Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test compound is neither mutagenic in bacterial nor in mammalian cells.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
equivalent or similar to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Version / remarks:
84/449/EWG, B.13, B.14 (Standard Platten-Test und Preincubations-Test)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
bacteria, other: TA1535, TA1537, TA1538, TA98, TA100, E.coli WP2uvrA
Metabolic activation:
with and without
Metabolic activation system:
S9-Mix from Rats and Hamster liver
Test concentrations with justification for top dose:
Concentration range in the first test (with and without metabolic activation): 4, 20, 100, 500, 2500 and 5000 µg/plate
Concentration range in the second test (with and without metabolic activation): 4, 20, 100, 500, 2500 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: none
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)
Species / strain:
bacteria, other: TA1535, TA1537, TA1538, TA98, TA100, E.coli WP2uvrA
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
up to 10'000 ug/plate
Vehicle controls validity:
not applicable
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'. Remarks: First experiment
Conclusions:
negative with metabolic activation
negative without metabolic activation

negative
Executive summary:

Reaktiv-Rot F-52 167 was tested for mutagenicity with the strains TA100, TA1535, TA1537, TA1538, TA98 of salmonella typhimurium and Escherichia coli WP2uvrA.

The mutagenicity studies were conducted in the absence and in the presence of a metabolizing system derived from rat liver homogenate. A dose range of 6 different dosees from 4 µg/plate to 5000 µg/plate was used.

Control plates without mutagen showed that the number of spontaneous revertant colonies was similar to that described in the literature. All the positiv control compounds gave the expected increase in the number of revertant colonies.

Toxicity: the test compound proved to be not toxic o f the bacterial strains. On the basis of the preliminary test results the top dose level did no exceed 5000 µg/plate.

Mutagenicity: In the absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Also in the presence of a metabolic activation system, treatment of the cells with Reaktiv-Rot F-52 167 did no result in relevant increases in the number of revertant colonies.

Sumarizing, it can be stated that Reaktiv-Rot F-52 167 is not mutagenic in these bacterial test systems niether with or without exogenous metabolic activation at the dose levels investigated.

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1989
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to
Guideline:
EPA OTS 798.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Metabolic activation system:
S9-Mix from rat and hamster liver
Test concentrations with justification for top dose:
Concentration range in the first test (with and without metabolic activation): 4, 20, 100, 500, 2500, 10000 µg/plate
Concentration range in the second test (with and without metabolic activation): 4, 20, 100, 500, 2500, 10000 µg/plate
Vehicle / solvent:
none
Details on test system and experimental conditions:
According to the modification proposed by Prival using 30 minutes preincubation in the presence of 30% hamster S-9 (Syrien golden hamster)
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
up to 10000 µg/plate
Vehicle controls validity:
not applicable
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'. Remarks: First experiment
Conclusions:
The test compound did not cause a significant increase in the number of revertant colonies with any of the tester strains either in the absence or in the presence of S9 Mix. No dose dependent effect was obtained.
Executive summary:

Reaktiv-Rot F-52 167 FW was tested for mutagenicity with the strains TA 100, TA 1535, TA 1537 and TA 98 of Salmonella typhimurium.

The mutagenicity studies were conducted in the standard plate test (Ames Test) and in a modified preincubation test (Prival Test). The studies were performed in the absence and in the presence of a metabolizing system derived from rat or hamster liver homogenate. A dose range of 6 different doses from 4 to 5000 µg/plate was used.

Control plates without mutagen showed that the number of spontaneous revertant colonies was similar to that described in the literature. All the positive control compounds gave the expected increase in the number of revertant colonies.

Toxicity: The test compound proved to be not toxic to the bacterial strains. 5000 µg/plate was chosen as top dose level for the mutagenicity study.

Ames Test:

Mutagenicity: In the absence of the metabolic activation system the test com-pound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Also, in the presence of a metabolic activation system, treatment of the cells with Reaktiv-Rot F-52 167 FW did not result in relevant increases in the number of revertant colonies.

Prival Test:

In the presence of hamster liver S-9 using the preincubation method according to Prival Reaktiv-Rot F-52 167 FW did not induce a significant increase in the number of revertant colonies, with any of the tester strains.

Summarizing, it can be stated that Reaktiv-Rot F-52 167 FW is not mutagenic in the standard plate test (Ames Test) and in the preincubation method according to Prival.

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From Nov. 17, 1999 to Apr. 10, 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test
Deviations:
no
Qualifier:
according to
Guideline:
other: EEC Directive 87/302, L133, pp. 61 - 63, March 1987
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Target gene:
HRPT locus (V79 Chinese hamster cells)

Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
Source: cell bank of "Genetic Toxicology", Aventis Pharma Deutschland GmbH, ProTox
Cell culture medium: MEM (minimal essential medium) with Hankssalts and 25 mM Hepes-buffer
pH values and osmolality of the treatment media: Determined before treatment
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9-mix
Test concentrations with justification for top dose:
Main mutation experiment (First):
- without S9-mix: 100, 250, 500, 1000, 1750, 2500, 3750 and 5000* µg/mL
- with S9-mix: 100, 250, 500, 1000, 1750, 2500, 3750 and 5000 µg/mL

* = because of high toxicity in the first main experiment no mutant selection was performed

Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle:
Untreated negative controls:
yes
Remarks:
(Untreated controls)
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
(without metabolic activation )
Positive control substance:
ethylmethanesulphonate
Remarks:
None Migrated to IUCLID6: (1.0 mg/mL = 8 mM)
Untreated negative controls:
yes
Remarks:
(Untreated controls)
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Remarks:
(with metabolic activation )
Positive control substance:
9,10-dimethylbenzanthracene
Remarks:
None Migrated to IUCLID6: (7.7 µg/mL = 30 µM)
Details on test system and experimental conditions:
METHOD OF APPLICATION: Seeded in each well of a microtiter plate

DURATION (Preliminary toxicity)
- Preincubation period: 12 h (overnight)
- Exposure duration: 4 h
- Incubation period: 24 h
- Fixation and staining: Crystal violet
NUMBER OF REPLICATIONS: 6 (For each concentration at least 6 wells)

DURATION (Main experiment)
- Preincubation period: 24 h
- Exposure duration: 4 h (Medium+ test substance+ buffer/S9-mix)
- Incubation period: 24 h
- Fixation and staining: Crystal violet

Evaluation criteria:
The test item was considered positive if
(a) It reproducibly induces with one of the test compound concentrations a mutation frequency that is three times higher than the spontaneous mutant frequency in this experiment.
(b) There is a reproducible concentration-related increase in the mutation frequency. Such an evaluation may be considered independently from the enhancement factor for induced mutants.
(c) Survival of the responding dose group is at least 30 %.
However, in a case by case evaluation both decisions depend on the level of the corresponding negative control data.

Statistics:
- The biometry of the results for the test compound is performed off-line with the Mann-Whitney- U-Test

Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No effects
- Effects of osmolality: No effects


RANGE-FINDING/SCREENING STUDIES:
Evaluation of the solubility of the suspension in cell culture medium showed that 5000 µg/mL was the highest concentration at which no visible precipitation was observed.

Preliminary toxicity study: Was carried out using a maximum concentration of 5000 µg/mL and a range of lower dose levels down to 10 µg/mL.

Based on the preliminary study results, 5000 µg/mL was selected as the maximum dose level for the main mutation experiments in both the absence and in the presence of S9-mix. Seven lower concentrations down to 100 µg/mL were also included.

COMPARISON WITH HISTORICAL CONTROL DATA: Yes

Remarks on result:
other: other: V79 Chinese hamster cell line
Remarks:
Migrated from field 'Test system'.

The sensitivity of the test system and efficacy of the S9-mix was demonstrated by the enhanced mutation frequency in the cell cultures treated with the positive control substances.

Mutation results:

Main experiment: A significant increase of the mutation frequency was observed in the presence of metabolic activation at the concentrations of 250, 2500 and 3750 µg/mL.

These results were not reproducible and not three fold higher than the corresponding controls and were therefore considered to be of no biological relevance.

No relevant reproducible increase in the mutant colonies or mutant frequency over the range of the solvent control was found with any other concentration, either with or without metabolic activation by S9-mix.

Conclusions:
Under the test conditions, the test substance is considered to be non-mutagenic in the HPRT-test with V79 Chinese hamster cells, either in the presence or in the absence of metabolic activation.
Executive summary:

The study was performed to investigate the potential of test substance to induce gene mutations at the HPRT locus in V 79 cells of the Chinese hamster in vitro according to OECD Guideline 476, EPA OPPTS 870.5300 and EEC Directive 87/302, L133 in compliance with GLP.

Two independent experiments were conducted both with and without an exogenous rat liver microsomal activation system (S9-mix). The test substance was dissolved in cell culture medium and tested at the following concentrations:

- without S9-mix: - without S9-mix: 100, 250, 500, 1000, 1750, 2500, 3750 and 5000 µg/mL

- with S9-mix: 100, 250, 500, 1000, 1750, 2500, 3750 and 5000 µg/mL

Due to the limitation of the OECD guideline 5000 µg/mL is the highest tolerated dose.

The concentration ranges were based on the results of preliminary tests for solubility and toxicity. The highest concentration showed toxic effects with and without metabolic activation.

Test substance produced no macroscopical precipitation. Up to the highest investigated dose no relevant and reproducible increase in mutant colony numbers was obtained in two independent experiments.

Appropriate reference mutagens used as positive controls showed a distinct increase in induced mutant colonies, thus indicating the sensitivity of the assay and the efficacy of the S9-mix.

Test substance does not induce gene mutations in the HPRT-test with V79 Chinese hamster cells, both in the presence as well as in the absence of a metabolic activation system.

Under the test conditions, the test substance is considered to be non-mutagenic in the HPRT-test with V79 Chinese hamster cells, either in the presence or absence of metabolic activation.

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
August 1999 to November 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
Version / remarks:
EEC Directive 87/302
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Target gene:
HPRT
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Details on mammalian cell type (if applicable):
Stocks of the CHO cell line (1-ml portions) were maintained at -196°C in liquid nitrogen using 7% DMSO in culture medium as a cryoprotectant. Each batch used for muta¬genicity testing was checked for mycoplasma contamination.
Additional strain / cell type characteristics:
other: substrain K1
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Test concentrations with justification for top dose:
According to an initial range-finding cytotoxicity test for the determination of the experimental doses and taking into account the cytotoxicity actually found in the main experiment, the following doses were evaluated in the 1st experiment after an exposure period of 4 hours:
1st experiment
without S-9 mix
0; 156.25; 312.5; 625; 1,250; 2,500; 5,000 μg/ml

with S-9 mix (S-9 fraction : cofactors = 3:7)
0; 156.25; 312.5; 625; 1,250; 2,500; 5,000 μg/ml

2nd experiment

In a 2nd experiment for confirmation of the results of the 1 st experiment, the following doses were evaluated after an exposure period of 4 hours with and without metabolic activation;

without S-9 mix
0; 156.25; 312.5; 625; 1,250; 2,500; 5,000 μg/ml

with S-9 mix (S-9 fraction : cofactors = 1 : 9)
0; 156.25; 312.5; 625; 1,250; 2,500; 5,000 μg/ml

Test substance precipitation in the culture medium was not observed.
Vehicle / solvent:
Culture media
Ham's F12 medium
Ham's F12 medium with glutamine and hypoxanthine supplemented with

- 10% (v/v) fetal calf serum (FCS)

During exposure to the test substance, Ham's F12 medium was used without FCS supplementation. In the case of continuous treatment Ham's F12 medium with FCS supplementation was used.

Pretreatment medium ("HAT medium")

FCS-supplemented Ham's F12 medium with glutamine and hypoxanthine containing per ml
- Hypoxanthine (13.6 x 10-3 mg)
- Aminopterin (0.18 x 10-3mg)
- Thymidine (3.88 x 10-3 mg)'

Selection medium ("TG medium")

Glutamine- and FCS-supplemented, hypoxanthine-free Ham's F12 medium with - 6-thioguanine at a final concentration of 10 ug/ml

All media were supplemented with

- 1% (v/v) penicillin/streptomycin (10,000 IU / 10,000 ug/ml)
- 1% (v/v) amphotericine B (250 ug/ml)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: x2 +ve control substances
Remarks:
+ve control substances detailed below
Details on test system and experimental conditions:
Preparation of test cultures: Logarithmically growing cells (after the 3rd passages both in the 1st and 2nd experiments respectively) in monolayers were trypsinized (2 ml Ca-Mg-free Hanks' balanced salt solution (HBSS) + 2 ml of a 0.25% trypsin solution) for each subculture. Optionally, prior to trypsin treatment, the cells were rinsed once with 5 ml Ca-Mg-free Phosphate buffered salt solution (PBS). This process was stopped by adding 6 ml Ham's F12 medium supplemented with 10% FCS.
A single cell suspension was prepared, and about 300,000 or 500,000 cells were seeded in 75-cm2 flasks (approx, 10 ml for 75-cm2 flasks) for the mutagenicity test. For the determination of the cloning efficiency, about 200 cells were seeded in 25-cm2 flasks and covered with medium (approx. 5 ml for 25-cm2 flasks). The flasks were incubated with 5% C02 at 37°C and > 90% humidity. 2 subcultures were prepared per week,

Pretreatment of cells: During the week prior to treatment, spontaneous HPRT-deficient mutants were eliminated from the stock cultures by growing the cells for 3 - 4 days in pretreatment medium (HAT medium).

Attachment period: For each test group, about 500,000 cells per flask were seeded into about 10 ml Ham's F12 medium supplemented with 10% FCS and incubated for about 24 hours with 5% C02 at 37°C and > 90% humidity. 2x2 flasks (2 flasks designated as A and two flasks designated as B) were used for each test group. A counter was used for cell counting. The numbers obtained in the untreated controls were cross-checked in a Neubauer chamber, yielding an internal correction factor.

Treatment of test cultures and expression period:
About 24 hours after seeding and incubating the cells, the medium was replaced by fresh medium. The test article, dissolved in 1 ml serum-free medium, was added to the culture medium:
Without S-9 mix
9 ml Ham's F12 medium without FCS and 1 ml test substance preparation
With S-9 mix
7 ml Ham's F12 medium without FCS, 1 ml test substance preparation and 2 ml S-9 mix (= 6% and 2% S-9 fraction in the 1st and 2nd experiment, respectively).

Concurrent negative and positive controls were tested in parallel.

After incubation (5% C02, 37°C and > 90% humidity) for 4 hours both without and with S-9 mix, the serum-free medium was replaced by 10 mf Ham's F12 medium with 10% FCS after having been rinsed twice with Hanks' balanced salt solution (HBSS). Subsequently, the flasks were incubated for another 17 - 24 hours; two flasks were pooled in each case and then subcultured (1st passage). After two further passages (duration of the expression period is about 1 week), cells were transferred into selection medium (TG medium) at the 4th passage.
Evaluation criteria:
The criteria for a positive response are:
Increases of the corrected mutation frequencies above the concurrent negative control values and above 15 mutants per 106 clonable cells and/or the evidence of a dose-response relationship in the increase in mutant frequencies.
Evidence of reproducibility of any increase in mutant frequencies.
A statistically significant increase in mutant frequencies and the evidence of a dose-response relationship.

Isolated increases of mutant frequencies above 15 mutants per 106 clonable ceils or isolated statistically significant increases without a dose-response relationship may indicate a biological effect but are not regarded as sufficient evidence of mutagenicity.

The test chemical is considered to be non-mutagenic according to the following criteria.
The corrected mutation frequency in all dose groups is within the historical control range and is not significantly above the concurrent negative control.
Statistics:
Due to the clearly negative findings, a statistical evaluation was not carried out.
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
According to the results of the present in vitro study, the test substance did not lead to an increase in the number of mutant colonies either without S-9 mix or after the addition of a metabolizing system in two experiments performed independently of each other. The mutant frequencies at any dose were nearly the range of that of the concurrent negative control values and within the range of the historical control data.

The increase in the frequencies of mutant colonies induced by the positive control agents EMS and MCA clearly demonstrated the sensitivity of the test method and of the metabolic activity of the S-9 mix employed.
Conclusions:
Under the experimental conditions chosen here, the substance does not induce forward mutations in vitro in the CHO/HPRT assay.
Executive summary:

Study conducted to EU test guidance 97/302/EEC and OECD test guideline 476 in compliance with GLP.

According to the results of the present in vitro study, the test substance did not lead to an increase in the number of mutant colonies either without S-9 mix or after the addition of a metabolizing system in two experiments performed independently of each other. The mutant frequencies at any dose were nearly the range of that of the concurrent negative control values and within the range of the historical control data.

The increase in the frequencies of mutant colonies induced by the positive control agents EMS and MCA clearly demonstrated the sensitivity of the test method and of the metabolic activity of the S-9 mix employed.

Under the experimental conditions chosen here, the substance does not induce forward mutations in vitro in the CHO/HPRT assay.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

The test substance did not cause any adverse effects in the mouse micronucleus test.

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
Version / remarks:
84/449/EWG, B.12
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
micronucleus assay
Species:
mouse
Strain:
NMRI
Sex:
male/female
Route of administration:
oral: gavage
Vehicle:
deionised water
Duration of treatment / exposure:
animals were killed after 24, 48 or 72 hours after administration of the test compound
Frequency of treatment:
The test compound was given in two equal parts within two hours
Remarks:
Doses / Concentrations:
6250 mg/kg
Basis:
nominal in water
No. of animals per sex per dose:
Male: 6250 mg/kg; No. of animals: 5; Sacrifice time: 24 hours
Male: 6250 mg/kg; No. of animals: 5; Sacrifice time: 48 hours
Male: 6250 mg/kg; No. of animals: 5; Sacrifice time: 72 hours
Female: 6250 mg/kg; No. of animals: 5; Sacrifice times: 24 hours
Female: 6250 mg/kg; No. of animals: 5; Sacrifice times: 48 hours
Female: 6250 mg/kg; No. of animals: 5; Sacrifice times: 72 hours
Positive control(s):
endoxane, 50 mg/kg BW
Additional information on results:
The incidence of micronucleated polychromatic erythrocytes in the dose groups of Reaktiv-Rot F-52167 FW was within the normal range of the negative control groups. No statistically significant increase of micronucleated polychromatic erythrocytes has been observed.
Due to the colour of the test compound urine, faeces and skin were red coloured.
Conclusions:
Interpretation of results (migrated information): negative
not mutagenic
Executive summary:

Reaktiv-Rot F-52 167 FW was tested in the micronucleus test. The test compound was administered orally by gavage to male and female mice. The following doses were tested: 0 and 6250 mg Reaktiv-Rot F-52 167 FW per kg bodyweight. The 6250 mg per kg bodyweight dose level was chosen since a preliminary study had shown it to be the maximal applicable dose.

The test compound was given in two equal parts within two hours and according to the test procedure the animals were killed 24, 48, 72 hours after administration of the test compound.

Endoxan was used as positive control substance and was administered orally at a dose of 50 mg per kg bodyweight.

The incidence of micronucleated polychromatic erythrocytes of the animals treated with Reaktiv-Rot F-52 167 FW was within the normal range of the negative control. The number of normochromatic erythrocytes containing micronuclei was not increased. The ratio of polychromatic/normochromatic erythrocytes in both male and female animals remained unaffected by the treatment with Reaktiv-Rot F-52 167 FW and was statistically not different form the control values.

Endoxan induced in both males and females a marked statistically significant increase in the number of polychromatic cells with micronuclei, indicating the sensitivity of the system. The ratio of polychromatic erythrocytes to normocytes was not changed to a significant extend.

The results indicated that, under the conditions of the present study, Reaktiv-Rot F-52 167 FW is not mutagenic in the micronucleus test.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Reaktiv-Rot F-52 167 FW was tested for mutagenicity with the strains TA100, TA1535, TA1537, TA1538, TA98 of Salmonella typhimurium and Escherichia coli WP2uvrA. Reaktiv-Rot F-52 167 is not mutagenic in these bacterial test systems neither with or without exogenous metabolic activation at the dose levels investigated.

Reaktiv-Rot F-52 167 FW was tested in the micronucleus test. The test compound was administered orally by gavage to male and female mice. The results indicated that, under the conditions of the present study, Reaktiv-Rot F-52 167 FW is not mutagenic in the micronucleus test.

Structural analogues were tested negative in the HPRT assay.

Justification for classification or non-classification