorders. Pioglitazone-NPs were developed and characterized physiochemically. Interaction studies, biopharmaceutical behavior, ex vivo corneal and scleral permeation, and in vivo bioavailability evaluations were conducted. Fibrillar diameter and interfibrillar corneal spacing of collagen was analyzed by synchrotron X-ray scattering techniques and stability studies at 4 °C and was carried out before and after the spray drying process. NPs showed physicochemical characteristics suitable for ocular administration. The release was sustained up to 46 h after drying; ex vivo corneal and scleral permeation profiles of pioglitazone-NPs before and after drying demonstrated higher retention and permeation through cornea than sclera. These results were correlated with an in vivo bioavailability study. Small-angle X-ray scattering (SAXS) analysis did not show a significant difference in the organization of the corneal collagen after the treatment with pioglitazone-NPs before and after the drying process, regarding the negative control. The stabilization process by Nano Spray Dryer B-90 was shown to be useful in preserving the activity of pioglitazone inside the NPs, maintaining their physicochemical characteristics, in vivo bioavailability, and non-damage to corneal collagen function after SAXS analysis was observed.