A mixture of: trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-(4-nitro-2-sulfonatoanilino)phenylazo)phenolato)ferrate(1-); trisodium bis(2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-nitro-2-sulfonatophenylazo)phenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(3-sulfonatophenylazo)phenolato)ferrate(1-); disodium 3,3'-(2,4-dihydroxy-1,3(or 1,5 or 3,5)-phenylenediazo)dibenzenesulfonate

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

From September 03, 1990 to November 12, 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

1983

Deviations:

no

GLP compliance:

no

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

A mixture of: trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-(4-nitro-2-sulfonatoanilino)phenylazo)phenolato)ferrate(1-); trisodium bis(2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-nitro-2-sulfonatophenylazo)phenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(3-sulfonatophenylazo)phenolato)ferrate(1-); disodium 3,3'-(2,4-dihydroxy-1,3(or 1,5 or 3,5)-phenylenediazo)dibenzenesulfonate

EC Number:

406-870-1

EC Name:

A mixture of: trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-(4-nitro-2-sulfonatoanilino)phenylazo)phenolato)ferrate(1-); trisodium bis(2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-nitro-2-sulfonatophenylazo)phenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(3-sulfonatophenylazo)phenolato)ferrate(1-); disodium 3,3'-(2,4-dihydroxy-1,3(or 1,5 or 3,5)-phenylenediazo)dibenzenesulfonate

Cas Number:

115100-55-5

Molecular formula:

not applicable

IUPAC Name:

triiron(3+) nonasodium 2-(2-{4-hydroxy-2-oxido-5-[2-(3-sulfophenyl)diazen-1-yl]phenyl}diazen-1-yl)-4,6-dinitrobenzen-1-olate 2-(2-{5-[2-(3,5-dinitro-2-oxidophenyl)diazen-1-yl]-2-hydroxy-4-oxidophenyl}diazen-1-yl)-4,6-dinitrobenzen-1-olate tris(2-(2-{5-[2-(3,5-dinitro-2-oxidophenyl)diazen-1-yl]-4-hydroxy-2-oxidophenyl}diazen-1-yl)-4,6-dinitrobenzen-1-olate) 2-{2-[4-hydroxy-5-(2-{4-[(4-nitro-2-sulfophenyl)amino]phenyl}diazen-1-yl)-2-oxidophenyl]diazen-1-yl}-4,6-dinitrobenzen-1-olate

Test material form:

solid: particulate/powder

Test animals

Species:

mouse

Strain:

NMRI

Details on species / strain selection:

The mouse is an animal which has been used for many years as suitable experimental animal in cytogenetic investigations. There are many data available from such investigations which may be
helpful in the interpretation of results from the micronucleus test. In addition, the mouse is an experimental animal in many physiological, pharmacological and toxicological studies. Data from such experiment also may be useful for the design and the performance of the micronucleus test.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: BRL Tierfarm Füllinsdorf
- Age at study initiation: mínimum 10 weeks
- Weight at study initiation: ~ 30 g
- Acclimation period: mínimum 5 days
- Fasting period before study: Approximately 18 hours before treatment
- Housing: single, Makrolon Type I, with wire mesh top (EHRET GmbH, D-7830 Emmendingen)
- Diet (e.g. ad libitum): pelleted standard diet (ALTROMIN, D-4937 Lage/Lippe), ad libitum.
- Water (e.g. ad libitum): tap water, ad libitum (Gemeindewerke, D-6101 Roßdorf)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 3 °C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m, - 6.00 p.m.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: water
- Concentration of test material in vehicle: 4000 mg/kg b.w.
- Amount of vehicle (if gavage or dermal): 20 ml/kg b.w.

Duration of treatment / exposure:

Single injection//Sampling of the bone marrow was done 24, 48 and 72 hours after treatment.

Post exposure period:

72 hours after treatment.

Doses / concentrations

Dose / conc.:

4 000 mg/kg bw/day

No. of animals per sex per dose:

Test article: 5 per sex per dose per sampling time (24/48/72 h) = 15 ♂ + 15 ♀
Negative control: 5 per sex per dose per sampling time (24/48/72 h)= 15 ♂ + 15 ♀
Positive control: 5 per sex per sampling time (24 h) = 5 ♂ + 5 ♀

Control animals:

yes, concurrent vehicle

Positive control(s):

Name: CPA; Cyclophosphamide
Dose: 40 mg/kg b.w
Route: orally
Frequency: single injection
Volume Administered: 10 ml/kg b.w.
Solution prepared on day of administration. The stability of CPA at room temperature is good. At 20°C only 1 % of CPA is hydrolysed per day in aqueous solution.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Any other information on results incl. tables

VALIDITY CRITERIA FULFILLED:

A test article producing neither a statistically significant dose-related increase in the number of micronucleated polychromatic erythrocytes nor a statistically significant and reproducible positive response at anyone of the test points is considered non-mutagenic in this system.

Applicant's summary and conclusion

Conclusions:

During the study described and under the experimental conditions reported, the test article did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse.

Executive summary:

This study was performed to investigate the potential of test substance to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse, according to OECD Guideline 474 (1983). The test article was dissolved in water. This solvent was used as negative control. The volume administered orally was 20 ml/kg b.w. 24 h, 48 h and 72 h after a single application of the test article the bone marrow cells were collected for micronuclei analysis. Ten animals (5 males, 5 females) per test group were evaluated for the occurrence of micronuclei. 1000 polychromatic erythrocytes (PCE) per animal were scored for micronuclei. To describe a cytotoxic effect due to the treatment with the test article the ratio between polychromatic and normochromatic erythrocytes (NCE) was determined in the same sample and reported as the number of NCE per 1000 PCE.

The following dose level of the test article was investigated: 24 h, 48 h, and 72 h preparation interval: 4000 mg/kg b.w.

In pre-experiments this dose level was estimated to be the maximum tolerated dose. The animals expressed toxic reactions. After treatment with the test article the ratio between PCEs and NCEs was not affected as compared to the corresponding negative controls thus indicating no cytotoxic effects.

In comparison to the corresponding negative controls there was no significant enhancement in the frequency of the detected micronuclei at any preparation interval after application of the test article.

An appropriate reference mutagen was used as positive control which showed a distinct increase of induced micronucleus frequency .

In conclusion, it can be stated that during the study described and under the experimental conditions reported, the test article did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse. Therefore, the substance is considered to be non-mutagenic in this micronucleus assay.