Mechanistic Similarities between 3D Human Bronchial Epithelium and Mice Lung, Exposed to Copper Oxide Nanoparticles, Support Non-Animal Methods for Hazard Assessment.

The diversity and increasing prevalence of products derived from engineered nanomaterials (ENM), warrants implementation of non-animal approaches to health hazard assessment for ethical and practical reasons. Although non-animal approaches are becoming increasingly popular, there are almost no studies of side-by-side comparisons with traditional in vivo assays. Here, transcriptomics is used to investigate mechanistic similarities between healthy/asthmatic models of 3D air-liquid interface (ALI) cultures of donor-derived human bronchial epithelia cells, and mouse lung tissue, following exposure to copper oxide ENM. Only 19% of mouse lung genes with human orthologues are not expressed in the human 3D ALI model. Despite differences in taxonomy and cellular complexity between the systems, a core subset of matching genes cluster mouse and human samples strictly based on ENM dose (exposure severity). Overlapping gene orthologue pairs are highly enriched for innate immune functions, suggesting an important and maybe underestimated role of epithelial cells. In conclusion, 3D ALI models based on epithelial cells, are primed to bridge the gap between traditional 2D in vitro assays and animal models of airway exposure, and transcriptomics appears to be a unifying dose metric that links in vivo and in vitro test systems.

A practical approach to assess inhalation toxicity of metal oxide nanoparticles in vitro.

Exposure of humans to metal oxide nanoparticles (NPs) occurs mainly via air, and inhaled metal oxide NPs may generate inflammation. The aim of this study was to investigate the proinflammatory potential of six metal oxide NPs (CeO2 , Mn2 O3 , CuO, ZnO, Co3 O4 and WO3 ; 27-108 µg ml-1 ) using human primary 3-dimensional airway epithelium (MucilAir™) and dendritic cell (DC) models. Metal oxide NPs were mainly aggregated/agglomerated in the cell media, as determined by dynamic light scattering, scanning electron microscopy and differential centrifugal sedimentation. WO3 and ZnO were highly soluble, both with and without respiratory mucus. Proinflammatory signalling by the epithelium was evaluated after a 24 hour exposure by increased interleukin-6 and -8 and monocyte chemoattractant protein 1 cytokine release, which occurred only for CuO. Moreover, maturation of immature human DCs, which play a key role in the lung immune system, were evaluated by expression of surface markers HLA-DR, CD80, CD83 and CD86 after a 48 hour exposure. Only Mn2 O3 consistently upregulated DC maturation markers. Furthermore, by addition of medium from metal oxide NP-exposed 3-dimensional airway cultures to metal oxide NP-exposed DC cultures, the interplay between lung epithelium and DCs was studied. Such an interplay was again only observed for Mn2 O3 and in one of five DC donors. Our results show that, even when using dosages that represent very high in vivo exposure levels, up to 27 hours of constant human airway exposure, metal oxide NPs cause minimal proinflammatory effects and that epithelial cells not necessarily interfere with DC maturation upon metal oxide NP exposure. The present approach exemplifies a relevant translation towards human safety assessment.

Physicochemical characterization of titanium dioxide pigments using various techniques for size determination and asymmetric flow field flow fractionation hyphenated with inductively coupled plasma mass spectrometry.

Seven commercial titanium dioxide pigments and two other well-defined TiO2 materials (TiMs) were physicochemically characterised using asymmetric flow field flow fractionation (aF4) for separation, various techniques to determine size distribution and inductively coupled plasma mass spectrometry (ICPMS) for chemical characterization. The aF4-ICPMS conditions were optimised and validated for linearity, limit of detection, recovery, repeatability and reproducibility, all indicating good performance. Multi-element detection with aF4-ICPMS showed that some commercial pigments contained zirconium co-eluting with titanium in aF4. The other two TiMs, NM103 and NM104, contained aluminium as integral part of the titanium peak eluting in aF4. The materials were characterised using various size determination techniques: retention time in aF4, aF4 hyphenated with multi-angle laser light spectrometry (MALS), single particle ICPMS (spICPMS), scanning electron microscopy (SEM) and particle tracking analysis (PTA). PTA appeared inappropriate. For the other techniques, size distribution patterns were quite similar, i.e. high polydispersity with diameters from 20 to >700 nm, a modal peak between 200 and 500 nm and a shoulder at 600 nm. Number-based size distribution techniques as spICPMS and SEM showed smaller modal diameters than aF4-UV, from which mass-based diameters are calculated. With aF4-MALS calculated, light-scattering-based "diameters of gyration" (Øg) are similar to hydrodynamic diameters (Øh) from aF4-UV analyses and diameters observed with SEM, but much larger than with spICPMS. A Øg/Øh ratio of about 1 indicates that the TiMs are oblate spheres or fractal aggregates. SEM observations confirm the latter structure. The rationale for differences in modal peak diameter is discussed.

Potential Release of Manufactured Nano Objects During Sanding of Nano-Coated Wood Surfaces.

Increasing production and applications of manufactured nano objects (MNOs) have become a source for human exposure and therefore raise concerns and questions about the possible health effects. In this study, the potential release of nano objects, their agglomerates, and aggregates (NOAA) as a result of sanding of hardwood treated with MNOs-containing coating was examined. Two types of MNO-containing coating were compared with untreated hardwood that allowed the evaluation of the influence of the chemical composition on the release of particles. Furthermore, the rotation speed of the sander and the grit size of the sanding paper were varied in order to assess their influence on the release of particles.Measurements were conducted in a gas-tight chamber with a volume of 19.5 m(3) in which ventilation was minimized during experiments. Particle size distributions were assessed by scanning mobility particle sizer , aerodynamic particle sizer, and electrical low pressure impactor. Furthermore, aerosol number concentrations (Nanotracer), active surface area (LQ1), and fractionated mass (Cascade Impactor) were measured before, during, and after sanding. Scanning electron microscope/energy dispersive X-ray (SEM/EDX) analysis was performed to adequately characterize the morphology, size, and chemical composition of released particles.SEM/EDX analysis indicated that sanding surfaces treated with MNO-containing coating did not release the designated MNO as free primary particles. In both coatings, clusters of MNO were perceived embedded in and attached to micro-sized wood and/or coating particles created by sanding the coated surface. Real-time measurements indicated a lower release of micro-sized particles from sanding of surfaces treated with Coating I than from sanding untreated surfaces or surfaces treated with Coating II. A substantial increase in nanosized and a slight increase in micro-sized particles was perceived as the rotation speed of the sander increased. However, most nanosized particles were most likely emitted by the sanding machine. No effect of the grit size on the release of particles was detected.

Occupational Exposure to Multi-Walled Carbon Nanotubes During Commercial Production Synthesis and Handling.

The world-wide production of carbon nanotubes (CNTs) has increased substantially in the last decade, leading to occupational exposures. There is a paucity of exposure data of workers involved in the commercial production of CNTs. The goals of this study were to assess personal exposure to multi-walled carbon nanotubes (MWCNTs) during the synthesis and handling of MWCNTs in a commercial production facility and to link these exposure levels to specific activities. Personal full-shift filter-based samples were collected, during commercial production and handling of MWCNTs, R&D activities, and office work. The concentrations of MWCNT were evaluated on the basis of EC concentrations. Associations were studied between observed MWCNT exposure levels and location and activities. SEM analyses showed MWCNTs, present as agglomerates ranging between 200 nm and 100 µm. Exposure levels of MWCNTs observed in the production area during the full scale synthesis of MWCNTs (N = 23) were comparable to levels observed during further handling of MWCNTs (N = 19): (GM (95% lower confidence limit-95% upper confidence limit)) 41 µg m(-3) (20-88) versus 43 µg m(-3) (22-86), respectively. In the R&D area (N = 11) and the office (N = 5), exposure levels of MWCNTs were significantly (P < 0.05) lower: 5 µg m(-3) (2-11) and 7 µg m(-3) (2-28), respectively. Bagging, maintenance of the reactor, and powder conditioning were associated with higher exposure levels in the production area, whereas increased exposure levels in the R&D area were related to handling of MWCNTs powder.

Sub-chronic toxicity study in rats orally exposed to nanostructured silica.

BACKGROUND: Synthetic Amorphous Silica (SAS) is commonly used in food and drugs. Recently, a consumer intake of silica from food was estimated at 9.4 mg/kg bw/day, of which 1.8 mg/kg bw/day was estimated to be in the nano-size range. Food products containing SAS have been shown to contain silica in the nanometer size range (i.e. 5-200 nm) up to 43% of the total silica content. Concerns have been raised about the possible adverse effects of chronic exposure to nanostructured silica. METHODS: Rats were orally exposed to 100, 1000 or 2500 mg/kg bw/day of SAS, or to 100, 500 or 1000 mg/kg bw/day of NM-202 (a representative nanostructured silica for OECD testing) for 28 days, or to the highest dose of SAS or NM-202 for 84 days. RESULTS: SAS and NM-202 were extensively characterized as pristine materials, but also in the feed matrix and gut content of the animals, and after in vitro digestion. The latter indicated that the intestinal content of the mid/high-dose groups had stronger gel-like properties than the low-dose groups, implying low gelation and high bioaccessibility of silica in the human intestine at realistic consumer exposure levels. Exposure to SAS or NM-202 did not result in clearly elevated tissue silica levels after 28-days of exposure. However, after 84-days of exposure to SAS, but not to NM-202, silica accumulated in the spleen. Biochemical and immunological markers in blood and isolated cells did not indicate toxicity, but histopathological analysis, showed an increased incidence of liver fibrosis after 84-days of exposure, which only reached significance in the NM-202 treated animals. This observation was accompanied by a moderate, but significant increase in the expression of fibrosis-related genes in liver samples. CONCLUSIONS: Although only few adverse effects were observed, additional studies are warranted to further evaluate the biological relevance of observed fibrosis in liver and possible accumulation of silica in the spleen in the NM-202 and SAS exposed animals respectively. In these studies, dose-effect relations should be studied at lower dosages, more representative of the current exposure of consumers, since only the highest dosages were used for the present 84-day exposure study.

Swimming pool attendance and respiratory symptoms and allergies among Dutch children.

OBJECTIVES: To describe associations among swimming, respiratory health, allergen sensitisation and Clara cell protein 16 (CC16) levels in Dutch schoolchildren. Trichloramine levels in swimming pool air were determined to assess potential exposure levels. METHODS: Respiratory health and pool attendance information was collected from 2359 children, aged 6-13 years. Serum from 419 children was tested for allergen sensitisation and CC16 levels. Trichloramine levels were assessed in nine swimming facilities. RESULTS: Trichloramine levels ranged from 0.03 to 0.78 mg/m3 (average 0.21 mg/m3). Reported swimming pool attendance and trichloramine exposure were both not associated with asthma, wheezing, rhinitis or CC16 levels. Birch and house dust mite sensitisation were associated with recent indoor swimming (OR>1.86), but not after considering recent swimming frequency multiplied by trichloramine levels. Sensitisation to house dust mites was associated with frequent baby swimming (ORs=1.75; 95% CI 1.09 to 2.79). Furthermore, sensitisation was associated with lower serum CC16 levels. CC16 levels were associated with average trichloramine concentrations in pools; however, not after considering swimming frequency multiplied by trichloramine levels. CONCLUSIONS: Measured trichloramine levels were comparable with other studies but lower than in an earlier Dutch study. Swimming pool attendance was not associated with respiratory symptoms. The association between sensitisation and swimming during the first 2 years of life suggests that early-life exposures might be important, although this needs further study. The interpretation of transient and chronic changes of CC16 and other inflammatory markers in relation to the pool environment and health impacts warrants further investigation. Detailed comparisons with other studies are limited as few studies have measured trichloramine levels.

The Development and Calibration of a Mechanistic Asbestos Removal Exposure Assessment Tool (AREAT).

Exposure to asbestos fibres is linked to numerous adverse health effects and the use of asbestos is currently banned in many countries. Still, asbestos applications are present in numerous residential and professional/industrial buildings or installations which need to be removed. Exposure measurements give good insight in exposure levels on the basis of which the required control regime is determined to ensure that workers are protected against adverse health effects. However, it is a costly and time-consuming process to measure all situations as working conditions and materials may vary greatly. Therefore, the mechanistic model 'Asbestos Removal Exposure Assessment Tool (AREAT)' was developed to estimate exposure to respirable asbestos fibres released during asbestos abatement processes where measurements are not available. In such instances tailored control regimes can be implemented based on modelled exposure levels. The mechanistic model was developed using scientific literature, an in-house asbestos abatement dataset, and knowledge with regard to previously developed models. Several exposure determinants such as the substance emission potential, activity emission potential, control measures, and dilution in air were identified and specific modifiers were developed for each category. Through an algorithm, AREAT calculates a dimensionless score based on the model inputs. The model was calibrated using a statistical model on an extensive measurement dataset containing a broad variety of exposure scenarios. This statistical model enabled the translation of dimensionless AREAT scores to actual estimated fibre concentrations in fibres m-3. In total, 370 personal inhalation exposure measurements from 71 different studies were used for calibration of AREAT. Of these measurements, in 191 cases (52%) with microscopic analysis (all asbestos fibre analyses were conducted with scanning electron microscopy/energy dispersive X-ray analysis in accordance with ISO 14966) no fibres were detected and the limit of detection values(LODs) were given. To assess the influence of the large number of measurements with exposures below LOD values on the performance of the model, calibrations were performed on the total dataset and the selection of data excluding measurements below LOD. The AREAT model correlated well with the datasets, with a Pearson correlation of 0.73 and 0.8 and Spearman rank correlation of 0.56 and 0.8. The model was fitted to estimate a typical exposure value [i.e. geometric mean (GM) exposures], but it is recommended to use a more conservative worst case higher percentile (for example the 90th percentile; which adds a factor of 17.3 based on the model uncertainty on the GM estimate), to account for variability in the measurements and uncertainty in model estimates. This work has shown the development and calibration of a mechanistic model, capable of estimating asbestos fibre exposures during asbestos abatement processes. The AREAT model will be implemented as a lower tier exposure model in a risk assessment tool used within the Netherlands to plan abatement processes and to develop control strategies.

Silicon dioxide and titanium dioxide particles found in human tissues.

Silicon dioxide (silica, SiO2, SAS) and titanium dioxide (TiO2) are produced in high volumes and applied in many consumer and food products. As a consequence, there is a potential human exposure and subsequent systemic uptake of these particles. In this study we show the characterization and quantification of both total silicon (Si) and titanium (Ti), and particulate SiO2 and TiO2 in postmortem tissue samples from 15 deceased persons. Included tissues are liver, spleen, kidney and the intestinal tissues jejunum and ileum. Low-level analysis was enabled by the use of fully validated sample digestion methods combined with (single particle) inductively coupled plasma high resolution mass spectrometry techniques (spICP-HRMS). The results show a total-Si concentration ranging from <2 to 191 mg Si/kg (median values of 5.8 (liver), 9.5 (spleen), 7.7 (kidney), 6.8 (jejunum), 7.6 (ileum) mg Si/kg) while the particulate SiO2 ranged from <0.2 to 25 mg Si/kg (median values of 0.4 (liver), 1.0 (spleen), 0.4 (kidney), 0.7 (jejunum, 0.6 (ileum) mg Si/kg), explaining about 10% of the total-Si concentration. Particle sizes ranged from 150 to 850 nm with a mode of 270 nm. For total-Ti the results show concentrations ranging from <0.01 to 2.0 mg Ti/kg (median values of 0.02 (liver), 0.04 (spleen), 0.05 (kidney), 0.13 (jejunum), 0.26 (ileum) mg Ti/kg) while particulate TiO2 concentrations ranged from 0.01 to 1.8 mg Ti/kg (median values of 0.02 (liver), 0.02 (spleen), 0.03 (kidney), 0.08 (jejunum), 0.25 (ileum) mg Ti/kg). In general, the particulate TiO2 explained 80% of the total-Ti concentration. This indicates that most Ti in these organ tissues is particulate material. The detected particles comprise primary particles, aggregates and agglomerates, and were in the range of 50-500 nm with a mode in the range of 100-160 nm. About 17% of the detected TiO2 particles had a size <100 nm. The presence of SiO2 and TiO2 particles in liver tissue was confirmed by scanning electron microscopy with energy dispersive X-ray spectrometry.

Calibration of polydimethylsiloxane and polyurethane foam passive air samplers for measuring semi volatile organic compounds using a novel exposure chamber design.

Passive air sampling is increasingly used for air quality monitoring and for personal sampling. In a novel experimental exposure chamber study, 3 types of polydimethylsiloxane (PDMS, including sheet and wristband) and 1 type of polyurethane foam (PUF) passive air samplers were tested for gas-phase uptake of 200 semi volatile organic compounds (SVOCs) during six months. For 155 SVOCs including PAH, PCB, phthalates, organophosphate esters, musk compounds, organochlorine- and other pesticides, a normalized generic uptake rate (Rs) of 7.6 ±â€¯1.3 m3 d-1 dm-2 and a generic mass transfer coefficient (MTC) of 0.87 ±â€¯0.15 cm s-1 at a wind speed of 1.3 m s-1 were determined. Variability of sampling rates within and between passive sampling media and analyte groups was not statistically significant, supporting the hypothesis of air-side controlled uptake regardless of sampling material. A statistical relationship was developed between the sampling rate and windspeed which can be used to obtain a sampling rate applicable to specific deployment conditions. For 98 SVOCs, partition coefficients (Ksampler-air) for PUF and PDMS were obtained, which determine the duration of linear uptake and capacity of the sampler for gas-phase uptake. Ksampler-air for PDMS were approximately 10 times higher than for PUF, suggesting that PDMS can be deployed for longer time per volume of sampler, while uptake remains in the linear phase. Statistical relationships were developed to estimate Kpuf-air and Kpdms-air from Koa. These results improve the understanding of the performance of PDMS and PUF passive samplers and contribute to the development of PDMS for the use as a promising personal sampler.

Molecular Signature of Asthma-Enhanced Sensitivity to CuO Nanoparticle Aerosols from 3D Cell Model.

More than 5% of any population suffers from asthma, and there are indications that these individuals are more sensitive to nanoparticle aerosols than the healthy population. We used an air-liquid interface model of inhalation exposure to investigate global transcriptomic responses in reconstituted three-dimensional airway epithelia of healthy and asthmatic subjects exposed to pristine (nCuO) and carboxylated (nCuOCOOH) copper oxide nanoparticle aerosols. A dose-dependent increase in cytotoxicity (highest in asthmatic donor cells) and pro-inflammatory signaling within 24 h confirmed the reliability and sensitivity of the system to detect acute inhalation toxicity. Gene expression changes between nanoparticle-exposed versus air-exposed cells were investigated. Hierarchical clustering based on the expression profiles of all differentially expressed genes (DEGs), cell-death-associated DEGs (567 genes), or a subset of 48 highly overlapping DEGs categorized all samples according to "exposure severity", wherein nanoparticle surface chemistry and asthma are incorporated into the dose-response axis. For example, asthmatics exposed to low and medium dose nCuO clustered with healthy donor cells exposed to medium and high dose nCuO, respectively. Of note, a set of genes with high relevance to mucociliary clearance were observed to distinctly differentiate asthmatic and healthy donor cells. These genes also responded differently to nCuO and nCuOCOOH nanoparticles. Additionally, because response to transition-metal nanoparticles was a highly enriched Gene Ontology term (FDR 8 × 10-13) from the subset of 48 highly overlapping DEGs, these genes may represent biomarkers to a potentially large variety of metal/metal oxide nanoparticles.

A New Approach Combining Analytical Methods for Workplace Exposure Assessment of Inhalable Multi-Walled Carbon Nanotubes.

To date there is no consensus about the most appropriate analytical method for measuring carbon nanotubes (CNTs), hampering the assessment and limiting the comparison of data. The goal of this study is to develop an approach for the assessment of the level and nature of inhalable multi-wall CNTs (MWCNTs) in an actual workplace setting by optimizing and evaluating existing analytical methods. In a company commercially producing MWCNTs, personal breathing zone samples were collected for the inhalable size fraction with IOM samplers; which were analyzed with carbon analysis, inductively coupled plasma mass spectrometry (ICP-MS) and scanning electron microscopy/energy dispersive X-ray spectroscopy (SEM/EDX). Analytical methods were optimized for carbon analysis and SEM/EDX. More specifically, methods were applied and evaluated for background correction using carbon analyses and SEM/EDX, CNT structure count with SEM/EDX and subsequent mass conversion based on both carbon analyses and SEM/EDX. A moderate-to-high concordance correlation coefficient (RC) between carbon analyses and SEM/EDX was observed [RC = 0.81, 95% confidence interval (CI): 0.59-0.92] with an absolute mean difference of 59 µg m-3. A low RC between carbon analyses and ICP-MS (RC = 0.41, 95% CI: 0.07-0.67) with an absolute mean difference of 570 µg m-3 was observed. The large absolute difference between EC and metals is due to the presence of non-embedded inhalable catalyst particles, as a result of which MWCNT concentrations were overestimated. Combining carbon analysis and SEM/EDX is the most suitable for quantitative exposure assessment of MWCNTs in an actual workplace situation.

Quartz in ash, and air in a high lung cancer incidence area in China.

Exposure to crystalline silica (quartz) has been implicated as a potential cause of the high lung cancer rates in the neighbouring counties of Xuanwei and Fuyuan, China, where the domestic combustion of locally sourced "smoky" coal (a bituminous coal) is responsible for some of the highest lung cancer rates in the nation, irrespective of gender or smoking status. Previous studies have shown that smoky coal contains approximately twice as much quartz when compared to alternative fuels in the area, although it is unclear how the quartz in coal relates to household air pollution. Samples of ash and fine particulate matter (PM2.5) were collected from 163 households and analysed for quartz content by Fourier transformed infrared spectroscopy (FT-IR). Additionally, air samples from 12 further households, were analysed by scanning electron microscopy (SEM) to evaluate particle structure and silica content. The majority (89%) of household air samples had undetectable quartz levels (<0.2 µg/m3) with no clear differences by fuel-type. SEM analyses indicated that there were higher amounts of silica in the smoke of smoky coal than smokeless coal (0.27 µg/m3 vs. 0.03 µg/m3). We also identified fibre-like particles in a higher concentration within the smoke of smoky coal than smokeless coal (5800 fibres/m3 vs. 550 fibres/m3). Ash analysis suggested that the bulk of the quartz in smoky coal went on to form part of the ash. These findings indicate that the quartz within smoky coal does not become adequately airborne during the combustion process to cause significant lung cancer risk, instead going on to form part of the ash. The identification of fibre-like particles in air samples is an interesting finding, although the clinical relevance of this finding remains unclear.

Is there evidence for man-made nanoparticles in the Dutch environment?

Only very limited information is available on measured environmental concentrations of nanoparticles. In this study, several environmental compartments in The Netherlands were probed for the presence of nanoparticles. Different types of water were screened for the presence of inorganic (Ag, Au, TiO2) and organic nanoparticles (C60, C70, [6,6]-phenyl-C61-butyric acid octyl ester, [6,6]-phenyl-C61-butyric acid butyl ester, [6,6]-phenyl-C61-butyric acid methyl ester, [6,6]-bis-phenyl-C61-butyric acid methyl ester, [6,6]-phenyl-C71-butyric acid methyl ester, [6,6]-thienyl-C61-butyric acid methyl ester). Air samples were analysed for the presence of nanoparticulate Mo, Ag, Ce, W, Pd, Pt, Rh, Zn, Ti, Si, B as well as Fe and Cu. ICP-MS, Orbitrap-HRMS, SEM and EDX were used for this survey. Water samples included dune and bank filtrates, surface waters and ground waters as well as influents, effluents and sludge of sewage treatment plants (STPs), and surface waters collected near airports and harbours. Air samples included both urban and rural samples. C60 was detected in air, sewage treatment plants, influents, effluents and sludge, but in no other aqueous samples despite the low detection limit of 0.1ng/L. C70 and functionalised fullerenes were not detected at all. In STP sludge and influent the occurrence of Ag and Au nanoparticles was verified by SEM/EDX and ICP-MS. In air up to about 25m% of certain metals was found in the nanosize fraction. Overall, between 1 and 6% of the total mass from metals in the air samples was found in the size fraction <100nm.

Transfer Study of Silver Nanoparticles in Poultry Production.

Silver nanoparticles (AgNPs) are of interest due to their antimicrobial activity and are seen as potential candidates to replace antibiotics in animal husbandry. A few studies have focused on this new application, but they lack any considerations about residual accumulation of AgNPs in edible animal tissues and animal products. In this research, a 22 day in vivo study was carried out by oral administration of 20 nm spherical PVP coated AgNPs to hens. Six doses of approximately 1 mg kg-1 of AgNPs-PVP each were administered to animals throughout the experimentation. Atomic absorption spectroscopy (AAS) was used for quantitative determination of residual total Ag in different organs and matrices. The analyses showed that Ag accumulates in livers (concentration ranging from 141 µg kg-1 to 269 µg kg-1) and yolks (concentration ranging from 20 µg kg-1 to 49 µg kg-1) but not in muscles, kidneys, and albumen belonging to hens of the treated group (tG2). Ag was not detected in animals of the control group (uG1) (i.e., total Ag < LOD = 10 µg kg-1). Single particle inductively coupled plasma mass spectrometry (spICP-MS) and scanning electron microscopy with energy dispersive X-ray detection (SEM-EDX) were employed to elucidate the presence of AgNPs in livers and yolks belonging to tG2 animals. spICP-MS highlighted that part of residual Ag found in livers (about 5-20%) is in NP form with an average dimension of approximately 20 nm. SEM-EDX technique confirmed the presence of AgNPs only in livers of treated animals. The results show that feeding AgNPs to hens may become a source of consumer exposure to AgNPs. As far as we know this is the first study showing transfer of AgNPs or reaction products thereof from animal feed to animal products.

Translocation of positively and negatively charged polystyrene nanoparticles in an in vitro placental model.

To obtain insight in translocation of nanoparticles across the placental barrier, translocation was studied for one positively and two negatively charged polystyrene nanoparticles (PS-NPs) of similar size in an in vitro model. The model consisted of BeWo b30 cells, derived from a human choriocarcinoma grown on a transwell insert forming a cell layer that separates an apical from a basolateral compartment. PS-NPs were characterized with respect to size, surface charge, morphology and protein corona. Translocation of PS-NPs was not related to PS-NP charge. Two PS-NPs were translocated across the BeWo transwell model to a lower extent than amoxicillin, a model compound known to be translocated over the placental barrier to only a limited extent, whereas one PS-NP showed a slightly higher translocation. Studies on the effect of transporter inhibitors on the translocation of the PS-NPs indicated that their translocation was not mediated by known transporters and mainly dependent on passive diffusion. It is concluded that the BeWo b30 model can be used as an efficient method to get an initial qualitative impression about the capacity of NPs to translocate across the placental barrier and set priorities in further in vivo studies on translocation of NPs to the fetus.

Translocation of differently sized and charged polystyrene nanoparticles in in vitro intestinal cell models of increasing complexity.

Intestinal translocation is a key factor for determining bioavailability of nanoparticles (NPs) after oral uptake. Therefore, we evaluated three in vitro intestinal cell models of increasing complexity which might affect the translocation of NPs: a mono-culture (Caco-2 cells), a co-culture with mucus secreting HT29-MTX cells and a tri-culture with M-cells. Cell models were exposed to well characterized differently sized (50 and 100 nm) and charged (neutral, positively and negatively) polystyrene NPs. In addition, two types of negatively charged NPs with different surface chemistries were used. Size strongly affected the translocation of NPs, ranging up to 7.8% for the 50 nm NPs and 0.8% for the 100 nm NPs. Surface charge of NPs affected the translocation, however, surface chemistry seems more important, as the two types of negatively charged 50 nm NPs had an over 30-fold difference in translocation. Compared with the Caco-2 mono-culture, presence of mucus significantly reduced the translocation of neutral 50 nm NPs, but significantly increased the translocation of one type of negatively charged NPs. Incorporation of M-cells shifted the translocation rates for both NPs closer to those in the mono-culture model. The relative pattern of NP translocation in all three models was similar, but the absolute amounts of translocated NPs differed per model. We conclude that for comparing the relative translocation of different NPs, using one intestinal model is sufficient. To choose the most representative model for risk assessment, in vivo experiments are now needed to determine the in vivo translocation rates of the used NPs.

Airborne manufactured nano-objects released from commercially available spray products: temporal and spatial influences.

This paper reports a study of the dispersion of manufactured nano-objects (MNOs) through the air, both in time and space, during the use of two commercially available nano-spray products and comparable products without MNOs. The main objective was to identify whether personal exposure can occur at a greater distance than the immediate proximity of the source (>1 m from the source), that is, in the "far field" (bystanders), or at a period after the emission occurred (re-entry). The spray experiments were conducted in an experimental room with well-controlled environmental and ventilation conditions (19.5 m(3)). The concentration of MNOs was investigated by measuring real-time size distribution, number, and active surface area concentration. For off-line analysis of the particles in the air, samples for scanning/transmission electron microscopy and elemental analysis were collected. The release of MNOs was measured at ∼30 and 290 cm from the source ("near field" and "far field", respectively). For all four spray products, the maximum number and surface area concentrations in the "near field" exceeded the maximum concentrations reached in the "far field". At 2 min after the emission occurred, the concentration in both the "near field" and "far field" reached a comparable steady-state level above background level. These steady-state concentrations remained elevated above background concentration throughout the entire measurement period (12 min). The results of the real-time measurement devices mainly reflect the liquid aerosols emitted by the spray process itself rather than only the MNO, which hampers the interpretation of the results. However, the combination of the off-line analysis and the results of the real-time devices indicates that after the use of nano-spray products, personal exposure to MNOs can occur not only in the near field, but also at a greater distance than the immediate proximity of the source and at a period after emission occurred.

Characterization of titanium dioxide nanoparticles in food products: analytical methods to define nanoparticles.

Titanium dioxide (TiO2) is a common food additive used to enhance the white color, brightness, and sometimes flavor of a variety of food products. In this study 7 food grade TiO2 materials (E171), 24 food products, and 3 personal care products were investigated for their TiO2 content and the number-based size distribution of TiO2 particles present in these products. Three principally different methods have been used to determine the number-based size distribution of TiO2 particles: electron microscopy, asymmetric flow field-flow fractionation combined with inductively coupled mass spectrometry, and single-particle inductively coupled mass spectrometry. The results show that all E171 materials have similar size distributions with primary particle sizes in the range of 60-300 nm. Depending on the analytical method used, 10-15% of the particles in these materials had sizes below 100 nm. In 24 of the 27 foods and personal care products detectable amounts of titanium were found ranging from 0.02 to 9.0 mg TiO2/g product. The number-based size distributions for TiO2 particles in the food and personal care products showed that 5-10% of the particles in these products had sizes below 100 nm, comparable to that found in the E171 materials. Comparable size distributions were found using the three principally different analytical methods. Although the applied methods are considered state of the art, they showed practical size limits for TiO2 particles in the range of 20-50 nm, which may introduce a significant bias in the size distribution because particles <20 nm are excluded. This shows the inability of current state of the art methods to support the European Union recommendation for the definition of nanomaterials.

Effects of silver nanoparticles (NM-300K) on Lumbricus rubellus earthworms and particle characterization in relevant test matrices including soil.

The impact of silver nanoparticles (AgNP; at 0 mg Ag/kg, 1.5 mg Ag/kg, 15.4 mg Ag/kg, and 154 mg Ag/kg soil) and silver nitrate (AgNO3 ; 15.4 mg Ag/kg soil) on earthworms, Lumbricus rubellus, was assessed. A 4-wk exposure to the highest AgNP treatment reduced growth and reproduction compared with the control. Silver nitrate (AgNO3 ) exposure also impaired reproduction, but not as much as the highest AgNP treatment. Long-term exposure to the highest AgNP treatment caused complete juvenile mortality. All AgNP treatments induced tissue pathology. Population modeling demonstrated reduced population growth rates for the AgNP and AgNO3 treatments, and no population growth at the highest AgNP treatment because of juvenile mortality. Analysis of AgNP treated soil samples revealed that single AgNP and AgNP clusters were present in the soil, and that the total Ag in soil porewater remained high throughout the long-term experiment. In addition, immune cells (coelomocytes) of earthworms showed sensitivity to both AgNP and AgNO3 in vitro. Overall, the present study indicates that AgNP exposure may affect earthworm populations and that the exposure may be prolonged because of the release of a dissolved Ag fraction to soil porewater.