Dialuminium nickel tetraoxide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP - Guideline study, tested with CAS number 1313-99-1. In accordance to the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Nickel monoxide

EC Number:

215-215-7

EC Name:

Nickel monoxide

Cas Number:

1313-99-1

IUPAC Name:

oxonickel

Details on test material:

- Name of test material (as cited in study report): NiO green (N112); Nickel oxide sinter
- Physical state: granules / dark grey
- Composition of test material, percentage of components: nickel oxide: 98%, cobaltous oxide: 1.5%
- Lot/batch No.: not provided by the sponsor
- Stability under test conditions: stable
- Storage condition of test material: at room temperature, closed container

Method

Target gene:

Thymidine kinase (TK)

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

Mammalian microsomal fraction S9 mix

Test concentrations with justification for top dose:

Experiment I (with and without metabolic activation): 1,3,5,6, 7, 8,9, 10 mM
Experiment II (with metabolic activation): 1.5,3.5,4.5,5.5, 7.5, 8.5, 9.5, 10 mM
Experiment II (without metabolic activation): 1,3,5,6, 7, 8, 9, 10 mM

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: [cell culture medium]

Controls

Controls 1

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

vehicle

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Controls 2

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

vehicle

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Controls 3

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

vehicle

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in suspension


DURATION
- Preincubation period: not applicable
- Exposure duration: 4 or 24 hr
- Expression time (cells in growth medium): 48-72 hr
- Selection time (if incubation with a selection agent): 11-14 days


SELECTION AGENT (mutation assays): TFT (Trifluorothymidine)



DETERMINATION OF CYTOTOXICITY
- Method: After the expression period the relative cloning efficiency (RCE; percentage cloning efficiency of the test group in relation to the negative control) of the cells was determined by seeding a statistical number of 1.6 cells/well in two 96-well-plates. The cells were incubated for 6 days at 37°C in a humidified atmosphere with 5% C02. Analysis of the results was based on the number of cultures with cell growth (positive cultures) and/or those without cell growth (negative cultures) compared to the total number of cultures seeded. Relative suspension and total growth
(RSG and RTG; RTG= [RSG x RCE]/100) of the treated cell cultures were
calculated according to the method of Clive and Spector.

Evaluation criteria:

There are several criteria for determining a positive result:
- clear and dose-related increase in the mutant frequency,
- biologically relevant response (at least a 2-fold increase of mutant frequencies related to the comparable negative control values and higher than the historical range of negative controls) for at least one of the dose groups,
- combined with a positive effect in the mutant frequency, an increased occurrence of small colonies (slow growth colonies) indicated by a low large/small colonies ratio (ratio of the clastogenic controls MMS and/or B[a]P with a coefficient of 1.5) is an indication for potential clastogenic effects and/or chromosomal aberrations.

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES: The selection of the concentrations was based on data from the pre-experiment.

ADDITIONAL INFORMATION ON CYTOTOXICITY: No growth inhibition was observed in experiment I and II with and without metabolic activation. In experiment I with metabolic activation the relative total growth (RTG) was 100.09% for the highest concentration (10 mM) evaluated. The highest concentration evaluated without metabolic activation was 10 mM with a
RTG of 115.77 %. In experiment II with metabolic activation the relative total growth (RTG) was 115.04% for the highest concentration (10 mM) evaluated. The highest concentration evaluated without metabolic activation was 10 mM with a RTG of79.90%.

Remarks on result:

other: strain/cell type:

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Toxicity:

No growth inhibition was observed in experiment I and II with and without metabolic activation.

Mutagenicity:

The mutation frequencies found in the groups treated with the test item did not show a biologically relevant increase as compared to the negative controls.

Clastogenicity (small colonies):

All dose groups were considered as not clastogenic.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The study authors concluded that in the described mutagenicity test under the experimental conditions reported, the test item Nickel oxide green (N112) is considered to be non-mutagenic in the mouse lymphoma thymidine kinase locus using the cell line L5178Y.

Executive summary:

BSL Bioservice Scientific Laboratories (BSL) GmbH conducted an in vitro mammalian cell gene mutation assay according to OECD Test #476 – In vitro Mammalian Cell Gene Mutation Test guidelines and using GLP standards. The study was completed on 2008-06-10 and the final report issued on 2008-11-27. The test substance, “Nickel oxide green (sample ID N112)”, was assessed for its potential to induce mutations at the mouse lymphoma thymidine kinase (TK) locus using the cell line L5178Y. The selection of the concentrations was based on data from the pre-experiment. Four sets of experiments were run where cells were exposed to NiO for 4 or 24 hr with or without metabolic activation. EMS and MMS were used as positive controls for studies without metabolic activation, and B[a]P was used in studies with metabolic activation. Exposure to vehicle or solvents alone served as the negative controls. In Experiment I (4 hours, with and without metabolic activation), the test concentrations were: 1, 3, 5, 6, 7, 8, 9, 10 mM. In Experiment II (24 hours, referred to as longterm exposure) with and without metabolic activation, the concentrations were respectively: 1.5, 3.5, 4.5, 5.5, 7.5, 8.5, 9.5, 10 mM and 1, 3, 5, 6, 7, 8, 9, 10 mM.

Findings indicated that i) no significant growth inhibition was observed at any concentrations in Experiments I and II, with and without metabolic activation, ii) mutation frequencies (in all experiments) did not show biologically relevant increases as compared to the negative controls, and iii) all dose groups were considered as not clastogenic based on colony size. The study authors concluded that Nickel oxide green (sample ID N112) was considered to be non-mutagenic in the mouse lymphoma thymidine kinase locus using the cell line L5178Y. STUDY RATED BY AN INDEPENDENT REVIEWER