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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Constituent 1
Chemical structure
Reference substance name:
1,4-butanediyl diacrylate
EC Number:
213-979-6
EC Name:
1,4-butanediyl diacrylate
Cas Number:
1070-70-8
Molecular formula:
C10H14O4
IUPAC Name:
1,4-butanediyl diacrylate
Test material form:
liquid

Test animals

Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland GmbH
- Age at study initiation: 5 - 8 weeks
- Assigned to test groups randomly: yes
- Housing: Makrolon cages
- Diet (e.g. ad libitum): ad lib.
- Water (e.g. ad libitum): tap water ad lib.
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: Limited solubility of the test substance in water
Duration of treatment / exposure:
24 (all treatments), 48 h (control and high dose)
Frequency of treatment:
once
Doses / concentrationsopen allclose all
Remarks:
Females: 1500, 750, 375mg/kg
Remarks:
Males: 750, 375, 187.5mg/kg
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide: 20 mg/kg

Examinations

Tissues and cell types examined:
Bone marrow
Details of tissue and slide preparation:
DETAILS OF SLIDE PREPARATION
The slides were stained in eosin and methylene blue (modified May-Gruenwald solution or Wrights solution) for about 5 minutes.
After having briefly been rinsed in purified water, the preparations were soaked in purified water for about 2 - 3 minutes.
Subsequently, the slides were stained in Giemsa solution (15 ml Giemsa, 185 ml purified water) for about 15 minutes.
After having been rinsed twice in purified water and clarified in xylene, the preparations were mounted in Gorbit-Balsam.
Evaluation criteria:
Acceptance criteria
The mouse micronucleus test is considered valid if the following criteria are met:
- The quality of the slides must allow the identification and evaluation of a sufficient number of analyzable cells, i.e. >= 2000 PCEs and a clear differentiation between PCEs and NCEs.
- The ratio of PCEs/NCEs in the untreated animals (negative control) has to be within the normal range for the animal strain selected.
- The number of cells containing micronuclei in negative control animals has to be within the range of the historical control data both for PCEs and for NCEs.
- The two positive control substances have to induce a significant increase in the number of PCEs containing small and large micronuclei within the range of the historical control data or above.


Assessment criteria
A finding is considered positive if the following criteria are met:
- Significant and dose-related increase in the number of PCEs containing micronuclei.
- The number of PCEs containing micronuclei has to exceed both the concurrent negative control and the highest value of the historical control range.

A test substance is considered negative if the following criteria are met:
- The number of cells containing micronuclei in the dose groups is not significantly above the negative control and is within the historical control data.

Results and discussion

Test resultsopen allclose all
Sex:
male
Genotoxicity:
negative
Toxicity:
yes
Remarks:
mortality at 1500mg/kg
Vehicle controls validity:
valid
Positive controls validity:
valid
Sex:
female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Since mortality occured in 1500mg/kg males, this dose could not be scored. An additional experiment was performed, adding a new low dose of 187.5mg/kg as well as the 48h evaluation for the new high dose of 750mg/kg for males. Concurrent controls were included in this experiment and valid.
The slight increase observed in the 48h control samples (females) and in the low dose (females) had no relation to dosing and was not confirmed when twice the number of PCEs were recounted.
There was no significant difference in the percentage of PCEs per erythrocytes between groups.

Any other information on results incl. tables

Males
Interval Exp. Micronuclei Range PCEs relative to control
Vehicle control 24 2 1.1 ‰ 2-6 100%
Positive control 24 2 16.9 ‰** 51-86 104%
187.5mg/kg 24 2 1.5 ‰ 2-10 104%
Vehicle control 24 1 1.9 ‰ 2-10 100%
Positive control 24 1 13.5 ‰** 34-71 130%
375mg/kg 24 1 1.7 ‰ 5-8 115%
750mg/kg 24 1 1.9 ‰ 3-12 125%
Vehicle control 48 2 1.1 ‰ 3-7 100%
750mg/kg 48 2 1.3 ‰ 1-9 97%
Females
Interval sample size Micronuclei Range PCEs relative to control
Vehicle control 24 4000 1.7 ‰ 3-7 100%
Vehicle control 24 8000 1.5 ‰ 10-14 100%
Positive control 24 4000 12.8 ‰** 28-79 94%
375mg/kg 24 4000 2.5 ‰ 5-16 85%
375mg/kg 24 8000 1.7 ‰ 8-21 89%
750mg/kg 24 4000 1.3 ‰ 3-7 97%
1500mg/kg 24 4000 1.5 ‰ 4-8 79%
Vehicle control 48 4000 2.4 ‰ 6-14 100%
1500mg/kg 48 4000 1.8 ‰ 2-11 94%

** significantly different from control

Applicant's summary and conclusion