Tag Archives: Lee L. Yu

Using sugar for a better way to clean nanoparticles from organisms

Researchers at the US National Institute of Standards and Technology (NIST) have found that a laboratory technique used for over 60 years is the best way to date to clean nanoparticles from organisms. From a Jan. 26, 2017 news item on ScienceDaily,

Sometimes old-school methods provide the best ways of studying cutting-edge tech and its effects on the modern world.

Giving a 65-year-old laboratory technique a new role, researchers at the National Institute of Standards and Technology (NIST) have performed the cleanest separation to date of synthetic nanoparticles from a living organism. The new NIST method is expected to significantly improve experiments looking at the potential environmental and health impacts of these manufactured entities. It will allow scientists to more accurately count how many nanoparticles have actually been ingested by organisms exposed to them.

A Jan. 26, 2017 NIST news release (also on EurekAlert), which originated the news item, offers more detail,

The common roundworm Caenorhabditis elegans has been used in recent years as a living model for laboratory studies of how biological and chemical compounds may affect multicellular organisms. These compounds include engineered nanoparticles (ENPs), bits of material between 1 and 100 nanometers (billionths of a meter, or about 1/10,000 the diameter of a red blood cell). Previous research has often focused on quantifying the amount and size of engineered nanoparticles ingested by C. elegans. Measuring the nanoparticles that actually make it into an organism is considered a more relevant indicator of potential toxicity than just the amount of ENPs to which the worms are exposed.

Traditional methods for counting ingested ENPs have produced questionable results. Currently, researchers expose C. elegans to metal ENPs such as silver or gold in solution, then rinse the excess particles away with water followed by centrifugation and freeze-drying. A portion of the “cleaned” sample produced is then typically examined by a technique that determines the amount of metal present, known as inductively coupled plasma mass spectrometry (ICP-MS). It often yields ENP counts in the tens of thousands per worm; however, those numbers always seem too high to NIST researchers working with C. elegans.

“Since ICP-MS will detect all of the nanoparticles associated with the worms, both those ingested and those that remain attached externally, we suspect that the latter is what makes the ‘ENPs’ per-worm counts so high,” said NIST analytical chemist Monique Johnson (link sends e-mail), the lead author on the ACS Nano paper. “Since we only wanted to quantify the ingested ENPs, a more robust and reliable separation method was needed.”

Luckily, the solution to the problem was already in the lab.

Cross section of the roundworm C. elegans

Scanning electron micrograph showing a cross section of the roundworm C. elegans with two ingested engineered nanoparticles (red dots just right of center). Images such as this provided NIST researchers with visual confirmation that nanoparticle consumption actually occurred. Credit: K. Scott/NIST

In the course of culturing C. elegans for ENP-exposure experiments, Johnson and her colleagues had used sucrose density gradient centrifugation, a decades-old and established system for cleanly separating cellular components, to isolate the worms from debris and bacteria. “We wondered if the same process would allow us to perform an organism-from-ENP separation as well, so I designed a study to find out,” Johnson said.

In their experiment, the NIST researchers first exposed separate samples of C. elegans to low and high concentrations of two sizes of gold nanospheres, 30 and 60 nanometers in diameter. The researchers put each of the samples into a centrifuge and removed the supernatant (liquid portion), leaving the worms and ENPs in the remaining pellets. These were centrifuged twice in a salt solution (rather than just water as in previous separation methods), and then centrifuged again, but this time, through a uniquely designed sucrose density gradient.

“From top to bottom, our gradient consisted of a salt solution layer to trap excess ENPs and three increasingly dense layers of sucrose [20, 40 and 50 percent] to isolate the C. elegans,” Johnson explained. “We followed up the gradient with three water rinses and with centrifugations to ensure that only worms with ingested ENPs, and not the sucrose separation medium with any excess ENPs, would make it into the final pellet.”

Analyzing the range of masses in the ultrapurified samples indicated gold levels more in line with what the researchers expected would be found as ingested ENPs. Experimental validation of the NIST separation method’s success came when the worms were examined in detail under a scanning electron microscope (SEM).

“For me, the eureka moment was when I first saw gold ENPs in the cross section images taken from the C. elegans samples that had been processed through the sucrose density gradient,” Johnson said. “I had been dreaming about finding ENPs in the worm’s digestive tract and now they were really there!”

The high-resolution SEM images also provided visual evidence that only ingested ENPs were counted. “No ENPs were attached to the cuticle, the exoskeleton of C. elegans, in any of the sucrose density gradient samples,” Johnson said. “When we examined worms from our control experiments [processed using the traditional no-gradient, water-rinse-only separation method], there were a number of nanospheres found attached to the cuticle.

Now that it has been successfully demonstrated, the NIST researchers plan to refine and further validate their system for evaluating the uptake of ENPs by C. elegans. “Hopefully, our method will become a useful and valuable tool for reducing the measurement variability and sampling bias that can plague environmental nanotoxicology studies,” Johnson said.

They’ve tested this technique on gold nanoparticles, which begs the question, What kinds of nanoparticles can this technique be used for? Metal nanoparticles only or all nanoparticles?

I’m sure the researchers have already asked these questions and started researching the answers. While the rest of us wait, here’s a link to and a citation for the paper about this promising new technique,

Separation, Sizing, and Quantitation of Engineered Nanoparticles in an Organism Model Using Inductively Coupled Plasma Mass Spectrometry and Image Analysis by Monique E. Johnson, Shannon K. Hanna, Antonio R. Montoro Bustos, Christopher M. Sims, Lindsay C. C. Elliott, Akshay Lingayat, Adrian C. Johnston, Babak Nikoobakht, John T. Elliott, R. David Holbrook, Keana C. K. Scott, Karen E. Murphy, Elijah J. Petersen, Lee L. Yu, and Bryant C. Nelson. ACS Nano, 2017, 11 (1), pp 526–540 DOI: 10.1021/acsnano.6b06582 Publication Date (Web): December 16, 2016

Copyright This article not subject to U.S. Copyright. Published 2016 by the American Chemical Society

This paper is behind a paywall.

Mimicking rain and sun to test plastic for nanoparticle release

One of Canada’s nanotechnology experts once informed a House of Commons Committee on Health that nanoparticles encased in plastic (he was talking about cell phones) weren’t likely to harm you except in two circumstances (when workers were using them in the manufacturing process and when the product was being disposed of). Apparently, under some circumstances, that isn’t true any more. From a Sept. 30, 2016 news item on Nanowerk,

If the 1967 film “The Graduate” were remade today, Mr. McGuire’s famous advice to young Benjamin Braddock would probably be updated to “Plastics … with nanoparticles.” These days, the mechanical, electrical and durability properties of polymers—the class of materials that includes plastics—are often enhanced by adding miniature particles (smaller than 100 nanometers or billionths of a meter) made of elements such as silicon or silver. But could those nanoparticles be released into the environment after the polymers are exposed to years of sun and water—and if so, what might be the health and ecological consequences?

A Sept. 30, 2016 US National Institute of Standards and Technology (NIST) news release, which originated the news item, describes how the research was conducted and its results (Note: Links have been removed),

In a recently published paper (link is external), researchers from the National Institute of Standards and Technology (NIST) describe how they subjected a commercial nanoparticle-infused coating to NIST-developed methods for accelerating the effects of weathering from ultraviolet (UV) radiation and simulated washings of rainwater. Their results indicate that humidity and exposure time are contributing factors for nanoparticle release, findings that may be useful in designing future studies to determine potential impacts.

In their recent experiment, the researchers exposed multiple samples of a commercially available polyurethane coating containing silicon dioxide nanoparticles to intense UV radiation for 100 days inside the NIST SPHERE (Simulated Photodegradation via High-Energy Radiant Exposure), a hollow, 2-meter (7-foot) diameter black aluminum chamber lined with highly UV reflective material that bears a casual resemblance to the Death Star in the film “Star Wars.” For this study, one day in the SPHERE was equivalent to 10 to 15 days outdoors. All samples were weathered at a constant temperature of 50 degrees Celsius (122 degrees Fahrenheit) with one group done in extremely dry conditions (approximately 0 percent humidity) and the other in humid conditions (75 percent humidity).

To determine if any nanoparticles were released from the polymer coating during UV exposure, the researchers used a technique they created and dubbed “NIST simulated rain.” Filtered water was converted into tiny droplets, sprayed under pressure onto the individual samples, and then the runoff—with any loose nanoparticles—was collected in a bottle. This procedure was conducted at the beginning of the UV exposure, at every two weeks during the weathering run and at the end. All of the runoff fluids were then analyzed by NIST chemists for the presence of silicon and in what amounts. Additionally, the weathered coatings were examined with atomic force microscopy (AFM) and scanning electron microscopy (SEM) to reveal surface changes resulting from UV exposure.

Both sets of coating samples—those weathered in very low humidity and the others in very humid conditions—degraded but released only small amounts of nanoparticles. The researchers found that more silicon was recovered from the samples weathered in humid conditions and that nanoparticle release increased as the UV exposure time increased. Microscopic examination showed that deformations in the coating surface became more numerous with longer exposure time, and that nanoparticles left behind after the coating degraded often bound together in clusters.

“These data, and the data from future experiments of this type, are valuable for developing computer models to predict the long-term release of nanoparticles from commercial coatings used outdoors, and in turn, help manufacturers, regulatory officials and others assess any health and environmental impacts from them,” said NIST research chemist Deborah Jacobs, lead author on the study published in the Journal of Coatings Technology and Research (link is external).

Here’s a link to and a citation for the paper,

Surface degradation and nanoparticle release of a commercial nanosilica/polyurethane coating under UV exposure by Deborah S. Jacobs, Sin-Ru Huang, Yu-Lun Cheng, Savelas A. Rabb, Justin M. Gorham, Peter J. Krommenhoek, Lee L. Yu, Tinh Nguyen, Lipiin Sung. J Coat Technol Res (2016) 13: 735. doi:10.1007/s11998-016-9796-2 First published online 13 July 2016

This paper is behind a paywall.

For anyone interested in the details about the House of Commons nano story I told at the start of this post, here’s the June 23, 2010 posting where I summarized the hearing on nanotechnology. If you scroll down about 50% of the way, you’ll find Dr. Nils Petersen’s (then director of Canada’s National Institute of Nanotechnology) comments about nanoparticles being encased. The topic had been nanosunscreens and he was describing the conditions under which he believed nanoparticles could be dangerous.