Interpenetrated biosurfactant-silk fibroin networks - a SANS study.

Silk fibroin (SF) based hydrogels have been exploited for years for their inherent biocompatibility and favorable mechanical properties which makes them interesting for biotechnology applications. In this study we investigate silk based composite hydrogels where pH-sensitive, anionic biosurfactant assemblies (sophorolipids SL-C18 : 1 and SL-C18 : 0), are employed to improve the present properties of SF. Results suggest that the presence of SL surfactant assemblies leads to faster gelling of SF by accelerating the refolding from random coil to ß-sheet as shown by infrared and UV-visible spectroscopy. Small angle neutron scattering (SANS) including contrast matching studies show that SF and SL assemblies coexist in a fibrillary network that is, in the case of SL-C18 : 0, interpenetrating. The resulting overall network structure in composite gels is slightly more affected by SL-C18 : 1 than by SL-C18 : 0, whereas the structure of both SF and surfactant assemblies remains unchanged. No disassembly of SL surfactant structures is observed, which gives a new perspective on SF-surfactant interactions. The hydrophobic effect within SF is favored in the presence of SL, leading to faster refolding of SF into ß-sheet conformation. The presented composite gels, being an interpenetrating network of which one compound (SL-C18 : 0) can be tweaked by pH, open an interesting option towards improved workability and stimuli responsive mechanical properties of SF based hydrogels with possible applications in controlled cell culture and tissue engineering or drug delivery. The presented SANS analysis approach has the potential to be expanded to other protein-surfactant systems and composite hydrogels.

Design Principles for Thermoresponsive Core-Shell Nanoparticles: Controlling Thermal Transitions by Brush Morphology.

In this feature article, we summarize our recent work on understanding and controlling the thermal behavior of nanoparticles grafted with thermoresponsive polymer shells. Precision synthesis of monodisperse superparamagnetic iron oxide nanocrystals was combined with irreversible dense grafting of nitrodopamide-anchored thermoresponsive polymer chains. We provide an overview of how the dense and stable grafting of biomedically relevant polymers, including poly(ethylene glycol), poly( N-isopropylacrylamide), polysarcosin, and polyoxazolines, can be achieved. This platform has made it possible for us to demonstrate that the polymer brush geometry, as defined by the nanoparticle core and relative polymer brush size, determines the thermal transitions of the polymer brush. We furthermore summarize our work on how the polymer shell transitions and nanoparticle aggregation can be tuned. With the independent variation of the core and the shell, we can optimize and precisely control the thermally controlled solubility of our system. Finally, our feature article gives examples relevant to current and future applications. We show how the thermal response of the shell influences the nanoparticle performance in biological fluids and interactions with proteins and cells, also under purely magnetic actuation of the nanoparticles through the superparamagnetic iron oxide core.

Biocompatible Glyconanoparticles by Grafting Sophorolipid Monolayers on Monodispersed Iron Oxide Nanoparticles.

This work presents the synthesis and characterization of sophorolipid-coated monodisperse iron oxide nanoparticles. Sophorolipids are biological glycosylated amphiphiles produced by the yeast S. bombicola. In their open acidic form, sophorolipids have been used as a surface stabilizing agent for metal and metal oxide nanoparticles but with a poor control over size and structural properties. In this work, the COOH function of sophorolipids (SL) was modified with nitrodopamine (NDA), a catechol known for its high affinity to iron ions. The resulting new form of sophorolipid-nitrodopamide (SL-NDA) was used as a surface ligand for monodisperse iron oxide nanoparticles. We show by a combination of thermogravimetric analysis and small-angle X-ray and neutron scattering that iron oxide nanoparticles (IONP) are stabilized by a single, high-density SL-NDA layer. This results in excellent colloidal stability under biologically relevant conditions, such as at high protein and salt concentrations. The IONP grafted with SL-NDA showed a negligible uptake by cells and no cytotoxicity, which was tested on two representative cell lines. Thus, they reveal the potential of sophorolipids as stable and nontoxic surface coatings for IONP-based biomedical and biotechnological applications.

Individually Stabilized, Superparamagnetic Nanoparticles with Controlled Shell and Size Leading to Exceptional Stealth Properties and High Relaxivities.

Superparamagnetic iron oxide nanoparticles (SPION) have received immense interest for biomedical applications, with the first clinical application as negative contrast agent in magnetic resonance imaging (MRI). However, the first generation MRI contrast agents with dextran-enwrapped, polydisperse iron oxide nanoparticle clusters are limited to imaging of the liver and spleen; this is related to their poor colloidal stability in biological media and inability to evade clearance by the reticuloendothelial system. We investigate the qualitatively different performance of a new generation of individually PEG-grafted core-shell SPION in terms of relaxivity and cell uptake and compare them to benchmark iron oxide contrast agents. These PEG-grafted SPION uniquely enable relaxivity measurements in aqueous suspension without aggregation even at 9.4 T magnetic fields due to their extraordinary colloidal stability. This allows for determination of the size-dependent scaling of relaxivity, which is shown to follow a d2 dependence for identical core-shell structures. The here introduced core-shell SPION with ∼15 nm core diameter yield a higher R2 relaxivity than previous clinically used contrast agents as well as previous generations of individually stabilized SPION. The colloidal stability extends to control over evasion of macrophage clearance and stimulated uptake by SPION functionalized with protein ligands, which is a key requirement for targeted MRI.

Interaction of Size-Tailored PEGylated Iron Oxide Nanoparticles with Lipid Membranes and Cells.

Targeted nanomedicine builds on the concept that nanoparticles can be directed to specific tissues while remaining inert to others organs. Many studies have been performed on the synthesis and cellular interactions of core-shell nanoparticles, in which a functional inorganic core is coated with a biocompatible polymer layer that should reduce nonspecific uptake and cytotoxicity. However, work is lacking that relates structural parameters of the core-shell structure and colloidal properties directly to interactions with cell membranes and further correlates these interactions to cell uptake. We have synthesized monodisperse (SD < 10%), single-crystalline, and superparamagnetic iron oxide nanoparticles (SPION) of different core size (3-8 nm) that are densely grafted with nitrodopamine-poly(ethylene glycol) (NDA-PEG(5 kDa)) brushes. We investigated the interactions of the PEGylated SPION with biomimetic membranes and cancer and kidney cells. It is shown that a dense homogeneous PEG shell suppresses membrane interactions and cell uptake but that nanoparticle curvature can influence membrane interactions for similarly grafted nanoparticles. Weak adsorption to anionic lipid membranes is shown to correlate with eukaryote cell uptake and is attributed to double-layer interactions without direct membrane penetration. This attraction is strongly suppressed during physiological conditions and leads to unprecedented low cell uptake and full cell viability when compared to those of traditional dextran-coated SPION. Less curved (larger core) PEGylated SPION show weaker membrane adsorption and lower cell uptake due to effectively denser shells. These results provide a better understanding of design criteria for core-shell nanoparticles in terms of avoiding nonspecific uptake by cells, reducing toxicity, and increasing circulation time.

Evaluation of High-Yield Purification Methods on Monodisperse PEG-Grafted Iron Oxide Nanoparticles.

Fundamental research on nanoparticle (NP) interactions and development of next-generation biomedical NP applications relies on synthesis of monodisperse, functional, core-shell nanoparticles free of residual dispersants with truly homogeneous and controlled physical properties. Still, synthesis and purification of e.g. such superparamagnetic iron oxide NPs remain a challenge. Comparing the success of different methods is marred by the sensitivity of analysis methods to the purity of the product. We synthesize monodisperse, oleic acid (OA)-capped, Fe3O4 NPs in the superparamagnetic size range (3-10 nm). Ligand exchange of OA for poly(ethylene glycol) (PEG) was performed with the PEG irreversibly grafted to the NP surface by a nitrodopamine (NDA) anchor. Four different methods were investigated to remove excess ligands and residual OA: membrane centrifugation, dialysis, size exclusion chromatography, and precipitation combined with magnetic decantation. Infrared spectroscopy and thermogravimetric analysis were used to determine the purity of samples after each purification step. Importantly, only magnetic decantation yielded pure NPs at high yields with sufficient grafting density for biomedical applications (∼1 NDA-PEG(5 kDa)/nm(2), irrespective of size). The purified NPs withstand challenging tests such as temperature cycling in serum and long-term storage in biological buffers. Dynamic light scattering, transmission electron microscopy, and small-angle X-ray scattering show stability over at least 4 months also in serum. The successful synthesis and purification route is compatible with any conceivable functionalization for biomedical or biomaterial applications of PEGylated Fe3O4 NPs.

Previous Homologous and Heterologous Stress Exposure Induces Tolerance Development to Pulsed Light in Listeria monocytogenes.

As one of the emerging non-thermal technologies, pulsed light (PL) facilitates rapid, mild and residue-free microbial surface decontamination of food and food contact materials. While notable progress has been made in the characterization of the inactivation potential of PL, experimental data available on the tolerance development to the same (homologous) stress or to different (heterologous) stresses commonly applied in food manufacturing (e.g., acid, heat, salt) is rather controversial. The findings of the present study clearly indicate that both the homologous tolerance development against PL as well as the heterologous tolerance development from heat to PL can be triggered in Listeria monocytogenes. Further, conducted kinetic analysis confirmed that the conventionally applied log-linear model is not well suited to describe the inactivation of L. monocytogenes, when exposed to PL. Instead, the Weibull model as well as the log-linear + tail model were identified as suitable models. Transmission electron microscopic (TEM) approaches allow suggestions on the morphological alterations in L. monocytogenes cells after being subjected to PL.

Core-Shell Structure of Monodisperse Poly(ethylene glycol)-Grafted Iron Oxide Nanoparticles Studied by Small-Angle X-ray Scattering.

The promising applications of core-shell nanoparticles in the biological and medical field have been well investigated in recent years. One remaining challenge is the characterization of the structure of the hydrated polymer shell. Here we use small-angle X-ray scattering (SAXS) to investigate iron oxide core-poly(ethylene glycol) brush shell nanoparticles with extremely high polymer grafting density. It is shown that the shell density profile can be described by a scaling model that takes into account the locally very high grafting density near the core. A good fit to a constant density region followed by a star-polymer-like, monotonously decaying density profile is shown, which could help explain the unique colloidal properties of such densely grafted core-shell nanoparticles. SAXS experiments probing the thermally induced dehydration of the shell and the response to dilution confirmed that the observed features are associated with the brush and not attributed to structure factors from particle aggregates. We thereby demonstrate that the structure of monodisperse core-shell nanoparticles with dense solvated shells can be well studied with SAXS and that different density models can be distinguished from each other.

Complete Exchange of the Hydrophobic Dispersant Shell on Monodisperse Superparamagnetic Iron Oxide Nanoparticles.

High-temperature synthesized monodisperse superparamagnetic iron oxide nanoparticles are obtained with a strongly bound ligand shell of oleic acid and its decomposition products. Most applications require a stable presentation of a defined surface chemistry; therefore, the native shell has to be completely exchanged for dispersants with irreversible affinity to the nanoparticle surface. We evaluate by attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) and thermogravimetric analysis/differential scanning calorimetry (TGA/DSC) the limitations of commonly used approaches. A mechanism and multiple exchange scheme that attains the goal of complete and irreversible ligand replacement on monodisperse nanoparticles of various sizes is presented. The obtained hydrophobic nanoparticles are ideally suited for magnetically controlled drug delivery and membrane applications and for the investigation of fundamental interfacial properties of ultrasmall core-shell architectures.

Melt-grafting for the synthesis of core-shell nanoparticles with ultra-high dispersant density.

Superparamagnetic iron oxide nanoparticles (NPs) are used in a rapidly expanding number of applications in e.g. the biomedical field, for which brushes of biocompatible polymers such as poly(ethylene glycol) (PEG) have to be densely grafted to the core. Grafting of such shells to monodisperse iron oxide NPs has remained a challenge mainly due to the conflicting requirements to replace the ligand shell of as-synthesized NPs with irreversibly bound PEG dispersants. We introduce a general two-step method to graft PEG dispersants from a melt to iron oxide NPs first functionalized with nitrodopamine (NDA). This method yields uniquely dense spherical PEG-brushes (∼3 chains per nm(2) of PEG(5 kDa)) compared to existing methods, and remarkably colloidally stable NPs also under challenging conditions.