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Even although quite a number of studies have been performed so far to demonstrate nanoparticle-specific effects of substances in living systems, clear evidence of these effects is still under debate. The present study was designed as a comparative proteomic analysis of human intestinal cells exposed to a commercial silver nanoparticle reference material and ions from AgNO3. A two-dimensional gel electrophoresis/MALDI mass spectrometry (MS)-based proteomic analysis was conducted after 24-h incubation of differentiated Caco-2 cells with non-cytotoxic and low cytotoxic silver concentrations (2.5 and 25 µg ml−1 nanosilver, 0.5 and 5 µg ml−1 AgNO3). Out of an overall number of 316 protein spots differentially expressed at a fold change of ≥ 1.4 or ≤ −1.4 in all treatments, 169 proteins could be identified. In total, 231 spots were specifically deregulated in particle-treated groups compared with 41 spots, which were limited to AgNO3-treatments. Forty-four spots (14 %) were commonly deregulated by both types of treatment. A considerable fraction of the proteins differentially expressed after treatment with nanoparticles is related to protein folding, synthesis or modification of proteins as well as cellular assembly and organization. Overlays of networks obtained for particulate and ionic treatments showed matches, indicating common mechanisms of combined particle and ionic silver exposure and exclusive ionic silver treatment. However, proteomic responses of Caco-2 cells treated with higher concentrations of silver species also showed some differences, for example regarding proteins related to fatty acid and energy metabolism, suggesting an induction of also some different molecular mechanisms for particle exposure and ionic treatment.
The mechanism of nanotriangle formation in multivesicular vesicles (MMV) is investigated by using time-dependent SAXS measurements in combination with UV−vis spectroscopy, light, and transmission electron microscopy. In the first time period 6.5 nm sized spherical gold nanoparticles are formed inside of the vesicles, which build up soft nanoparticle aggregates. In situ SAXS experiments show a linear increase of the volume and molar mass of nanotriangles in the second time period. The volume growth rate of the triangles is 16.1 nm3/min, and the growth rate in the vertical direction is only 0.02 nm/min. Therefore, flat nanotriangles with a thickness of 7 nm and a diameter of 23 nm are formed. This process can be described by a diffusionlimited Ostwald ripening growth mechanism. TEM micrographs visualize soft coral-like structures with thin nanoplatelets at the periphery of the aggregates, which disaggregate in the third time period into nanotriangles and spherical particles. The 16 times faster growth of nanotriangles in the lateral than that in the vertical direction is related to the adsorption of symmetry breaking components, i.e., AOT and the polyampholyte PalPhBisCarb, on the {111} facets of the gold nanoplatelets in combination with confinement effects of the vesicular template phase.
Characterization of silver nanoparticles in cell culture medium containing fetal bovine serum
(2016)
Nanoparticles are being increasingly used in consumer products worldwide, and their toxicological effects are currently being intensely debated. In vitro tests play a significant role in nanoparticle risk assessment, but reliable particle characterization in the cell culture medium with added fetal bovine serum (CCM) used in these tests is not available. As a step toward filling this gap, we report on silver ion release by silver nanoparticles, and changes in the particle radii and in their protein corona when incubated in CCM. Particles of a certified reference material (CRM), p1, and particles of a commercial silver nanoparticle material, p2, were investigated. The colloidal stability of p1 is provided by the surfactants polyethylene glycole-25 glyceryl trioleate and polyethylene glycole-20 sorbitan monolaurate, whereas p2 is stabilized by polyvinylpyrrolidone (PVP). Dialysis of p1 and p2 reveal that their silver ion release rates in CCM are much larger than in water. Particle characterization was performed with asymmetrical flow field-flow fractionation (FFF), small-angle X-ray scattering (SAXS), dynamic light scattering (DLS) and electron microscopy. p1 and p2 have similar hydrodynamic radii of 15 nm and 16 nm, respectively. The silver core radii are 9.2 and 10.2 nm. Gel electrophoresis and subsequent peptide identification reveal that albumin is the main corona component of p1 and p2 after incubation in CCM, which consists of Dulbeccos Modified Eagle Medium with 10% fetal bovine serum added.
Aluminum has gathered toxicological Attention based on relevant human exposure and its suspected hazardous potential. Nanoparticles from food supplements or Food contact materials may reach the human gastrointestinal tract.
Here, we monitored the physicochemical fate of aluminum containing nanoparticles and aluminum ions when passaging an in vitro model of the human gastrointestinal tract. Smallangle X-ray scattering (SAXS), transmission electron microscopy (TEM), ion beam microscopy (IBM), secondary ion beam mass spectrometry (TOF-SIMS), and inductively coupled plasma mass spectrometry (ICP-MS) in the singleparticle mode were employed to characterize two aluminumcontaining nanomaterials with different particle core materials (Al0, γAl2O3) and soluble AlCl3. Particle size and shape remained unchanged in saliva, whereas strong Agglomeration of both aluminum nanoparticle species was observed at low pH in gastric fluid together with an increased ion release. The levels of free aluminum ions decreased in intestinal fluid and the particles deagglomerated, thus liberating primary particles again. Dissolution of nanoparticles was limited and substantial changes of their shape and size were not detected. The amounts of particle-associated phosphorus, chlorine, potassium, and calcium increased in intestinal fluid, as compared to nanoparticles in standard dispersion.
Interestingly, nanoparticles were found in the intestinal fluid after addition of ionic aluminum. We provide a comprehensive characterization of the fate of aluminum nanoparticles in simulated gastrointestinal fluids, demonstrating that orally ingested nanoparticles probably reach the intestinal epithelium. The balance between dissolution and de novo complex formation should be considered when evaluating nanotoxicological experiments.
Size and shape are crucial parameters which have impact on the potential of nanoparticles to penetrate cell membranes and epithelial barriers. Current research in nanotoxicology additionally focuses on particle coating. To distinguish between core- and coating-related effects in nanoparticle uptake and translocation, two nanoparticles equal in size, coating and charge but different in core material were investigated.
Silver and iron oxide nanoparticles coated with poly(acrylic acid) were chosen and extensively characterized by small-angle x-ray scattering, nanoparticle tracing analysis and transmission electron microscopy (TEM). Uptake and transport were studied in the intestinal Caco-2 model in a Transwell System with subsequent elemental analysis. TEM and ion beam microscopy were conducted for particle visualization.
Although equal in size, charge and coating, the behavior of the two particles in Caco-2 cells was different: while the internalized amount was comparable, only iron oxide nanoparticles additionally passed the epithelium. Our findings suggest that the coating material influenced only the uptake of the nanoparticles whereas the translocation was determined by the core material.
Knowledge about the different roles of the particle coating and core materials in crossing biological barriers will facilitate toxicological risk assessment of nanoparticles and contribute to the optimization of pharmacokinetic properties of nano-scaled pharmaceuticals.
tNegatively charged ultrathin gold nanotriangles (AuNTs) were synthesized in a vesicular dioctyl sodiumsulfosuccinate (AOT)/phospholipid-based template phase. These “naked” AuNTs with localized surfaceplasmon resonances in the NIR region at about 1300 nm and special photothermal properties are ofparticular interest for imaging and hyperthermia of cancerous tissues. For these kinds of applicationsthe toxicity and the cellular uptake of the AuNTs is of outstanding importance. Therefore, this studyfocuses on the toxicity of “naked” AOT-stabilized AuNTs compared to polymer-coated AuNTs. Poly-meric coating consisted of non-modified hyperbranched poly(ethyleneimine) (PEI), maltose-modifiedpoly(ethyleneimine) (PEI-Mal) and heparin. The toxicological experiments were carried out with twodifferent cell lines (embryonic kidney carcinoma cell line HEK293T and NK-cell leukemia cell line YTS).This study revealed that the heparin-coating of AuNTs improved biocompatibility by a factor of 50 whencompared to naked AuNTs. Of note, the highest nontoxic concentration of the AuNTs coated with PEI andPEI-Mal is drastically decreased. Overall, this is mainly triggered by the different surface charges of poly-meric coatings. Therefore, AuNTs coated with heparin were selected to carry out uptake studies. Theirpromising high biocompatibility and cellular uptake may open future studies in the field of biomedicalapplications.
Core-shell nanoparticles are widespread in nature, industrial applications and nanotechnology research. Facile ways of modern synthesis will be discussed and possibilities to reveal their structures with small-angle X-ray scattering (SAXS). A recent review on using block copolymer templates as one of the most reliable routes for tuning size and shape of nanoparticles is provided by Li et al.1 Ferritin and apoferritin are archetypical examples for protein-based core-shell nanoparticles. Their structures are easily accessed by synchrotron SAXS2 but also with commercial instruments and allow fast performance tests.3 SASfit4 is a suitable program tool based on classical curve fitting and McSAS5 is a complementary program based on a Monte Carlo technique. Detailed refinements of SAXS data evaluation are on the way for better data analysis.6 A sub nanometer resolution is state-of-the-art for quantification of the size distribution of polyacrylic acid stabilized silver nanoparticles.3 Such particles are useful in catalysis.7 It was observed that the catalytic activity can be tuned easily by varying the shell material of the particles.
Fate of fluorescence labels - Their adsorption and desorption kinetics to silver nanoparticles
(2018)
Silver nanoparticles are among the most widely used and produced nanoparticles. Because of their frequent application in consumer products, the assessment of their toxicological potential has seen a renewed importance. A Major difficulty is the traceability of nanoparticles in in vitro and in vivo experiments. Even if the particles are labeled, for example, by a fluorescent marker, the dynamic exchange of ligands often prohibits their spatial localization. Our study provides an insight into the adsorption and desorption kinetics of two different fluorescent labels on silver nanoparticles with a core radius of 3 nm by dynamic light scattering, small-angle X-ray scattering, and fluorescence spectroscopy. We used BSA-FITC and tyrosine as examples for common fluorescent ligands. It is shown that the adsorption of BSA-FITC takes at least 3 days, whereas tyrosine adsorbs immediately. The quantitative amount of stabilizer on the particle surface was determined by fluorescence spectroscopy and revealed that the particles are stabilized by a monolayer of BSA-FITC (corresponding to 20 ± 9 molecules), whereas tyrosine forms a multilayered structure consisting of 15900 ± 200 molecules. Desorption experiments show that the BSA-FITC-stabilized particles are ideally suited for application in in vitro and in vivo experiments because the ligand desorption takes several days. Depending on the BSA concentration in the particles surroundings, the rate constant is k = 0.2 per day or lower when applying first order kinetics, that is, 50% of the BSAFITC molecules are released from the particle’s surface within 3.4 days. For illustration, we provide a first application of the fluorescence-labeled particles in an uptake study with two different commonly used cell lines, the human liver cell model HepG2 and the human intestinal cell model of differentiated Caco-2 cells.
Polymeric core-shell particles were synthesized in a semi-batch emulsion polymerization process. The shell of the particles consist of PVDF with a high amount of beta-phase. Small-angle X-ray scattering (SAXS) was used to quantify the size of the cores of the particles and the thickness of the shell.
In contrast to microplastics, little is known about nanoplastics (1 to 100 nm). In order to make the dectecability of nanoplasics more reliable, we started to develop nanoplastic reference materials. This project also aims to anser the question of how the single chain conformation of bio(polymers) changes in contact with nanoplastics. Small-angle X-ray and neutron scattering methods are suitable methods for studing this topic. Recently the soft and hard interactions between polystyrene nanoplasics and human serum albumin corona was investigated with small-angle neutron scattering. Here we concentrate on small-angle X-ray scattering as our favorite method to study how (bio)polymers change their conformation in contact with nanoplastics. The scattering of bovine serum albumin in its native state can be detected easily. The scattering pattern of this biopolymer changes dramatically when its globular stucture changes to a coil structure. Modeling of chain conformations and the calculation of the scattering pattern is relatively easy to perform. Numerous model calculations will be provided to predict the changes of conformation of single bio(polymer) chains when in conatact with nanoplastics. These predictions will be compared with recent experimenal results from in situ measurments of bio(polymers) in contact with nanoplastics. The impact of temperature, polymer concentration and salt on the single-chain conformation changes will be discussed.