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Toxicological information

Acute Toxicity: other routes

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Administrative data

Endpoint:
acute toxicity: other routes
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Meets generally accepted scientific standards, well documented, GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
Freshly isolated primary rat alveolar macrophages were treated for 24 hours with coated nanoscaled ZnO (Z-COTE HP1), non-coated nanoscaled ZnO (Z-COTE) and microscaled ZnO as well as the reference compounds Aluminium oxide, Silicon dioxide DQ12 and Zymosan. ROS formation was measured using a chemiluminescence-based assay while cytotoxicity and the production of pro-inflammatory cytokines were determined by ELISA systems.
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
TEST ITEM
- Name of test material (as cited in study report): Z-COTE HP1
- Molecular weight (if other than submission substance): 81.38g/mol
- Physical state: solid
- Composition of test material, percentage of components: Z-COTE HP1 (98%), coated with triethoxycaprylylsilane (CAS # 2943-75-1; 2%)
- Lot/batch No.: NPL Ref#: ZB250#65
- Expiration date of the lot/batch: June 2014
- Storage condition of test material: room temperature, dry, exclusion of light
- Other: MMAD of the aerosol < 3.0 µm, GSD about 1.5

NANOSCALED REFERENCE ITEM
- Name of test material (as cited in study report): Z-COTE
- Molecular weight (if other than submission substance): 81.38g/mol
- Physical state: solid
- Composition of test material: no coating of surface
- Lot/batch No.: NPL Ref#: ZC250#32
- Expiration date of the lot/batch: June 2014
- Storage condition of test material: room temperature, dry, exclusion of light
- Other: MMAD of the aerosol < 3.0 µm, GSD about 1.5

MICROSCALED REFERENCE ITEM
- Name of test material (as cited in study report): Zinc Oxide 205532, Micron Size Powder
- Molecular weight (if other than submission substance): 81.38g/mol
- Physical state: solid
- Composition of test material, percentage of components: non-coated ZnO
- Lot/batch No.: NPL Ref#: ZrA250#60
- Expiration date of the lot/batch: May 2014
- Storage condition of test material: room temperature, dry, exclusion of light
- Other: MMAD of the aerosol < 3.0 µm, GSD about 1.5

Test animals

Species:
other: in vitro, primary rat alveolar macrophages
Strain:
Wistar
Sex:
male
Details on test animals and environmental conditions:
in vitro Test

Administration / exposure

Route of administration:
other: in vitro
Vehicle:
not specified
Details on exposure:
in vitro test, 24 h exposure
Doses:
15, 150, 1500, and 15000 µg/ml
No. of animals per sex per dose:
in vitro test
Control animals:
other: technical replicates as concurrent positive and negative controls
Details on study design:
Fresh primary alveolar macrophages were isolated via ex vivo bronchoalveolar lavage of 12 weeks old untreated Wistar rats. Cells were recovered by centrifugation and seeded in 96 well plates with a density of 2x 105cells per well. The cells were treated for 24 hours with ready-to-use suspensions of nanoscaled coated ZnO (Z-COTE HP1), nanoscaled non-coated ZnO (Z-COTE), and microscaled non-coated ZnO. Further technical replicates were treated for 24 hours with the reference substances Aluminium oxide and Silicon dioxide as well as the positive controls Lipopopolysaccharide (LPS) and Zymosan. The release of reactive oxygen species (ROS) was measured with a lucigenin-based chemiluminescence assay using a kinetic method over a period of 30 minutes. LDH release of the treated cells was determined to assess cell viability using a photometrical assay based on the conversion of a tetrazolium salt to formazan. Furthermore the release of cytokines (IL-6, TNFα, CINC-1) and prostaglandin E2 was measured by ELISA.
Statistics:
Statistical analysis was performed by non-parametric Dunnett test (Software: GraphPad Prism 4, Version 4.03). Diferences between exposed samples and controls were considered as statistically significant at the level of p<0.05.

Results and discussion

Effect levelsopen allclose all
Remarks on result:
other: Silicium dioxide but not Z-COTE HP1, Z-COTE, microscaled ZnO and Aluminium oxide induced ROS production of alveolar macrophages in vitro.
Remarks on result:
other: Aluminium oxide and Silicium dioxide induced cellular production of IL-6 and CINC-1, respectively, while Z-COTE HP1 and Z-COTE did not.
Remarks on result:
other: Cellular production of prostaglandin E2 was increased by the highest concentrations used for Z-COTE HP1, Z-COTE and microscaled ZnO. These concentrations were also cytotoxic.
Mortality:
in vitro test
Clinical signs:
in vitro test
Body weight:
in vitro test
Gross pathology:
in vitro test
Other findings:
Exposure of rat alveolar macrophages to Z-COTE HP1, Z-COTE, microscaled ZnO as well as Aluminium oxide did not result in a significant increase in of ROS while Silicium dioxide slightly increased ROS production of the treated macrophages.Furthermore Z-COTE HP1, Z-COTE, microscaled ZnO
did not affect or decrease cytokine production while Aluminium oxide induced IL-6 and Silicium dioxide CINC-1 production. The highest concentration of all ZnO particles tested during this study induced the production of prostaglandin E2 but were also cytotoxic to the treated cells.

Any other information on results incl. tables

Exposure of rat alveolar macrophages to Z-COTE HP1, Z-COTE, microscaled ZnO as well as Aluminium oxide did not result in a significant increase in of ROS while Silicium dioxide slightly increased ROS production of the treated macrophages.Furthermore Z-COTE HP1, Z-COTE, microscaled ZnO

did not affect or decrease cytokine production while Aluminium oxide induced IL-6 and Silicium dioxide CINC-1 production. The highest concentration of all ZnO particles tested during this study induced the production of prostaglandin E2 but were also cytotoxic to the treated cells.

Applicant's summary and conclusion

Executive summary:

Exposure of rat alveolar macrophages to Z-COTE HP1, Z-COTE, microscaled ZnO as well as Aluminium oxide did not result in a significant increase in of ROS while Silicium dioxide slightly increased ROS production of the treated macrophages.Furthermore Z-COTE HP1, Z-COTE, microscaled ZnO

did not affect or decrease cytokine production while Aluminium oxide induced IL-6 and Silicium dioxide CINC-1 production. The highest concentration of all ZnO particles tested during this study induced the production of prostaglandin E2 but were also cytotoxic to the treated cells.