Analysis Of Small-angle X-ray Research Articles

Sodium dodecyl sulfate micelles: Accurate analysis of small-angle X-ray scattering data through form factor and atomistic molecular dynamics modelling

The structure of micelles of sodium dodecyl sulfate (SDS) is probed via analysis of small-angle x-ray scattering (SAXS) data with the aim to fit the data over an extended wavenumber q range. This provides detailed information on the micelle shape, which can be described as a polydisperse triaxial ellipsoidal core-shell structure according to model form factor fitting. This model was necessary to fit the data over a wide q range, which is not accurately represented by simpler models such as biaxial ellipsoidal core-shell structures. Data for SDS (at fixed concentration) in a NaCl concentration series revealed increasing structure factor effects with decreasing salt concentration. This reflects decreased charge screening on the headgroups. The structure factor could be modelled using a simple hard sphere structure factor. The analysis of form factor was complemented by atomistic molecular dynamics (MD) simulations, starting from an unbiased initial configuration of a defined number of molecules in a box. The MD configurations were used to calculate the form factor using the software CRYSOL (for small-angle scattering analysis of solution scattering, traditionally for proteins, here for micelles) and accounting for the boundary layer hydration effects. Good agreement with experimental data was found for systems with association numbers close to p = 60. This association number is consistent with that obtained from analysis of the form factor (in the case where structure factor effects could be neglected) and from model-free analysis of the forward scattering intensity. It is also in agreement with prior literature and our findings in regard to form factor parameters are also compared to previous reports.

Effects of wheat protein on hot-extrusion 3D-printing performance and the release behaviours of caffeic acid-loaded wheat starch

In this study, the effects of wheat protein (WP) on the hot-extrusion 3D-printing (HE-3DP) performance of wheat starch (WS) gels, as well as effects of such gels on the encapsulation of caffeic acid, were investigated for the first time. The HE-3DP results show that the addition of WP can reduce print-line width and improve printing accuracy and fidelity, and the best printing results were achieved when using gels with 10 % WP. The rheological results show that WP reduced the gels' linear viscoelastic region (LVR), yield stress (τy), flow stress (τf) and consistency factor (K) but increased their structural recovery rate, which facilitated smooth extrusion during 3D printing and, thus, improved printing accuracy. The analysis of X-ray diffraction and small-angle X-ray scattering indicates that adding WP to WS could increase the mass fractal dimension and lead to denser gel network structures. The results regarding release kinetics demonstrate that the maximum release of caffeic acid from gels decreased by 28 % with the addition of WP, indicating slow-release behaviour. This study provided valuable information about processing wheat products via 3D printing.

Absolute scattering length density profile of liposome bilayers obtained by SAXS combined with GIXOS: a tool to determine model biomembrane structure

Lipid membranes play an essential role in biology, acting as host matrices for biomolecules like proteins and facilitating their functions. Their structures and structural responses to physiologically relevant interactions (i.e. with membrane proteins) provide key information for understanding biophysical mechanisms. Hence, there is a crucial need of methods to understand the effects of membrane host molecules on the lipid bilayer structure. Here, a purely experimental method is presented for obtaining the absolute scattering length density profile and the area per lipid of liposomal bilayers, by aiding the analysis of small-angle X-ray scattering (SAXS) data with the volume of bare headgroups obtained from grazing-incidence X-ray off-specular scattering (GIXOS) data of monolayers of the same model membrane lipid composition. The GIXOS data experimentally demonstrate that the variation of the bare headgroup volume upon change in lipid packing density is small enough to allow its usage as a reference value without knowing the lipid packing stage in a bilayer. This approach also has the advantage that the reference volume is obtained in the same aqueous environment as used for the model membrane bilayers. The validity of this method is demonstrated using several typical membrane compositions, as well as one example of a phospholipid membrane with an incorporated transmembrane peptide. This methodology allows us to obtain absolute scale rather than relative scale values using solely X-ray-based instrumentation, retaining a similar resolution to SAXS experiments. The method presented has high potential for understanding the structural effects of membrane proteins on the biomembrane structure.

High unimer concentration and asymmetric bilayer observed in small unilamellar vesicles of mixed surfactants TDMAO/LiPFO.

The mixed surfactant system of tetradecyldimethylamine oxide (TDMAO) and lithium perfluorooctanoate (LiPFO) is known to spontaneously self-assemble into well-defined small unilamellar vesicles. For a quantitative analysis of small-angle x-ray scattering on this model system, we complemented the measurements with densitometry, conductimetry, and contrast-variation small-angle neutron scattering. The analysis points to two main findings: first, the vesicles formed to contain a much higher mole fraction (0.61-0.64) of TDMAO than the bulk sample (0.43) and predicted by Regular Solution Theory (RST, 0.46). In consequence, the unimer concentration of LiPFO is more than 5 times higher than predicted by RST. Second, the vesicle bilayer is asymmetric with a higher fraction of LiPFO on the outside. These findings on a model system should be of broader relevance for the understanding of similar mixed surfactant vesicle systems and thereby also be of importance for their use in a number of applications.

Lyotropic liquid crystals of tetradecyldimethylaminoxide in water and the in situ formation of gold nanomaterials

Lyotropic liquid crystals (LLCs) produced by the self-assembly of surfactant in water represent an important class of highly ordered soft materials that have a wide range of applications. This study investigates the LLCs formed by a zwitterionic surfactant (tetradecyldimethylaminoxide, C14DMAO) in water. The organization of C14DMAO within the LLCs was determined based on a detailed analysis of small-angle X-ray scattering measurements and polarized microscopy observations of a typical sample. Additional to the singe-phase region, which has a hexagonal organization, several two-phase regions were observed, exhibiting the coexistence of hexagonal/cubic, cubic/lamellar, and hexagonal/lamellar phases. The phase behavior showed an obvious dependence on temperature, with more pronounced two-phase regions at lower temperatures. Using the LLCs as a matrix, Au nanospheres, nanoellipsoids, and nanorods were prepared without requiring additional reducing reagents. These three- and one-dimensional Au nanomaterials could be converted to two-dimensional plates via the introduction of a small amount of cationic surfactant to the LLCs, such as cetyltrimethylammonium bromide (CTAB) and 1-hexadecyl-3-methylimidazolium bromide ([C16MIm]B), which showed pronounced surface-enhanced Raman scattering activity towards solid rhodamine. The LLCs loaded with CTAB (or [C16MIm]B) and HAuCl4 exhibited slightly different structures and mechanical strength from the original LLCs, thereby forming a new class of highly crowded colloidal materials.

Extracting structural insights from soft X-ray scattering of biological assemblies.

Resonant soft X-ray scattering (RSoXS), a technique that combines X-ray absorption spectroscopy and X-ray scattering, can probe the nano- and meso-scale structure of biological assemblies with chemical specificity. RSoXS experiments yield scattering data collected at several photon energies, for example across an elemental absorption edge of interest. Collecting a near-edge X-ray absorption fine structure (NEXAFS) spectrum complements RSoXS experiments and determines X-ray energies that are best suited for RSoXS measurements. The analysis of RSoXS data is similar in many ways to analysis of small angle X-ray scattering using hard X-rays, with an added dimension that includes an X-ray energy dependence. This chapter discusses procedures for predicting scattering contrast and thereby identifying energies suitable for RSoXS measurements using NEXAFS spectra, analyses of 2D RSoXS images through integration into 1D profiles, and strategies for elucidating the origin of RSoXS scattering features. It also discusses existing and potential methods for interpretation of RSoXS data to gain detailed structural insights into biological systems.

Computational Reverse-Engineering Analysis for Scattering Experiments (CREASE) with Machine Learning Enhancement to Determine Structure of Nanoparticle Mixtures and Solutions.

We present a new open-source, machine learning (ML) enhanced computational method for experimentalists to quickly analyze high-throughput small-angle scattering results from multicomponent nanoparticle mixtures and solutions at varying compositions and concentrations to obtain reconstructed 3D structures of the sample. This new method is an improvement over our original computational reverse-engineering analysis for scattering experiments (CREASE) method (ACS Materials Au2021, 1 (2 (2), ), 140−156), which takes as input the experimental scattering profiles and outputs a 3D visualization and structural characterization (e.g., real space pair-correlation functions, domain sizes, and extent of mixing in binary nanoparticle mixtures) of the nanoparticle mixtures. The new gene-based CREASE method reduces the computational running time by >95% as compared to the original CREASE and performs better in scenarios where the original CREASE method performed poorly. Furthermore, the ML model linking features of nanoparticle solutions (e.g., concentration, nanoparticles’ tendency to aggregate) to a computed scattering profile is generic enough to analyze scattering profiles for nanoparticle solutions at conditions (nanoparticle chemistry and size) beyond those that were used for the ML training. Finally, we demonstrate application of this new gene-based CREASE method for analysis of small-angle X-ray scattering results from a nanoparticle solution with unknown nanoparticle aggregation and small-angle neutron scattering results from a binary nanoparticle assembly with unknown mixing/segregation among the nanoparticles.

Performance of the new biological small- and wide-angle X-ray scattering beamline 13A at the Taiwan Photon Source.

Recent developments in the instrumentation and data analysis of synchrotron small-angle X-ray scattering (SAXS) on biomolecules in solution have made biological SAXS (BioSAXS) a mature and popular tool in structural biology. This article reports on an advanced endstation developed at beamline 13A of the 3.0 GeV Taiwan Photon Source for biological small- and wide-angle X-ray scattering (SAXS-WAXS or SWAXS). The endstation features an in-vacuum SWAXS detection system comprising two mobile area detectors (Eiger X 9M/1M) and an online size-exclusion chromatography system incorporating several optical probes including a UV-Vis absorption spectrometer and refractometer. The instrumentation and automation allow simultaneous SAXS-WAXS data collection and data reduction for high-throughput biomolecular conformation and composition determinations. The performance of the endstation is illustrated with the SWAXS data collected for several model proteins in solution, covering a scattering vector magnitude q across three orders of magnitude. The crystal-model fittings to the data in the q range ∼0.005-2.0 Å-1 indicate high similarity of the solution structures of the proteins to their crystalline forms, except for some subtle hydration-dependent local details. These results open up new horizons of SWAXS in studying correlated local and global structures of biomolecules in solution.

A Lipid-Based Depot Formulation with a Novel Non-lamellar Liquid Crystal Forming Lipid.

PurposeNon-lamellar liquid crystal (NLLC)-forming lipids have gained attention as a novel component because of their ability to self-assemble upon contact with body fluids. In this study, a novel NLLC-forming lipid, mono-O-(5, 9, 13-trimethyl-4-tetradecenyl) glycerol ester (C17MGE), and a model drug with a middle molecule weight, leuprolide acetate (LA), were used to confirm the usefulness of C17MGE as an excipient for depot formulations with sustained release properties.MethodsA self-constructed depot formulation was prepared by mixing C17MGE and different types of phospholipids. The constructed NLLC structure was evaluated using small angle X-ray analysis and cryo-transmission electron microscopy. In vitro release and blood concentration profiles of LA were investigated.ResultsThe NLLC structure was confirmed by small angle X-ray analysis. LA release was able to be modified by adding different ratios of various phospholipids to C17MGE. Formulations containing 1, 2-dioleoyl-sn-glycero-3-phosphoglycerol sodium salt with a mixing ratio of 12% or 24% (MDOPG12 or MDOPG24, respectively) exhibited sustained release profiles of LA. In addition, the blood concentration of LA was detected over 21 days or more after administration of MDOPG12, and the absolute bioavailability was calculated to be about 100%.ConclusionsA depot formulation using C17MGE was useful to achieve sustained release of LA.

Activated Carbon from Agricultural Wastes for Adsorption of Organic Pollutants.

Agricultural waste materials (strawberry seeds and pistachio shells) were used for preparation of activated carbons by two various methods. Chemical activation using acetic acid and physical activation with gaseous agents (carbon dioxide and water vapor) were chosen as mild and environmentally friendly methods. The effect of type of raw material, temperature, and activation agent on the porous structure characteristics of the materials was discussed applying various methods of analysis. The best obtained activated carbons were characterized by high values of specific surface area (555–685 m2/g). The Guinier analysis of small-angle X-ray scattering (SAXS) curves showed that a time of activation affects pore size. The samples activated using carbon dioxide were characterized mostly by the spherical morphology of pores. Adsorbents were utilized for removal of the model organic pollutants from the single- and multicomponent systems. The adsorption capacities for the 4-chloro-2-methyphenoxyacetic acid (MCPA) removal were equal to 1.43–1.56 mmol/g; however, for adsorbent from strawberry seeds it was much lower. Slight effect of crystal violet presence on the MCPA adsorption and inversely was noticed as a result of adsorption in different types of pores. For similar herbicides strong competition in capacity and adsorption rate was observed. For analysis of kinetic data various equations were used.

The N-terminus region of Drp1, a Rint1 family protein is essential for cell survival and its interaction with Rad50 protein in fission yeast S.pombe

BackgroundDefects in DNA repair pathway can lead to double-strand breaks leading to genomic instability. Earlier we have shown that S.pombe Drp1, a Rint1/Tip1 family protein is required for the recovery from DNA damage. MethodsVarious truncations of Drp1 protein were constructed and their role in DNA damage response and interaction with Rad50 protein has been studied by co-immunoprecipitation and pull-down assays. ResultsThe structural and functional analysis of Drp1 protein revealed that the N-terminus region of Drp1 is indispensable for the survival. The C-terminus truncation mutants, drp1C1Δ and drp1C2Δ exhibit temperature sensitive phenotype and are hypersensitive against DNA damaging agents with elevated level of Rad52-YFP foci at non-permissive temperature indicating the impairment for DNA damage repair pathway. The essential N-terminus region of Drp1 interacts with the C-terminus region of Rad50 and might be involved in influencing the MRN/X function. Small-angle X-ray (SAXS) analysis revealed three-domain like shapes in Drp1 protein while the C-terminus region of Rad50 exhibit unusual bulges. Computational docking studies revealed the amino acid residues at the C-terminus region of Rad50 that are involved in the interaction with the residues present at the N-terminal region of Drp1 indicating the importance of the N-terminal region of Drp1 protein. ConclusionsWe have identified the region of Drp1 and Rad50 proteins that are involved in the interaction and their role in the DNA damage response pathway has been analyzed. General significanceThe functional and structural aspects of fission yeast Drp1 protein and its interaction with Rad50 have been elucidated.

In-Situ saxs/waxd analysis on structural evolution in peek irradiated by 1 MeV electrons during tensile deformation

Polyether ether ketone (PEEK) is widely used as an electrical insulation material in aerospace field due to its excellent thermal and electrical properties. The changes in micro structure of the PEEK irradiated by 1 MeV electrons are studied by in-situ analysis of synchronous small-angle X-ray scattering (SAXS) and wide-angle X-ray diffraction (WAXD) during tensile deformation at room temperature. The tensile strength and the elongation at break of the irradiated PEEK decrease obviously, which is compared with the pristine one. The differential scanning calorimetry (DSC) analysis shows that the pristine PEEK begins to melt at 334.8 °C, the melting peak Tm appears at 339.7 °C and a deeper exothermic crystallization peak Tc appears at 297.8 °C during the cooling process. The crystallization peak temperature decreases with the increase of the irradiation fluences. The pristine PEEK has a significant scattering peak along the meridian direction of the SAXS pattern, which indicates that the crystal lamellae of the material exhibits a certain orientation, and the position of the scattering peak changes little after irradiation. The intensity of SAXS decreases with the increasing of stretching strains, and the descent rate increases with the increasing of the irradiation fluences. According to the WAXD analysis, the width at half height of the diffraction peak increases with the increasing of the stretching strain, and the peak tends to disappear during tensile deformation. The diffraction peak disappearance rate increases with the increasing of the irradiation fluences.

The Cold-Unfolded State Is Expanded but Contains Long- and Medium-Range Contacts and Is Poorly Described by Homopolymer Models.

Cold unfolding of proteins is predicted by the Gibbs-Helmholtz equation and is thought to be driven by a strongly temperature-dependent interaction of protein nonpolar groups with water. Studies of the cold-unfolded state provide insight into protein energetics, partially structured states, and folding cooperativity and are of practical interest in biotechnology. However, structural characterization of the cold-unfolded state is much less extensive than studies of thermally or chemically denatured unfolded states, in large part because the midpoint of the cold unfolding transition is usually below freezing. We exploit a rationally designed point mutation (I98A) in the hydrophobic core of the C-terminal domain of the ribosomal protein L9 that allows the cold denatured state ensemble to be observed above 0 °C at near neutral pH and ambient pressure in the absence of added denaturants. A combined approach consisting of paramagnetic relaxation enhancement measurements, analysis of small-angle X-ray scattering data, all-atom simulations, and polymer theory provides a detailed description of the cold-unfolded state. Despite a globally expanded ensemble, as determined by small-angle X-ray scattering, sequence-specific medium- and long-range interactions in the cold-unfolded state give rise to deviations from homopolymer-like behavior. Our results reveal that the cold-denatured state is heterogeneous with local and long-range intramolecular interactions that may prime the folded state and also demonstrate that significant long-range interactions are compatible with expanded unfolded ensembles. The work also highlights the limitations of homopolymer-based descriptions of unfolded states of proteins.

Mechanisms of Directional Polymer Crystallization.

Zone annealing, a directional crystallization technique originally used for the purification of semiconductors, is applied here to crystalline polymers. Tight control over the final lamellar orientation and thickness of semicrystalline polymers can be obtained by directionally solidifying the material under optimal conditions. It has previously been postulated by Lovinger and Gryte that, at steady state, the crystal growth rate of a polymer undergoing zone annealing is equal to the velocity at which the sample is drawn through the temperature gradient. These researchers further implied that directional crystallization only occurs below a critical velocity, when crystal growth rate dominates over nucleation. Here, we perform an analysis of small-angle X-ray scattering, differential scanning calorimetry, and cross-polarized optical microscopy of zone-annealed poly(ethylene oxide) to examine these conjectures. Our long period data validate the steady-state ansatz, while an analysis of Herman’s orientation function confirms the existence of a transitional region around a critical velocity, vcrit, where there is a coexistence of oriented and isotropic domains. Below vcrit, directional crystallization is achieved, while above vcrit, the mechanism more closely resembles that of conventional isotropic isothermal crystallization.

Comparative characterisation of non-monodisperse gold nanoparticle populations by X-ray scattering and electron microscopy.

Accurate nanoparticle size determination is essential across various research domains, with many functionalities in nanoscience and biomedical research being size-dependent. Although electron microscopy is capable of resolving a single particle down to the sub-nm scale, the reliable representation of entire populations is plagued by challenges in providing statistical significance, suboptimal preparation procedures and operator bias. While alternative techniques exist that provide ensemble information in solution, their implementation is generally challenging for non-monodisperse populations. Herein, we explore the use of small-angle X-ray scattering in combination with form-free Monte Carlo fitting of scattering profiles as an alternative to conventional electron microscopy imaging in providing access to any type of core size distribution. We report on a cross-method comparison for quasi-monodisperse, polydisperse and bimodal gold nanoparticles of 2-7 nm in diameter and discuss advantages and limitations of both techniques.

Magainin 2 and PGLa in Bacterial Membrane Mimics I: Peptide-Peptide and Lipid-Peptide Interactions

We addressed the onset of synergistic activity of the two well-studied antimicrobial peptides magainin 2 (MG2a) and PGLa using lipid-only mimics of Gram-negative cytoplasmic membranes. Specifically, we coupled a joint analysis of small-angle x-ray and neutron scattering experiments on fully hydrated lipid vesicles in the presence of MG2a and L18W-PGLa to all-atom and coarse-grained molecular dynamics simulations. In agreement with previous studies, both peptides, as well as their equimolar mixture, were found to remain upon adsorption in a surface-aligned topology and to induce significant membrane perturbation, as evidenced by membrane thinning and hydrocarbon order parameter changes in the vicinity of the inserted peptide. These effects were particularly pronounced for the so-called synergistic mixture of 1:1 (mol/mol) L18W-PGLa/MG2a and cannot be accounted for by a linear combination of the membrane perturbations of two peptides individually. Our data are consistent with the formation of parallel heterodimers at concentrations below a synergistic increase of dye leakage from vesicles. Our simulations further show that the heterodimers interact via salt bridges and hydrophobic forces, which apparently makes them more stable than putatively formed antiparallel L18W-PGLa and MG2a homodimers. Moreover, dimerization of L18W-PGLa and MG2a leads to a relocation of the peptides within the lipid headgroup region as compared to the individual peptides. The early onset of dimerization of L18W-PGLa and MG2a at low peptide concentrations consequently appears to be key to their synergistic dye-releasing activity from lipid vesicles at high concentrations.

The Underestimated Effect of Intracrystalline Chain Dynamics on the Morphology and Stability of Semicrystalline Polymers

Semicrystalline polymers have been classified into crystal-mobile and crystal-fixed polymers, depending on the existence or absence of intracrystalline chain dynamics. Although it was claimed that polymers with intracrystalline chain dynamics generally have a higher crystallinity, its effect on the semicrystalline morphology is not known in detail. Using a new approach for the quantitative analysis of small-angle X-ray scattering data, we compare the structural characteristics of fully crystallized samples for two model polymers with and without chain motion in the crystallites. Whereas for crystal-fixed polymers the semicrystalline morphology is characterized by lamellar crystals of well-defined thickness and marginal stability, the intracrystalline dynamics leads to additional stabilization of the crystals during growth and a minimization of the amorphous layers characterized by a well-defined thickness. Results of 1H solid-state NMR experiments enable us to determine the time scale of intracrystalline chain dynamics in the relevant temperature range and to relate it to the time scale of crystal growth. If both processes act on the same time scale, the crystallization process is an interplay between crystal growth and stabilization by reorganization enabled by intracrystalline mobility. Viewing this competition as fundamental for the formation of the semicrystalline morphology, seemingly contradictory models suggested in the past to describe polymer crystallization can at least be partially reconciled.

Deep Eutectic Solvent-Induced Structural Transition of Microemulsions Explored with Small-Angle X-ray Scattering

Microemulsions (MEs) containing deep eutectic solvents (DESs) and water in the inner phase for use in transdermal delivery of poorly soluble drugs were prepared using a mixture of polyoxyethylene sorbitan monooleate (Tween-80) and sorbitan laurate (Span-20) as surfactants. We investigated the effects of the ratios of surfactant (Tween-80/Span-20) and solvents (DES components/water) on the ME structure determined by the analysis of small-angle X-ray scattering profiles with the core-corona model. Tween-80 with an unsaturated long alkyl chain induced a structural transition of MEs from a sphere to a cylinder. DESs caused the aggregation of surfactants due to the solvophobic interactions between DESs and the alkyl chains of surfactants. Transmittance electron microscopy images of MEs indicated the presence of aggregates of the dispersed ME particles with each shape.

Model-free classification of X-ray scattering signals applied to image segmentation.

In most cases, the analysis of small-angle and wide-angle X-ray scattering (SAXS and WAXS, respectively) requires a theoretical model to describe the sample's scattering, complicating the interpretation of the scattering resulting from complex heterogeneous samples. This is the reason why, in general, the analysis of a large number of scattering patterns, such as are generated by time-resolved and scanning methods, remains challenging. Here, a model-free classification method to separate SAXS/WAXS signals on the basis of their inflection points is introduced and demonstrated. This article focuses on the segmentation of scanning SAXS/WAXS maps for which each pixel corresponds to an azimuthally integrated scattering curve. In such a way, the sample composition distribution can be segmented through signal classification without applying a model or previous sample knowledge. Dimensionality reduction and clustering algorithms are employed to classify SAXS/WAXS signals according to their similarity. The number of clusters, i.e. the main sample regions detected by SAXS/WAXS signal similarity, is automatically estimated. From each cluster, a main representative SAXS/WAXS signal is extracted to uncover the spatial distribution of the mixtures of phases that form the sample. As examples of applications, a mudrock sample and two breast tissue lesions are segmented.

Laponite-graphene oxide hybrid particulate filler enhances mechanical properties of cross-linked epoxy

A new hybrid of Laponite and graphene oxide (LGO), prepared in aqueous media by ultrasonication followed by solvent evaporation was used to reinforce epoxy matrix. The hybrid system was dispersed in liquid epoxy using a two-step solvent-assisted process. The suspensions showed negligible enhancements in processing barrier as revealed by rheology. A combinatorial analysis of small-angle x-ray scattering (SAXS) and microscopy suggested uniform dispersion of nanofillers in the matrix. The fillers showed fractal dimensions in polymer matrix as inferred from SAXS studies. Below 0.5 wt% LGO, the structure showed surface fractal and above 0.5 wt% the composites showed mass fractal, indicating a transformation from well-dispersed to agglomerated composites as the filler content increases. The composites exhibited substantial improvements in various mechanical properties. Notably, the flexural strength and modulus increased by ~23% and ~29%, respectively, with only 0.5 wt% LGO and the fracture toughness showed an increment of ~23% with 0.3 wt% LGO in epoxy matrix. A bimodal distribution of glass transition temperature (T g ) with improved T g was obtained for the composites. The simultaneous strengthening and toughening effects of nanofillers are explained by means of fractography.