This research was supported by the "Development of technologies for safety evaluation and standardization of nanomaterials and nanoproducts" (10059135)" through the Korea Evaluation Institute of Industrial Technology by the Korean Ministry of Trade, Industry &amp; Energy.

1. Preparation of Instruments and Specimens
<ol>
	<li>Abrasor
	<ol>
		<li>Based on an abrasion tester, use an abrasor with one specimen rotation stage (140 mm diameter), two abrasion wheel holders, and a rotation speed of 30 - 80 rpm.</li>
		<li>Use a weight to secure the abrasion wheel to the abrasion wheel holder, which also applies load to the test specimen.</li>
		<li>Install an additional air inlet to provide better suspension for the abrased particles, as shown in Figure 3. Use a 1/8&#34;-diameter tube located 15 mm above and 40 mm away from the center of the test specimen.</li>
	</ol>
	</li>
	<li>Abrasion wheel
	<ol>
		<li>Wrap the abrasion wheel (55 mm diameter, 13 mm thick) with sand paper (100 grit and brand new).</li>
	</ol>
	</li>
	<li>Specimen
	<ol>
		<li>Specimen is a composite containing nanomaterial for abrasion test. To installed at abrasor, the specimen should be prepared with 140 mm diameter.</li>
	</ol>
	</li>
	<li>Chamber
	<ol>
		<li>Use stainless steel for the chamber walls to avoid particle deposition due to electrostatic force. Place the abrasor inside the chamber (volume 1 m3) (Table 1), and locate the air inlet and outlet in the upper and lower part of the chamber, respectively. Use a mixer, consisting of three perforated plates, at the air outlet to achieve a uniformly mixed particle flow.</li>
	</ol>
	</li>
	<li>Neutralizer
	<ol>
		<li>As electro-statically charged particles enhance particle deposition on the chamber walls, use a neutralizer (soft X-ray ionizer) to minimize the charged state of the particles.</li>
	</ol>
	</li>
	<li>Online measuring instruments 12, 13
	<ol>
		<li>Use a CPC and OPC to measure the particle number concentration and particle size distribution as per manufacturer&#39;s instructions.</li>
		<li>Install the CPC and OPC at the outlet of the chamber to measure the particle number concentration and particle size distribution.</li>
	</ol>
	</li>
	<li>Particle sampling instruments
	<ol>
		<li>Sample the released particles using a particle sampler containing filter media or a TEM grid to analyze the particle morphology and components.</li>
		<li>Install the particle sampler containing filter media or a TEM grid at the outlet of the chamber to analyze the morphology of the release particles.</li>
	</ol>
	</li>
</ol>
2. Abrasion Test for Nanoparticle Release Using Chamber System
NOTE: The abrasion test conditions are described in Table 2.
<ol>
	<li>Locate the abrasor in the center of the chamber.</li>
	<li>Install the test specimen on the specimen rotation stage of the abrasor.</li>
	<li>Secure the abrasion wheels in the abrasion wheel holders with a 1,000 g weight to apply load to the test specimen.</li>
	<li>Locate the neutralizer (soft X-ray ionizer) 28 cm away from the center of the test specimen at a 45&#176; angle, as seen in Figure 2, to reduce the electro-static particle deposition on the chamber walls. 
	NOTE: The neutralizer removes the electrostatic force by beam exposure. However, since the air inlet and abrasion wheels are located above the specimen rotation stage, this restricts the access of the neutralizer beam to the surface of the test specimen. Therefore, the neutralizer is located diagonally to allow the beam to reach as much of the specimen surface as possible.</li>
	<li>Operate the blower installed at the outlet of the chamber at a 50 L/min flow rate.</li>
	<li>Supply 25 L/min additional particle-free suspension air using an air compressor through the additional air inlet. 
	NOTE: The particles, which are generated by abrasion, were deposited on surface of the specimen and abrasion wheels, strongly. Therefore, it is hard to measure the abrased particles. The additional air inlet can help to solve this problem to particle suspension.</li>
	<li>Check the background particle number concentration inside the chamber to reach an average particle number concentration for 1 h of below 1 #/cc using CPC, as described in Figure 4.</li>
	<li>Operate the specimen rotation stage of the abrasor using a step motor that rotates the specimen rotation stage at 72 rpm with 1,000 rotations.</li>
	<li>Measure and record the released particle number concentration and particle size distribution using the CPC and OPC. 
	NOTE: The particles released from the nanocomposites are suspended and carried by the air that is being pumped. These suspended particles are eventually transported to the outlet following the airstream. The released particles are then detected by the CPC and OPC at the outlet of the chamber. A CPC and OPC are most frequently used for measuring the particle number concentration, while an OPC can also measure the particle size distribution.</li>
	<li>Sample the released particles using a particle sampler containing filter media or a TEM grid. 
	NOTE: The particles released from nanocomposites by abrasion move to the outlet of the chamber following the airstream. At the outlet of the chamber, the released particles can be sampled using a particle sampler. The released particles collected on filter media or a TEM grid can then be analyzed using TEM or SEM (scanning electron microscopy).</li>
	<li>Stop the measurement and sampling when the particle number concentration reaches below 0.1% of the peak particle number concentration.</li>
	<li>Save the all data (CPC, OPC) and remove all the samples (test specimens).</li>
	<li>Use a new specimen and new abrasion wheels for each test, and wash the chamber and abrasor with kimwipes and IPA (iso-propyl alcohol) after each abrasion test to confirm repeatability.</li>
</ol>

<ol>
	<li>Froggett, S. J., Clancy, S. F., Boverhof, D. R., Canady, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+and+perspectives+of+existing+research+on+the+release+of+nanomaterials+from+solid+nanocomposites.">A review and perspectives of existing research on the release of nanomaterials from solid nanocomposites.</a> Part Fibre Toxicol. 11, (2014).</li><li>Kingston, C., Zepp, R., Andrady, A., Boverhof, D., Fehir, R., Hawkins, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Release+characteristics+of+selected+carbon+nanotube+polymer+composites.">Release characteristics of selected carbon nanotube polymer composites.</a> Carbon. 68, 33-57 (2014).</li><li>Kaiser, D., Stefaniak, A., Scott, K., Nguyen, T., Schutz, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Methods for the Measurement of Release of MWCNTs from MWCNT-Polymer Composites, NIST. , (2014).</li><li>Nowack, B., David, R. M., Fissan, H., Morris, H., Shatkin, J. A., Stintz, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Potential+release+scenarios+for+carbon+nanotubes+used+in+composites.">Potential release scenarios for carbon nanotubes used in composites.</a> Environ. Int. 59, 1-11 (2013).</li><li>Kim, E., Lee, J. H., Kim, J. K., Lee, G. H., Ahn, K., Park, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Case+Study+on+Risk+Evaluation+of+Silver+Nanoparticle+Exposure+from+Antibacterial+Sprays+Containing+Silver+Nanoparticles.">Case Study on Risk Evaluation of Silver Nanoparticle Exposure from Antibacterial Sprays Containing Silver Nanoparticles.</a> J of Nanomaterial. , 346586 (2015).</li><li>Kim, E., Lee, J. H., Kim, J. K., Lee, G. H., Ahn, K., Park, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Case+study+on+risk+evaluation+of+printed+electronics+using+nanosilver+ink.">Case study on risk evaluation of printed electronics using nanosilver ink.</a> Nano Convergence. , (2016).</li><li>Vorbau, M., Hillemann, L., Stintz, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Method+for+the+characterization+of+the+abrasion+induced+nanoparticle+release+into+air+from+surface+coatings.">Method for the characterization of the abrasion induced nanoparticle release into air from surface coatings.</a> J. Aerosol Sci. 40, 209-217 (2009).</li><li>Golanski, L., Gaborieau, A., Guiot, A., Uzu, G., Chatenet, J., Tardif, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+abrasion-induced+nanoparticle+release+from+paints+into+liquids+and+air.">Characterization of abrasion-induced nanoparticle release from paints into liquids and air.</a> J. Phys. Conf. Ser. 304, 012062 (2011).</li><li>Wohlleben, W., Brill, S., Meier, M. W., Mertler, M., Cox, G., Hirth, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+the+lifecycle+of+nanocomposites:+Comparing+released+fragments+and+their+in-vivo+hazards+from+three+release+mechanisms+and+four+nanocomposites.">On the lifecycle of nanocomposites: Comparing released fragments and their in-vivo hazards from three release mechanisms and four nanocomposites.</a> Small. 7, 2384-2395 (2011).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> ISO 7784-1, Paints and varnishes -- Determination of resistance to abrasion -- Part 1: Rotating abrasive-paper-covered wheel method. , (1997).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> ISO 5470-1, Rubber- or plastics-coated fabrics -- Determination of abrasion resistance -- Part 1: Taber abrader. , (1999).</li><li>Schlagenhauf, L., Chu, B. T. T., Buha, J., N&#252;sch, F., Wang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Release+of+carbon+nanotubes+from+an+epoxy-based+nanocomposites+during+an+abrasion+process.">Release of carbon nanotubes from an epoxy-based nanocomposites during an abrasion process.</a> Enviorn. Sci. Tech. 46, 7366-7372 (2012).</li><li>Bello, D., Wardle, B. L., Yamamoto, N., deVilloria, R. G., Garcia, E. J., Hart, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exposure+to+nanoscale+particles+and+fibers+during+machining+of+hybrid+advanced+composites+containing+carbon+nanotubes.">Exposure to nanoscale particles and fibers during machining of hybrid advanced composites containing carbon nanotubes.</a> J. Nanopart. Res. 11, 231-249 (2009).</li><li>Cena, L. G., Peters, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+and+control+of+airborne+particles+emitted+during+production+of+epoxy/carbon+nanotube+nanocomposites.">Characterization and control of airborne particles emitted during production of epoxy/carbon nanotube nanocomposites.</a> J. Occup. Environ. Hyg. 8, 86-92 (2011).</li></ol>

The authors have nothing to disclose.

The most critical steps when conducting the nanorelease test from nanocomposite materials using an abrasion test were: 1) using a chamber system made of stainless steel with a neutralizer to remove the electrostatic charge generated by abrasion and reduce the deposition of particles on the chamber walls; 2) supplying additional air to provide better particle suspension; and 3) sampling the released particles and online monitoring using a CPC and OPC from the outlet that contained a mixer consisting of three perforated pl...

Nanomaterial exposure has mostly been studied in relation to workers in workplaces manufacturing, handling, fabricating, and packaging nanomaterials, while consumer exposure has not been studied extensively. A recent analysis of the environmental and health literature database created by the International Council of Nanotechnology (ICON) also indicated that most nanomaterial safety research has focused on hazards (83%) and potential exposure (16%), with the release from nanocomposites, representing consumer exposure, only representing 0.8% 1. Thus, very little is known about consumer exposure to nanomaterials.
Nanoparticle release has been used to estimate consumer exposure in simulation studies, including the abrasion and weathering of nanocomposites, washing textiles, or dustiness testing methods, such as the rotating drum method, vortex shaking method, and other shaker methods 2-3. Plus, several international attempts, such as the ILSI (International Life Science Institute) nanorelease and EU NanoReg, have been made to develop technology to understand the release of nanomaterials used in consumer products. The ILSI nanorelease consumer product launched in 2011 represents a life-cycle approach to nanomaterial release from consumer products, where phase 1 involves nanomaterial selection, phase 2 covers evaluation methods, and phase 3 implements interlaboratory studies. Several monographs and publications on the safety of nanomaterials in consumer products have also been published 4-6.
Meanwhile, NanoReg represents a common European approach to the regulatory testing of manufactured nanomaterials and provides a program of methods for use in simulation approaches to nanorelease from consumer products 2. ISO TC 229 is also trying to develop standards relevant to consumer safety and submit a new working item proposal for consumer safety. The OECD WPMN (working party on nanomaterials), especially SG8 (steering group on exposure assessment and exposure mitigation), recently conducted a survey on the direction of future work, especially consumer and environmental exposure assessment. Therefore, in light of these international activities, the Korean Ministries of Trade, Industry and Energy launched a tiered project in 2013 focused on the &#34;Development of technologies for the safety evaluation and standardization of nanomaterials and nanoproducts&#34;. Plus, several consumer safety-relevant studies to standardize nanomaterial release from consumer products have also been published 7-8.
An abrasion test is one of the simulation approaches included in the ILSI nanorelease and NanoReg 2-3 for determining the potential emission level of nanoparticles from different commercial composite products. The mass weight loss is deduced based on the difference in the specimen weight before and after abrasion using an abrasor. The nanocomposite sample is abraded at a constant speed, a sampler sucks up the aerosol, and the particles are then analyzed using particle counting devices, such as a Condensation Particle Counter (CPC) or optical particle counter (OPC), and collected on a TEM (transmission electron microscopy) grid or membrane for further visual analysis. However, conducting an abrasion test for nanocomposite materials requires a consistent nanoparticle release, which is difficult due to particle charging as a result of abrasion and when the particle sampling is conducted near the emission point 2-3, 9-11.
Accordingly, this paper presents a chamber system as a new method for evaluating nanomaterial release in the case of abrasion of nanocomposite materials. When compared with other abrasion and simulation tests, the proposed chamber system provides consistent nanoparticle release data in the case of abrasion. Moreover, this new test method has been used widely in the field of indoor air quality and semi-conduct industry as total particle number counting method 12, 13. Therefore, it is anticipated that the proposed method can be developed into a standardized method for testing nanoparticle release from consumer products containing nanomaterials.

<table><tbody><tr><td>Foamex</td><td>Taeyoung, R. of Korea</td><td></td><td></td></tr><tr><td>MWCNT (multiwalled carbon nanotube) composite</td><td>Hanwha, Incheon, R. of Korea</td><td></td><td>2% MWCNTs in low density polyethylene</td></tr><tr><td>Abrasion Paper</td><td>Derfos, R. of Korea</td><td>#100</td><td>100 grit sand paper</td></tr><tr><td>Condensation Particle Counter (CPC)</td><td>TSI Inc, Shoreview, MN</td><td>UCPC 3775</td><td></td></tr><tr><td>Optical Paritcle Counter (OPC)</td><td>Grimm, Ainring, Germany</td><td>1.109</td><td></td></tr><tr><td>Mini Particle Sampler</td><td>Ecomesure, Saclay, France</td><td></td><td></td></tr><tr><td>Quantifoil Holey Carbon Film</td><td>TED PELLA Inc. USA</td><td>1.2/1.3</td><td></td></tr><tr><td>Filter Holder</td><td></td><td></td><td>custom made</td></tr><tr><td>Polycarbonate Filter&amp;nbsp;</td><td>Millipore, USA</td><td>CAT No. GTTP02500</td><td></td></tr><tr><td>Soft X-ray Ionizer (Neutralizer)</td><td>SUNJE, R. of Korea</td><td>SXN-05U</td><td></td></tr><tr><td>Field Emission-Scanning Electron Microscope (FE-SEM)</td><td>Hitachi</td><td>S-4300</td><td></td></tr></tbody></table>

testing of nanoparticle release from a composite containing nanomaterial using a chamber system

With the rapid development of nanotechnology as one of the most important technologies in the 21st century, interest in the safety of consumer products containing nanomaterials is also increasing. Evaluating the nanomaterial release from products containing nanomaterials is a crucial step in assessing the safety of these products, and has resulted in several international efforts to develop consistent and reliable technologies for standardizing the evaluation of nanomaterial release. In this study, the release of nanomaterials from products containing nanomaterials is evaluated using a chamber system that includes a condensation particle counter, optical particle counter, and sampling ports to collect filter samples for electron microscopy analysis. The proposed chamber system is tested using an abrasor and disc-type nanocomposite material specimens to determine whether the nanomaterial release is repeatable and consistent within an acceptable range. The test results indicate that the total number of particles in each test is within 20% from the average after several trials. The release trends are similar and they show very good repeatability. Therefore, the proposed chamber system can be effectively used for nanomaterial release testing of products containing nanomaterials.

Abrasion Test Repeatability Using Chamber System
The total particle numbers were consistent for 8 abrasion tests, as shown in Table 3. The CPC measured an average of 3.67 x109 particles, while the OPC counted an average of 1.98 x 109 particles (&#62; 0.3 &#181;m). The deviations were within 20%, which represented a consistent release of particles during abrasion.
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Watch this Scientific Journal Video about Testing of Nanoparticle Release from a Composite Containing Nanomaterial Using a Chamber System at JoVE.com

Testing of Nanoparticle Release from a Composite Containing Nanomaterial Using a Chamber System

Nanoparticle release is tested using a chamber system that includes a condensation particle counter, an optical particle counter and sampling ports to collect filter samples for microscopy analysis. The proposed chamber system can be effectively used for nanomaterial release testing with a repeatable and consistent data range.

With the rapid development of nanotechnology as one of the most important technologies in the 21st century, interest in the safety of consumer products ...

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6.7K Views.  Hanyang University. Nanoparticle release is tested using a chamber system that includes a condensation particle counter, an optical particle counter and sampling ports to collect filter samples for microscopy analysis. The proposed chamber system can be effectively used for nanomaterial release testing with a repeatable and consistent data range.

Video: Testing of Nanoparticle Release from a Composite Containing Nanomaterial Using a Chamber System

Optical Trapping of Nanoparticles

Optical trapping is a technique for immobilizing and manipulating small objects in a gentle way using light, and it has been widely applied in trapping and manipulating small biological particles. Ashkin and co-workers first demonstrated optical tweezers using a single focused beam1. The single beam trap can be described accurately using the perturbative gradient force formulation in the case of small Rayleigh regime particles1. In the perturbative regime, the optical power required for trapping a particle scales as the inverse fourth power of the particle size. High optical powers can damage dielectric particles and cause heating. For instance, trapped latex spheres of 109 nm in diameter were destroyed by a 15 mW beam in 25 sec1, which has serious implications for biological matter2,3.
A self-induced back-action (SIBA) optical trapping was proposed to trap 50 nm polystyrene spheres in the non-perturbative regime4. In a non-perturbative regime, even a small particle with little permittivity contrast to the background can influence significantly the ambient electromagnetic field and induce a large optical force. As a particle enters an illuminated aperture, light transmission increases dramatically because of dielectric loading. If the particle attempts to leave the aperture, decreased transmission causes a change in momentum outwards from the hole and, by Newton's Third Law, results in a force on the particle inwards into the hole, trapping the particle. The light transmission can be monitored; hence, the trap can become a sensor. The SIBA trapping technique can be further improved by using a double-nanohole structure.
The double-nanohole structure has been shown to give a strong local field enhancement5,6. Between the two sharp tips of the double-nanohole, a small particle can cause a large change in optical transmission, thereby inducing a large optical force. As a result, smaller nanoparticles can be trapped, such as 12 nm silicate spheres7 and 3.4 nm hydrodynamic radius bovine serum albumin proteins8. In this work, the experimental configuration used for nanoparticle trapping is outlined. First, we detail the assembly of the trapping setup which is based on a Thorlabs Optical Tweezer Kit. Next, we explain the nanofabrication procedure of the double-nanohole in a metal film, the fabrication of the microfluidic chamber and the sample preparation. Finally, we detail the data acquisition procedure and provide typical results for trapping 20 nm polystyrene nanospheres.

The following setup approach details low power optical trapping of dielectric nanoparticles using a double-nanohole in metal film.

Optical trapping is a technique for immobilizing and manipulating small objects in a gentle way using light, and it has been widely applied in trapping ...

Combinatorial Synthesis of and High-throughput Protein Release from Polymer Film and Nanoparticle Libraries

Polyanhydrides are a class of biomaterials with excellent biocompatibility and drug delivery capabilities. While they have been studied extensively with conventional one-sample-at-a-time synthesis techniques, a more recent high-throughput approach has been developed enabling the synthesis and testing of large libraries of polyanhydrides1. This will facilitate more efficient optimization and design process of these biomaterials for drug and vaccine delivery applications. The method in this work describes the combinatorial synthesis of biodegradable polyanhydride film and nanoparticle libraries and the high-throughput detection of protein release from these libraries. In this robotically operated method (Figure 1), linear actuators and syringe pumps are controlled by LabVIEW, which enables a hands-free automated protocol, eliminating user error. Furthermore, this method enables the rapid fabrication of micro-scale polymer libraries, reducing the batch size while resulting in the creation of multivariant polymer systems. This combinatorial approach to polymer synthesis facilitates the synthesis of up to 15 different polymers in an equivalent amount of time it would take to synthesize one polymer conventionally. In addition, the combinatorial polymer library can be fabricated into blank or protein-loaded geometries including films or nanoparticles upon dissolution of the polymer library in a solvent and precipitation into a non-solvent (for nanoparticles) or by vacuum drying (for films). Upon loading a fluorochrome-conjugated protein into the polymer libraries, protein release kinetics can be assessed at high-throughput using a fluorescence-based detection method (Figures 2 and 3) as described previously1. This combinatorial platform has been validated with conventional methods2 and the polyanhydride film and nanoparticle libraries have been characterized with 1H NMR and FTIR. The libraries have been screened for protein release kinetics, stability and antigenicity; in vitro cellular toxicity, cytokine production, surface marker expression, adhesion, proliferation and differentiation; and in vivo biodistribution and mucoadhesion1-11. The combinatorial method developed herein enables high-throughput polymer synthesis and fabrication of protein-loaded nanoparticle and film libraries, which can, in turn, be screened in vitro and in vivo for optimization of biomaterial performance.

This method describes the combinatorial synthesis of biodegradable polyanhydride film and nanoparticle libraries and the high-throughput detection of protein release from these libraries.

Polyanhydrides are a class of biomaterials with excellent biocompatibility and drug delivery capabilities. While they have been studied extensively with ...

Bioengineering

Experimental Protocol to Investigate Particle Aerosolization of a Product Under Abrasion and Under Environmental Weathering

The present article presents an experimental protocol to investigate particle aerosolization of a product under abrasion and under environmental weathering, which is a fundamental element to the approach of nanosafety-by-design of nanostructured products for their durable development. This approach is basically a preemptive one in which the focus is put on minimizing the emission of engineered nanomaterials' aerosols during the usage phase of the product's life cycle. This can be attained by altering its material properties during its design phase without compromising with any of its added benefits. In this article, an experimental protocol is presented to investigate the nanosafety-by-design of three commercial nanostructured products with respect to their mechanical solicitation and environmental weathering. The means chosen for applying the mechanical solicitation is an abrasion process and for the environmental weathering, it is an accelerated UV exposure in the presence of humidity and heat. The eventual emission of engineered nanomaterials is studied in terms of their number concentration, size distribution, morphology and chemical composition. The purpose of the protocol is to study the emission for test samples and experimental conditions which are corresponding to real life situations. It was found that the application of the mechanical stresses alone emits the engineered nanomaterials' aerosols in which the engineered nanomaterial is always embedded inside the product matrix, thus, a representative product element. In such a case, the emitted aerosols comprise of both nanoparticles as well as microparticles. But if the mechanical stresses are coupled with the environmental weathering, the experimental protocol reveals then the eventual deterioration of the product, after a certain weathering duration, may lead to the emission of the free engineered nanomaterial aerosols too.

In this article, an experimental protocol to investigate particle aerosolization of a product under abrasion and under environmental weathering is presented. Results on the emission of engineered nanomaterials, in the form of aerosols are presented. The specific experimental set-up is described in detail.

The present article presents an experimental protocol to investigate particle aerosolization of a product under abrasion and under environmental ...

Evaluation of Polymeric Gene Delivery Nanoparticles by Nanoparticle Tracking Analysis and High-throughput Flow Cytometry

Non-viral gene delivery using polymeric nanoparticles has emerged as an attractive approach for gene therapy to treat genetic diseases1 and as a technology for regenerative medicine2. Unlike viruses, which have significant safety issues, polymeric nanoparticles can be designed to be non-toxic, non-immunogenic, non-mutagenic, easier to synthesize, chemically versatile, capable of carrying larger nucleic acid cargo and biodegradable and/or environmentally responsive. Cationic polymers self-assemble with negatively charged DNA via electrostatic interaction to form complexes on the order of 100 nm that are commonly termed polymeric nanoparticles. Examples of biomaterials used to form nanoscale polycationic gene delivery nanoparticles include polylysine, polyphosphoesters, poly(amidoamines)s and polyethylenimine (PEI), which is a non-degradable off-the-shelf cationic polymer commonly used for nucleic acid delivery1,3 . Poly(beta-amino ester)s (PBAEs) are a newer class of cationic polymers4 that are hydrolytically degradable5,6 and have been shown to be effective at gene delivery to hard-to-transfect cell types such as human retinal endothelial cells (HRECs)7, mouse mammary epithelial cells8, human brain cancer cells9 and macrovascular (human umbilical vein, HUVECs) endothelial cells10.
A new protocol to characterize polymeric nanoparticles utilizing nanoparticle tracking analysis (NTA) is described. In this approach, both the particle size distribution and the distribution of the number of plasmids per particle are obtained11. In addition, a high-throughput 96-well plate transfection assay for rapid screening of the transfection efficacy of polymeric nanoparticles is presented. In this protocol, poly(beta-amino ester)s (PBAEs) are used as model polymers and human retinal endothelial cells (HRECs) are used as model human cells. This protocol can be easily adapted to evaluate any polymeric nanoparticle and any cell type of interest in a multi-well plate format.

A protocol for nanoparticle tracking analysis (NTA) and high-throughput flow cytometry to evaluate polymeric gene delivery nanoparticles is described. NTA is utilized to characterize the nanoparticle particle size distribution and the plasmid per particle distribution. High-throughput flow cytometry enables quantitative transfection efficacy evaluation for a library of gene delivery biomaterials.

Non-viral gene delivery using polymeric nanoparticles has emerged as an attractive approach for gene therapy to treat genetic diseases1 and as a ...

Advanced Compositional Analysis of Nanoparticle-polymer Composites Using Direct Fluorescence Imaging

The fabrication of polymer-nanoparticle composites is extremely important in the development of many functional materials. Identifying the precise composition of these materials is essential, especially in the design of surface catalysts, where the surface concentration of the active component determines the activity of the material. Antimicrobial materials which utilize nanoparticles are a particular focus of this technology. Recently swell encapsulation has emerged as a technique for inserting antimicrobial nanoparticles into a host polymer matrix. Swell encapsulation provides the advantage of localizing the incorporation to the external surfaces of materials, which act as the active sites of these materials. However, quantification of this nanoparticle uptake is challenging. Previous studies explore the link between antimicrobial activity and surface concentration of the active component, but this is not directly visualized. Here we show a reliable method to monitor the incorporation of nanoparticles into a polymer host matrix via swell encapsulation. We show that the surface concentration of CdSe/ZnS nanoparticles can be accurately visualized through cross-sectional fluorescence imaging. Using this method, we can quantify the uptake of nanoparticles via swell encapsulation and measure the surface concentration of encapsulated particles, which is key in optimizing the activity of functional materials.

Here we present a reliable method to monitor the incorporation of nanoparticles into a polymer host matrix via swell encapsulation. We show that the surface concentration of cadmium selenide quantum dots can be accurately visualized through cross-sectional fluorescence imaging.

The fabrication of polymer-nanoparticle composites is extremely important in the development of many functional materials. Identifying the precise ...

A System to Create Stable Nanoparticle Aerosols from Nanopowders

Nanoparticle aerosols released from nanopowders in workplaces are associated with human exposure and health risks. We developed a novel system, requiring minimal amounts of test materials (min. 200 mg), for studying powder aerosolization behavior and aerosol properties. The aerosolization procedure follows the concept of the fluidized-bed process, but occurs in the modified volume of a V-shaped aerosol generator. The airborne particle number concentration is adjustable by controlling the air flow rate. The system supplied stable aerosol generation rates and particle size distributions over long periods (0.5-2 hr and possibly longer), which are important, for example, to study aerosol behavior, but also for toxicological studies. Strict adherence to the operating procedures during the aerosolization experiments ensures the generation of reproducible test results. The critical steps in the standard protocol are the preparation of the material and setup, and the aerosolization operations themselves. The system can be used for experiments requiring stable aerosol concentrations and may also be an alternative method for testing dustiness. The controlled aerosolization made possible with this setup occurs using energy inputs (may be characterized by aerosolization air velocity) that are within the ranges commonly found in occupational environments where nanomaterial powders are handled. This setup and its operating protocol are thus helpful for human exposure and risk assessment.

We designed and developed an effective nanopowder aerosolization setup and operating protocol. The system generated nanoparticle aerosols with stable number concentrations and size distributions for long durations, requiring only small quantities of test material (min. 200 mg).

Nanoparticle aerosols released from nanopowders in workplaces are associated with human exposure and health risks. We developed a novel system, requiring ...

A Continuous-flow Photocatalytic Reactor for the Precisely Controlled Deposition of Metallic Nanoparticles

In this work, a novel photocatalytic reactor for the pulsed and controlled excitation of the photocatalyst and the precise deposition of metallic nanoparticles is developed. Guidelines for the replication of the reactor and its operation are provided in detail. Three different composite systems (Pt/graphene, Pt/TiO2, and Au/TiO2) with monodisperse and uniformly distributed particles are produced by this reactor, and the photodeposition mechanism, as well as the synthesis optimization strategy, are discussed. The synthesis methods and their technical aspects are described comprehensively. The role of the ultraviolet (UV) dose (in each excitation pulse) on the photodeposition process is investigated and the optimum values for each composite system are provided.

For a continuous and scalable synthesis of noble-metal-based nanocomposites, a novel photocatalytic reactor is developed and its structure, operation principles, and product quality optimization strategies are described.

In this work, a novel photocatalytic reactor for the pulsed and controlled excitation of the photocatalyst and the precise deposition of metallic ...

Chemistry

Development of a Nose-only Inhalation Toxicity Test Chamber That Provides Four Exposure Concentrations of Nano-sized Particles

Using a numerical analysis based on computerized fluid dynamics, a nose-only inhalation toxicity chamber with four different exposure concentrations is designed and validated for flow field uniformity and cross-contamination among the exposure ports for each concentration. The designed flow field values are compared with the measured values from exposure ports located horizontally and vertically. For this purpose, nanoscale sodium chloride particles are generated as test particles and introduced to the inhalation chamber to evaluate the cross-contamination and concentration maintenance among the chambers, for each concentration group. The results indicate that the designed multiconcentration inhalation chamber can be used in animal inhalation toxicity testing without cross-contamination among concentration groups. Moreover, the designed multiconcentration inhalation toxicity chamber can also be converted to a single-concentration inhalation chamber. Further testing with gas, organic vapor, or non-nanoscale particles will ensure the use of the chamber in the inhalation testing of other test articles.

A nose-only inhalation toxicity chamber capable of testing inhalation toxicity at four different exposure concentrations was designed and validated for flow field uniformity and cross-contamination between the exposure ports for each concentration. Here, we present a protocol to confirm that the designed chamber is effective for inhalation toxicity testing.

Using a numerical analysis based on computerized fluid dynamics, a nose-only inhalation toxicity chamber with four different exposure concentrations is ...

Whole-Body Nanoparticle Aerosol Inhalation Exposures

Inhalation is the most likely exposure route for individuals working with aerosolizable engineered nano-materials (ENM). To properly perform nanoparticle inhalation toxicology studies, the aerosols in a chamber housing the experimental animals must have: 1) a steady concentration maintained at a desired level for the entire exposure period; 2) a homogenous composition free of contaminants; and 3) a stable size distribution with a geometric mean diameter &lt; 200 nm and a geometric standard deviation &sigma;g &lt; 2.5 5. The generation of aerosols containing nanoparticles is quite challenging because nanoparticles easily agglomerate. This is largely due to very strong inter-particle forces and the formation of large fractal structures in tens or hundreds of microns in size 6, which are difficult to be broken up. Several common aerosol generators, including nebulizers, fluidized beds, Venturi aspirators and the Wright dust feed, were tested; however, none were able to produce nanoparticle aerosols which satisfy all criteria 5.
A whole-body nanoparticle aerosol inhalation exposure system was fabricated, validated and utilized for nano-TiO2 inhalation toxicology studies. Critical components: 1) novel nano-TiO2 aerosol generator; 2) 0.5 m3 whole-body inhalation exposure chamber; and 3) monitor and control system. Nano-TiO2 aerosols generated from bulk dry nano-TiO2 powders (primary diameter of 21 nm, bulk density of 3.8 g/cm3) were delivered into the exposure chamber at a flow rate of 90 LPM (10.8 air changes/hr). Particle size distribution and mass concentration profiles were measured continuously with a scanning mobility particle sizer (SMPS), and an electric low pressure impactor (ELPI). The aerosol mass concentration (C) was verified gravimetrically (mg/m3). The mass (M) of the collected particles was determined as M = (Mpost-Mpre), where Mpre and Mpost are masses of the filter before and after sampling (mg). The mass concentration was calculated as C = M/(Q*t), where Q is sampling flowrate (m3/min), and t is the sampling time (minute). The chamber pressure, temperature, relative humidity (RH), O2 and CO2 concentrations were monitored and controlled continuously. Nano-TiO2 aerosols collected on Nuclepore filters were analyzed with a scanning electron microscope (SEM) and energy dispersive X-ray (EDX) analysis.
In summary, we report that the nano-particle aerosols generated and delivered to our exposure chamber have: 1) steady mass concentration; 2) homogenous composition free of contaminants; 3) stable particle size distributions with a count-median aerodynamic diameter of 157 nm during aerosol generation. This system reliably and repeatedly creates test atmospheres that simulate occupational, environmental or domestic ENM aerosol exposures.

A whole-body nanoparticle aerosol inhalation exposure facility was constructed for nano-sized titanium dioxide (TiO2) inhalation toxicology studies. This system provides nano-TiO2 aerosol test atmospheres that have: 1) a steady mass concentration; 2) a homogenous composition free of contaminants; and 3) a stable particle size distribution during aerosol generation.

Inhalation is the most likely exposure route for individuals working with aerosolizable engineered nano-materials (ENM). To properly perform nanoparticle ...

Biology

An Air-liquid Interface Bronchial Epithelial Model for Realistic, Repeated Inhalation Exposure to Airborne Particles for Toxicity Testing

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic exposure conditions. This puts specific demands on the cell culture models. Many cell types are negatively affected by exposure to air (e.g., drying out) and only remain viable for a few days. This limits the exposure conditions that can be used in these models: usually relatively high concentrations are applied as a cloud (i.e., droplets containing particles, which settle down rapidly) within a short period of time. Such experimental conditions do not reflect realistic long-term exposure to low concentrations of particles. To overcome these limitations the use of a human bronchial epithelial cell line, Calu-3 was investigated. These cells can be cultured at ALI conditions for several weeks while retaining a healthy morphology and a stable monolayer with tight junctions. In addition, this bronchial model is suitable for testing the effects of repeated exposures to low, realistic concentrations of airborne particles using an ALI exposure system. This system uses a continuous airflow in contrast to other ALI exposure systems that use a single nebulization producing a cloud. Therefore, the continuous flow system is suitable for repeated and prolonged exposure to airborne particles while continuously monitoring the particle characteristics, exposure concentration, and delivered dose. Taken together, this bronchial model, in combination with the continuous flow exposure system, is able to mimic realistic, repeated inhalation exposure conditions that can be used for toxicity testing.

Described is the cell culture and exposure method of an in vitro bronchial model for realistic, repeated inhalation exposure to particles for toxicity testing.

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic ...