Registration Dossier

Administrative data

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
Study period:
August - October 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes (incl. certificate)
Remarks:
Harlan Cytotest Cell Research GmbH, 64380 Rossdorf, Germany
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): N-(3-Aminopropyl) imidazol
- Physical state: liquide
- Analytical purity: 99.2 %
- Lot/batch No.: O 2894

Test animals

Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Sulzfeld, Germany
- Age at study initiation: 8 - 9 weeks
- Weight at study initiation: mean value 35.6 g (SD+/_ 1.9 g)
- Assigned to test groups randomly: yes
- Housing: single
- Diet (e.g. ad libitum): pelleted standard diet, ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 45 - 110 %
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: sterile water

Duration of treatment / exposure:
24 and 48 hours
Frequency of treatment:
once
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
1500, 750 and 375 mg/kg (24h)
Basis:
nominal in water
Remarks:
Doses / Concentrations:
1500 mg/kg (48h)
Basis:
nominal in water
No. of animals per sex per dose:
7 animals (except the vehicle and positive control grous with 5 males only)
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide (CPA)

Examinations

Tissues and cell types examined:
Bone marrow cells were collected for micronuclei analysis.
Evaluation criteria:
A test item is classified as mutagenic if it induces either a dose-related increase or a clear increase in the number of micronucleated polychromatic erythrocytes in a single dose group.
Statistics:
Statistical methods (nonparametric Mann-Whitney test (8)) are used as an aid in evaluating the results, if necessary. However, the primary point of consideration is the biological relevance of the results.

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
yes
Remarks:
see additional information below
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Pre-Experiment for Toxicity
Two animals of each sex treated in the pre-experiments received the test item N-(3-Aminopropyl) imidazol dissolved in sterile water once orally. The volume administered was 10 mL/kg b.w.. The following dose levels were tested and the expressed toxic reactions are shown:
1st pre-experiment: 2000 mg/kg; males/females
2nd pre-experiment: 1000 mg/kg; males/females
3rd pre-experiment: 1500 mg/kg; males/females
Following the 1st pre-experiment severe symptoms were observed during autopsy of one male and one female animal. Visible necrotic damage was observed e.g. on liver, inner abdominal wall, spleen, pancreas and gastrointestinal tract. The stomach of the animals showed extremely severe symptoms such as enlargement and stomach rupture.
On the basis of these data 1500 mg/kg b.w. were estimated to be suitable as highest dose.
No substantial sex specific differences were observed with regard to clinical signs. In agreement with the sponsor the main study was performed using males only.

Toxic Symptoms in the Main Experiment
In the main experiment for each test item dose groups 7 males received once orally administrations of N-(3-Aminopropyl) imidazol dissolved in sterile water. The volume administered was 10 mL/kg b.w.. The syndromes of toxicity observed following treatment are:
375 mg/kg: No toxic reaction was observed.
750 kg/kg: reduction of spontaneous activity, ruffled fur
1500 mg/kg: reduction of spontaneous acitvity, fuffled fur, Hunchback
The animals treated with the negative control (sterile water) did not express any toxic reaction.

After treatment with the test item the number of PCEs was not decreased as compared to the mean value of PCEs of the vehicle control thus indicating that N-(3-Aminopropyl) imidazol did not exert any cytotoxic effects in the bone marrow.
In comparison to the corresponding vehicle controls there was no statistically significant or biologically relevant enhancement in the frequency of thedetected micronuclei at any preparation interval and dose level after administration of the test item. The mean values of micronuclei observed after treatment with N-(3-Aminopropyl) imidazol were near or below to the value of the vehicle control group.
Cyclophosphamide administered orally [40 mg/kg b.w.; 10 mL/kg b.w.; once] was used as positive control which showed a substantial and biologically relevant increase of induced micronucleus frequency.

In conclusion, it can be stated that during the study described and under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test in the bone marrow cells of the mouse.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse. Therefore, N-(3-Aminopropyl)-imidazol is considered to be non-mutagenic in this micronucleus assay.
Executive summary:

N-(3-Aminopropyl)-imidazol was tested in a MNT in mice in concentrations between 375, 750 and 1500 mg/kg (OECD474; BASF SE, 2011). After treatment with the test item the number of PCEs was not decreased as compared to the mean value of PCEs of the vehicle control thus indicating that N-(3-Aminopropyl)-imidazol did not exert any cytotoxic effects in the bone marrow. In comparison to the corresponding vehicle controls there was no statistically significant or biologically relevant enhancement in the frequency of thedetected micronuclei at any preparation interval and dose level after administration of the test item. The mean values of micronuclei observed after treatment with N-(3-Aminopropyl)-imidazol were near or below to the value of the vehicle control group. Cyclophosphamide administered orally [40 mg/kg b.w.; 10 mL/kg b.w.; once] was used as positive control which showed a substantial and biologically relevant increase of induced micronucleus frequency. The test item did not induce micronuclei as determined by the micronucleus test in the bone marrow cells of the mouse.