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Toxicological information

Genetic toxicity: in vivo

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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:
1995-05-02 to 1995-07-10
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP Guideline study (OECD 474)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Version / remarks:
May 26,1983
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: flakes

Test animals

Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann, Borchen, Germany
- Age at study initiation: young adults
- Body weight at study initiation: 29.9 ±6.0 g
- Housing: conventional, 5 mice/sex in Macrolon cages type III
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 3°C
- Humidity:30 - 70 %
- Air changes: 15 per h
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 1995-05-03 To: 1995-05-12 (toxicity test); 1995-05-15 To: 1995-05-22 (main study)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle used: physiol. saline
- Amount of vehicle: 10 mL/kg bw
Frequency of treatment:
single exposure
Post exposure period:
24 or 48 hrs
Doses / concentrations
Remarks:
Doses / Concentrations:
1200 mg/kg bw (MTD)
Basis:
nominal conc.
No. of animals per sex per dose:
5 males and 5 females
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Route of administration: oral gavage
- Doses / concentrations: 100 mg/kg bw in 10 mL/kg bw

Examinations

Tissues and cell types examined:
femoral bone marrow erythrocytes
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION:
MTD (limit dose: 2000 mg/kg bw): highest dosage causing signs of toxicity but no mortality within 48 hrs of test compound administration), determined in pre-test

DETAILS OF SLIDE PREPARATION:
After sacrifice, bone marrow was suspended in FCS and erythrocytes were purified in a cellulose chromatography column. The eluate was centrifuged and the pellet was re-suspended in FCS/EDTA. Two slides were prepared per animal and stained with May-Grünwald Giemsa.

METHOD OF ANALYSIS:
Fully automated scoring: counting micronuclei in a total of appr. 2000 PCE (i.e. 100000 PCE per treatment group). Based on 2000 PCE scored, the PCE/NCE index was calculated.
Evaluation criteria:
A test compound is considered an inducer of micronuclei in PCE, if at least one group of mice treated with the test compound reveals a statistically and biologically relevant increase in micronucleated PCE (as compared to the negative control group of the same sampling time).
Statistics:
Heterogeneity Chi square calculated first. If homogenous: Pearson's contingency 2x2 chi square test (DF = 1), including a Yates correction factor. If inhomogenous: transformation and t-test (one sided, two sample). The latter also for evaluation of PCE/NCE ratios.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 400, 750, 1000, 1250, 1500, 1750 or 2000 mg/kg bw
- Solubility: soluble in vehicle up to limit concentration (2000 mg/kg bw)
- Clinical signs of toxicity in test animals: clinical symptoms (hunched posture, sedation, piloerrection and closed eyes), death at high dosages
- Evidence of cytotoxicity in tissue analyzed: not reported

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei: No
- Ratio of PCE/NCE: unaltered compared to controls

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
The genetic toxicity in vivo of 4-Hydroxy TEMPO was tested in a GLP compliant OECD 474 guideline study (mouse micronucleus test). Based on the results of the study the substance is not considered mutagenic.