Static Light Scattering Research Articles

Small angle X-ray scattering based structure, modeling and molecular dynamics analyses of family 43 glycoside hydrolase α-L-arabinofuranosidase from Clostridium thermocellum

Enzymes that participate in the hydrolysis of complex carbohydrates display a modular architecture, although the significance of enzyme modularity to flexibility and catalytic efficacy is not fully understood. α-L-arabinofuranosidase from Clostridium thermocellum (CtAraf43) catalyzes the release of α-1,2-, α-1,3-, or α-1,5- linked L-arabinose from arabinose decorated polysaccharides. CtAraf43 comprises an N-terminal catalytic domain (CtAbf43A) connected with two family 6 carbohydrate-binding modules (CBMs), termed as CtCBM6A and CtCBM6B, through flexible linker peptides. Here, we modeled the structure of CtAraf43 revealing that the module, CtAbf43A displays a 5-fold β-propeller fold and the CBMs the typical jellyroll type β-sandwich folds. Ramachandran plot showed 98.5% residues in the favored region and 1.5% residues in the disallowed region. Molecular dynamics simulation analysis of CtAraf43 revealed significant flexibility that is more expressive in the C-terminal CtCBM6B module in terms of structure and orientation. Small angle X-ray scattering analysis of CtAraf43 revealed its elongated structure. CtAraf43 at 1.2 mg/mL demonstrated the monomeric nature and a multi-modular shaped molecular envelope in solution with a Dmax of 12 nm. However, at 4.7 mg/mL, CtAraf43 displayed a dimeric structure and elongated molecular envelope. Kratky plot analysis revealed the folded state of CtAraf43 with limited flexibility at both concentrations. The data revealed higher flexibility at the C-terminal of CtAraf43 suggesting a coordinated action of the N-terminal catalytic module CtAbf43A and the internal CtCBM6A.AbbreviationCBMsCarbohydrate Binding ModulesCtAraf43α-L-arabinofuranosidaseGHsGlycoside HydrolasesMDMolecular DynamicsRMSDRoot Mean Square DeviationRMSFRoot Mean Square FluctuationSAXSSmall angle X-ray scatteringCommunicated by Ramaswamy H. Sarma

Structural and electrical properties of BaZr0.7Ce0.2Y0.1O3−δ proton conducting ceramic fabricated by spark plasma sintering

Spark plasma sintering (SPS) is an attractive method to obtain fully densified polycrystalline ceramics at rapid sintering rates. Cerium- and yttrium-doped barium zirconate, BaZr0.7Ce0.2Y0.1O3−δ, was prepared by a SPS route, resulting in high values of density of above 98%. Investigations of microstructural and crystallographic properties with determination of grain sizes were performed by means of Scanning Electron Microscopy, X-ray Diffraction and Static Light Scattering and discussed in correlation to bulk and grain boundary conductivities obtained by AC impedance spectroscopy. Samples prepared by SPS were generally found to consist of small grains that exhibit high bulk conductivities, whereas their grain boundary conductivities were comparably small, possibly also dominated by the presence of minor phases.

Large and Giant Unilamellar Vesicle(s) Obtained by Self-Assembly of Poly(dimethylsiloxane)-b-poly(ethylene oxide) Diblock Copolymers, Membrane Properties and Preliminary Investigation of their Ability to Form Hybrid Polymer/Lipid Vesicles.

In the emerging field of hybrid polymer/lipid vesicles, relatively few copolymers have been evaluated regarding their ability to form these structures and the resulting membrane properties have been scarcely studied. Here, we present the synthesis and self-assembly in solution of poly(dimethylsiloxane)-block-poly(ethylene oxide) diblock copolymers (PDMS-b-PEO). A library of different PDMS-b-PEO diblock copolymers was synthesized using ring-opening polymerization of hexamethylcyclotrisiloxane (D3) and further coupling with PEO chains via click chemistry. Self-assembly of the copolymers in water was studied using Dynamic Light Scattering (DLS), Static Light Scattering (SLS), Small Angle Neutron Scattering (SANS), and Cryo-Transmission Electron Microscopy (Cryo-TEM). Giant polymersomes obtained by electroformation present high toughness compared to those obtained from triblock copolymer in previous studies, for similar membrane thickness. Interestingly, these copolymers can be associated to phospholipids to form Giant Hybrid Unilamellar Vesicles (GHUV); preliminary investigations of their mechanical properties show that tough hybrid vesicles can be obtained.

Analysis of Tertiary Structural Features of Branched DNA Nanostructures with Partially Common Sequences Using Small-Angle X-ray Scattering.

We demonstrated that polypod-like structured DNA, composed of 3-8 kinds of oligodeoxynucleotides (ODNs), exhibit accelerated cellular uptake depending on their structural properties. The biological activities, including immunostimulation and unwanted adverse effects of ODNs depend on their sequences. Therefore, as the number of different types of ODNs within a polypod-like structured DNA increases, the possible risks and concerns regarding its future clinical applications also increase. To minimize this risk and to explore the relationship between structural properties and cellular uptake, we designed Tet(id12), a tetrapod-like structured DNA with 12 identical palindrome sequences in the center of ODNs, and compared it with other tetrapod-like structures without palindrome sequences, symmetric Tet(sym) and asymmetric Tet(asym12). The thermal stability analysis of Tet(id12) revealed a two-step dissociation process. Polyacrylamide gel electrophoresis, small-angle X-ray scattering profile, and the Guinier plot also revealed that the conformation of Tet(id12) was close to that of Tet(sym). No significant differences were observed between Tet(id12) and others in terms of cellular uptake. These results indicate that ODNs with palindrome sequences can be used to design polypod-like structured DNAs without significant changes to their physicochemical and biological properties, although their thermal stability is somewhat different from the ones without palindrome sequences.

Highly versatile laboratory X-ray scattering instrument enabling (nano-)material structure analysis on multiple length scales by covering a scattering vector range of almost five decades.

A compact laboratory X-ray scattering platform that uniquely enables for high-performance ultra-small-angle X-ray scattering (USAXS), small- and wide-angle X-ray scattering (SAXS/WAXS), and total scattering (atomic pair distribution function analysis; PDF) experiments was developed. It covers Bragg spacings from sub-Angstroms to 1.7 μm, thus allowing the analysis of dimensions and complex structures in (nano-)materials on multiple length scales. The accessible scattering vector q-range spans over almost five decades (qmin = 0.0036 nm-1, qmax = 215 nm-1), without any gaps. Whereas SAXS is suitable to characterize materials on a length scale of 1-100 nm, with USAXS, this range can be significantly extended to the micrometer range. On the other end, from WAXS and particularly from PDF measurements, information about the local atomic order and disorder can be obtained. The high performance, exceptional versatility, and ease-of-use of the instrument are enabled by a high-resolution 2-circle goniometer with kinematic mounts, a modular concept based on prealigned, quickly interchangeable X-ray components, and advanced detector technology. For USAXS measurements, a modified Bonse-Hart experimental setup with single crystal collimator and analyzer optics is used. SAXS/WAXS measurements are enabled by focusing optics, an evacuated beam path, and a 2D detector. For total scattering experiments, a high-energy X-ray source is used in combination with a hybrid pixel array detector that is based on a CdTe sensor for the highest counting efficiency. To ensure high resolution and sensitivity in these various applications, special care is taken to suppress any type of background scattering signal. The high resolution that can be achieved with the USAXS collimation system is demonstrated on a set of monodisperse, colloidal silica dispersions and derived colloidal crystals, with particle diameters in the range of hundreds of nanometers up to 1.6 µm. USAXS and SAXS results are shown to be consistent with those obtained by static light scattering (SLS) and dynamic light scattering. It is demonstrated that the obtainable USAXS data bridge the gap in q between SAXS and SLS. The capabilities of the instrument to acquire high-quality total scattering data for PDF analysis are demonstrated on amorphous SiO2 nanoparticles as well as on NaYF4 upconversion nanocrystals. To the best of our knowledge, it is for the first time that we present a single laboratory instrument that enables measurements of high-quality X-ray scattering data within such a wide q-range, by combining four complementary elastic X-ray scattering techniques. The modular design concept of the instrument allows for incremental improvements as well as to add more applications in the future.

Preparation and characterization of water-based magnetorheological fluid using wormlike surfactant micelles

Magnetorheological fluids (MRFs) are smart materials whose rheological properties can be controlled via external magnetic field. One of the most important challenges of MRFs is their sedimentation problem. In this paper, worm-like micelles (WLMs) has been used to improve the properties of water based MRFs. Cetyltrimethyl ammonium bromide (CTAB) was used as a dispersant in the presence of KBr. The steady state and dynamic rheological measurements were performed to study the effect of CTAB concentration on the properties and the internal structure of the fluids. The static light scattering (SLS) and cryo-scanning electron microscope were done to study the structure of WLMs. The magnetorheological properties was studied under external magnetic field up to 76 kA/m. The stability of the suspensions was investigated by sedimentation test. The WLMs are formed in water at CTAB and KBr concentration of 0.2 M and 0.1 M respectively. It was found that MRFs with improved stability and yield stress can be obtained using WLMs in water.

Aggregation morphology of planar engineered nanomaterials

In this investigation, the utility of a static light scattering (SLS) technique to characterize aggregate morphology of two-dimensional engineered nanomaterials (2D ENMs) was systematically evaluated. The aggregation of graphene oxide (GO) and lithiated-molybdenum disulfide (Li-MoS2) were measured and compared to that of a spherical reference colloid, carboxylate-modified latex (CML) nanoparticles. The critical coagulation concentration (CCC) for all dispersions was determined via analysis of aggregation kinetics using time-resolved dynamic light scattering. This technique allowed for the elucidation of the transition from the reaction-limited aggregation (RLA) regime to diffusion-limited aggregation (DLA). The findings of this study support the aggregation trends predicted by Derjaguin-Landau-Verwey-Overbeek (DLVO) theory and recent computer simulations of aggregation kinetics. For all nanomaterials, as ionic strength increased towards the respective the CCC, fractal dimension decreased; any increase in ionic strength beyond the CCC did not yield significant change in fractal dimension. Across comparable primary particle sizes and using both carbonaceous (GO) and inorganic (Li-MoS2) 2D ENMs, this study further supports the use of SLS for the measurement of fractal dimension for 2D materials. To further support this claim, the aggregate morphology of GO in both RLA and DLA regimes was measured via cryogenic transmission electron microscopy.

Penetration of phospholipid membranes by poly-l-lysine depends on cholesterol and phospholipid composition

Clusters of positively-charged basic amino acid residues, particularly lysine, are known to promote the interaction of many peripheral membrane proteins with the cytoplasmic surface of the plasma membrane via electrostatic interactions. In this work, cholesterol's effects on the interaction between lysine residues and membranes have been studied. Using poly-l-lysine (PLL) and vesicles as models to mimic the interaction between lysine-rich protein domains and the plasma membrane, light scattering measurements indicated cholesterol enhanced the electrostatic interaction through indirectly affecting the negatively charged phospholipid dioleoylphosphatidylserine, DOPS. Addition of PLL to lipid vesicles containing DOPS showed an initial increase in static light scattering (SLS), attributed to binding of PLL to the vesicle surface, followed by a slower continuously declining SLS signal, which, from comparison with fluorescent dye leakage studies could be attributed to vesicle lysis. Although electrostatic interactions between PLL and the membrane were not necessary for penetration to occur, cholesterol promoted membrane disruption of negatively charged vesicles, possibly by increasing the electrostatic interactions between PLL and the membrane. In contrast, cholesterol lowered the susceptibility of uncharged vesicles (formed using dioleoylphosphatidylcholine, DOPC) to PLL penetration. This can be explained by the absence of electrostatic interactions and cholesterol's known ability to increase membrane thickness and mechanical strength. Thus, the ability of cationic peptides to penetrate membranes including cholesterol is likely to depend on the membrane's PS:PC ratio.

Investigating the Effect of Added Salt on the Chain Dimensions of Poly(ethylene oxide) through Small-Angle Neutron Scattering

We have measured the effect of added salt on the chain dimensions of mixtures of poly(ethylene oxide) (PEO) and lithium bis(trifluoromethanesulfonyl)imide salt (LiTFSI) in the melt state through small-angle neutron scattering (SANS) experiments. Scattering profiles from blends of hydrogenated and deuterated PEO mixed with LiTFSI were measured as a function of salt concentration. Scattering profiles from pure deuterated PEO/LiTFSI mixtures were used for background-subtraction purposes. The densities of PEO/LiTFSI mixtures of varying salt concentrations were measured to calculate partial molar monomer volumes of PEO and LiTFSI to account for nonideal mixing. Kratky plots of the scattering profiles were used to calculate the salt concentration dependence of statistical segment length. At low salt concentrations, segment length decreases with increasing salt concentration, before increasing with increasing salt concentration in the high salt concentration regime. The random phase approximation was used to predict theoretical scattering profiles from the calculated segment lengths and partial molar volumes; there is excellent agreement between the theoretical and measured scattering profiles at all salt concentrations. There appears to be a correlation between chain dimensions and coordination between lithium ions and EO monomers. The scattering profiles of the pure deuterated PEO/LiTFSI mixtures suggested the presence of ion clusters of characteristic size of 0.6 nm at high salt concentrations.

Synthesis of a Smart Conductive Block Copolymer Responsive to Heat and Near Infrared Light.

A method for the synthesis of a linear block copolymer (PNIPAM-b-PANI), containing a thermoresponsive block (poly(N-isopropylacrylamide), PNIPAM) and a Near Infrared (NIR) light-absorbing block (polyaniline, PANI), is reported. The synthetic approach involves a two-step successive polymerization reaction. First, the radical polymerization of NIPAM is done using 4-aminothiophenol as a chain transfer agent for the obtention of thermosensitive block terminated with an aniline (ANI) moiety. Second, the oxidative polymerization of ANI is initiated in ANI moiety of thermosensitive block to grow the second conductive PANI block. 1H nuclear magnetic resonance (NMR) and FT-IR spectroscopy shows the characteristics peaks of both polymeric blocks revealing the successful copolymerization process. Static Light Scattering (SLS) and UV-Visible combined measurements allowed the determination of the Mw for PNIPAM-b-PANI macromolecule: 5.5 × 105 g mol−1. The resulting copolymer is soluble in water (8.3 g L−1) and in non-aqueous solvents, such as ethanol, formic acid, acetonitrile, and others. Both polymer blocks chains show the properties of the polymer chains. The block copolymer shows a lower critical solution temperature (LCST) at the same temperature (32–34 °C) than PNIPAM, while the copolymer shows pH dependent UV-vis-NIR absorption similar to PANI. The PNIPAM block suffers a coil to globule transition upon NIR light irradiation (785 nm, 100 mW), as shown by turbidimetry and Atomic Force Microscopy (AFM), due to local heating (more than 9 °C in 12 min) induced by the NIR absorption at the PANI block. Furthermore, the electrical conductivity of PNIPAM-b-PANI thin films is demonstrated (resistivity of 5.3 × 10−4 Ω−1 cm−1), indicating that the PANI block is present in its conductive form.

Small-angle X-ray scattering based structure, modeling and molecular dynamics analyses of a family 5 glycoside hydrolase first endo-mannanase named as RfGH5_7 from Ruminococcus flavefaciens

Endo-β-1,4-mannanase named as RfGH5_7 from Ruminococcus flavefaciens cloned, expressed and purified earlier was structurally characterized in present study. The RaptorX modeled structure of RfGH5_7 showed a (β/α)8 Triose-phosphate Isomerase (TIM) barrel fold. The Ramachandran plot assessment of RfGH5_7 showed that all amino acids fall in allowed region except one, Asn22 in the disallowed region. The superposition of RfGH5_7 modeled structure with its nearest homologues revealed that Glu154 acts as proton donor while Glu249 acts as nucleophile. Secondary structure of RfGH5_7 through Circular Dichroism (CD) analysis revealed 33.5% α-helices, 17% β-strands and 49.5% random coils. Molecular Dynamic (MD) simulation showed Root Mean Square Deviation (RMSD), 0.67 nm and radius of gyration (Rg) between 1.9 nm and 1.85 nm. The binding interaction of mannotetraose on the surface of RfGH5_7 structure displayed polar interactions with His219, Tyr221, Trp278, Ser279 and Gly282 residues. Small-angle X-ray scattering (SAXS) analysis displayed the intact and monodispersed nature of the enzyme RfGH5_7. The radius of gyration (Rg) by Guinier analysis for globular shape was found to be 2.29 ± 0.09 nm and for rod-shape it was 0.95 ± 0.02 nm. Kratky plot confirmed that RfGH5_7 structure is compact and folded in solution. The ab initio derived dummy model of RfGH5_7 displayed single domain structure of yellow humped fish like shape. The RfGH5_7 modeled structure was well fitted with ab initio derived model from SAXS data. Communicated by Ramaswamy H. Sarma

Determining the molecular-weight and interfacial properties of chitosan built nanohydrogel for controlled drug delivery applications

Recent development in polymeric nanohydrogel demands surface modification, interfacial properties and determination of molecular weight for effective drug loading and release studies. Therefore, chitosan built nanohydrogel were synthesized via free radical copolymerization and functionalized with hydrophilic ligands to act as amphiphilic diblock copolymers in aqueous medium. The investigated properties like porosity (61- 73 %) , cross-linking density (20.38- 25.22), critical micelle concentration (CMC≈ 0.41- 0.62 g/L), thermodynamic of adsorption followed by micellization, and the molecular weight of chitosan built nanohydrogel (Mw ≈11.59- 13.05 x 105 (g/mol, determined by applying Debye plot) have been investigated as a new tool to understand the art and chemical identity of hydrogel at air- water interface in aqueous medium regarding controlled drug delivery applications.

Antimicrobial activity of N-acetylmuramoyl-L-alanine amidase produced by Lactobacillus sakei, against Leuconostoc mesenteroides and Weissella viridescens

Glucuronoxylan-β-1,4-xylanohydrolase from Clostridium thermocellum (CtXynGH30) hydrolyzes β-1,4-xylosidic linkages in 4-O-Methyl-D-glucuronoxylan. CtXynGH30 comprises an N-terminal catalytic domain, CtXyn30A, joined by a typical linker sequence to a family 6 carbohydrate-binding module, termed CtCBM6. ITC, mass spectrometric and enzyme activity analyses of CtXyn30A:CtCBM6 (1:1 M ratio), CtXyn30A and CtXynGH30 showed that the linker peptide plays a key role in connecting and orienting CtXyn30A and CtCBM6 modules resulting in the enhanced activity of CtXynGH30. To visualize the disposition of the two protein domains of CtXynGH30, SAXS analysis revealed that CtXynGH30 is monomeric and has a boot-shaped molecular envelope in solution with a Dmax of 18 nm and Rg of 3.6 nm. Kratky plot displayed the protein in a fully folded and flexible state. The ab initio derived dummy atom model of CtXynGH30 superposed well with the modelled structure.

Nanoparticle Measurement in Liquid by Static and Dynamic Light Scattering

Nanoparticle Measurement in Liquid by Static and Dynamic Light Scattering

Structure and biochemical characterization of glucose tolerant β-1,4 glucosidase (HtBgl) of family 1 glycoside hydrolase from Hungateiclostridium thermocellum

β-1,4-glucosidase (HtBgl) of family 1 glycoside hydrolase from Hungateiclostridium thermocellum was cloned in pET28a(+) vector, expressed, biochemically and structurally characterized. HtBgl displayed 67 U/mg activity against 4-nitrophenyl-β-d-glucopyranoside, followed by 180 U/mg against cellobiose and 42 U/mg activity against 4-nitrophenyl-β-d-galactopyranoside. HtBgl displayed an optimum temperature of 65 °C and an optimum pH of 6.0. HtBgl was stable in the pH range, 4.0–8.0 and displayed the thermostability up to 60 °C for 1 h. HtBgl displayed the glucose tolerance up to 750 mM and retained ~70% activity after 20 h. HtBgl crystal structure submitted (PDB id 5OGZ) by others exhibited a classical Triosephosphate Isomerase, (β/α)8-barrel fold. Protein melting analysis of HtBgl exhibited a single peak at 78 °C and the addition of 5 mM Mg2+ shifted the peak to 82 °C. Molecular dynamics studies showed that the amino acid residues from 351 to 375 exhibit the flexibility due to the presence of the catalytic acid residue. The structure comparison of HtBgl with homologous proteins and its docking analysis with probable ligands revealed that the residues, E166 and E355 are involved in the catalysis. The SAXS analysis of HtBgl showed that the protein is monomeric and present in a fully folded state. The radius of gyration (Rg) found was 2.15–2.26 nm. The bell-shaped curve obtained by Kratky plot analysis displayed the globular shape and fully folded state with flexibility in the N-terminal region. The HtBgl crystal structure superposed well with the SAXS derived dummy atom model.

Disulfide-Mediated Bioconjugation: Disulfide Formation and Restructuring on the Surface of Nanomanufactured (Microfluidics) Nanoparticles.

This study is about (1) nanomanufacturing (focusing on microfluidic-assisted nanoprecipitation), (2) advanced colloid characterization (focusing on field flow fractionation), and (3) the possible restructuring of surface disulfides. Disulfides are dynamic and exchangeable groups, and here we specifically focus, first, on their use to introduce biofunctional groups and, second, on their re-organization, which may lead to variable surface chemistries and uncontrolled cell interactions. The particles were obtained via microfluidic-assisted (flow-focused) nanoprecipitation of poly(ethylene glycol)-b-poly(ε-caprolactone) bearing or not a 2-pyridyl disulfide (PDS) terminal group, which quantitatively exchanges with thiols in solution. In this study, we have paid specific attention to size characterization, thereby also demonstrating the limitations of dynamic light scattering (DLS) as a stand-alone technique. By using asymmetric flow field flow fractionation coupled with DLS, static light scattering (SLS), and refractive index detectors, we show that relatively small amounts of >100 nm aggregates (cryogenic transmission electron microscopy and SLS/DLS comparison suggesting them to be wormlike micelles) dominated the stand-alone DLS results, whereas the "real" size distributions picked <50 nm. Our key result is that the kinetics of the conjugation based on PDS-thiol exchange was controlled by the thiol pKa, and this also determined the rate of the exchange between the resulting disulfides and glutathione (GSH). In particular, more acidic thiols (e.g., peptides, where a cysteine is flanked by cationic residues) react faster with PDS, but their disulfides hardly exchange with GSH; the reverse applies to thiols with a higher pKa. Disulfides that resist against restructuring via thiol-disulfide exchange allow for a stable bioconjugation, although they may be bad news for payload release under reducing conditions. However, experiments of both thiol release and nanoparticles uptake in cells (HCT116) show that also the disulfides formed from less-acidic and, therefore, less-reactive, and more exchangeable thiols were stable for at least a few hours even in a GSH-rich (10 mM) environment; this suggests a sufficiently long stability of surface groups to achieve, for example, a cell-targeting effect.

Size-exclusion chromatography combined with solution X-ray scattering measurement of the heat-induced aggregates of water-soluble proteins at low ionic strength in a neutral solution

The heat-induced (80 ± 1 ℃, 1 h) aggregates of bovine serum albumin and ovalbumin at neutral pH and low ionic strength (10 mM sodium phosphate buffer, pH 6.9) were characterized using size-exclusion chromatography combined with small-angle X-ray scattering measurement. The values calculated for the radius of gyration and molecular weight of the eluted aggregates of the bovine serum albumin were 9–11 nm and 540,000–820,000, respectively. Those of ovalbumin were 11–16 nm and 500,000–1,820,000, respectively. The overall linear conformation of the bovine serum albumin aggregates slightly differed from that of the ovalbumin aggregates since the mass fractal dimensions were found from calculation to be 1.63–1.7 for the bovine serum albumin and 1.36–1.51 for the ovalbumin. The surface property of the aggregates of both proteins was suggested to be similar to that of their native monomer since all the surface fractal dimensions were almost equivalent. The dimensionless Kratky plots of the eluted aggregates indicated that the non-globular conformation of the bovine serum albumin aggregates differs from that of the ovalbumin aggregates. These analyses using size-exclusion chromatography combined with the solution X-ray scattering measurement will be helpful for characterization of the components of the denatured protein aggregation in solution.

Self-assembly and nanostructure of poly(vinyl alcohol)-graft-poly(methyl methacrylate) amphiphilic nanoparticles

HypothesisPoly(vinyl alcohol)-g-poly(methyl methacrylate) (PVA-g-PMMA) amphiphilic copolymers self-assemble to form multi-micellar colloidal systems. ExperimentsA PVA-g-PMMA copolymer containing 16–17% w/w of PMMA was synthesized by the free radical graft polymerization of methyl methacrylate on a PVA backbone by utilizing cerium(IV) ammonium nitrate as initiator and tetramethylethylenediamine (TEMED) as initiation activator. The aggregation behavior of the copolymer in water was comprehensively characterized by dynamic light scattering (DLS) and static light scattering (SLS), small angle neutron scattering (SANS), asymmetrical flow field-flow fractionation (A4F) and transmission electron microscopy (TEM). The colloidal stability before and after non-covalent crosslinking of PVA domains with boric acid was assessed by DLS. Finally, nanoparticles were nano spray-dried. FindingsThis copolymer self-assembles in water to form a complex nanostructure of low aggregation number particles (ca. 12–15 nm in diameter) that aggregate into larger ones with size ca. 60–80 nm, as determined by SANS and TEM. In addition, boric acid-crosslinking preserves the nanoparticle size, while conferring full physical stability under extreme dilution conditions. Nano spray-drying consolidates the crosslnking and enables the production of a dry flowing powder that upon re-dispersion in water regenerates the nanoparticles without major size changes.

Bovine beta-casein micelles as delivery systems for hydrophobic flavonoids

The milk protein β-casein (β-CN) is an intrinsically unstructured amphiphilic protein that self-assembles into micelles. Naringenin is the main hydrophobic flavanone in grapefruit and has several beneficial biological effects: it exhibits, for example, antioxidant, anticancer and anti-inflammatory activity. This paper shows that naringenin can be encapsulated in β-CN micelles. Fluorescence spectroscopy, molecular docking modelling, dynamic light scattering (DLS), static light scattering (SLS) and isothermal titration calorimetry (ITC) were applied to characterize the effect of naringenin on the protein association behavior and properties of the resulting micelles. Naringenin binds to β-CN at both pH 7 and pH 2, promotes the formation of micelles with a well-defined size distribution and stabilizes the micelles. It was found that naringenin-containing β-CN micelles have a lower critical micelle concentration (CMC) and a larger aggregation number (Nagg) compared to pure β-CN micelles. SLS and multi-angle DLS results suggest considerable differences between the structures of pure β-CN micelles and naringenin-containing β-CN micelles. In the presence of naringenin spherical micelles were formed with a relatively loose core (“hollow sphere”), while the pure β-CN micelles are smaller and seem to be elliptic. Notably, by uptake of naringenin in the micelles, the concentration of naringenin in aqueous solution could be raised considerably. These findings lead to the conclusion that β-CN micelles are very promising as effective delivery nano-vehicles for hydrophobic bioactive compounds.

Do depletant stabilized water-in-oil microemulsions have implications for nanoencapsulation?

The physico-chemical mechanisms associated with surfactant-free depletant stabilized water-in-oil microemulsion based shell encapsulation processes merit detailed investigation owing to the possibility of a direct correlation between encapsulation and the stable water-in-oil microemulsion system. The presence of core nanoparticles is expected to influence the formation of stable water-in-oil microemulsion system wherein polyethylene glycol (PEG) is used as depletant material. The depletant is expected to play a dual role: on one hand, it reduces the surface tension of de-ionized water during the synthesis of core while on the other hand, it helps to form stable water-in-oil microemulsion system (surfactant free) and the subsequent organization of silica coating over magnetite and systematic measurements have implications on nano encapsulation. The water-in-oil microemulsion system stabilized by depletant material is formulated with the presence of core-nanoparticles. The stability of the water-in-oil microemulsion is quantified by measuring the average hydrodynamic diameter. The presence of silica coating on core nanoparticles is robustly demonstrated using (a) small angle X-ray scattering data and Guinier plot analysis, (b) isoelectric point measurement, (c) high-resolution transmission electron microscopy, and (d) infrared spectroscopy. When the concentration of PEG and water-particles mixture (WP mixture) increases, the average hydrodynamic diameter of water-in-oil microemulsion decreases. Multiple magnetite nanoparticles are accommodated within each water droplet, without the occurrence of instability/agglomeration event. This ensures a higher yield via this process. Breakdown of stable water-in-oil microemulsion hinders the shell coating process and this is a first attempt to form the surfactant-free depletant stabilized water-in-oil microemulsion systems. The effects observed in this work have important implications for the development of new approaches to the formation of water-in-oil microemulsion systems and subsequent encapsulation processes.