1. science
2. physics
3. nanotechnology

Svenska myndigheter och nanomatrial
PM 3/15
Nanomaterials upptag och spridning i
kroppen och miljön
kemikalieinspektionen.se
Kemikalieinspektionen är en myndighet under regeringen. Vi arbetar i Sverige, inom EU och
internationellt för att utveckla lagstiftning och andra styrmedel som främjar god hälsa och bättre
miljö. Vi har tillsyn över reglerna för kemiska produkter, bekämpningsmedel och ämnen i varor
och gör inspektioner. Vi granskar och godkänner bekämpningsmedel innan de får användas.
Vårt miljökvalitetsmål är Giftfri miljö.
© Kemikalieinspektionen, Stockholm 2015.
Artikelnummer: 511 153.
Förord
Kemikalieinspektionen har på uppdrag av regeringen tagit fram en handlingsplan för en
giftfrivardag Handlingsplan för en giftfri vardag 2011– 2014 – Skydda barnen bättre. Insatser
sker nu på flera områden både nationellt, inom EU och internationellt och ofta i samarbete
med andra myndigheter. Att minska kemiska risker i vardagen är ett steg på vägen att nå
riksdagens miljökvalitetsmål Giftfri miljö – det mål Kemikalieinspektionen ansvarar för.
Inom ramen för handlingsplanen tar KemI fram sammanställningar som publiceras i
Kemikalieinspektionens rapport respektive PM-serie. Publikationerna, som är kostnadsfria,
finns på webbplatsen www.kemikalieinspektionen.se
I Kemikalieinspektionens handlingsplan för en Giftfri vardag 2011-2014 framhålls behovet av
insatser för att nå en hög skyddsnivå för eventuella hälso- och miljörisker orsakade av
nanomaterial. Som ett led i arbetetet sammankallades myndigheter/statliga aktörer vars
verksamheter är berörda av nanoteknik och nanomaterial till ett fjärde möte för kunskaps- och
erfarenhetsutbyte samt diskussion. Myndighetsmötet ordnades denna gång i samarbete med
Nordiska Nanomaterial Gruppen sponsrat av Nordiska Kemikalie Gruppen (NKG). Mötet
bestod av en gemensam workshop på förmiddagen och ett myndighetsmöte med nordiska
deltagare på eftermiddagen.
Föreläsarna har själva bidragit med sammanfattningar från sina presentationer. I detta PM
ingår bidrag från Kungliga Tekniska högskolan, Danska Miljöstyrelsen, Totalförsvarets
forskningsinstitut, Lunds Universitet, representanter för OECD och EU kommissionen,
Läkemedelsverket, Kemikalieinspektionen och Miljödepartementet.
Samordning av arbetet och sammanställning från mötet har gjorts av Lena Hellmér,
Kemikalieinspektionen. Ansvarig för arbetet var Lisa Anfält, chef för enheten EU
koordinering på Kemikalieinspektionen.
Innehåll
Inledning............................................................................................................ 5
Summary ........................................................................................................... 6
1
Sammanfattningar från föredragen ..................................................... 9
Nanomaterials in a Life Cycle Perspective .................................................................................... 9
Dermal absorption of nanomaterials ............................................................................................. 9
Interactions of nanoparticles with organs protected by internal biological barriers ..................... 10
Measurement Techniques for Airborne Nanoparticles ................................................................ 11
Effekter efter inandning av metalloxider i friska och känsliga individer ....................................... 12
Nanosäkerhet ur ett EU- och OECD perspektiv .......................................................................... 13
Update on NM projects for the method, guidance and testing strategy development in the OECD
Test Guidelines programme .......................................................................................... 14
Nytt om nano från Kemikalieinspektionen ................................................................................... 15
Nanomaterial i kosmetiska produkter – vad har hänt sedan de nya reglerna infördes? ............. 15
Danske aktiviteter vedr. nanomaterialer ...................................................................................... 16
Nanocellulose and NANoREG- project ....................................................................................... 17
2
Bilagor ................................................................................................. 19
Bilaga 1. Deltagarlista.................................................................................................................. 19
Bilaga 2. Föreläsarnas power-point presentationer .................................................................... 21
Inledning
Detta PM är en sammanställning av det fjärde myndighetsmötet om nanoteknik och nanomaterial Syftet med mötet som redovisas i detta PM var att få kunskap om aktuell forskning
och de frågor som svenska och statliga aktörer arbetar med inom nanoområdet samt att
fortsätta utveckla myndighetsnätverket.
Denna gång deltog även forskare och myndighetsrepresentanter från övriga Norden. En
inbjudan till myndighetsmötet skickades till samtliga departement inom Regeringskansliet
och de statliga myndigheter eller offentliga finansiärer som bedömdes ha aktiviteter eller
intressen med anknytning till nanomaterial och nanoteknik i Sverige. Totalt deltog 49
personer från 24 aktörer på seminariet den 27 november 2014.
Dessa var Bioforsk, Danmarks tekniska universitet, Europeiska kommissionen, Finska
Arbetshälsoinstitutet, Totalförsvarets forskningsinstitut (FOI), Formas, Försvarets materielverk, Försvarsmakten, Generalläkaren, Kemikalieinspektionen, Kommerskollegium, Kungliga
Tekniska högskolan (KTH), Livsmedelsverket, Lund universitet, Läkemedelsverket, Naturvårdsverket, Norwegian institute for water research, norska Miljødirektoratet, OECD
sekretariat, Regeringskansliet, SINTEF Materials and Chemistry, Socialstyrelsen, Danska
miljöstyrelsen och Trafikverket.
Workshopen/seminariet hölls omväxlande på engelska och ”skandinaviska” och därför
varierar språket även i rapporten. Som moderator för mötet fungerade Gregory Moore från
Kemikalieinspektionen.
Introduction
This PM is a compilation of speeches made at the fourth network meeting with authorities on
nanotechnology and nanomaterials. The purpose of the meeting accounted for in this PM was
to acquire knowledge about current research and topical questions at Swedish and government
authorities within the nano area and to continue developing the authority network on
nanotechnology and nanomaterials.
Participants at this meeting were researchers and authority representatives from the Nordic
countries and the invitation was sent to all Swedish ministries and official financiers
estimated to carry out activities or having interests in nanotechnology and nanomaterials in
Sweden. 50 participants from 26 actors participated at the seminar on 27 November 2014,
which was held alternately in English and a Scandinavian language and that is why the
language varies in the report too. Moderator of the meeting was Dr. Gregory Moore, the
Swedish Chemicals Agency.
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Summary
David Lazarevic, from KTH gave an introductory lecture on Nanomaterials in a Life Cycle
Perspective. Engineered nanomaterials are being used in a growing number of products. The
assessment of the impacts, or benefits, upon human health and the environment should be
conducted from a life cycle perspective. However, current assessments are inadequate due to
the lack of data on the emissions from engineered nanomaterials and nanoparticles during the
production, use and disposal phases of a product life cycle, and the lack of characterisation
factors for nanomaterials in life cycle impact assessment models.
From the Danish EPA Anne Mette Zenner Boisen, presented two projects on dermal
absorption of nanomaterials. The literary project from 2013 performed a scientific appraisal of
the reliability and relevance of available studies on dermal absorption using Klimisch criteria
and nanomaterial characterisation, database available from the Danish EPA.
Recommendations include that study designs (in vivo and in vitro) should consist of a
sufficient post-exposure duration to account for potential lag-times of dermal absorption. An
experimental project on dermal absorption of TiO2 and ZnO in a realistic exposure scenario
using in vitro and in vivo test methods is ongoing and will be published in 2015.
Anders Bucht introduced us to the area of interactions of nanoparticles with organs protected
by internal biological barriers. Inhalation of nanoparticles leads to surface adsorption of
biomolecules in the lung lining fluid that influence responses of cells in the lung epithelium
and uptake in the body. When translocated into the blood circulation a corona of blood plasma
proteins will be formed which can activate the contact dependent coagulation system. This
activation may elicit tromboinflammation, blood clot formation, as well as activate innate and
adaptive immune responses.
The important topic of Measurement Techniques for Airborne Nanoparticles was presented
by Jakob Löndahl, Lund University. A major pathway for exposure to nanoparticles is
through the air during breathing. Thus, it is necessary to have adequate monitoring techniques
that ideally are able to separate engineered nanoparticles from the background. There is a
range of instrumental options for measurement of airborne nanoparticles, but often it is
necessary to choose between simplicity and accuracy (or high time resolution and relevant
particle characteristics). It is still also difficult to measure relevant exposure metrics such as
only the engineered nanoparticles. However, new methods are emerging that might overcome
this problem at least partly.
Åsa Gustafsson held a presentation about her thesis entitled ”Nanomaterials, respiratory and
immunological effects following inhalation of engineered nanoparticles”. It shows that
exposure to nano sized titanium dioxide (TiO2) induced long-lasting immune response in the
airways. The long-term activation of the immune system could potentially trigger the
development of diseases. The particles that were deposited in the alveolar region were also
retained in the lung for a long time, up to three months after exposure. By comparing different
inbred rat strains it was demonstrated that genetically determined factors influence the
immune and respiratory responses to TiO2. An important issue of this thesis was to study the
responses in sensitive individuals. These sensitive individuals were represented by groups of
animals with an allergic airway inflammation or by rat strains that were genetically
susceptible to inflammatory disorders. One study showed that when the mice were exposed to
particles and an allergen during the same period, a decline in general health was observed.
Altogether, this thesis emphasises the complexity of assessing health risks associated with
nanoparticle exposure and the importance of including sensitive populations when evaluating
adverse health effects of ENMs.
6
Eva Hellsten, former Head of the Chemicals unit at the European Commission, vice
president in the OECD WPMN, and member of the Swedish Nano Commission, informed
about nanosafety from an EU and OECD perspective and her experiences in the area. Within
the EU, work on health and environment safety aspects of nanomaterials started in 2004. Via
the EU research programmes, 25 million Euros have been invested each year in this area. The
EU legislation has been reviewed to analyse in which ways legislation needs to be adapted to
better cover nanomaterials. For legislative purposes adaptation of existing test methods is
needed, e.g. testing of (eco)toxicology, exposure and measurements. Work to ensure that test
methods are standardised and internationally harmonised is on-going within the OECD
WPMN since 2006. In Sweden, a Government Nano Commission was initiated in 2012. The
conclusions from this investigation (SOU 2013:70) stresses the importance of additional
Swedish efforts in participating in EU and OECD work, as well as improved co-ordination
and communication between Swedish authorities, academia industry and civil society. A
Nano Council, with a Nano Centre as secretariat, was proposed to be established for this
purpose.
Jukka Ahtiainen, presently Senior researcher at the OECD secretariat continued with the topic
and gave an update on NM projects for the method, guidance and testing strategy
development in the OECD Test Guidelines programme. The OECD test guidelines for testing
chemicals have been widely used for regulatory purposes all over the world since the
establishment of the MAD principle in 1984. This Mutual Acceptance of Data ensures that if
a chemical is tested under the GLP conditions according to an OECD Test Guideline, the data
should be accepted in all OECD countries. The rationale behind this agreement is to save
resources and avoid especially duplicate vertebrate testing. Eventually the OECD test
methods are referred to or taken into the national chemicals legislation such as the test method
regulation (440/2008/EU) in the European Union.
Sofia Tapper from the Swedish Ministry of the Environment and Energy informed among
other things about the Nano Commission and an action plan for safe use and handling of
nanomaterials. The new Swedish government has shown interest in the commission and its
conclusions and intends to continue pressing the European Commission for the importance of
adequate EU-rules on nano materials. Work on establishing a possible Nano Centre or Nano
Council continues.
Elin Simonsson, the Swedish Chemicals Agency, briefly presented ongoing work at the
agency on nano materials, i.e. amendments of REACH annexes, overview of the EU
Commission recommended definition of nanomaterials and its survey of consequences with
regard to measures for increased transparency of nanomaterials on the market.
Tomas Byström, the Swedish Medical Products Agency, informed about the rules on
nanomaterials in cosmetic products, which came into force in 2013. Some regulatory support
is, however, lacking with regard to approved substances in nano form. Within the EU; 25 000
cosmetic products and nanomaterials have been notified. The European Commission will
within short publish a catalogue listing nanomaterials used in cosmetic products.
Flemming Ingerslev, gave an account of the activities of the Danish Environmental Protection
Agency
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6 million DKK have been allocated per year during 2012-2015 to ”Bedre styr på
nano”, with the overall aim to get an overview of where in Denmark there may be a
risk to consumers and the environment from the use of nano products.
7
-
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One part of the effort included to set up a register of consumer products releasing
nanomaterials. Danish manufacturers have to report their production and import of
nano products aimed for consumers to this register.
A number of other projects are also included in the work, from which reports are
published continuously.
Denmark also contributes to the OECD work on developing ecotoxicological methods
for testing nano materials.
How can authorities make use of the results of nano research and how can authority work have
an effect on research?
Jukka Ahtiainen opened the discussion by presenting the Finish participation in the
NANoREG project.The safety of nanomaterials is investigated in the EU’s just-beginning
extensive NANoREG project as a joint effort by the authorities and the industry. Finland’s
goal in the project is to continue to study the safety of microfibrillar and nanofibrillar
cellulose. In Finland, participation in the NANoREG project is coordinated by the Finnish
Safety and Chemicals Agency (Tukes) that is also responsible for oversight and guidance
concerning the REACH regulation. Also included in the project are the Finnish Institute of
Occupational Health, responsible for the experimental in vitro and in vivo studies, and Stora
Enso and UPM as a joint Nordic Cellulosa consortium.
Finland’s national research portion of the Nanoreg project focuses on microfibrillar and
nanofibrillar cellulose materials that have several potential industrial applications in different
products. Microcellulose and nanocellulose comprise wood fibre and fibre bundles originating
from wood cellulose. The intention is to study the safety of biodegradable microcellulose and
nanocellulose experimentally by means of biological testing. This is important in order for it
to be possible to use nanocellulose, for example, as raw material for cosmetics, food additives
or packaging.
The ensuing discussion raised the cooperation between the OECD and NANoREG, the design
of a possible Nano centre in Sweden and ways in which Nordic colleagues treat different
aspects of nanomaterials. In conclusion, it was agreed that a combination of a Nordic
workshop and authority meeting was fruitful, particularly with respect to the discussions. The
moderator extended his thanks to speakers and participants.
8
1
Sammanfattningar från föredragen
Nanomaterials in a Life Cycle Perspective
David Lazarevic, PhD, Researcher, KTH
Nanotechnology and nanomaterials have been promoted as having the potential to bring
benefits to many areas of research, and to positively contribute to sustainable development.
As such, this rapidly growing field is increasing attracting investments from governments and
businesses worldwide. However, it is also recognised that engineered nanomaterials may pose
a risk to human health and the environment.
There is a general consensus that the potential health and environmental risks of engineered
nanomaterial should be evaluated over their entire life cycle. Life cycle assessment is one tool
which has been promoted for such an evaluation. This work reviewed the literature on the
application of life cycle assessment to engineered nanomaterials to identify current research
and difficulties in applying life cycle assessment in this field.
Twenty five LCA studies of nanomaterials were identified, including nanomaterial such as
cadmium telluride, calcium carbonate, carbon black, carbon nanofibres, carbon nanotubes,
nanoclay, nanoscale platinum-group metals, silica, silver, silicon, titanium and titanium oxide.
Product systems studied include: auto-body panels, biopolymers, coatings, electronic
displays, electronic sensors, lithium-ion batteries, photo voltaic systems, packaging and
agriculture polymer films, nanomaterial production processes, textiles and wind turbine
blades. These studies only looked at parts of the life cycle, with no quantitative studies
addressing the impact of nanomaterials to human health and the environment from the cradle
to the grave. Results from these studies showed the potential for a significant cumulative
energy demand in the production of nanomaterials such as carbon nanotubes and carbon
nanofibres. However, this is reduced when taking into consideration the small amounts of
nanomaterials in products and the potential benefits during the use phase, such as weight
reduction.
Due to the different properties and functions of engineered nanomaterials when compared to
conventional materials and products, special attention is required during the goal and scope
definition phase in order to obtain meaningful results. The life cycle inventories of current
LCA studies cannot be classified as comprehensive as they often lack nanomaterial specific
data related to the outputs of processes. Hence, populating life cycle inventory databases with
nanomaterial specific information, such as size and shape, is of critical importance. Although
the UNEP/SETAC framework for toxic impacts can in principle be used for specific impacts
causes by nanoparticles, life cycle impact assessment methods currently lack characterisation
factors for the release of nanoparticles indoors and outdoors. Hence, no LCA studies to date
have considered the human toxicity and eco-toxicity of nanomaterials from a life cycle
perspective with consideration of the nano-specific properties.
Dermal absorption of nanomaterials
Anne Mette Zenner Boisen,PhD Danish Ministry of the Environment
Under the Danish Nano Initiative ‘Better Control of Nano’ the Danish Environmental
Protection Agency (EPA) has initiated a series of projects with the aim of further clarifying
possible risks to consumers and the environment from nanomaterials. Two of these projects
investigating dermal absorption of nanomaterials were presented at the workshop on 27th
November 2014.
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The first, Danish EPA Environmental Project No. 1504, 2013 was a literary project conducted
in 2013 by the Institute of Occupational Medicine IOM, UK. The project performed a
scientific appraisal of the reliability and relevance of available studies on dermal absorption
using Klimisch criteria and nanomaterial characterization (as described in Card and
Magnuson 2010). The appraised studies are available in a database, which can be found in a
link within the project report (original version). A link to an updated version of the database,
which included more studies were also performed in 2013 and can be found on the Danish
EPA website (mst.dk). This update did not change any of the conclusions found in the project
report. Results from the project showed that a very low dermal absorption of nanomaterials is
possible in some cases; Nano-specific characteristics that may influence dermal absorption
include size, shape and surface chemistry; test method(s) which most closely simulate the
transport of nanomaterials through human skin are evaluated in the report and research gaps
concerning dermal absorption of nanomaterials are described. It has been noted in several
studies that there can be a considerable lag time between application of a test substance and
appearance in the circulation. Gulson et al. 2010 noted a lag time of ~30 h before the first
detection of 68Zn in the blood and urine of human volunteers. Therefore one recommendation
from the project is that study designs (in vivo and in vitro) should consist of a sufficient postexposure duration to account for potential lag-times.
The second project, which is experimental is ongoing and will be concluded in the summer of
2015. This project is conducted by Aarhus University in Denmark. The main goal of this
project is to investigate if TiO2 and ZnO nanoparticles are able to penetrate the skin in a
realistic exposure scenario. Sun screen is used as the vehicle and 2.5% nanoparticles by
weight are added to the sunscreen. The Main goal of the project is to investigate if TiO2 and
ZnO nanoparticles are able to penetrate the skin in a realistic exposure scenario. Nanospecific characteristics investigated in the project are size and coating. Test methods include:
The in vitro Epiderm model; An in vivo mouse inflammation model and an in vivo mouse
xenograft transplantation model with human skin. The results from this project will be
published in peer-reviewed scientific articles and in a Danish EPA project report in 2015.
Interactions of nanoparticles with organs protected by internal
biological barriers
Anders Bucht, Division of CBRN Defence and Security, Swedish Defence Research Agency,
and Department of Respiratory Medicine, Umeå University.
The widespread exploitation of new types of nanoparticles in a variety of industrial
applications and cosmetics has raised concerns about toxicity when dispersed in occupational
and public environments, especially when humans are exposed by the inhalation route. We
and others have demonstrated cellular uptake of titanium dioxide nanoparticles (TiO2 NP)
when exposed to lung epithelial cells, and that the NP interactions with the cells results in
inflammatory responses and oxidative stress ( 1 2). The ability of NPs to translocate
intracellularly and activate cells is, however, highly dependent on particle properties (e.g.
size, crystal structure surface chemistry and agglomeration), cell culture conditions and type
of responder cells. For example, addition of serum proteins to cell cultures will greatly
influence the responses to the NPs, most likely due to formation of a protein corona on the
surface of the particles. The composition of biomolecules adsorbed on the surface will
1
Ekstrand-Hammarström B, Akfur CM, Andersson PO, Lejon C, Österlund L and Bucht A. Nanotoxicology
2012, 6:623-634
2
Andersson PO, Lejon C, Ekström Hammarström B, Akfur C, Bucht A and Österlund L. Small 2011,7:514-23
10
gradually shift when particles translocate into the cells, i.e. when exposed through the airways
the NP corona will initially consist of components of the lung lining fluid and after cellular
uptake biomolecules from the cell interior will attach to the particles. When NPs are further
translocated into the blood circulation, a corona of blood plasma proteins will be formed,
including components of the complement and the coagulation systems.
Detailed analysis of adsorbed plasma proteins bound to TiO2 particles after incubation with
human plasma has shown enrichment in proteins of the contact dependent coagulation system
on the surface. Using a whole-blood model utilizing fresh non-anticoagulated human blood, it
was shown that TiO2 NPs at very low concentrations (50 ng/mL) induce strong activation of
the contact coagulation system, which in this model elicits thromboinflammation and blood clot
formation ( 3). These data are in line with the finding of components of the contact system in
the protein corona of the TiO2 NPs after exposure to blood. From that study it was concluded
that TiO2 NPs, generally considered to be relatively harmless, are highly thrombogenic when
they enter the body and cross epithelial and endothelial borders. Such activation may potentially
induce immune activation, inflammation and tissues damage in vivo.
Measurement Techniques for Airborne Nanoparticles
Jakob Löndahl, Div. of Ergonomics and Aerosol Technology, Lund University
The rapidly growing nanotechnology sector has resulted in an increasing need to understand
health risks associated with exposure to the new materials emerging. A major pathway for
exposure is through the air during breathing. To investigate, control and limit airborne
exposure to nanoparticles it is necessary to have adequate measurement techniques. Such
techniques should ideally be able to separate engineered nanoparticles from the background
and to measure the most relevant particle properties. Relevant particle properties may for
instance be size, shape, surface area, chemical composition, solubility and biological activity.
Among the easiest real time techniques to use are the instruments based on electrical charge.
These measure particle number and/or surface area with reasonable accuracy, but are
generally not able to sort out particles smaller than 100 nm. More advanced instruments
include for instance the scanning mobility particle sizer (SMPS), the electrical low pressure
impactor (ELPI), the aerosol particle mass analyser (APM) and the aerosol mass spectrometer
(AMS). The latter being the most informative in terms of time resolved chemical composition
with simultaneous particle size distribution. To measure certain particle properties it is
necessary to perform sample collection with subsequent analysis by for instance electron
microscopy, x-ray, PCR etc.
There is a range of possibilities to measure a wide range of properties of airborne
(nano)particles. However, due to instrumental limitations and cost effectivity it is often
necessary to choose between simplicity and accuracy (or high time resolution and relevant
particle characteristics). It is still also difficult to measure relevant exposure metrics such as
only the engineered nanoparticles, but new methods are emerging.
3
Ekstrand-Hammarström B, Hong J, Davoodpour P, Sandholm K, Ekdahl KN, Bucht A, Nilsson B. Manuscript.
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Effekter efter inandning av metalloxider i friska och känsliga
individer
Åsa Gustafsson, Forskningsingenjör PhD student, FOI
Syftet med denna avhandling har varit att få en mer detaljerad förståelse av lungornas och
immunsystemets påverkan efter inandning av nanopartiklar. En viktig del av arbetet har varit
att skapa en förståelse om hur känsliga grupper i samhället påverkas av nanopartiklar. I detta
arbete representeras speciellt känsliga individer av försöksdjur med inducerad allergisk luftvägsinflammation samt av djur med särskilt stor benägenhet att utveckla inflammatoriska
sjukdomar. Dessa exponeringar är tänkta att efterlikna de som förekommer i arbetsmiljöer vid
framställning av sådana nanomaterial. I denna avhandling har friska möss och råttor samt djur
med en allergisk luftvägsinflammation andats in nanopartiklar av titandioxid eller järnoxid
varefter de respiratoriska, inflammatoriska och immunologiska svaren studerats.
Studierna visade att kroppen har svårt att göra sig av med titandioxidpartiklar som hamnar i
lungblåsorna. Partiklarna inducerade en tidig ökning av celler i lungan redan efter 1 dag, och
fortfarande efter tre månader kunde en förhöjd ökning av inflammatoriska celler observeras i
lungblåsorna. Histologisk analys visade att det även fanns partiklar kvar i lungvävnaden. De
djur som har en genetisk benägenhet för autoimmun-liknande sjukdomar utvecklade ett
kraftigare immunologiskt svar efter partikelexponering jämfört med de djur som har medfödd
benägenhet för allergiska sjukdomar. Djur med en allergisk luftvägsinflammation fick inga
förvärrade andningsbesvär eller förvärrade inflammationer i lungan efter partikelexponering.
Däremot var cellsammansättningen i lungan annorlunda jämfört med de allergiska djuren som
inte fick partiklar. Dessutom påverkade tidpunkten för partikelexponeringen det inflammatoriska och immunologiska svaret i djuren beroende på om de ges vid allergensensibilisering
eller senare då redan sensibiliserade djur utsätts för allergenet på nytt.
Studien av järnoxidexponeringar visade att allergiska och friska möss som fick partiklarna i
lungorna fick helt olika inflammatoriska svar. De friska mössen utvecklade en inflammation i
lungan och i de lymfkörtlar som dränerar lungorna. Däremot observerade vi färre inflammatoriska celler hos möss med en pågående allergisk luftvägsinflammation en dag efter exponering för partiklar. Minskningen kunde även noteras i både luftvägar och lymfkörtlar. Cellminskningen kan bero på att lungor har förhöjda nivåer av fria syreradikaler vid pågående
inflammation samt att järnoxid kan generera ytterligare reaktiva syreradikaler. Detta tillsammans kan leda till ökad oxidativ stress som i sig kan leda till celldöd.
Studierna visade att titandioxidpartiklar ligger kvar i lungblåsorna under lång tid samt att en
långvarig aktivering av immunsystemet kan uppstå vid lungexponering för nanopartiklar. En
sådan immunaktivering skulle kunna leda till utveckling av immunmedierade sjukdomar. I
råtta visades att nedärvda faktorer har betydelse för hur immunsystemet aktiveras efter inandning av titandioxidpartiklar. Allmäntillståndet hos allergiska möss försämrades efter titandioxidexponering men detta observerades inte i allergiska råttor. Däremot kunde en ökning av
neutrofiler konstateras i möss och den råttstam som är benägen för autoimmuna sjukdomar.
Den stora skillnaden mellan friska och allergiska djur vid lungexponering för nanopartiklar
pekar på hur viktigt det är att inkludera känsliga individer vid hälsoriskbedömning av
nanomaterial.
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Nanosäkerhet ur ett EU- och OECD perspektiv
Eva Hellsten, tidigare avdelningschef vid EU-kommissionen, vice ordf i OECD WPMN samt
medverkan i svenska Nanoutredningen
EU-kommissionen publicerade 2004 ”Towards an European Strategy for Nanotechnology” som
framhöll nödvändigheten av att studera nanomaterialens miljö- och hälsoeffekter i samband
med planerat utökat forsknings- och innovationsstöd till nanoteknologi via EU:s
ramforskningsprogram. Året efter publicerades ”EU Action plan for Nanotechnology 20052009” som listar ett stort antal åtgärder inom forskning, etik, lagstiftning, internationellt
samarbete etc. som EU-kommissionen och medlemsstaterna borde genomföra för att säkerställa
en ”säker, integrerad och ansvarsfull” utveckling av nanoteknologi. Aktionsplanen
slutredovisades av EU-kommissionen i en skrivelse 2009.
Forskningssatsningarna har ökat radikalt i EU:s ramprogram sedan år 2000. Genom sjunde
ramprogrammet har totalt flera miljarder Euro satsats. Vad gäller miljö- och hälsoforskningen
ligger budgeten på cirka 25 miljoner Euro per år. Detta utgör mellan 5 och 10 % av den totala
nano-budgeten (uppgifter från kommissionen varierar beroende på hur den totala nanoforskningen definieras inom ramprogrammen).
Nano-området kräver långtgående samordning mellan olika politikområden. Inom EUkommissionen skapades en horisontell grupp med representanter från forskning, industri,
miljö, konsumenthälsa och arbetsmiljö tidigt i processen för att koordinera åtgärder av olika
slag, främst samordna lagstiftning med satsningar på forskning och kunskapsuppbyggnad om
nano-säkerhet. Ett flertal medlemsländer har skapat statliga tvärgående organisationer för att
säkerställa samordning på nationell nivå, inte minst när det gäller landets ställningstagande i
olika nano-relevanta EU-frågor.
EU-kommissionen har genomfört två översyner av lagstiftningen för nano-säkerhet (2008
respektive 2012). Kemikalielagstiftningen, Reach, är den viktigaste lagen för att se till att
grundläggande information om potentiella risker tas fram. Dock krävs en anpassning av de
metoder som används för ”vanliga” kemikalier till nanomaterial. Arbete med detta har därför
pågått sedan ett antal år inom EU och internationellt. I arbetet har OECD en nyckelroll genom
att länder världen över här enas om internationella standarder för testning inom kemikalieområdet. År 2006 etablerades OECD Working Party on Manufactured Nanomaterial (OECDWPMN) i vilket EU:s medlemsstater, EU-kommissionen, USA, Kanada, Australien, Japan,
Korea m.fl. arbetar tillsammans för att se över hur testmetoder för kemikalier ska anpassas till
att gälla även för nanomaterial. Även om lagstiftningen kan skilja sig åt mellan olika delar av
världen bör de underliggande vetenskapliga testerna av farlighet inte ge olika resultat
beroende på var i världen de är utförda.
Det svenska regeringsuppdraget att utveckla nationell handlingsplan för nanosäkerhet samt
säkerställa en god samordning av det nationella arbetet med nanosäkerhet redovisades i
oktober 2013 (SOU 2013:70, www.regeringen.se ). Rapporten ger en bred översikt av
området. Huvudförslagen innebär en ökad satsning på forskning om säkerhetsaspekter, ökade
insatser inom EU och OECD samt en samordning av svenska myndigheter, forskare och
intressenter genom bildandet av ett Nanoråd med ett operativt sekretariat, Nanocentrum.
13
Update on NM projects for the method, guidance and testing
strategy development in the OECD Test Guidelines programme
Jukka Ahtiainen, Senior researcher, OECD secretariat
Historically the test guidelines have been developed and validated to be used for hazard
identification and risk assessment of various chemicals. But are these test guidelines
applicable for the regulatory testing of nanoforms of a substances or chemicals? In principle,
most of the existing "endpoints" or more precisely measurement variables are applicable.
However, the dosage of the test material and the characterization of the exposure need
specific guidance in order to gain regulatory relevant data. The Guidance on sample
preparation and dosimetry has been developed in the OECD (OECD 2012). As testing has to
be adapted, and in a worst case for testing of each type of nanomaterial, will this challenge
also the principle of MAD?
The test conditions e.g. organic matter content during the test will affect the form and
bioavailability of the nanomaterial, and detailed guidance is needed on the test conditions,
Also, the conditions during the test should be documented carefully in order to achieve
comparable and understandable results. The additional guidance for testing of nanomaterials
with OECD test guidelines shall be seen as a refinement of the methods. This approach
enables proper use of test guidelines and production of good quality data under the MAD
principle for the regulatory purposes.
The way forward
Currently the harmonized guidance on testing the fate and effects of nanomaterials is being
developed at various stages. New harmonized and validated OECD test guidelines are needed
especially for the physical-chemical characterization (e.g. size, shape, surface chemistry and
charge, zeta-potential) as well as for physic-chemical interactions (dissolution of ions and
dispersion stability) of nanomaterials. This basic knowledge of the nanomaterial under
assessment is crucial to guide further (eco)toxicity and fate testing – which tests in which
compartment are relevant, but also to better understand and interpret the results. The basic
reactions and transformations of the nanomaterial for example in aquatic media should be
tested at various conditions e.g. pH, ion strength and organic matter content.
Several OECD guidance documents are under development e.g. for aquatic toxicity testing
(http://www.oecd.org/env/ehs/nanosafety). These comprise of decision frameworks based
on the basic physical-chemical properties of the nanomaterial to be tested. One specific
example in the environment fate testing is to develop technical guidance for dietary
bioaccumulation of nanomaterials in fish using the OECD 305 test guideline.
There are regulatory needs for harmonized guidance for testing the effects and fate of
nanomaterials, however at the same time OECD is also developing guidance for grouping and
categorization of nanomaterials. The grouping of nanomaterials will be based on their
chemical composition including coatings, size, shape, basic physical-chemical reactions and
biological effects. This will hopefully help to understand some basic mechanisms of the
effects and provide possibilities for read-across interpretation between some nanomaterials.
References
OECD 2012 Guidance on sample preparation and dosimetry for the safety testing of manufactured
nanomaterials. Series on the Safety of Manufactured Nanomaterials No. 36, OECD Paris, France
14
Nytt om nano från Kemikalieinspektionen
Elin Simonsson, Kemikalieinspektionen
Ändringar av bilagorna till EU:s kemikalieförordning Reach har planerats på att genomföras
sedan 2013. Ändringarna är tänkta att anpassa informationskraven i bilagorna till nanomaterials speciella egenskaper. Arbetet har blivit ytterligare försenat och tidigaste tidpunkten
för ett förslag från Europeiska kommissionen är nu satt till sommaren 2015. Diskussioner förs
i en av Europeiska kommissionens expertgrupper (CASG Nano).
En annan viktig fråga är Europeiska kommissionens rekommenderade definition som är
avsedd att användas i lagstiftningssammanhang. Under 2014 har en översyn av definitionen
pågått som förväntas bli färdig under 2015. Definitionen ska särskilt ses över med hänsyn till
kravet på att minst 50 % av partiklarna ska vara inom nanoskalan (1-100 nanometer). Översynen är uppdelad i en vetenskaplig del och en policy del. Den vetenskapliga delen består av
tre rapporter från Europeiska kommissionens vetenskapliga gren (JRC). Policydelen kommer
bl.a. bestå av ett offentligt samråd där det kommer att vara möjligt lämna synpunkter på hur
definitionen bör se ut.
Den tredje frågan som är aktuell i EU just nu är Europeiska kommissionens konsekvensutredning om åtgärder för ökad transparens på marknaden. Den åtgärd som främst diskuteras är
att eventuellt införa ett EU-register för nanomaterial. Kemikalieinspektionen har svarat på ett
offentligt samråd i frågan. Konsekvensutredningen färdigställs under 2015 varpå Europeiska
kommissionen kommer att avgöra om och vilka åtgärder de tänker vidta.
Nanomaterial i kosmetiska produkter – vad har hänt sedan de nya
reglerna infördes?
Tomas Byström, Läkemedelsverket
Reglerna för kosmetiska produkter har ändrats från att vara EU-direktiv som implementerades
nationellt till att vara en direkt gällande EU-förordning. Förordning (EG) nr 1223/2009 om
kosmetiska produkter trädde i kraft i slutet av 2009, men den tillämpades fullt ut först den 11
juli 2013.
Bland de tydligaste nyheterna i EU-förordningen var särskilda regler för nanomaterial i
kosmetiska produkter. Nanomaterial definieras som: ett olösligt eller biopersistent material
som är avsiktligt tillverkat, med en eller fler yttre dimensioner, eller en inre struktur, med ett
spann på mellan 1 och 100 nm.
De regler som gäller för nanomaterial i kosmetiska produkter är:
·
·
Varje användning av nanomaterial måste anmälas till EU-kommissionen 6 månader
före produkter släpps ut på marknaden
EU-kommissionen avgör från fall till fall om den anmälda användningen behöver
granskas av en vetenskaplig kommitté
Vissa ämneskategorier förhandsgranskade innan tillåtande (UV-filter, färgämnen,
konserveringsmedel)
Övriga nanomaterial granskas specifikt för den anmälda användningen, anmälan
ska innehålla uppgifter om nanomaterialets egenskaper
·
Innehåll av nanomaterial måste framgå av innehållsförteckningen på förpackningen
Exempel: Titanium Dioxide (nano)
15
Sedan reglerna för nanomaterial började tillämpas fullt ut har det inom EU anmälts cirka
25000 kosmetiska produkter med nanomaterial. Vanligaste kategorierna är solskyddsprodukter (7300 st.), sminkprodukter (7000 st.) och ansiktsprodukter (2700 st.). De anmälda
ansiktsprodukterna är av allt att döma dagkrämer som innehåller UV-filter.
Från tillverkare/importörer Sverige är cirka 150 produkter anmälda, varav 80 st. sminkprodukter och 60 st. solskyddsprodukter. Läkemedelsverket kan dock konstatera att flera aktörer
som sätter solskyddsprodukter på marknaden inte anmält dessa på rätt sätt.
Delar av det regulatoriska arbetet med nanomaterial har dock inte varit i tid. Rent formellt är
de vanligaste nanomaterialen (Titanium dioxide, CI 77266) ännu inte tillåtna i nanoform.
Detta delvis på grund av oklara definitioner av produkttyper som innebär inhalationsrisk
(”spayprodukter”). EU-kommissionen skulle senast den 11 januari 2014 ha publicerat en
katalog över de nanomaterial som används. Denna publicering har dock dröjt på grund av
felaktiga anmälningar från företagen. Katalogen är sammanställd, och ska publiceras så snart
den är översatt.
Danske aktiviteter vedr. nanomaterialer
Flemming Ingerslev, The Danish Environmental Protection Agency
Nanomaterialer (NM) indgik i den danske kemikaliehandlingsplan fra 2010-2013, hvor fokus
især var på at skabe overblik viden over de vigtigste nanomaterialer 4 samt at bidrage til EUarbejde med nanomaterialer 5. I den opfølgende kemikalieindsats (2014-2017) er der fokus på
at sikre, at dansk viden om nanoteknologi bidrager til udviklingen af en fælles EU-løsning
(http://kemikalieindsatsen.dk).
Den ny danske regering vedtog i 2011 initiativet ”bedre styr på nano”, som har til formål at
skabe øget klarhed over eksponeringsveje og konsekvenserne for forbrugere og miljø ved
anvendelse af nanomaterialer. Den styrkede indsats på nanoområdet omfatter blandt andet
udvikling af et nanoproduktregister. Der er i årene 2012-2015 afsat 6 millioner danske kroner
til indsatsen. Samlet skal initiativet skabe overblik over situationen med hensyn miljø- og
forbrugersikkerhed i forhold til nanomaterialer i Danmark. Initiativet omfatter således
projekter som skal 1) skabe overblik over eksisterende viden vedr. nanomaterialers
sundhedsegenskaber og deres miljøegenskaber, 2) bidrage til ny viden om nanomaterialers
hudgennemtrængelighed og nanomaterialers opløselighedshastighed i miljøet, 3) undersøge
udbredelsen, anvendelse og risici af konkrete nanoprodukter, samt 4) skabe generelt overblik
over anvendelsen af nanomaterialer i Danmark.
Et vigtigt element til at skabe overblik over nanomaterialer er det danske nanoproduktregister,
som blev oprettet i juni 2014 i forbindelse med offentliggørelse af bekendtgørelsen om det
danske nanoproduktregister (BEK nr 644 af 13/06/2014). Denne bekendtgørelse beskriver den
registreringspligt som gælder alle virksomheder der sætter nanoprodukter på
forbrugermarkedet i Danmark. Bekendtgørelsen omfatter således både producenter og
importører af nanoprodukter. Nanoprodukter defineres som udgangspunkt som artikler eller
kemiske blandinger, der indeholder nanomaterialer (jf. EU’s definition) og som frigiver disse
nanomaterialer. Bekendtgørelsen undtager dog produkter, som reguleres under reglerne for
fødevarer, fødevarekontaktmaterialer, medicinsk udstyr, kosmetik, pesticider og affald.
4
The Danish Environmental Protection Agency. Survey on basic knowledge about exposure and potential
environmental and health risks for selected nanomaterials. Environmental Project 1370, 2011.
5
The Danish Environmental Protection Agency. Information Requirements for nanomaterials – IRNANO.
Environmental Project 1469, 2013.
16
Endvidere undtager den produkter, hvor nanomaterialerne ikke bevidst er fremstillet i
nanostørrelse og endelig undtager bekendtgørelsen visse konkrete produkttyper (bl.a. maling
og gummiprodukter) med indhold af titandioksid, carbon black og silicium dioksid. Første
indberetningsår slutter d. 30. august 2015.
I Nanoproduktregisteret er der dels nogle tvungne informationskrav og dels nogle frivillige
krav. De tvungne krav omfatter information om den indberettende virksomhed, om produktet,
om nanomaterialets kemiske sammensætning og om det er registreret under REACH.
Endvidere er det frivilligt at indberette en række oplysninger om selve nanomaterialet (blandt
andet partikel størrelse, størrelsesfordeling, oplysninger om aggregering m.m.). De frivillige
oplysninger svarer til dem, som OECD har vedtaget harmoniserede standard formater for
(OHTs). Som hjælp til de virksomheder der skal indberette, har Miljøstyrelsen udarbejdet
vejledninger på dansk og engelsk 6 7. Der er endvidere oprettet en help-desk og en FAQ-side,
hvor virksomheder kan få svar på spørgsmål (http://mst.dk/virksomhedmyndighed/kemikalier/miljoestyrelsens-nanoindsats/).
Nanocellulose and NANoREG- project
Finnish Safety and Chemicals Agency
The safety of nanomaterials is investigated in the EU’s just-beginning extensive NANoREG
project as a joint effort by the authorities and the industry. Finland’s goal in the project is to
continue to study the safety of microfibrillar and nanofibrillar cellulose. To date, only a little
is known about the health and environmental hazard impacts of nanoparticles, although the
industrial use of nanomaterials has increased rapidly. For example, they are already common
in sports equipment, sunscreens and other cosmetics products.
Nanotechnology is a technique for building nanometre-scale structures (one nanometre is one
millionth of a millimetre). Interest in the use of nanoparticles is great, as they can be used to
improve product characteristics; for example, nanoparticles help make a coat of paint more
scratch-resistant. On the other hand, the safety of nanoparticles raises questions.
It is difficult to identify risks, as matter can have an unknown behaviour in nanoscale. The
effect on cellular level can also vary according to what impurities have attached to the
nanoparticle or which substance has been purposefully used to coat it.
The industrial use of nanomaterials has stirred up animated discussions in the EU. The
REACH Regulation regulating the registration, evaluation, authorisation and restriction of
chemicals plays a central role in legislation. The idea of safety-promoting dialogue between
the industry and the authorities has come up to complement the idea of just using regulatory
measures. This forms the basis of the NANoREG project to be implemented in the years
2013 to 2017.
Authorities, research institutes and companies and consortiums from fourteen European
countries will participate in NANoREG. The project aims at developing guidelines for safe
usage risk management and safety instructions, while assessing the need for new legislation.
6
Miljøstyrelsen. Vejledning om indberetning til det danske nanoproduktregister. Vejledning fra Miljøstyrelsen
nr. 5, 2014.
7
The Danish Environmental Protection Agency. Guideline for the Danish Inventory of Nanoproducts. Guidance
from the Danish Environmental Protection Agency No. 5, 2014
17
A stable and safe operating environment is a benefit that is shared by both the authorities and
the industry. It is necessary for new applications and innovations, and the resulting
investments, to be possible in the first place.
In Finland, participation in the NANoREG project is coordinated by the Finnish Safety and
Chemicals Agency (Tukes) that is also responsible for oversight and guidance concerning the
REACH regulation. Also included in the project are the Finnish Institute of Occupational
Health, responsible for the experimental in vitro and in vivo studies, and Stora Enso and UPM
as a joint Nordic Cellulosa consortium.
Finland’s national research portion of the Nanoreg project focuses on microfibrillar and
nanofibrillar cellulose materials that have several potential industrial applications in different
products. Microcellulose and nanocellulose comprise wood fibre and fibre bundles originating
from wood cellulose. The intention is to study the safety of biodegradable microcellulose and
nanocellulose experimentally by means of biological testing. This is important in order for it
to be possible to use nanocellulose, for example, as raw material for cosmetics, food additives
or packaging.
In the responsible development of products manufactured from micro and nanocellulose,
research plays a key role. Research works aim to find and confirm potential benefits in
microcellulose and assess the safety measures required for manufacturing and utilization of
the materials. For this reason, the Nordic Cellulosa consortium acts in close cooperation with
other fields and authorities, and promotes research in safe applications of microfibrillar and
nanofibrillar cellulose.
18
2
Bilagor
Bilaga 1. Deltagarlista
Bioforsk
Joner Erik
Danmarks Tekniske Universitet
Bloch Hartmann Nanna
Cupi Denisa
Europeiska kommissionen
Hellsten Eva
Finnish Institute of Occupational Health
Stockmann Juvala Helene
Hyytinen Eija-Riitta
Einola Juha
Finnish Safety and Chemicals Agency
Palomäki Jaana
Bucht Anders
FOI
Ekstrand-Hammarström Barbro
Gustafsson Åsa
Formas
Vikström Anna
Försvarets Materielverk
Henningsson Kenth
Ramfjord Birgit
Westlund Robert
Försvarsmakten
Jalalian Nazli
Generalläkaren
Duf Jessica
Kängström Marianne
Andersson Yvonne
Kemikalieinspektionen
Anfält Lisa
Gellerstedt Therese
Hellmér Lena
Moore Gregory
Simonsson Elin
Wendt - Rasch Lina
Virefjord Tania
Kommerskollegium
Housset Cedric
KTH
Lazarevic David
Livsmedelsverket
Pihlström Tuija
Svensson Kettil
Löndahl Jakob
Lund University
Nilsson Annika
Bohgard Mats
19
Byström Tomas
Läkemedelsverket
Hillgren Anna
Salin Kia
Hedlund Britta
Naturvårdsverket
Mattsson Cecilia
Norwegian Institute for water research
Macken Ailbhe
Norwegian Environment Agency
Andersen Sjur
Gudbrandsen Marius
OECD Secretariate
Ahtiainen Jukka
Regeringskansliet
Tapper Sofia
SINTEF Materials and Chemistry
Roman Netzer
Socialstyrelsen
Domeij Helena
The Danish Environmental Protection Agency
Zenner Boisen Anne Mette
Ingerslev Flemming
Bengtsson Malena
Trafikverket
Reuithe Anna
20
Bilaga 2. Föreläsarnas power-point presentationer
21
Nanomaterials in a Life Cycle
Perspective
David Lazarevic
[email protected]
Division of Industrial Ecology
Department of Sustainable Development, Environmental Science &
Engineering
KTH – Royal Institute of Technology
•
National action plan for the safe use and handling of
nanomaterials (2013)
•
As part of this action plan:
• Lazarevic and Finnveden, 2013. Life cycle aspects of
nanomaterials. Environmental Strategies Research,
KTH, Stockholm. (In English)
• Finnveden and Lazarevic, 2013. Livscykelaspekter och
nanomaterial. Avdelningen för miljöstrategisk analys,
KTH, Stockholm. (In Swedish)
•
Liljenström, C., Lazarevic, D., & Finnveden, G. 2013.
Silicon-based nanomaterials in a life-cycle perspective:
including a case study on self-cleaning coatings. KTH,
Stockholm.
2
Benefits and Risks of Nanomaterials
“To some it represents the miracle cure for all that ails us. To others,
it could be the end of the world as we know it” (Maynard, 2010).
Potential Benefits
• Expected significant impact on virtually all industrial
sectors including healthcare, agrifood, transport, energy,
materials, and ICT.
• Potential areas
• Monitoring
• Remediation and pollution
• Resource saving (energy)
3
Benefits and Risks of Nanomaterials
Potential impacts
• Toxicological risks to humans and the environment
• Increase in the extraction of raw materials
• Increase energy use during production phases
• Higher material requirement (better specifications)
• Increased waste production during production (hazardous
waste)
• End-of-life: what happens to waste products?
4
Nanomaterials in a Life cycle perspective
The general consensus among scientists, researchers, and
regulatory agencies is that the potential health and
environmental risks of engineered nanomaterials (ENMs)
should be evaluated over their entire life-cycle (Grieger et al.
2012).
•
•
Life cycle Assessment
Risk Assessment
5
Life Cycle Assessment
•
A Tool for assessing the
potential environmental
impacts associated with a
product/service system by:
o compiling an inventory of
relevant inputs and
outputs
o evaluating potential
impacts associated with
inputs and outputs
o interpreting results of
inventory and impact
assessment
•
LCA follows a cradle-tograve approach
(Lazarevic & Finnveden, 2013)
6
Resources
and materials
Energy
Components and
semi -products
Manufacture
Use and
maintenance
Recycling
and disposal
Raw
materials
Process
Waste
Products
Emissions
LCA Phases
• Goal and Scope Definition
• Life Cycle Inventory (LCI)
• Life Cycle Impact
Assessment (LCIA)
• Interpretation
Hazardous waste
Waste (volume)
Land use
Exotoxicity
Human toxicity
Eutrphoication
Photochemical ozone formation
Acidification
Global Warming
B
A
0
20
40
60
80
100 120
7
Life Cycle Assessment
•
Enables the study and comparison of different options to
supply a given function
•
Enables the identification of environmental ‘hotspots’
throughout the product/service life cycle
•
LCA attempts to be comprehensive with respect to the
environmental interventions and environmental issues
considered.
8
•
LCA’s focus on the
product/service system
and RA’s focus on the
emissions of a single
substance
•
The results of LCA are
comparative
whereas
the results of RA are
absolute
•
LCA covers a range of
environmental impacts
whereas RA primarily
cover toxicological and
(eco)toxicological
impacts.
(Grieger et al. 2012)
9
Raw
Materials
Manufacturing
Use
E-O-L
Babaizadeh and Hassan (2013): Comparison of TiO2 coated class with float glass
Bauer et al. (2008)
Greijer et al. (2011) Nanocrystaline dye sensitized solar cell (from nano-TiO2 and carbon black)
Griffiths and O’Byrne (2013) MWCNT formation via catalytic chemical vapour deposition.
Grubb and Bakshi (2011a, 2011b) Life cycle energy use of nano TiO2 production and e.g. steel,
aluminium, polysilicon production
Isaacs et al. (2010) Production of single wall carbon nanotubes
Joshi (2008) Comparison of nanoclay composite biopolymer with biobased polymers
Khanna et al. (2008) & Khanna and Bakshi (2009) polymer nanocomposites compared to steel,
aluminum and PP
Kushnir and Sandén (2008) Energy requirements for fullerene and nanotube synthesis
Lloyd and Lave (2003) Clay polypropylene ENM's instead of steel or aluminum in vehicle body panels
Lloyd et al. (2005) Nanofabrication technique of platinum group metal (PGM) particles in automotive
catalysts
Merugula et al. (2010) Comparison of Glass fibre reinforced plastics an vapour grown CNts for wind
turbine blades
Meyer et al. (2011) Socks with and without Ag ENM's
Moign et al. (2010) Manufacturing of yttria-stabilized-zirconia ENM coating
Osterwalder et al. (2006) Energy comparison of wet and dry synthesis methods for oxide nanoparticle
production
Roes et al. (2007) Comparison of polypropylene nanocomposite with conventional polypropylene
Şengül and Theis (2011) LCA of quantum dot photovoltaic (QDPV) module compared to silicon and thin
film PV’s
Singh et al. (2008) SWCNT production
Walser et al. (2011) Comparison of nanosilver t-shirts and conventional t-shirts
Environmental impact categories
Toxicological impact categories (non-nanoparticle release based)
10
Energy use
Nanoparticle Toxicity impact (nanoparticle release based)
Nanomaterials in a Life cycle perspective
Incineration &
Landfill
Environmental Burden
•
Potentially energy intensive raw
material extraction, processing, and
nanomanufacturing processes
•
Potential depletion of non-renewable
resources
•
Possible release of toxic emissions
& unconventional liquid streams
Extraction of
Raw Materials
Recovery
Recycling
Release of ENMs
to the
Environment
End of life
Reuse
Production
Release of
ENMs to the
Environment
Distribution
Use
Missing Information
•
Impacts of raw material use on
supply chains of other products
•
Potential release, fate and transport
of ENMs
•
Performance of ENM products
•
Amount & characterisation of waste
streams
Uncertainties
•
Waste & emissions from ENM
production
•
ENM exposure & toxicity
•
Preferred method of ENM disposal
(Lazarevic & Finnveden, 2013)
Engineered nanomaterial
Product life cycle
11
Current use of LCA: Cradle to Gate
CED for CNFs compared to aluminium, steel and polypropylene
(Reproduced from Khanna et al. (2008))
12
Current use of LCA: Cradle to Gate
CED of polymer nanocomposites that provide equal
stiffness to a steel component
(Reproduced from Khanna and Bakshi (2009))
13
Current use of LCA: Cradle to Use
as part of a
car, driving
280000 km
(Reproduced from Hischier and Walser (2012))
14
Current limitations of Applying LCA to
nanomaterials
CNF emissions and impacts not included
Khanna et al. (2007)
15
Life Cycle Assessment of Nanomaterials
Goal and Scope Definition
• ENMs may have specific functions and material properties
that provide additional gains when used as a substitute for
traditional materials
Life Cycle Inventory
• Lack ENM specific data related to the outputs of the
processes
o Data on Input side, no data on output side
o It is important to know if ENMs change their form during
their life cycle, due to aging and other influences such as
weather, mechanical stress/pressure, etc.
• LCI databases do not distinguish between bulk materials and
ENMs
• Populating LCI databases with ENM specific information,
such as size and shape, is of critical importance
16
Life Cycle Assessment of Nanomaterials
Life Cycle Impact Assessment
•
•
•
•
ENMs may exhibit unconventional behaviour, leading to
unexpected fate, transport, and toxicity mechanisms in human
and ecological systems
A complete lack of characterization factors for release of
nanoparticles indoors and outdoors
LCIA methods such as CML 2001, Eco-Indicator 1999 or Impact
2002 do not cover toxicological effects of nanoparticles
The current understanding of effect mechanisms, dose-response
relationships, as well as transport and transformations in the
environment may not be sufficient to ascertain a representative
characterization of ENMs.
17
Conclusions
•
•
•
•
LCA of nanomaterials is still in the very early stages
Consideration of the impacts of ENMs in LCA is currently
inadequate, due to:
– Lack of data on nanomaterial production
– Lack of data and uncertainties on what should be
included in the LCI
– Uncertainties on how to assess the impacts of ENMs
(LCIA)
Need for cooperation between the LCA community and the
nanotoxicology community
LCA needs to be complemented by RA to assess risk
related to specific life cycle stages
18
References
Greijer, H., Karlson, L., Lindquist, S.E., Hagfeldt, A., 2001.
Environmental aspects of electricity generation from a nanocrystalline
dye sensitized solar cell system. Renewable Energy 23, 27–39.
Hischier, R., Walser, T., 2012. Life cycle assessment of engineered
nanomaterials: State of the art and strategies to overcome existing
gaps. Science of The Total Environment 425, 271–282.
Khanna, V., Bakshi, B.R., 2009. Carbon nanofiber polymer composites:
evaluation of life cycle energy use. Environmental Science &
Technology 43, 2078–2084.
Khanna, V., Bakshi, B.R., Lee, L.J., 2007. Life cycle energy analysis and
environmental life cycle assessment of carbon nanofibers production,
in: Electronics & the Environment, Proceedings of the 2007 IEEE
International Symposium On. IEEE, pp. 128–133.
Khanna, V., Bakshi, B.R., Lee, L.J., 2008. Carbon nanofiber production:
Life Cycle Energy Consumption and Environmental Impact. Journal of
Industrial Ecology 12, 394–410.
Lazarevic, D. and Finnveden, G., 2013. Life cycle aspects of
nanomaterials. Environmental Strategies Research, KTH, Stockholm.
19
Dermal absorption of
nanomaterials
Projects from
the Danish EPA
Anne Mette Boisen
Chemicals Unit,
Danish Environmental
Protection Agency (EPA)
Presentation outline
 Introduction to the Danish Nano Initiative
Better Control of Nano
 Dermal absorption of nanomaterials – Literary project
(conducted by IOM from UK)
 Dermal absorption of nanomaterials – Eksperimental
project
(conducted by Aarhus University from DK)
Better control of Nano initiative
(24 mio. DKK in 2012-15):
Dermal absorption –Literary project

Danish EPA Environmental Project No. 1504, 2013
Report and database
Scientific assessment of available studies
on dermal absorption
Aims
• assessment of the extent of absorption of
nanomaterials;
• identification of nano-specific characteristics that may
influence the absorption of nanomaterials;
• evaluation of which test method(s) would most closely
simulate the transport of nanomaterials through
human skin
• candidates for testing and assessment of the specific
research areas that require more knowledge.
Search strategy
Assessment of reliability and relevance of studies
-Klimisch criteria and NM characterization (Card and
Magnuson 2010)
Results and recommendations
 Absorption of NM into systemic circulation:
Possible (small fraction) few robust, well performed studies, no
clear guidelines (hard to compare/interpret)
 Parameters:
Size, surface chemistry (coating), shape
Results and recommendations
 Test methods:
Harmonised testing approaches (specific technical guidance
given in the report). Human golden standard (but ethics and
cost)
 Candidates for testing:
Systematic test of priority candidate properties (size,
surface chemistry)
 Research gaps:
Effect of flexed skin, follicular penetration, children,
extending the study period
Dermal absorption –Experimental project
Project manager Christiane Beer, Aarhus University
Started spring 2014 ends summer 2015
Overview
• Candidates for testing:
Nanomaterials used in sunscreen:
Titanium dioxide and Zinc Oxide
• Investigation of characteristics:
Size and coating
• Test methods:
In vitro Epiderm model
In vivo mouse inflammation model
In vivo mouse xenograft model
Candidates for testing

Titanium Dioxide
Size:
(TiO2, rutile, high purity 99.9%, 30 nm)
(TiO2, rutile, high purity 99.9%, 100 nm)
Coating:
(TiO2, rutile, 30 nm) coated: Silicone Oil (hydrophobe)
(TiO2, rutile, 30 nm) coated: Silicon and Aluminium (hydrophile)
UV-Titan M 161: TiO2 nanoparticles, rutile, 17 nm, coated: Alumina, stearic acid
UV-Titan M 262: TiO2 nanoparticles, rutile, ~20 nm, coated: Alumina, silicone

Zinc Oxide:
Coating: (ZnO, high purity 99.95%, 18 nm, uncoated)
(ZnO, 99+%, 20 nm, coated: 1 wt% Silane Coupling Agent)
(hydrophile)
Vehicle
Wt%
61.90
%
0.20
%
0.10
%
1.00
%
2.00
%
2.00
%
10.00
%
2.80
%
4.00
%
3.00
%
3.00
%
10.00
%
Trade name
INCI
Phase
Water
1
Aqua
Keltrol AP
1
Xanthan Gum
Dermofeel PA-3
1
Sodium Phytate, Aqua, Alcohol
Verstatil PC
1
Phenoxyethanol, Caprylyl Glycol
Propylen Glycol
1
Propylene Glycol
Glycerin
1
Glycerin
Waglinol AB
1215
2
C12-C15 Alkyl Benzoate
Parsol 340
2
Octocrylene
Parsol MCX
2
Ethylhexyl Methoxycinnamate
Parsol 1789
2
Butyl Methoxydibenzoylmethane
Mulsifan RT 11
2
Ceteareth-22
Soldoc EB 29
2
Isostearyle Isostearate
C12-C15 Alkyl Benzoate is used
to pre-disperse nanoparticles
before adding them
to the sun cream.
2.5 wt% NP
In vitro model
in vitro EpiDerm™ system (MatTek)
3D in vitro skin model
Analyses
•
•
•
•
•
•
TEER (trans epithelial electric resistance)
electron microscopy (TEM)
Histology
skin corrosion/irritation tests (MTT assay)
ICP-MS
Cytokine analysis of medium (11 cytokines by
multiplex flow cytometer assay)
Set-up
Each sample will be treated with 30 μl of the test substances (All 8 NP are
tested).

Day 1 – Arrival of the cells

Day 2 – TEER measurement
Exposure to sunscreen +/- nanoparticles and untreated control for 20 hours

Day 3 – Washing of the EpiDerm models (Medium samples for ICP-MS and
cytokine analysis)

Day 5 – Medium exchange (Medium samples for cytokine analysis)

Day 7 – TEER measurement
Sample preparation for EM and histology
Medium samples for cytokine analysis
MTT assay skin irritation/skin corrosion
In vivo Inflammation model
 Acute irritant contact dermatitis model
TPA:
12-O-tetradecanoylphorbol-13 acetate
Induced:
Erythema
Oedema
Scaling
Epidermal hyperplasia
Infiltrates of
monocytes
lymphocytes
neutrophils
IL-1b, TNFa, IL-6
0 hours
4 hours
8 hours
24 hours
Set-up
A = Untreated (no sun spray, no nanoparticles)
B = Control sun screen
C = NP1 : (TiO2, rutile, high purity 99.9%, 30 nm)
D = NP2 : (TiO2, rutile, high purity 99.9%, 100 nm)
E = NP3 : (TiO2, rutile, 30 nm coated with Silicone Oil )
F = NP4 : (TiO2, rutile, 30 nm Coated with Silicon and Aluminium )
Set-up
Day
1
2
3
4
8
TPA
treatment
X
X
X
Sun screen
application
X
X
X
Ear thickness
Ear thickness
Ear thickness
Ear thickness
Ear thickness
EM, histology and ICP-MS sample
preparation
Storage blood and organs
10 µl sun screen per cm2
2 x 3 mm biopsies are taken from each ear for TEM and
histology; leftover from the ear is frozen and stored for ICP-MS
In vivo
xenograft transplantation model
Human skin
Mouse skin
Transplantation
Engraftment
In vivo xenograft model

Vehicle and NP1-4 are applied on day 1, 2, and 3

(10 µl sun screen per cm2).

On day 8, mice are killed (4 days post last treatment with
NPs):

A = Untreated (no sun screen, no nanoparticles)

B = vehicle control sun screen no NPs

C = NP1 : (TiO2, rutile, high purity 99.9%, 30 nm)

D = NP2 : (TiO2, rutile, high purity 99.9%, 100 nm)

E = NP3 : (TiO2, rutile, 30 nm coated with Silicone Oil )

F = NP4 : (TiO2, rutile, 30 nm coated with Silicon and Aluminium )
Analyses
• ICP-MS
• TEM
• histology
Expected Outcome
Experimental project
• Project report (summer 2015)
• Articles in peer reviewed scientific journals
(summer 2015)
Acknowledgements
Literary project

Dr. Craig Poland, Steve Hankin, Craig Poland, Sheona Read, Julia Varet,
Gillian Carse, Steven M. Hankin, IOM, United Kingdom,

Frans M. Christensen, COWI Denmark.

Reference group: Maxine McCall, CSIRO, Australia,

Jesper Bo Nielsen, Southern University of Denmark.
Experimental project

Christiane Beer, Herman Autrup , Karin Stenderup, Lars Iversen, Duncan
Sutherland, Jing Wang, Jens Randel Nyengaard, Torben Sigsgaard, Jakob
Bønløkke, Aarhus University, Denmark.

Reference group: Craig Poland, Steve Hankin, IOM, United Kingdom.
Uptake of TiO2-nanoparticles in lung epithelial cells
ER
TiO2-nanoparticles
Mitochondria
TiO2-nanoparticles
30nm
Nucleus
Interactions of nanoparticles with organs
protected by internal biological barriers
Anders Bucht, Swedish Defence research Agency
Division of CBRN Defence and Security
Raman imaging measures polymorph and size specific NP
uptake and distribution in living lung epithelial cells
a)
P25/R9
-10
y /m
-5
0
10
2200
P25
w vib=144
150
cm-1
0
100
5
50
c)
y /m
-5
-5
0
5
10
R9
w vib=448 cm-1
b) Raman mapping performed within
the rectangular area visualized in
(a) of P25 anatase characterized
Leakage into
nucleus region
by the Eg vibrational mode at 145
cm-1.
0
50
440
30
0
220
5
c) Raman mapping of rutile
characterized by the Eg vibrational
mode at 450 cm-1.
Small, soft NP agglomerates (such as anatase
”P25”) penetrates membranes more easily
than hard agglommerates (such as rutile)
10
-5
0
5
Ekstrand-Hammarström B, Akfur CM, Andersson PO, Lejon C, Österlund L and Bucht A. Human primary
bronchial epithelial cells respond differently to titanium dioxide nanoparticles than the lung epithelial cell
lines A549 and BEAS-2B. Nanotoxicology 2011.
TiO2 induce oxidative stress and
pro-inflammatory response in lung epithelial cells
a) Optical micrograph of A549 cell
exposed to 50/50 wt% mixture of
P25/R9 TiO2 nanoparticles.
10
-10
120.000 x magnfication
P25 NP ontop of nucleus
0
b)
Nucleus
80.000 x magnification
10
x /m
Andersson PO, Lejon C, Ekström Hammarström B, Akfur C, Bucht A and Österlund L. Polymorph- and
size-dependent uptake and toxicity of TiO2 nanoparticles in living lung epithelial cells. Small 2011
Changed properties of TiO2 NPs in serum
Intracellular Superoxide production
IL-8
[pg ml-1]
MCP-1
[pg ml-1]
control
150  4
1254  19
A14
256  20*
1209  133
A60
411  59*
1076  48
R5
487  73*
1193  121
R9
424  25*
1414  383
P25
840  126**
2178  131**
Sample
Exposure of A549 cells for 24 h exposure to 50 g
TiO2 nanoparticles.
Andersson PO, Lejon C, Ekström Hammarström B, Akfur C, Bucht A and Österlund L.
Polymorph- and size-dependent uptake and toxicity of TiO2 nanoparticles in living
lung epithelial cells. Small 2011.
From studies of lung cells we know that:
• Nanoparticles can penetrate the cell membrane and
nucleus, and the penetration depends on size and
particle structure.
Z-potencial -24mV
• NPs can induce oxidative stress and inflammatory
response in cells, and this response depends on type of
particle, cell type and exposure conditions.
Analaysis of blood coagulation in vitro
From studies of blood coagulation we know that:
• TiO2 NPs at very low concentrations (50 ng/mL)
induce strong activation of the contact system, which
in this model elicits thromboinflammation.
• this is in line with the finding of components of the
contact system in the protein corona of the TiO2 NPs
after exposure to blood.
Tubing loops coated with heparin. Freshly drawn human blood is added to the loop
together with the test nanoparticles. Rotated at 37°C in a heat cabinet. Aliquots of
blood are removed at different time points during incubation for analysis
Bo Nilsson and colleagues at Uppsala University
Overall conclusions
Remaining issues
• A corona of biomolecules will be formed around the NPs
before meeting the epithelial cell layer.
• Do the adaptive immune system recognize biomolecules
bound to the NPs as foreign bodies, thereby providing a
risk for autoimmune reaction?
• The corona will influence translocation through the
epithelial barrier and the subsequent systemic response.
• In the blood circulation, nanoparticles may have an
impact on coagulation cascade, as well as induce innate
and adaptive immune responses.
Contributors and Collaborators
Swedish Defence Research Agency
The Toxicology team
Barbro Ekstrand-Hammarström
Åsa Gustafsson
Sofia Jonasson
Elisabeth Wigenstam
Linda Elfsmark
Bo Koch
Christine Akfur
Mona Koch
Lina Thors
Lina Ågren
Ulrika Bergström
Raman spectroscopy at FOI
Per-Ola Andersson
Christian Lejon
Linnea Ahlinder
Collaborators
Dep. of Respiratory Medicine and Allergy,
Umeå University
Thomas Sandström and Anders Blomberg
Dep.of Engineering Sciences and
solid phase physics, Uppsala University
Lars Österlund and colleagues
Dep. of Immunology, Genetics and Pathology,
Uppsala University
Bo Nilsson and colleagues
[email protected]
• Do NPs in complex with biomolecules penetrate the
blood-brain-barrier in concentrations that may harm the
brain?
Research on Nanosafety in Three Competence
Centres at Lund University
Measurement Techniques for
Airborne Nanoparticles
METALUND, Medicine and
Technology for Working Life and
Society
JAKOB LÖNDAHL
nmC@LU, Nanometer Structure
Consortium
CAST, Consortium for Aerosol
Science and Technology
The Lund Aerosol Group –
One laboratory, two departments
Ergonomics and Aerosol Technology
Mats Bohgard, Ville Berg, Anders Gudmundsson, Christina Isaxon,
Jonas Jakobsson, Jakob Löndahl, Patrik Nilsson, Erik Nordin, Joakim
Pagels, Jenny Rissler, Christian Svensson, Aneta Wierzbicka, et al.
Nuclear Physics
Erik Swietlicki, Birgitta Svenningsson, Adam Kristensson, Göran
Frank, Emelie Hermansson, Erik Ahlberg, Johan Martinsson, Pontus
Roldin, Moa Sporre, Axel Eriksson, Cerina Wittbom et al.
CAST
Nanoparticles: what should we
measure?
Physical properties
Size
Number / surface area / volume
Surface structure
Electrical charge
Radioactivity
Shape
Avoid measuring
background particles
What we want to
measure is not similar
to what we are able to
measure
Chemical properties
Chemical composition (metals, toxins, …)
Solubility
Biological activity
(viruses)
Nanoparticles: how could we measure?
Particle surface area monitors
They are only a small fraction of the air (by mass)
Nanoparticles smaller than the wavelength of visible light
Examples: miniDiSC, NanoTracer, AeroTrak, NSAM, Partector
Advantages: small, cheap, easy to use, suitable for personal
exposure, hith time resolution (seconds),
lung deposited surface area (??)
Disadvantages: total concentration (10-300 nm), limited precision
Particle size distribution, SMPS (FMPS)
DMA
3E+5
CPC
dN/dlogDp (cm‐3)
Particle surface area monitors
2E+5
Idle engine
Transient driving
1E+5
0E+0
10
100
1000
Dry mobility diameter (nm)
Advantages: precise, high time resolution (~1 min)
Disadvantages: complex, expensive
Particle size distribution, ELPI
Particle mass (and surface area) distribution
APM, aerosol particle mass analyzer
Advantages: high time resolution
(seconds), some size information,
possibility to analyse particles afterwards
Disadvantages: complicated, not ideal for
nanoparticles, lower precision than SMPS
Particle mass (and surface area) distribution
APM, aerosol particle mass analyzer (10-18 grams)
Effective density [gcm‐³]
DMA
APM
CPC
1.20
Why? A way to assess surface
area, mass and lung deposition
Cigarette smoke
1.00
Waterpipe smoke
Fitted density
0.80
10
100
Diameter [nm]
1000
Aerosol Mass Spectrometer
Particle composition with high time and size resolution (30-1000
nm), but requires a dedicated operator
Analysis of sampled nanoparticles
Lung deposition measurements
No online methods available to obtain many important particle
characteristics. Analysis of sampled particles needed:
•
Particle shape: electron microscopy (TEM, SEM)
•
Detailed chemical composition: e.g. x-ray, MS
•
Toxicity: in vivo (animals, humans) or in vitro (cell culture)
•
Biological properties: e.g. PCR
Deposition Fraction
Lung deposition measurements
1
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0
0.001
Total
Extrathoracic
Summary
Biological analysis
PCR, proteomics,
biomarkers…
Slow
Tracheobronchial
Alveolar
Particle mass
TEOM, gravimetric…
0.01
0.1
1
Particle diameter [µm]
10
100
Fast
Particle number and
surface area
CPC, electrostatic…
Cheap
Simple
(Unspecific)
Summary
•
Possibilities to measure a wide range of properties of
airborne (nano)particles
•
Sometimes a choice between simplicity and accuracy (or
high time resolution and relevant particle characteristics)
•
Difficult to measure relevant exposure:
- background particles vs produced particles
- appropriate health metric unclear
•
New methods are emerging…
Chemical analysis
x-ray (PIXE, XRF,
MAX-lab…), MS…
Particle shape
Electron microscopy,
APM
Particle size distribution
(SMPS, ELPI, FMPS…)
AMS, particle size and chemistry
Expensive
Complex
(Specific)
Background
Nanomaterial
Respiratory and immunological effects following
inhalation of engineered nanoparticles
Åsa Gustafsson
Åsa Gustafsson
Particle properties
Studied effects
Agglomeration
Concentration
Material composition
Cytokines and
chemokines in
Lung lavage and in
blood
Shape
NP
Lung function
Size
Size distribution
Surface charge, ζ-potential
Cells in lung lavage and
in lymph nodes
Surface functionality
Åsa Gustafsson
Åsa Gustafsson
How are susceptible individuals
affected?
Gustafsson et al (2011) J Immunotoxicol
Study I
Inflammatory and immunological responses in the
airways following one exposure to titanium dioxide.
Genetics
Åsa Gustafsson
•
•
•
•
•
In vivo, Rat
Administration
Dose
Particle size
Analysis





Åsa Gustafsson
Dark Aguoti
Intratracheal
5 mg/kg body weight
200 nm and 2 µm
1, 2, 8, 16, 30 and 90 days
following exposure
Gustafsson et al (2011) J Immunotoxicol
Gustafsson et al (2011) J Immunotoxicol
I. Innate and adaptive immune system
Nanoparticle
30
90 days after exp.
Alveolar macrophages
Lung epithelial cells
Pro-inflammatory
cytokines IL-1β, IL-10,
IFN-γ, TNF-α, IL-8
Lymph
nodes
Pro-inflammatory
cytokines IL-1β, IL-6,
GM-CSF, TNF-α etc.
250
Neutrophils
Neutrofiler
**
200
4
16
Antal celler
(x10 ) (x104)
Number
of cells
8
Number
Antal celler
of cells
(x104) (x104)
Instillation
0 1 2
150
**
100
**
50
0
0
1
**
**
2
#
8
16
30
17.5
Lymphocytes
Lymfocyter
12.5
**
10.0
90 90 (C)
7.5
**
5.0
##
2.5
0.0
0
Dagar
efter
instillering
Days
after
exposure
Dendritic cell
**
15.0
1
2
8
16
30
90 90 (C)
Dagar
efterexposure
instillering
Days
after
Lymphocyte
- activation
Åsa Gustafsson
Åsa Gustafsson
Gustafsson et al (2011) J Immunotoxicol
Gustafsson et al (2011) J Immunotoxicol
I. Titanium dioxide particles in
lungtissue
I. Cell differentiation
TNF-α, IL-6, IL-1, GM-CSF, IL-8
Neu
M1
TH1
IFN-
GM-CSF
TH1: Virus and bacteria
TH2: Parasites
IL-4, IL-10, IL-13
Neu: Neutrophil
IL-6
IL-4
GCS-F
IL-12
IL-10
TH17: Bacteria and fungal
TH2
IL-13
IL-7
TH9: Reglation of parasites?
TNF-α
IL-5
IL-2
M2
IL-17
TREG
IL-4, IL-5, IL-13
IL-1
20x
10x
40x
IL-18
IL-9
TH9
TREG: T-regulatory lymphocyte
M1: Macrophage type 1 – Virus and bacteria
100x
TH17
M2: Macrophage type 2 - Parasite
100x
2 days
IL-17, IL-6
Åsa Gustafsson
Jonasson et al (2013) Inhal Toxicol
II. Allergy schedule
Study II
Responses on the immune system and lung function following
exposure to titanium dioxide in mice at different time points
during the developement of allergic airway inflammation.
In vivo
Nanomaterial
Particle administration
Number of exposures
Deposition, each exposure
Dose, each exposure
Allergen
Åsa Gustafsson
90 days
Åsa Gustafsson
Jonasson et al (2013) Inhal Toxicol
•
•
•
•
•
•
•
64x
30 days







Mice Balb/c
TiO2
Aerosol
8 times á 2h
32±1 µg
1.5-1.8 mg/kg body weight
Ovalbumin
OVA
Immunisation of allergen
Allergen provocation
Immunized
OVA group
OVA allergen
1.
Allergen specific
IgE antibodies
3.
Airway reactivity
OVA allergen
2.
4.
Eosinophil
inflammation
TH2 immune activation
IL-4, IL-5, IL-13
Åsa Gustafsson
2015-03-10
Jonasson et al (2013) Inhal Toxicol
Jonasson et al (2013) Inhal Toxicol
II. OVA-specific antibodies
Airway inflammation
II. Allergy schedule-Particle exposure
OVA
TiO2
Immunisation of allergen
Allergen provocation
Immunized
OVA group
Åsa Gustafsson
4
2
G
r.
3
G
r.
1
0
G
r.
2
2
3
G
r.
r.
1
0
***
6
O
VA
20
Neutrophils
Neutrofiler
8
ko
nt
ro
ll
***
G
.3
Gr
.1
.2
Gr
Gr
ll
nt
ro
ko
O
VA
0.0
Immunized
TiO2/OVA
(gr. 3)
***
***
G
r.
1.0
0.5
***
40
ll
O.D
*
***
O
VA
***
1.5
Immunized
TiO2/OVA
(gr. 2)
Eosinophils
Eosinofiler
60
nt
ro
2.0
5
Number
cells
in BALF
Antalof
celler
i BAL
(x105) (x10 )
2. Airway inflammation
1. OVA specific
IgE antibodies
Immunized
TiO2/OVA
(gr. 1)
ko
OVA allergen
5
Number
cells
in BALF
Antalof
celler
i BAL
(x105) (x10 )
OVA allergen
Åsa Gustafsson
Jonasson et al (2013) Inhal Toxicol
Gustafsson et al (2014) Toxicol
Study III
II. Airway reactivity and TH2 immune activation
Genetic influence on immunologic responses and
lung physiology following exposure to titanium
dioxide.
4. TH2 immune activation (IL-4, IL-5, IL-13)
pg/ml in BAL
4
2
**
***
* *
*** *********
***
IL
-4
r.
3
r.
2
Gr. 1
Gr. 2
Gr. 3
***
***
2500
2000
1500
1000
500
0 ***
G
G
G
r.
1
O
VA
0
Kontroll
OVA
25000
20000
15000
10000
5000
IL
-1
3
*
**
IL
-5
6
ko
nt
ro
ll
RRS (cmH2Os/ml)
3. Airway reactivity
• In vivo, Rat
•
•
•
•
•
•
Nanomaterial
Particle administration
Number of exposures
Deposition, each exposure
Dose each exposure
Allergen
Åsa Gustafsson
 Dark Aguoti (DA)
Brown Norwegian (BN)
 TiO2
 Aerosol
 10 times á 2h
 160-170 µg
 0.64-0.8 mg/kg body weight
 Ovalbumin
Åsa Gustafsson
Gustafsson et al (2014) Toxicol
Gustafsson et al (2014) Toxicol
III. Allergen specific antibodies
Airway inflammation
Immunisation of allergen
Immunized
TiO2/OVA
III. Airway reactivity
Allergen provocation
DA. Central airways
Åsa Gustafsson
0.4
Kontroll n=8
OVA n=7
TiO2+ OVA n=7
***
0.3
0.2
0.1
G (cmH2Os×mL-1)
RRS (cmH2Os×mL-1)
*
3
A
A
V
2 /O
O
O
V
A
K
on
t ro
ll
2 /O
0.00
Ti
Ti
O
O
V
V
A
0.0
0.5
0.0
Saline
5
2.25
2.00
1.75
1.50
1.25
1.00
0.75
0.50
0.25
0.00
Kontroll n=8
OVA n=7
OVA+TiO2 n=7
**
Saline
MCh (mg×mL -1)
5
MCh (mg×mL -1)
*
***
Eosinofiler Neutrofiler Lymfocyter
3
*
2
0.1
1.25
TiO2/OVA
PBS
OVA
1.50
1.00
1.25
0.75
1.00
0.50
0.75
*
0.25
*
0.50
*
0.00
*
Eosinofiler
Neutrofiler
Lymfocyter
Neutrophils Lymphocytes
0.25 Eosinophils
**
2
0.2
K
on
t ro
ll
O.D
0.3
TiO2/OVA
OVA
DA Råtta
*
1
DA rat
BN rat
0.6
DARåtta
Rat
DA
Control
Kontroll
1.50
1
0.4
DA. Small airways
2. Airway inflammation
OVA specific
IgE antibodies
Antal
celler
(x106(x10
) 6)
Number
of cells
in6BALF
Antal
celler
(x10
)
1.
Åsa Gustafsson
3
Gustafsson et al (2014) Toxicol
Gustafsson et al. Manuskript
III. Mediators in healthy DA rats after
titanium dioxid exposures
Immunologic responses in airways and lymph nodes
following inhalation to iron oxide (hematite) in healthy
and in mice with allergic airway inflammation.
IL-6
Healthy (control)
50
VEGF
Healthy (TiO2)
25 000
40
50
40
30
15 000
125
100
10 000
100
80
75
25
10
30
40
50
10
30
20
3 000
20 25
20
IL-13
20
40
50
40
•
•
•
•
•
•
GM-CSF
T H2
10
20
40
60
T H1
30
20
5 000 10
50
Proinflammatoric
GCS-F
20 000
IL-7
Study IV
T- , B-cell mature
6 000
9 000
50
12 000
15 000
Cardiovascular
MIP-3α
75
60
80
IFN-γ
100
100
125
In vivo, Mice
Nanomaterial
Particle administration
Dose
Analysis
Allergen
•
•
•
•
•
•
Balb/c
Iron oxide (Hematite)
Intratracheally
5 mg/kg body weight
1, 2, och 7 days after exposure
Ovalbumin
IL-12 (p70)
IL-18
Åsa Gustafsson
Åsa Gustafsson
IV. Cellular reduction i mice with an
allergic airway inflammation
IV. Inflammation in healthy mice
0.1
0.0
***
0.15
0.10
0.05
0.00
Day 1
Day 2
Day 7
Number of cells (x106)
Number of cells (x106)
p=0.09
**
0.015
**
0.010
0.005
0.000
Day 1
Day 2
Day 2
Day 7
0.10
0.05
Day 1
0.04
0.03
***
0.01
0.00
Day 7
**
0.02
p=0.08
6
4
2
Åsa Gustafsson
IV. Production of oxygen species
Day 2
Day 7
Day 1
Day 2
Day 7
Eosinophils
Macrophages
*
1.0
0.5
Day 7
8
0
Day 1
Day 2
Eosinophils
0.05
1.5
0.0
0.00
Lymphocytes
Eosinophils
0.020
Day 1
0.15
Number of cells (x106)
0.2
Hematite
Hematit
Neutrophils
p=0.06
Number of cells (x106)
0.3
Vehicle
Vehikel
Macrophages
0.20
Number of cells (x106)
0.4
Hematit
Hematite
Number of cells (x106)
Number of cells (x106)
*
Number of cells (x106)
Vehikel
Vehicle
Neutrophils
0.5
Day 1
Day 2
Day 7
Lymphocytes
2.0
*
1.5
1.0
0.5
0.0
Day 1
Day 2
Day 7
Åsa Gustafsson
Conclusions
 TiO2-particles that are deposited in the alveolar region may remain over a long period of time
(Study I and III).
 TiO2-particles induce a long term activation of the innate- and adaptive immune system (Study I
and III).
 Inhalation of TiO2-particles before and during allergen provocation in immunized mice induced a
neutrophil inflammation, body weight reduction and impaired general condition (Study II).
 Inhalation of TiO2-particles does not exacerbate characteristics of alleric airway inflammation in
rat, although there is a genetic variety regarding cellular composition (Study III).
 Rats with inhereted predisposition towards developement of TH1 immune responses was more
sensitive and expressed more mediators (Study III).
 The cellular response in a lung with established allergic airway inflammation and associated
draining lymph nodes does probably induce cell death following exposure to hematite NPs (Study
IV).
Åsa Gustafsson
 It is important to include sensitive individuals when evaluating risk assessments of
nanomaterials.
Åsa Gustafsson
Förväntad tillväxt i världen år 2000-2020
2020 – marknadsvärde
3 000 miljarder $
Nanosäkerhet ur ett EU- och
OECD perspektiv
Erfarenheter från EU-kommissionen, OECD och svenska
regeringsuppdraget om nanosäkerhet
KemI myndighetsmöte, 27 November 2014
Eva Hellsten
Årlig tillväxt 2000 – 2008 ekonomi, sysselsättning, patent,
vetenskapliga publikationer, investering i R&D ~ 20-35 %
År 2000 – marknadsvärde
30 miljoner $
Adapted from M. Rocco et al 2011
EU Kommissionen år 2004…
”….Overall spending on R&D should be
increased in Europe to balance the heavy
investments that have been initiated by our
main competitors”
“….Any negative impact on public health,
safety or the environment must be addressed
upfront and as an integral part of
technological development”
“Towards a European Strategy for Nanotechnology, 2004”
EU:s Aktionsplan 2005-2009
R&D finansiering av Nanotech från EU
FP5
1998-2002
FP6
2002-2006
FP7
2007-2014

50-tal åtgärder som Kommissionen och
medlemsländerna skulle genomföra för att
säkerställa en “säker, integrerad och ansvarsfull”
utveckling av nanoteknologi

Helhetsgrepp om forskning & innovation,
industriell utveckling, etik, kunskapsuppbyggnad
och lagstiftning för skydd av miljö och hälsa,
internationellt samarbete etc.

Antogs inom EU 2005
280 miljoner €
1,4 miljarder €
> flera miljarder € (?)
Satsningen på miljö och hälsa har under åren ökat till
dagens nivå på cirka 25 miljoner € per år
“Nanosafety” som ett integrerat policy område
Forskning
och
Innovation
Inom EU kommissionen…..
samordning mellan generaldirektoraten
DG
ENV
Nano
safety
forskning
Koordinering
DG
RTD
DG
ENTR
DG
INFSO
DG
SANCO
Kommunikation
Lagstiftning
för
nanomateria
l
DG
EMPL
Nano och REACH – en kapplöpning i tiden
2002
Nanotech R&D ökar i EU forskning
2003
REACH antas i EU Kommissionen
(ton-gräns för registrering)
2004
”Towards”-strategin
2005
”Action Plan 2005-2009”
2006
REACH-lagstiftningen antas av EU
Parlamentet och Rådet
Vetenskapliga frågeställningar om
nano och REACH
Flertalet lagstiftningar för nanomaterial
Kemikalier – REACH, Biocider, Pesticider
Arbetsmiljö
Konsument produkter – Läkemedel, Livsmedel,
Kosmetika m.m.
Miljö– Luft, Vatten, Avfall m.m.
EU Kommissionens översyn av EU lagstifning
2008 resp 2012
EU:s Rekommendation av en definition för
nanomaterial 2011
Risk-bedömning och nanomaterial
Hazards
Nya
och/eller modifierade testmetoder och guidelines
Validering
Metoder
av testmetoder.
(eco)toxicity tests
fate, transport
characterization, standards,
reference materials,
metrics, dosimetry,
validation
Exposures
exposure
assessment
Scientific uncertainty
för att mäta och bedöma exponering
Risk assessment
Precautionary
principle
Proportionality
principle
Risk management
Testmetoder
Internationell samordning och harmonisering
In vitro test
Test
Guidelines
Databas
OECD
Working Party on Manufactured
Nanomaterials (WPMN) – utveckla
internationellt harmoniserade test metoder och
test guidelines att användas i lagstiftning
Exponerings
ana
lys
OECD WPMN
start 2006
Risk
assesment
Forskning
– skapa internationella standards,
nomenklatur, mät- och analytiska metoder
ISO
Livscykel
analys
Lagstiftning
Samordning i medlemsländerna
Myndigheter
Forskare
Industri
NGO
Central koordinering
genom myndigheter &
departement:
Tyskland
UK
Nederländerna
Jfr USA där NNI
koordinerar
Huvudförslagen – i linje med vad som görs inom
EU och andra medlemsstater
Satsning på forskning för att öka kunskap om miljö och
hälsoeffekter I ett livscykelperspektiv.
Anpassning av lagstiftning och testmetoder genom att
förstärka arbetet inom EU och OECD
Bygga upp ett Nanoråd och Nanocentrum för att bättre
koordinera svenska insatser
Sverige 2012 –
regeringsuppdrag om nanosäkerhet

Utveckla nationell handlingsplan.
Säkerställa en god samordning av
arbetet med nanosäkerhet

Redovisades I oktober 2013. SOU
2013:70, www.regeringen.se

Remissrunda under våren 2014 med
ca 70 svar
OECD‐arbetet för nanomaterialsäkerhet idag –
Innehåll:
Ahtiainen Jukka | 27.6.2013
Myndighetsmöte Nanomaterials upptag och spridning i kroppen och miljön •
OECD‐arbetet för nanomaterialsäkerhet idag
•
Jukka Ahtiainen, senior forskare, PhD
OECD Sekretariat och
Säkerhets‐ och kemikalieverket (Tukes) •
•
OECD Working Party on Manufactured Nanomaterials (WPMN)
Säkerhetsbedömning och testning
Testmetodernas lämplighet
Riskhantering I arbetsmiljön
•
•
•
OECD testmetoder (TG) och nanomaterialer
Test Guidelines (TG) utveckling och WNT
TG tillämplighet att testa nanomateraler or behov för nya metoder och
riktlinjer (OECD GD) t.ex. testningstrategi
•
•
Idag viktiga OECD projekten
Fysikalisk‐ kemisk‐ egenskaper
Miljöspridning och akkumulering
Ekotoksisitet
Hälsoeffekter (med toksikokinetik)
•
•
•
•
Slutsatser och sammanfatning
27.11.2014, Kemi myndighetsmöte‐ Jukka Ahtiainen OECD Working Party on Manufactured
Nanomaterials (WPMN)

OECD Working Party on Manufactured
Nanomaterials (WPMN) ‐ Vad har gjort hittills?
Working Party on Manufactured Nanomaterials (WPMN) under OECD:s kemikaliekomitteen var inträttade i September 2006 med uppgiften att verka för internationell samverkan om hälso‐ och miljöriskrelaterade frågor när det gäller avsiktligt tillverkade nanomaterialer.


För detta ändamål har ett antal styrgrupper bildats under WPMN
 Systematisk analysering av resultat från “Sponsorship Programme” har börjat
befintlig forskning (SG1 och SG3), testar globalt och systematiskt givna representativa nanomaterialgrupper (SG3), bedömer och förbättrar kemikalieprovningsmetodernas tillämpbarhet (SG4 och SG7)
och 
 Guidance on sample preparation and dosimetry for the safety testing of
manufactured nanomaterials (2012)
I vilka man samlar information om:



 Publikationer och riktlinjer för testning:
uppgör anvisningar för riskbedömning och riskhanering (SG6 och SG8).
redan 2012 och det andra projektet (phase 2) för verklig risk bedömning skall
börjas snart (linken till EU projekten NANoREG som startade 2013).
 Några riktlinjer och metoder måste förbättras, men i allmänhet test metoder
och riktlinjer för kemikalier kan följas
http://www.OECD.org/env/chemicalsafetyandbiosafety/safetyofmanufacturednanomaterials/
Varför måste OECD testmetoder utnytjas?


OECD test guideline
Good Laboratory Practise
Mutual Acceptance of data
For testing intrinsic properties of chemicals (substances)
Binding OECD countries and
selected non‐members
►Avoids duplication of testing
►Reduces use of animals
►160 million euros saved each
year (2010)
►“easily” adopted to EU regulation (440/2008) for EU regulatory needs
27.11.2014, Kemi myndighetsmöte, Jukka Ahtiainen
Stakeholders (authority, NGO, industry, academia) initiative
TG proposal (SPSF) by MC
Expert Group
National Coordinators of the test guideline programme
(WNT)
Commenting rounds
Validation package
Draft TGs
OECD secretariat
Final approval by WNT at the meeting or written procedure
Joint Meeting , policy level and publication Ahtiainen et al 2009/SETAc Göteborg
27.9.2012 | Nordisk råds Miljö‐och naturesursutvalg, Ahtiainen Jukka
3/9/2015
27.11.2014 Kemi myndighetsmöte, Ahtiainen Jukka
OECD‐metoders tillämplighet att testa nanomaterialer (NM)

Biologiska “endpoints” (vad man observerar) är relevanta och tillämpliga
OECD Test Guideline Programme – Hur att handla nya
nanospesifika riklinjer och instrukrioner för NM testning

för NM testing till ex:






Number of offspring in reproduction tests‐ hur många babysar
If the guidance given in the separate GDs can be seen as refinement of

Or should the NM specific guidance be inserted as annex in the TG?
Bioaccumulation into tissues‐ akkumulering och spridning i kroppen
the test guideline, should the result be still under MAD?
CO2 production in biodegradation test‐ biologisk nedbrytning

Many apical and other endpoints in mammal tests‐ inflammationer
Några nya “nanorelevant” Phys‐Chem‐ metoder behövs
Guidance for similar matrices and test or for nanomaterial groups?
Metals Metal
CNTs
Fullerenes
NFC NCC
oxides
Soil tests
Dosing and dosimetry of the test material and NM detection and
Sediment tests
characterization very important – Riktlinjer måste utveklas

TG development and guidance documents (GDs) and MAD

Aquatic tests
Med harmoniserade riktlinjer oh metoder MAD‐principen torde hållas
Bioaccumulation
Degradation
MAD = Mutual Acceptance of Data
27.11.2014 Kemi myndihetsmöte, Jukka Ahtiainen 27.11.2014 Kemi myndighetsmöte, Jukka Ahtiainen OECD Test Guideline Programme – Hur att handla
nya nanospesifika riklinjer och instrukrioner för NM testning
 Inhalation toxicity testing as an example
 Minor changes in Test Guidelines
 More extensive revisions in Guidance Documents (e.g. GD 29)
 New Guidance Document for NM testing
 Guidance for similar tests or for nanomaterial groups?
Metals
Metal
oxides
CNTs
Fullerenes
NFC
Current OECD guidance on NM testing
 Preliminary review of OECD test guidelines for their applicability to
manufactured nanomaterials (2009)
 Guidance manual for the testing of manufactured nanomaterials: OECD
sponsorship programme – rev1(2009)
 Guidance on sample preparation and dosimetry for the safety testing of
NCC
manufactured nanomaterials revision published 2012
Inhalation tox
Oral tox
 Dispersion protocols for in vitro testing (on-going work)
Genotox
 Development of integrated testing strategy for NM testing (started)
Immunotox
In vitro
27.11.2014 Kemi myndighetsmöte, Jukka Ahtiainen OECD Horizontala mötena för detta (WNT & WPMN)
OECD metoder och instruktioner –
phys‐chem characterization methods
Aims: Review in terms of
•
…the need to amend existing or develop new OECD Test guidelines,
…the need to develop specific Guidance for the testing of NM
… identification of knowledge gaps regarding regulatory need
Topic
Venue
The Hague, NL
Inhalation Toxicology
Environmental Fate and Ecotoxicology Berlin, DE
Date
19‐20 October 2011
Physical‐Chemical Properties
Querétaro, MEX
Genotoxicity
Ottawa, CAN
19‐21 November 2013
Toxicokinetics
Seoul, COR
26‐28 February 2014
Physical‐Chemical Parameters: Measurements and Methods
Washington DC, USA
18‐19 June 2014
Categorization
Washington DC, USA
17‐19 September 2014
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen •
29‐31 January 2013
28 February – 01 March 2013
•
The phys‐chem data is crucial for substance ID, to guide further
testing and understand the results
• Overlaps partly to env.fate methods (e.g. dissolution)
• Reliable methods (MAD?) will enable triggering or waiwing of some
testing‐ verkligen viktiga och nyttiga och behövs snart!
Proposed methods
Size, form, fiber rigidity, surface charge, zeta‐potential, surface
chemistry…
• Approved list of WPMN‐ men skulle vara på WNT listan!
•
Current status
Not any included in the WNT project list!
Cooperation with ISO TC 229 – samarbetet borde förbättras!
•
•
27.11.2014, Kemi myndighetsmöte‐ Jukka Ahtiainen OECD‐ metoder och instruktioner–
ecotoxicity and environmental fate methods
OECD metoder och instruktioner–
ecotoxicity and environmental fate methods
TG: dissolution rate of nanomaterials in the aquatic environment
Lead: US (US Army Corp) in cooperation with DTU
TG: agglomeration behaviour of nanomaterials in different aquatic media
Lead: GER (UBA), contracted to University of Vienna
GD: agglomeration and dissolution of nanomaterials in aquatic media –
decision tree
GD: Assessing the Apparent Accumulation Potential of Nanomaterials
Lead: UK (Uni Plymouth) with ES, NL, DE, FI
TG: Nanomaterial Removal from Wastewater
Lead: US (EPA)
GD: Leaching in soil column Cooperation: CAN (ENV CAN), US (EPA), GER (UBA)
Lead: GER (UBA), contracted to University of Vienna
GD Aquatic (and Sediment) Toxicology Testing of Nanomaterials
Lead: US (US Army Corp) and UK (Uni Plymoth & Heriot‐Watt
‐ Alt dessa är viktiga, men mera behövs till ex. för jordtestning
27.11.2014, Kemi myndighetsmöte‐ Jukka Ahtiainen 27.11.2014, Kemi myndighetsmöte‐ Jukka Ahtiainen Current OECD activities –
ecotoxicity and environmental fate methods
Current OECD activities –
ecotoxicity and environmental fate methods
GD: agglomeration and dissolution of nanomaterials in aquatic media – decision tree
GD: agglomeration and dissolution of nanomaterials in aquatic media – decision tree
• Lead: Germany (UBA); contracted to University of Vienna with EC, DTU(DK)
• Should support decision making regarding environment fate& behaviour test performance/test strategy e.g. Identification whether
nanospecific test is needed,
• First information on mobility or possible target compartments
• Same timeframe like TG on agglomeration behaviour
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen 26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen Current OECD activities –
ecotoxicity and environmental fate methods
Current OECD activities –
ecotoxicity and environmental fate methods
GD Aquatic (and Sediment) Toxicology Testing of Nanomaterials
GD Aquatic (and Sediment) Toxicology Testing of Nanomaterials
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen 26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen Current OECD activities –
ecotoxicity and environmental fate methods
GD Aquatic (and Sediment) Toxicology Testing of Nanomaterials
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen Current OECD activities –
human health
Current OECD activities –
ecotoxicity and environmental fate methods
GD: Guidance on Fish Dietary Accumulation Studies for Engineered Nanomaterials (content)
• Introduction
• scope of the guidance, • limitations of TG 305, • Bioaccumulation/ digestability
• kinetics with nanomaterials…)
• Decision tree and triggers for needing to do the test
• Fate indications
• In vitro digestability
• Mixing into food
• Practical in vivo testing:
• Test designs, verification of exposure, detection and analytics
• Calculations and interpretations of the results
• Use in the risk assessment
• Not BCF or BMF as such but useful information
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen Current OECD activities –
human health
GD: Guidance on nanomaterial inhalation testing
Guidances on nanomaterial genotoxicity testing
 Project approved into the WNT work programme:
 “Amendments to the Inhalation TGs and GD to
Accommodate Nanomaterial Safety Testing (lead: US and
NL)
 Expertise to be broadened to WNT experts
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen OECD‐ arbetet för nano‐säkerhet idag‐
slutsatser och sammanfattning  Mera phys‐chem‐metoder (OECD TGs) behövs
 Många instruktioner (guidance documents, GDs) måste utvecklas för att testa NM med OECD metoder
 Alt detta menar att ECHA och dess instruktioner (ECHA guidance for registration and fulfilling the information
requirements) spelar mycket viktig roll
Tusen tack‐ några frågor?
27.11.2014, Kemi myndighetsdag‐ Jukka Ahtiainen  Not yet included in the WNT work programme
 OECD Workshop in Canada 2013‐ report available late 2014
 Modifications of 487 (In vitro Micronucleus Test to
accommodate NMs)
 Comet assay
 Not to use bacterial assays
26.11.2014, Nordic Nano WS‐ Jukka Ahtiainen Ändringar av bilagorna till EU:s
kemikalieförordning Reach
Nanomaterial
- en uppdatering om KemI:s arbete
Elin Simonsson, Stockholm
27 november 2014
Översyn av kommissionens
rekommenderade definition
• Definitionen ses över särskilt med hänsyn till kravet på
att minst 50 % av partiklarna ska vara inom nanoskalan
(1-100 nm).
• Kommissionens arbete med bilagorna blir ytterligare
försenade.
•
Tidigare utlovades ändringar innan utgången av 2014
senaste beskeden är nu våren 2015.
• Arbetet pågår i CASG Nano (Reach Competent
Authorities subgroup on Nanomaterials )
Åtgärder för ökad transparens på
marknaden
• Kommissionen arbetar med en konsekvensutredning om
åtgärder för ökad transparens på marknaden bl.a. ett
EU-register för nanomaterial
• Ett offentlig samråd har varit en del av arbete
• Översynen ska vara klar senast i december 2014 men
kommer sannolikt att försenas till våren 2015.
• Översynen är uppdelad i en vetenskaplig del och en
policy del.
Frågor?
• Konsekvensutredningen kommer att färdigställas under
2015 varpå kommissionen kan välja att föreslå åtgärder
Snabb regelrepetition
• Förordning (EG) nr 1223/2009 om Kosmetiska
Produkter
Nanomaterial i kosmetiska produkter –
vad har hänt sedan de nya reglerna?
Tomas Byström
Enheten för Kosmetiska Produkter
Läkemedelsverket
• Egen definition
‘nanomaterial: ett olösligt eller biopersistent material som är
avsiktligt tillverkat, med en eller fler yttre dimensioner, eller
en inre struktur, med ett spann på mellan 1 och 100 nm
• Arbetsgrupp har diskuterat justering av definitionen
– Ingen ändring aktuell
– Avvaktar hur definitionen i livsmedelsreglerna fungerar
Snabb regelrepetition
Vad har hänt?
•
Varje användning av nanomaterial måste anmälas till EU-kommissionen 6 månader
före produkter släpps ut på marknaden
•
EU-kommissionen avgör från fall till fall om användningen behöver granskas av en
vetenskaplig kommitté
• 25000 produkter anmälda i hela EU
– Spritt över så gott som alla kategorier
– 7300 solskyddsprodukter
– 7000 sminkprodukter (Carbon Black/TiO2)
– 2700 ansiktsprodukter (troligen stor del dagkräm med SPF)
• 150 produkter anmälda med hemvist i Sverige
– 60 solskyddsprodukter
– 80 sminkprodukter
• Dålig kunskap hos en del företag som anmäler produkter (t ex
har vatten och etanol anmälts som nanomaterial) – dock inte
noterat i Sverige
–
–
–
Förhandsgranskade och tillåtna nanomaterial (UV-filter, Färgämnen, Konserveringsmedel)
Granskning av produktspecifik användning (information om nanomaterialets egenskaper ska
ingå i anmälan)
Vägledningsdokument om säkerhetsbedömning av nanomaterial
•
•
Safety assessment of nanomaterials in Cosmetic Products
http://ec.europa.eu/health/scientific_committees/consumer_safety/docs/sccs_s_005.pdf
Relevance, Adequacy and Quality of Data in Safety Dossiers on Nanomaterials
http://ec.europa.eu/health/scientific_committees/consumer_safety/docs/sccs_o_142.pdf
•
Innehåll av nanomaterial måste framgå av innehållsförteckningen på förpackningen
•
EU-kommisionen ska publicera en katalog över de nanomaterial som används i
kosmetiska produkter
–
Exempel: Titanium Dioxide (nano)
• Ett ämne (UV-filter) godkänt som nanomaterial – flera släpar
Utmaningar för regelutveckling
Vilka nanomaterial har utvärderats
• Arbetet med säkerhetsvärderingar ligger aningen
sent
• Ett nanomaterial tillåtet
– Tris-biphenyl triazine (UV-filter)
• Nanomaterial som har utvärderats av vetenskapliga kommittén,
regeländring om tillåtande kvarstår
– Titanium Dioxide (UV-filter)
– Zinc Oxide (UV-filter)
– Carbon black (CI 77266, Färgämne)
– Methylene Bis-Benzotriazolyl Tetramethylbutylphenol (UV filter) –
utvärdering av kompletterande data kvarstår
• Pågående utvärdering
– Silica med derivat
• Annan utvärdering
– Säkerheten hos sprayprodukter (när finns inhalationsrisk?)
• Data saknas vilket gör att värderingarna inte kan
fullföljas
• Oklara definitioner som berör restriktioner i
användning av nanomaterial (t ex sprayprodukter
och inhalationsrisk)
• Trög process för regeländringar som resultat av
ovanstående
Kommande händelser
• Katalogen över nanomaterial är på översättning för
snar publicering
• Fullfölja regeluppdateringarna rörande
förhandsgranskade och tillåtna nanomaterial
• EU-kommissionen har uppmanat medlemsstaterna
att utöva tillsyn för att säkerställa att företagen gör
rätt
– Detta med anledning av de många tokiga anmälningarna
Definition av Nanomaterial i livsmedel
•
Förordning (EU) nr 1169/2011 om tillhandahållande av livsmedelsinformation till konsumenterna
•
Definition
konstruerat nanomaterial: avsiktligt tillverkat material som har en eller fler
dimensioner i storleksordningen 100 nm eller mindre eller som består av åtskilda
funktionella delar, antingen i sitt inre eller på ytan, varav många har en eller flera
dimensioner i storleksordningen 100 nm eller mindre, inbegripet strukturer,
agglomerat eller aggregat, som kan vara i en storleksordning över 100 nm men
behåller egenskaper som är utmärkande för nanonivån.
Egenskaper som är utmärkande för nanonivå inbegriper
i) de egenskaper som avser de stora särskilda ytorna hos materialet i fråga,
och/eller
ii) särskilda fysisk-kemiska egenskaper som skiljer sig från egenskaperna hos
samma materials icke-nanoform.
Tack för att ni
lyssnade!
2015‐03‐26
Overview
Danish initiatives on
nanomaterials
• Research initiatives, consumer concern and general collaboration
• Action plans for chemicals
• “Better control of nanomaterials”
• Overview of initiatives
• The nano product register
• Future
Flemming Ingerslev,
Section of Chemicals
The Danish Environmental Protection Agency
The Danish Environmental Protection Agency
PAGE 2
Danish Action Plans for chemicals
DK outside DK-EPA
(in total around 2 mio DKK for nano)
• Research
• 2010-2013:
• Environmental Chemistry, DTU and Aarhus University
• Overview of important nanomaterials
• National Research Center for the Working Environment (Danish Nanosafety Center)
• Proposal for REACH-annexes (IR-nano!)
• University of Copenhagen and others
• 2014-16:
• Industry
• Specific products
• Consumers
• International work
• Politicians
• Danish EPA network group for nanomaterials!
The Danish Environmental Protection Agency
PAGE 3
The Danish Environmental Protection Agency
PAGE 4
Projects on NMs under the DK nano-initiative
Better control of nanomaterials
NP - register
Use/occurrence of
NM
• New government i Denmark 3. October 2011 with a green
Government Programme
• National Budget Agreement 2012 on “Better Control of
Nanomaterials and their Safety” (~ 3.2 mio. € 2012-2015, )
Assessment of
specific products
• Subproject 1:
Knowledge building with focus on exposure pathways and
implications for consumers and the environment with regard to
the use of nanomaterials (approx. 3/4 of budget)
1. NM’s in food, food contact
materials, cosmetics, pesticides
2. NM’s in pigment
3. NM’s in waste
1. Products with photocatalytic
titanium dioxide
2. Aerosol products with
nanomaterials
3. Textiles with nanosilver
1. Oral and dermal uptake
Processes and fate
of NM
• Subproject 2:
Development of a nano product register in cooperation with
other countries (approx. 1/4 of budget)
2. Fate processes in the enviroment
3. Human exposure
4. OECD test methods
Effects of NM
The Danish Environmental Protection Agency
Supplementary
surveys
Potential
Environmental and
health risks due to
nanomaterials in DK
1. Ecotoxicology
2. Toxicology
PAGE 5
1
2015‐03‐26
Projects on NMs under the DK nano-initiative
NP - register
Use/occurrence of
NM
Assessment of
specific products
Supplementary
surveys
1. NM’s in food, food contact
materials, cosmetics, pesticides
2. NM’s in pigment
3. NM’s in waste
1. Products with photocatalytic
titanium dioxide
2. Aerosol products with
nanomaterials
3. Textiles with nanosilver
1. Oral and dermal uptake
Processes and fate
of NM
2. Fate processes in the enviroment
3. Human exposure
4. OECD test methods
Effects of NM
The Danish Environmental Protection Agency
Potential
Environmental and
health risks due to
nanomaterials in DK
1. Ecotoxicology
2. Toxicology
PAGE 7
Projects on NMs under the DK nano-initiative
NP - register
Use/occurrence of
NM
Assessment of
specific products
Supplementary
surveys
1. NM’s in food, food contact
materials, cosmetics, pesticides
2. NM’s in pigment
3. NM’s in waste
1. Products with photocatalytic
titanium dioxide
2. Aerosol products with
nanomaterials
3. Textiles with nanosilver
1. Oral and dermal uptake
Processes and fate
of NM
2. Fate processes in the enviroment
3. Human exposure
4. OECD test methods
Effects of NM
Potential
Environmental and
health risks due to
nanomaterials in DK
1. Ecotoxicology
2. Toxicology
Projects on NMs under the DK nano-initiative
NP - register
Use/occurrence of
NM
Assessment of
specific products
Supplementary
surveys
1. NM’s in food, food contact
materials, cosmetics, pesticides
2. NM’s in pigment
3. NM’s in waste
1. Products with photocatalytic
titanium dioxide
2. Aerosol products with
nanomaterials
3. Textiles with nanosilver
1. Oral and dermal uptake
Processes and fate
of NM
2. Fate processes in the enviroment
3. Human exposure
4. OECD test methods
Effects of NM
Potential
Environmental and
health risks due to
nanomaterials in DK
1. Ecotoxicology
2. Toxicology
2
2015‐03‐26
The Ministerial Order no. 644 of 13/06/2014
/ Bekendtgørelse nr. 644 of 13/06/2014
Nano Product Register
Objective:
• To provide an overview of the nanoproducts that are on the Danish market, the extent of
use and the purposes they are used for.
• To provide information to sub-project 1 (knowledge building).
• To inspire to an EU-solution for registration of nano products
The Danish Environmental Protection Agency
PAGE 13
The Danish Environmental Protection Agency
Purpose and scope of the nanoproduct register
PAGE 14
§ 3. Excemptions
§ 1. The purpose of this Order is to establish a register of mixtures and
articles that contain nanomaterials (nano products) and which are
intended for sale to the general public, as well as require producers
and importers of these mixtures and articles to report to the nano
product register information on said mixtures and articles and the
nano materials they contain.
§ 2. The reporting requirement to the nano product register includes
mixtures and articles that are intended for sale to the general public
and which contain nanomaterials, where the nanomaterial itself is
released under normal or reasonably foreseeable use of the mixture or
article or where the nanomaterial itself is not released but substances
in soluble form that are classified as CMRs or environmentally
dangerous substances are released from the nanomaterial; except as
provided for in § 3.
1. Foodstuffs and food contact
materials.
8. Nanomaterials listed Annex IV or V of
REACH (Natural substances).
2. Feed.
9. Not intentionally produced at the nanoscale.
3. Medicinal products.
10. Nanomaterial is part of a fixed matrix
4. Medical devices.
11. Articles or their labels on which the
nanomaterial is used directly as ink, in
newspapers etc.
5. Cosmetic products.
6. Pesticides.
7. Waste.
12. Textiles with nanomaterial used as ink or for
dyeing.
13. Paint, wood preservative, glue and filler that
contains nanopigment added solely for
colouring the mixture.
14. Articles of rubber, or rubber parts of articles,
that contain the nanomaterials carbon black
or silicon dioxide
The Danish Environmental Protection Agency
PAGE 15
• Registration is mandatory - not all information
Voluntary
Information on company: ID#,
adress, contakt person etc.
REACH: Use descriptor categories
(PC, PROC. ERC, AC)
Productinformation: name,
amount, use, professional use (Y/N)
Content of nanomaterial: in
product or mixture (gram or %)
PAGE 16
Important features/characteristics
Product information
Mandatory
The Danish Environmental Protection Agency
• Registrants are those introducing the nanoproduct to the Danish
marked (i.e. no intention of traceability)
• Registration is not triggered by hazards or risk of nanomaterials
• Re-use of information from the Danish product register
• Data only available for authorities and registrant (own data)
Information on the nanomaterial: Fhysical information on
Name, REACH-registration, occurrence nanomaterial: particle size, number
in product
size distribution, aggregation,
agglomeratino, form, specific surface
Chemical information: IUPAC, CAS area, crystalline state, surface
no., EU-number, formula
chemistry, surface charge
The Danish Environmental Protection Agency
PAGE 17
The Danish Environmental Protection Agency
PAGE 18
3
2015‐03‐26
Guidance and help for registrants
Ideas for future collaboration
• Guidance in Danish and English
Danish EPA network group for nanomaterials
•
Should I register my product (nano or not?)
OECD test methods
•
18 illustrative examples of products
Shared knowledge (reports, advisory groups)
•
How to use IT-system?
•
Example of letter to supplier
•
List of common nanomaterials
•
List of product-groups that may be subject to registration
Research projects
EU-work (IR-nano)
• FAQ on www
• Help-desk
The Danish Environmental Protection Agency
PAGE 19
The Danish Environmental Protection Agency
PAGE 20
4
NANoREG ‐ A common European approach to the regulatory testing of nanomaterials
EU‐projekt NANoREG och nano‐cellulosa

Vad och varför?
Vetenskapliga svar för administrativa frågor
Snabbare och bättre
Tillförlitligt omgivning för industrin och myndigheterna
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen Allt med samma takt med innovationer och inte att förhindra dom
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen NANoREG project‐ facts and figures:
A common European approach to the regulatory testing of nanomaterials
NANoREG ‐ målsättning
The NANoREG project is funded by the EU Framework 7 Programme with € 10.000.000, total budget € 50.000.000
The aims of this project are:
(I) to provide tools for risk assessment and decision making instruments;
The project started on March 1st, 2013 and runs until August 31st, 2016 (42 months)
(II) to develop, for the long term, new testing strategies adapted to innovation requirements;
(III) to establish a close collaboration among authorities and industry to come to 61 partners from 15 countries (Austria, Belgium, Denmark, Finland, France, Germany, Ireland, Italy, the Netherlands, Norway, Portugal, Spain, Sweden, Switzerland, United Kingdom)
efficient Risk Management approach and to bring in ”safe by design” in the application development phase.
verktyg
Coordination:Ministry of Infrastructure and the Environment, The Netherlands, Tom van Teunenbroek (coordinator)
strategi
tillsammans med industrin
”safe by design” – ”value chains”‐ tänkandet inbildat
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen 3
NANoREG‐ Work package list‐ vad det innehåller
Work package
No
Work package title
Type of activity
Lead partici
pant
No
Lead participant short name
Person‐
months
Start
month
End
month
RTD
2
JRC
174,6
1
42
WP1
Scientific answers to regulatory issues
WP2
Synthesis, supplying and characterization
RTD
4
NRCWE
621,20
1
42
WP3
Exposure through life cycle analysis
RTD
7
CEREGE
535,90
1
42
WP4
Biokinetics and toxicity testing in vivo
RTD
3
BAuA
680,90
1
42
WP5
Advancement of Regulatory Risk Assessment and Testing
RTD
12
NIA
1231,60
1
42
RTD
5
RIVM
124
1
42
OTH
10
TEMAS
170
1
42
MGT
1
Min I&M
30
1
42
WP6
WP7
WP8
Keeping pace with innovation
Interaction, Dissemination and Exploitation
Project Management
total
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen 3568,80
Nanocellulosa och REACH

Bleached pulp cellulose (CAS‐nro:65996‐61‐4):








Exempted from REACH registration according to REACH Annex IV
If grinded mechanically to NFC (without chemical modifications), should probably (?) stay exempted. The rationale should be given by the manufacturer.
If chemically modified e.g. with covalent bounds, would be considered
as a new substance, and this would lead to obligation to register
NFC nanocellulose is nanomaterial by definition
Om så här nanocellulosa behövs inte registreras
Men om den formuleras med kemikaliska metoden?
Och vad ska man då registreras när det är en polymer?
Inget nytta att registrera monomer
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen NANoREG och nanocellulosa (Nano Fibril Cellulose)
 Finska koordinering på Säkerhets‐ och kemikalieverket (Tukes)
 Samarbetet med industrin, forskning och myndigheten
 UPM Corporation och Stora Enso (pengar finns här)
 Toksikologiska (andningsorgan) studier på Arbetshälsoinstitutet i
Helsingfors
 Målet: “… to assess the in vivo and in vitro genotoxic and
immunotoxic effects of nanofibrillar cellulose (NFC)”
27.11.2014 | Kemi myndighetsmöte, Jukka Ahtiainen Tack!
Box 2, 172 13 Sundbyberg
08-519 41 100
Besöks- och leveransadress
Esplanaden 3A, Sundbyberg
[email protected]
www.kemikalieinspektionen.se