registration of zno in reach is it sufficient for safety ... · is it sufficient for safety...
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Registration of ZnO in REACH – is it sufficient for safety evaluation
of nano ZnO?
Project realized within European Toxicology Risk Assessment Training (TRISK) http://trisk-project.eu/
Author: Katarzyna Malkiewicz Supervisors from KemI: Emma Wikstad and Agneta Falk-Filipsson
TRISK mentor: Annika Handberg
Changed from originally planned: Regulation of NMs in REACH - discussion and
recommendations based on the SKEP- ERA NET research project
Royal Institute of Technology, Stockholm Katarzyna Malkiewicz, Sven Ove Hansson University of Birmingham Michala Pettitt, Jamie Lead University College Dublin Iseult Lynch, Kenneth A. Dawson University of Helsinki Arho Toikka, Janne Hukkinen
Project report available upon request: contact: [email protected] Project funded by SKEP ERA-NET
Relevance? ”Nano” in REACH now:
• There are no “nano” specific provisions in REACH but general provisions apply
• Ultimately it is registrant interpretation of the existing provisions
which determine the approach in registration for “nano” forms • Interpretation ambiguity leads to extremely diverse opinions on a
number of issues! e.g. is the “nano-scale” the “identifier” (“nano“ different substance than “bulk” or is it “characterizer” (“nano” same substance, included in the same dossier but considered as different “form” or perhaps no consideration at all?
• Difficulties in reaching the consensus within RIP-on1 project –
Major obstacle in development of concrete proposals for REACH TGD!
Why ZnO?
• HPV, registration deadline before 30/11/2010
• Produced and used in different forms: bulk and nano-sized
• It is physiologically essential element and can lead to toxic effects
• Scientific literature on toxicological aspects of and differences between bulk and nano forms existent (but still very limited)
Why ZnO?
• HPV, registration deadline before 30/11/2010
• Produced and used in different forms: bulk and nano-sized
• It is physiologically essential element and can lead to toxic effects
• Scientific literature on toxicological aspects of and differences between bulk and nano forms existent (but still very limited)
produced and used as a “bulk” …
• bulk ZnO powder: additive to: plastics, ceramics, glass, cement, rubber, lubricants, paints, ointments, adhesives, sealants, pigments, foods, batteries, ferries, fire retardants, first aid tapes
and as ….“nano” • Novel uses in material science and electronics
(why?) wide-bandgap semiconductor, transparent, luminescent
(for what?) transparent electrodes, liquid crystals displays, energy saving and heat-protecting windows, electronics: thin-film transistors and light-emitting diodes
• Specifically attractive - One-dimensional ZnO nanostructures of the high aspect ratios (e.g. wires, rods, tubes)
(why?) enhances photoluminescence and lasing, novel optoelectronic properties,
(for what?) nanosensors, nanotransducers, photocatalist, generators
• Other common uses of ZnO nano powders:
(why?) UV blocking, self-cleaning properties, anti-bacteria properties
(for what?) sunscreen lotions, textiles, food and beverages packaging, glass, paints or film coatings, animal feed, cosmetics
Why ZnO?
• HPV, registration deadline before 30/11/2010
• Produced and used in different forms: bulk and nano-sized
• It is physiologically essential element but can be toxic
• Scientific literature on toxicological aspects of and differences between bulk and nano forms existent (but still very limited)
Inhalatory exposure to ZnO fumes (particles < 1 uM) during dwelling or smelting resulted in respiratory syndrome called metal fume fever (MFF): 600 mg/Zn/m3 for 10 minutes, or 77mg/m3 for 30 minutes. OSHA PEL 5mg/m3 for ZnO
Involved in: cell proliferation, differentiation, signal transduction, gene expression, function of over 300 enzymes RDA 2-11 mg/day
Figure 1. Comparison of the effects of zinc intoxication versus deficiency (Plum, Rink, & Haase, 2010).
Why ZnO?
• HPV, registration deadline before 30/11/2010
• Produced and used in different forms: bulk and nano-sized
• It is physiologically essential element but can be toxic
• Scientific literature on toxicological aspects of and differences between bulk and nano forms exist (but still very limited)
The aims were to:
1. Clarify how nano ZnO is registered (same or different substance to bulk?)
2. Analyze the extend of data registered
3. Analyze the approaches in hazard /exposure /risk assessment
4. Discuss them, based on independently collected evidence concerning nano vs bulk toxicologically relevant properties
Finding (1)
Registration of ZnO seems to concern both nano and bulk but with inconsistent approach:
• The consortium for the registration include entities dealing with ZnO in bulk and /or nanoforms (as an example one manufacturer of nano ZnO has stated the registration within 100-1000 t/y/m)
• For the phys-chem, the granulomentry represents “typical sample of the LEAD” (not in nano size range)
• No considerations for differential classification
• Exposure information – over 60 types of uses listed, majority specific for workers, and few overlapping between workers and consumers – but no indication on which concerns “nano”, and no exposure scenarios
• Several phys-chem and environmental fate endpoints waived (no info on stability, dissociation constant, hydrolysis, photo transformation) as considered in REACH not relevant for metals/inorganic – however some maybe relevant for nano e.g increased surface higher solubility!
• Toxicological studies with different forms bulk and nano have been registered (Environmental part was not analyzed within the project)
Finding (2)
Many data gaps in registration for “nano” • Limited number of studies (74) concerning human health hazard of
ZnO were registered (majority performed in 70s-90s). • Scarce number (14?) of studies concerns nano ZnO, and some lacking
also particle characterization (valid?) • The endpoints covered by nano ZnO based studies: inhalatory kinetics
(1 study), inhalatory acute (9 studies) and repeated (4 studies) toxicity • Broader spectrum of endpoints was covered by the studies with the
bulk ZnO, including those required by REACH for tonnage band up till 1000 tonnes / year/ manufacturer or importer. No carcinogenicity study was registered.
Statistics of the studies on ZnO registered at ECHA (accessed March 2011) with respect to the type of endpoint and form of ZnO studied. h- human, r- rat, ra-rabbit, m – mouse, gp – guinea pig, s –sheep, f-ferret, uf – ultra fine, fu – fumes
Endpoint No studies for bulk No of studies for fume/ultra
fine/ nano REACH tonnage trigger for phase-in
In vitro In vivo In vitro In vivo
Pharmaco/ Toxico-kinetics
oral 1 (h) Not required
inhalatory 2 (r) 1 (r)
dermal 3 8 (h,r,ra) 1
Acute
oral 3 (r,m) > 10 (>1 for prioritized)
inhalatory 2 (r,m) 4 (h, fu) 5 (r,gp, ra)
> 10
dermal waived
Skin irritation 5 (h,m,ra,gp) > 10
Eye irritation 3 (ra) > 10
Skin sensitization 3 (h) > 10 (>1 for prioritized)
Repeated oral 7 (f,r,m,s) > 100
inhalatory 1 (h,fu) 3 (x, uf)
Mutagenic / Genotoxic 10 1 (h,x) > 10 (>1 for prioritized)
Carcinogen > 1000
Toxic to reproduction 2 (r) > 100
Developmental 6 (r,ha,m,ra)
Finding (3) The studies registered within the dossier does not include all available and relevant studies concerning the effects of nano ZnO • Over 30 additional studies with nano ZnO were collected with the
following endpoints: dermal penetration (9 studies), tissue distribution (1 study), acute oral (3 study), acute inhalatory (2 studies), repeated inhalatory (6 studies), mutagenicity (8 studies), phototoxicity (1 study), other (4 studies).
• The acute oral mice exposure study (Wang et al., 2008) indicated that
nano ZnO at doses (1g/kg) caused changes in biochemical and enzymatic parameters indicating cardiovascular, hepatic and /or thrombotic effects. The relevance of those findings is not fully understood however this level meets the criteria for STOT (cat 2) classification (oral rat 2000mg≥ C >300) and should be further investigated.
Finding (4)
The registrant questions / dismisses the relevance of the inhalatory studies on the ultrafine ZnO with the argument that the commercial grade ZnO is of much larger particle size and ”can have different toxicological characteristics”
• The experimental, based on the registered study, inhalatory NOAEC = 2.7 mg/m3 was proposed by the registrant but was not used for the derivation of inhalatory DNELs (instead oral study and route to route extrapolation was used)
• Based on all available repeated inhalatory studies with nano ZnO effect levels were
identified: LOAEC = 0.55 mg/m3 for 5 days (BASF, 2009), EL 5 mg/m3 3h per day for 6 days (Lam et al., 1985), 5.9 mg/m3 3h/day for 1-3 days (Conner et al., 1988)
• In general the effect level of nano ZnO in inhalatory studies have been observed at
concentrations similar to or much below OSHA PEL 5 mg/m3 (ZnO fumes and dust) 8h/day and 40 h/ week.
• The human evidence indicated no effects after 14 mg /m3 for 8h
Finding (5)
Registrant derives DNELs for different exposure routes based on extrapolation from oral dietary human study (study not registered!) (see table next slide)
• Critical study on human volunteers with diet supplementation with zinc gluconate NOAEL 50 mg zinc / day (0.83 mg/kg bw/day)
• For extrapolation from this study to the general population or workers overall AF = 1 was used by the registrant
• The route to route extrapolation for inhalatory and dermal DNELs was used using the estimates of different bioavailability after different exposure routes and dissolution rates for different zinc compounds (oral bioavailability soluble compound 20%, dermal 2% or 0.2%, inhalatory 40% or 20%.
endpoint Type of exposure for consumers (c) and workers (w)
Zinc compound NOEL Corrected external dose descriptor for systemic exposure
AF Endpoint-specific DNEL
Consumers/workers
Acute exposure – systemic and local effects
Oral (c) Long-term systemic oral / dermal / inhalation DNEL considered sufficient to ensure that theses effects do not occur
-
Dermal (w,c)
Inhalation (w,c)
Consumers/workers
Acute exposure – systemic and local effects
Dermal (w,c) Long-term systemic oral / dermal / inhalation DNEL considered sufficient to ensure that theses effects do not occur
-
Inhalation (w,c)
Consumers Repeated dose toxicity
Oral soluble
50
mg
Zn
/ d
ay
0.8
3 m
g /k
g b
w/
day
Not required 1 50 mg Zn / day 0.83 mg /kg bw/ day
insoluble Not required 50 mg Zn / day 0.83 mg /kg bw/ day
Dermal soluble 500 mg Zn / day 8.3 mg /kg bw/ day
500 mg Zn / day 8.3 mg /kg bw/ day
insoluble 5000 mg Zn / day 83 mg /kg bw/ day
5000 mg Zn / day 83 mg /kg bw/ day
Inhalation soluble 1.3 mg Zn / m3 1.3 mg Zn / m3
insoluble 2.5 mg Zn / m3 2.5 mg Zn / m3
Workers Repeated dose toxicity
Oral soluble 5
0 m
g Z
n /
day
0
.83
mg
/kg
bw
/ d
ay
Not required 1 50 mg Zn / day 0.83 mg /kg bw/ day
insoluble Not required 50 mg Zn / day 0.83 mg /kg bw/ day
Dermal soluble 500 mg Zn / day 8.3 mg /kg bw/ day
500 mg Zn / day 8.3 mg /kg bw/ day
insoluble 5000 mg Zn / day 83 mg /kg bw/ day
5000 mg Zn / day 83 mg /kg bw/ day
Inhalation soluble 2.5 mg Zn / m3 2.5 mg Zn / m3
insoluble 5 mg Zn / m3 5 mg Zn / m3
Finding (6)
Several assumptions were used by the registrant within RA.
1. The inhalatory studies on ultra-fine ZnO considered not relevant for the commercial uses
2. Bioaccumulation was considered not relevant due to homeostatic control considered typical for essential element
3. Zn ions were considered the only active component of all zinc compounds, and the dissolution rate determined the bioavailability.
4. Oral absorption has been assessed by the registrant (dependent on the solubility of compound) to be 12-20%. No consideration of increased bioavailability due to particle size has been given.
Discussion (1)
The inhalatory studies on ultra-fine ZnO were considered by the registrant not relevant for the commercial uses
• My personal view: The relevance of the inhalatory studies on nano ZnO can not be rule out for some scenarios.
• Nano ZnO based products that could potentially result in inhalatory exposure are already on the market (sunscreen sprays, deodorant, shaving sprays) and a new types of products based on the anti UV, antibacterial, antifungal or water-resistence properties could potentially result in other spray type products for treatment of surfaces, clothes, etc.
Discussion (2)
Bioaccumulation was considered not relevant due to homeostatic control considered typical for essential element
Figure 2. Cellular zinc homeostasis is mediated by three main mechanisms (Plum, Rink, & Haase, 2010).
• My personal view: This can be questioned with respect to nano ZnO
• The commonly recognized homeostatic mechanism builds on the transport of Zn2+ through the plasma membrane by protein importers and vesicular sequestration by export proteins (Plum et al., 2010)
• This does not cover the uptake of nano ZnO particles. The active uptake of particulate forms followed by the rapid dissolution of Zn2+ promoted by the acidic pH of cellular compartments has been reported (Muller et al, 2010)
Discussion (3)
Zn2+ considered by the registrant the only active component of all zinc compounds, and the dissolution rate determine the bioavailability and biological activity
• Again this assumption does not consider the particulate cellular uptake
• Some evidence suggest that for nano ZnO biological activity may not be solely based on externally released Zn2+ e.g.:
Study indicated that for toxicity in human colon cancer cells, the direct particle – cell contact was required (Moos et al., 2010) Study on macrophage response to ZnO nanorods indicated that cell adhesion and viability correlated to both nanotopography and dissolved Zn ions (Zaveri et al., 2010) Gene expression study on Daphnia (Poynton et al., 2011) suggest nano specific gene expression profile
Discussion (4)
Oral absorption has been assessed by the registrant (dependent on the solubility of compound) to be 12-20%. No consideration of increased bioavailability due to particle size has been given.
• My personal view: this can be questioned.
• The patent application on the nano ZnO supplementation of feed stuff for piglets indicated that the use of nano ZnO for this application instead of soluble compounds results in reduction of dose about 10-30 times (suggesting increased bioavailability?)
Summary and personal reflection • It appears that nanoZnO has been considered as the same substance as
bulk and registered in the same dossier (nanoZnO producers were represented in the consortium and examples of studies with nanoZnO were registered)
• However only scarce number of nanoZnO studies were registered with endpoint limited to inhalatory toxicity. Not all available study on nano ZnO were registered.
• The inhalatory studies on nano ZnO were considered not relevant for the commercially relevant ZnO and the hazard assessment, derivation of DNELs for different exposure routes (inhalatory, dermal, oral) was based on the human dietary exposure study to soluble zinc gluconate, with the AF = 1, and the route to route extrapolation based on the assumed differences in bioavailability. No consideration to nano differences in bioavailability.
• Several assumptions were used in hazard, exposure, risk assessment without considerations of the relevance / appropriateness to “nano”
• Personal view: The outcome of the registration of data in this case does
not support the motion that the “registration approach of nano should be left to the registrant” as it is far from being comprehensive and transparent when it comes to RA of nano ZnO
RA for inhalatory exposure to nano ZnO in spray preparations
My personal view: It should be recommended to avoid indoors use of spray applications containing high concentration (25%) of nano ZnO, until targeted to this application RA has been performed
• The ConsExpo modeling of inhalatory exposure to nano ZnO following 1 min use of the spray application containing 25% of nano ZnO in a small room with a middle ventilation rate resulted in high picks of estimated concentrations in a range of 100 mg/m3 of ZnO
• It is likely that this value is overestimated due to the conservative character of the model, and experimental confirmation would be desired
• This value exceeds the DNELs for consumers ca 40 times (DNEL 2.5 mg/m3) and is in a range of levels associated with health effects in human short occupational exposure.
