Some months ago, my colleague John Balbus posted here about studies finding that when multi-walled carbon nanotubes (MWCNTs) are injected into the abdominal cavities of mice, they induce inflammation and mesothelioma-like reactions similar to those caused by asbestos.  He appropriately cautioned that – among other critical questions – these studies had not demonstrated that inhaled MWCNTs could actually move out of the lung and into the tissues where asbestos gives rise to its effects.  Well, that particular dot now appears to have been connected.

We learned about the new findings via a blog post by Liz Borkowski at The Pump Handle.  She noted a disturbing item on the NIOSH blog posted by Vince Castranova and his colleagues late last week, in which they are seeking to share more broadly results they first presented at the recently-concluded Society of Toxicology meeting in Baltimore.

The NIOSH researchers reported new data showing for the first time that MWCNTs can migrate intact from the alveoli out of the lungs of mice and into the pleura, the tissue surrounding the lungs.  And it is in the pleura (as well as the abdominal cavity) where asbestos induces its signature form of cancer, mesothelioma.

In this case, the MWCNTs were introduced into the lungs using pharyngeal aspiration, a procedure by which mice are induced to inhale a droplet of liquid in which the MWCNTs are suspended.  While this procedure is thought to mimic direct inhalation, the NIOSH researchers note this and other limitations of the study, and caution that the results are preliminary and have not yet been peer-reviewed.

They also note that it's possible that the mice used in the study are unique and may not accurately portray what would happen in people, say, workers exposed to MWCNTs.  And, as my colleague pointed out in his earlier post, whether sufficient material could or would be suspended in the air to result in inhalation exposure also remains an open question.

Nonetheless, these new findings strongly suggest that, like asbestos, MWCNTs behave as stable fibers capable of penetrating and migrating through the lung.  And together with the earlier studies showing that introducing MWCNTs into the tissues surrounding the lung induces mesothelioma-like reactions, it's fair to say the alert level on MWCNTs just went up significantly.

In 2004, Gunter Oberdorster and colleagues demonstrated that upon inhalation, ultrafine particles, the dimensions of which are measured in nanometers, can move from the nasal passages of rodents to the brain via a specialized nerve called the olfactory bulb.  The evolutionary purpose of the olfactory bulb is to relay information about odors directly and rapidly from the nose to the brain. 

The extent to which rapid transit via the olfactory bulb is a significant potential route of exposure to engineered nanomaterials is still an open question.  But two new papers add support for the relevance of this intriguing exposure pathway, raising important questions regarding the safety of inhaled nanoparticles. 

The first paper, by Jiangxue Wang and colleagues, followed the movement of nanoscale titanium dioxide (TiO₂) particles placed directly in the nasal passages of mice to the brain via the olfactory bulb.  When they looked to see where in the brain the TiO₂ went, they found it went pretty much everywhere, although after 30 days the highest concentrations were found in the olfactory bulb and hippocampus.  Moreover, the brain tissue of the exposed mice exhibited changes in structure and biochemistry consistent with damage from reactive oxygen compounds.

Nanoscale silver was the subject of the second paper by Jae Hyuck Sung and colleagues.  Instead of a single exposure, rats were exposed to nanosilver in the air for 13 weeks.  Like an earlier 28-day inhalation study, this one also found widespread distribution of nanosilver in the rats. 

This study also sought to determine if there were any health effects associated with longer-term exposures.  And they did find effects:  inflammation in the lungs, and subtle cellular changes in the livers that are sometimes indicative of pre-cancerous conditions. 

Nanosilver was detected in both the olfactory bulb and the brain but unfortunately the paper did not report on any effects that might have been associated with the presence of nanosilver.  It is not clear from the description provided if the extent of examination of the brain would have been able to identify subtle effects if they were present.

So what should the next steps be?  Elucidating the possible impacts of nanoscale materials on the brain is tricky, in part because the effects could be very diverse, requiring lots of different types of tests to capture them.  While damage to brain cells may be relatively easy to discern, other effects, such as those altering brain development or biochemistry, may require more sophisticated testing. 

While it is often assumed that inhalation exposure to nanomaterials will be limited primarily to workplaces, such materials are also being used in consumer products that can be widely dispersed.  This is particularly true of nanosilver, which can be found in sprays that release nanosilver into the air.  This is why we have repeatedly advocated that such dispersive uses be avoided until more is known about the potential adverse effects that could come with these kinds of exposures. 

My last post identified two Section 8(e) "substantial risk" notices pertaining to carbon nanotubes, one submitted by BASF, the other by Arkema.  I have in my files one additional Section 8(e) notice for a single-walled carbon nanotube (SWCNT), submitted by DuPont.  With three Section 8(e) notices submitted for different rat pulmonary toxicity studies on carbon nanotubes, it's interesting to compare their results.

The DuPont Section 8(e) notice was submitted to EPA on April 10, 2003 (#8EHQ-0403-15319), but appears not to be available anywhere online.  It reports the results of an intratracheal instillation study in rats, which was among the first to identify the formation of lung granulomas.  The full study was published in Toxicological Sciences in 2004.

Here are a few details of each of the three studies reported in the Section 8(e) notices:

DuPont, both in its Section 8(e) notice and in the published paper, questioned the physiological significance of its finding of granulomas, suggesting it was likely an artifact of the method of administration.  Indeed, the instillation procedure introduced clumps or ropes, rather than individual CNTs, and some heavily dosed animals died of apparent suffocation.  DuPont based its conclusion on a variety of factors, including the apparent lack of dose-response and the absence of other signs of lung toxicity.  It called for inhalation toxicity studies to be conducted to resolve whether the effects it observed were real.

What do inhaled carbon nanotubes do to the lungs?

Via the more recent Section 8(e) notices, we now appear to have confirmation of the ability of CNTs to produce lung granulomas when inhaled.  While there are differences in the nature of the CNT material administered and other study details, both Arkema and BASF report the dose-dependent formation of granulomas. 

They also report numerous other signs of lung toxicity, including increased lung weights.  Arkema established 0.1 milligrams per cubic meter (mg/m³) as its short-term no-effect level, while BASF's longer-term study found granulomas at that same dose – the lowest tested – and hence could not establish a no-effect level.

This level of toxicity exhibited in a 90-day repeated dose inhalation study is very high.  To judge toxicity, EPA uses internationally accepted toxicity criteria set forth under the Globally Harmonized System (GHS) of Classification and Labeling.  The GHS "high-concern" classification criterion for a 90-day inhalation toxicity study involving exposure to a dust is a lowest-observed adverse effect level (LOAEL) of less than 0.02 milligrams per liter (mg/L) per day (see Appendix 2, Table 5 of this EPA document).

In the BASF study, the LOAEL was the lowest dose tested, 0.1 mg/m³ = 0.0001 mg/L, which is 200 times lower than the GHS' "high-concern" cutoff value!!

For further context, consider the LOAEL values found in rat subchronic and chronic inhalation studies of another particulate material, respirable crystalline silica.  LOAELs ranged from 1.0 mg/m³ in a 24-month inhalation study to 2.0 mg/m³ in 6-month studies (see Table 18 of this document).  That means the CNTs studied by BASF are at least an order of magnitude more toxic than silica.

Also note that inhaled silica is much more toxic to humans than to rats:  the human LOAEL for inhaled silica for chronic lung disease is far lower than the rat LOAEL, on the order of 0.02-0.05 mg/m³ (see Table 16 of this document).  So if humans are also more susceptible to the effects of CNTs than are rats, the human toxicity level could be far lower than what BASF has observed in rats.

Remember that other recent studies suggest certain MWCNTs can behave biologically rather like asbestos.  Together with these inhalation studies, it's no wonder EPA is requiring inhalation toxicity studies and workplace inhalation exposure controls for CNT producers, as I discussed in a previous post. 

But these findings beg the question:  Will that be enough?

[UPDATE:  Thanks to Andrew Maynard's blog post today, I have learned of a new published study by Shvedova et al. that found that administration of SWCNTs by inhalation was actually more effective than administration by pharyngeal aspiration in causing inflammatory responses and other signs of lung toxicity in the lungs of mice.]

Since my first post concerning EPA's Consent Order, I've been reflecting further on the management conditions it imposes – or, more accurately, on what conditions it doesn't impose.  The Order's only such conditions address potential worker exposure.  What about the rest of the nanomaterial's lifecycle?

The need to consider the full lifecycle and the full range of potential release and exposure pathways is a basic tenet of sound and responsible management of nanotechnology.  That's the backbone of the EDF-DuPont Nano Risk Framework, and it's also a key principle in EPA's own Nanotechnology White Paper and Nanotechnology Research Strategy.

Yet the Consent Order lacks conditions to address any potential releases or exposures beyond requiring gloves and other personal protective equipment for workers handling the nanomaterial.

Remember, the Consent Order notes that no test data were included in the producer's premanufacture notification (PMN).  On what basis, then, has EPA concluded that no other potential risks exist?  What about potential releases:

• from the manufacturing facility to the ambient air or water?
• from disposal or other management of wastes?
• from downstream transport, storage or processing?
• from post-use management (e.g., aging, weathering, repair, recycling) and disposal of products (electronics, polymer composites) containing the nanomaterial?

Nothing in the Consent Order addresses these questions – not, for example, a requirement to test products for potential releases, not even provisions to require reporting of waste management information or measurement or monitoring of releases.

Now, it may be that EPA has somehow managed to fully evaluate these and related questions and has determined that all of these risks are – and will remain, no matter what quantity of the nanomaterial is produced and used in the future – negligible.  If so, it should disclose how and on what basis it did so. 

EPA's failure to make public the decision framework it uses to evaluate new chemical submissions for nanomaterials – something we and other stakeholders have been requesting for some time – is a major impediment to building public trust in its process.

EPA's only bite at the apple

Some might argue that all this is premature and that EPA should wait until manufacture and use of this nanomaterial has ramped up even to consider such questions.  But here's the problem with that:  Under TSCA, the PMN review and conditions imposed through the Consent Order are essentially EPA's only bite at the apple.

Once Swan Chemical commences manufacture of its multiwalled carbon nanotubes (MWCNTs), they will be listed on the TSCA Inventory and will no longer be a "new" chemical.  At that point, anyone may manufacture and use the substance without even having to notify EPA.  And no matter what the quantity of the nanomaterial being produced and used, no further review by EPA would be triggered.

It should be noted that EPA is likely developing a Significant New Use Rule (SNUR) to accompany this Consent Order, and there are hints of that in the Order itself.  But all that a SNUR will do is to extend the same conditions that apply to the submitter of the PMN to other producers; it would require them to notify EPA only if they don't comply with these conditions.

Should a concern later develop about some other type of release or exposure not addressed by the Order and SNUR, EPA's only recourse would be to seek to use its authority under TSCA Section 6 to regulate the MWCNTs as an "existing" chemical – something EPA was unable to do, ironically, even for asbestos.  Yet that would be the only way that EPA could impose further conditions on production, processing, use, distribution or disposal of this nanomaterial.

Absent reform of TSCA to provide EPA with greater authority to regulate "existing" chemicals, EPA's new chemical review is, practically speaking, the only chance to ensure that potential risks across a new substance's full lifecycle are addressed.  An examination of EPA's Consent Order suggests that this opportunity has been lost for this nanomaterial.

[Part II of this post is available here.] 

Word hit the street today that EPA intends to make public a "sanitized" version of a Consent Order it has negotiated with a producer of multiwalled carbon nanotubes (MWCNTs).  [A link will be provided once available.]  We obtained a copy of the Order, which has redacted all information claimed confidential by the company involved.  What can we learn from this well-scrubbed Order?

The Order was triggered by EPA's review of a premanufacturing notification (PMN) – which, of course, is only required of companies producing the subset of nanomaterials EPA has decided to regard as "new."  This is the first public glimpse, albeit limited, into both EPA's thinking and its regulatory approach to "new" nanomaterials. 

EPA redacted the name of the company that has agreed to this Order – even though a simple Google search revealed that it almost certainly is Swan Chemical, Inc., of Lyndhurst, NJ.  That's because the company recently issued a press release announcing the Order.

So what does the Order call for?  The company is to:

• conduct a 90-day inhalation toxicity test in rats;
• supply EPA with a 1-gram sample of its MWCNTs and its Material Safety Data Sheet;
• submit certain characterization data within 6 months after commencing full manufacture;
• require its workers to wear protective gloves and clothing shown to be impermeable and NIOSH-approved respirators;
• use the substance only for a particular use, claimed confidential but generically identified as a "property modifier" in electronics and polymer composites; and
• provide the nanomaterial only to entities that agree to the same use restrictions and worker protection conditions.

Given recent evidence that MWCNTs can behave biologically rather like asbestos, such measures are more than called for.

But what's equally interesting are the details, including those that are missing from the sanitized Order because they were deemed confidential:

• The inhalation toxicity study doesn't need to be submitted before manufacture commences, but rather 14 weeks before either:  a) manufacture reaches a level of [BLEEP] kilograms, or a period of [BLEEP] years and [BLEEP] months after manufacture passes, whichever comes first. 
       From this, it's not at all clear:  a) why such details are secrets, and b) just how long it will be before the study is submitted.
• Adding to this mystery, the clock doesn't even start clicking on the time limit until two years after the Order is signed.
• The Order notes that no – zero, none, nada – test data were submitted with the company's PMN.
       This is actually not unusual for PMNs that come in under the Toxic Substances Control Act (TSCA), since EPA cannot require development and submission of such data up front, even for conventional chemicals. 
• Despite this lack of data, the Order states that EPA has determined that no significant environmental effects are expected.  Given how little environmental data exist on nanomaterials in general, let alone this particular MWCNT, it's hard to imagine how EPA reached this conclusion.
• The Order does state that "EPA is unable to determine the potential for human health effects" from exposure to this nanomaterial, and hence that it "may present an unreasonable risk of injury to human health."
       Presumably this is because of those recent findings that MWCNTs may act like asbestos.

So, to summarize these last two points:  Only if EPA already has evidence of a potential effect can it conclude that it is unable to determine whether there is an effect and call for testing.  If EPA doesn't have evidence of a potential effect – even if it has no data at all – it's ready to conclude that no significant effects are expected.  Welcome to life under TSCA!

Other very interesting tidbits:

The Order encourages the company to sign up for the in-depth phase of EPA's voluntary Nanoscale Materials Stewardship Program (NMSP), stating that if it does, EPA may waive the Order's requirement to conduct an inhalation toxicity test.  We can only wish that EPA gets something more than a non-binding commitment from the company to do some testing before vacating the Order!

Assuming the Order stays in place, if EPA finds the results of the required study to be "equivocal," the company can expand production beyond the production limit.  (I won't even discuss the perverse incentive this provision could create.)  Only if the company wants to get out from under the Order's other requirements (the use restriction or the worker protection provisions) need it reconduct the study.

What if the test results are invalid?

The Order appears to state that, if EPA finds the test data to be invalid, the company cannot expand production.  But there's a catch:  If there's not enough time to reconduct the study and submit it 14 weeks before exceeding the production limit, then the company can go ahead and exceed the limit as long as it submits the study "within a reasonable period of time."  The company can also exceed the production limit if it challenges EPA's determination that the data are invalid in writing.

If the company decides the test data are invalid before submitting them to EPA, as long as it informs EPA of this determination, EPA can still decide to allow expanded production.  As before, if EPA does decide to require the company to reconduct the study, but there's not enough time to reconduct and submit it 14 weeks before exceeding the production limit, then the company can go ahead and exceed the limit as long as it submits the study "within a reasonable period of time."

What if the test results show significant risk?

The Order states that if EPA determines that the data are valid and unequivocal and indicate the nanomaterial "will or may present an unreasonable risk," EPA may but is not required to notify the company, and may but is not required to impose additional conditions.  If EPA does issue such a notice, the company must either comply with the new conditions or cease production, use and distribution.  But again there's a catch:  If the company challenges EPA's determination in writing, it can continue these activities while the dispute is resolved.

All these allowances for the very type of nanomaterial that is #1 on just about everyone's concern list.

[UPDATE:  It should be noted that the above allowances are actually fairly standard practice for new chemical consent orders and are not limited to those issued for nanomaterials; indeed, EPA has developed "boilerplates" for its consent orders that contain very similar language.]

I have been among those calling on EPA for greater transparency in how it assesses new nanomaterials.  This is a step in the right direction in that regard.  But I have to say that getting this glimpse at EPA's inner workings doesn't exactly bolster my confidence in what they're doing.

John Balbus, M.D., M.P.H., is Chief Health Scientist.

A new study published today in Nature Nanotechnology finds that multi-walled carbon nanotubes (MWCNTs) cause inflammatory changes in mice that closely resemble those caused by asbestos.  This is the second study in a few months to make this finding.  (I posted on the first, Takagi et al., a few weeks ago.)  So is the case closed on multi-walled carbon nanotubes?  Or is too early to draw conclusions?

These two studies used different approaches to compare the effects of MWCNTs and asbestos.  Although both directly injected the nanotubes into the peritoneal cavities of mice (the peritoneum is the lining of the abdominal organs and walls, and is the part of the body – along with the pleura lining the lungs – where asbestos causes mesotheliomas to arise), the new study used a 60-fold lower dose and measured only inflammatory changes, not the actual development of mesotheliomas.  Takagi et al., on the other hand, used special mice that were genetically highly susceptible to cancer formation, and observed the mice until they developed tumors. 

Despite these differences, these studies together make a strong case that multi-walled carbon nanotubes cause inflammation and cancers in a similar fashion to asbestos, at least if they have been deposited directly within the peritoneum. 

But the studies only tell a part of the story.  In order to have a better handle on the risks from inhaling MWCNTs, studies need to be done to determine whether there could be substantial concentrations in the air that workers (or other exposed persons) breathe.  Then we need to find out whether inhaled MWCNTs will make their way from the airspaces of the lungs, through the lung tissue, to the lung and abdominal cavity linings – something asbestos does with relative ease. 

If these first two steps in causing disease are demonstrated, it is likely that MWCNTs will persist and continue to cause inflammation as long as asbestos does, but this needs further investigation as well.  So the case isn’t closed, but the evidence is stacking up.

Until these questions are answered, companies using MWCNTs should carefully characterize their materials and refrain from exposing workers to nanotubes longer than 5 microns with fibrous shapes.  And researchers developing applications for MWCNTs should look to develop forms that don’t have those size and shape characteristics.  These studies don’t show that other forms are harmless, but it can’t hurt to sidestep the trouble you can see.

John Balbus, M.D., M.P.H., is Chief Health Scientist.

It’s been a worry for engineered nanoparticles. Now, a new study from the Harvard School of Public Health (Suglia et al., 2008) is the first to suggest that particulate air pollution not only damages the lungs and heart, but also may damage the developing brain.

Researchers measured cognitive function in over 200 children in Boston in relation to their residential exposure to traffic-related air pollution by measuring airborne carbon black particles. They found the IQ-lowering effect of higher exposure is comparable to a pregnant mother smoking 10 cigarettes a day or moderate lead exposure.

It’s important to note that carbon black is only used as an indicator for traffic-related particle pollution in this study, and the authors stress that it should not be concluded that pure carbon black is the causative agent here. The actual particles inhaled are likely to be contaminated by heavy metals and other toxins, and these may well be the culprits or at least greatly contribute to the actual damage to brain tissues. But it does suggest that some of the ultrafine fraction of air pollution finds its way into the brain and causes harm.

This study may not be making headlines in the nanoworld, but the nanoworld should be taking notice.

The authors hypothesize that the combustion particles make their way via the olfactory nerves from the children’s nasal passages directly into the children’s brains, as has been demonstrated for a variety of engineered nanoparticles in animal models (e.g., Oberdorster et al., 2004; Elder et al., 2006). Once there, the particles, including attached contaminants, may cause oxidative stress, ultimately leading to inflammation and cellular malfunction and/or damage. The study did control for potentially confounding factors like maternal cigarette smoking, lead levels in the children’s blood, mother’s educational level, low birth weight, and other factors.

This study should give pause to those who are making nanoparticles containing known neurotoxins, such as lead and manganese. And with growing understanding of the linkages between developmental and degenerative diseases of the brain, there’s reason to think that processes that lead to IQ loss in growing kids can impair brain functioning in aging adults. One more reason to avoid inhalation of nanoparticles, especially those with known capacity to cause oxidative stress.