2,2'-[ethylenebis(oxymethylene)]bisoxirane

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

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)

Version / remarks:

Adopted: 29 July 2016

GLP compliance:

yes

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-[ethylenebis(oxymethylene)]bisoxirane

EC Number:

218-746-2

EC Name:

2,2'-[ethylenebis(oxymethylene)]bisoxirane

Cas Number:

2224-15-9

Molecular formula:

C8H14O4

IUPAC Name:

2,2'-[ethylenebis(oxymethylene)]bisoxirane

Test material form:

liquid

Test animals

Species:

mouse

Strain:

ICR

Details on species / strain selection:

ICR mice are commonly used in toxicity studies with a large historical control data base.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: ORIENTBIO INC., Republic of Korea
- Age at the start of administration:
Dose range finding study: 8 weeks old
Additional dose range finding study: 8 weeks old
Main study: 8 weeks old
- Weight at the start of administration:
Dose range finding study: males: 33.5–38.2 g ; females: 27.7–31.0 g
Additional dose range finding study: females: 26.9–30.4 g
Main study: males: 33.8–38.2 g ; females: 26.9–30.4 g
- Assigned to test groups randomly: yes
- Housing: polycarbonate cage
- Diet: ad libitum
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): permissible range: 19–25
- Humidity (%): permissible range: 40–70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: preliminary solubility test was conducted
- Concentration of test material in vehicle:
dose-range finding study: 25.0, 50.0, 100 and 200 mg/mL
additional dose-range finding and main study: 3.13, 6.25, 12.5, 25.0 mg/mL

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day

Remarks:

males

Doses / concentrations 2

Dose / conc.:

62.5 mg/kg bw/day

Remarks:

males

Doses / concentrations 3

Dose / conc.:

125 mg/kg bw/day

Remarks:

males

Doses / concentrations 4

Dose / conc.:

250 mg/kg bw/day

Remarks:

males

Doses / concentrations 5

Dose / conc.:

0 mg/kg bw/day

Remarks:

females

Doses / concentrations 6

Dose / conc.:

31.3 mg/kg bw/day

Remarks:

females

Doses / concentrations 7

Dose / conc.:

62.5 mg/kg bw/day

Remarks:

females

Doses / concentrations 8

Dose / conc.:

125 mg/kg bw/day

Remarks:

females

Control animals:

yes

Positive control(s):

mitomycin C
- Route of administration: intraperitoneal
- Doses / concentrations: 2 mg/kg bw

Examinations

Evaluation criteria:

The results of the study were considered to be positive when the following conditions were met:
- at least one of the test substance groups exhibits a statistically significant increase in the incidence of MNPCE compared to the negative control group
- the incidence of MNPCE is increased in a dose-dependent manner
- the incidence of MNPCE is increased outside the range of the distribution of the historical control data

Statistics:

Statistical analysis on the incidence of MNPCE, ratio of PCE to total erythrocytes and body weight was performed using SAS Program (version 9.3, SAS Institute Inc., U.S.A.).
For the incidence of MNPCE data, Mann-Whitney test was used for the comparison of the negative control group to each test substance group or the positive control group (significance levels: 0.05 and 0.01, two-tailed). Then, when significant in the test substance group, Cochran-Armitage trend test was employed for the comparison of the negative control group to each test substance group (significance levels: 0.05 and 0.01, two-tailed).
For the ratio of PCE to total erythrocytes and body weight data, Bartlett’s test was used for the comparison of homogeneity of variance of the negative control group to each test substance group (significance level: 0.05). One-way analysis of variance (ANOVA) was employed for homogeneous data (significance level: 0.05). In the comparison of the negative control group to the positive control group, Folded-F test was used for homogeneity of variance (significance level: 0.05).

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

positive

Toxicity:

no effects

Negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

As a result of the dose range finding study, deaths were observed at 500 mg/kg or more in males. In addition, deaths were observed at all dose levels in females. Therefore, an additional dose range finding study was conducted in females to determine the high dose of the main study.

An additional dose range finding study was conducted using the same method and conditions as the dose range finding study. The high dose of the test substance was set at 250 mg/kg and it was sequentially diluted by applying a geometric ratio of 2 to produce 3 lower dose levels (125, 62.5 and 31.3 mg/kg).

Applicant's summary and conclusion

Executive summary:

This study was designed to evaluate the potential of the test substance, 2,2’-[ethylenebis(oxymethylene)]bisoxirane, to induce micronuclei in bone marrow cells of mice when the test substance was administered twice to 8-week-old ICR mice via gastric intubation at 24 h intervals.

The dose range finding study was conducted at dose levels of 250, 500, 1,000 and 2,000 mg/kg to determine the high dose level for the main study and differences in clinical signs and mortality between the sexes. As a result of the dose range finding study, deaths were observed at 500 mg/kg or more in males. In addition, deaths were observed at all dose levels in females. Therefore, an additional dose range finding study was conducted to determine the high dose of the main study in females. The high dose of the test substance was set at 250 mg/kg and it was sequentially diluted by applying a geometric ratio of 2 to produce 3 lower dose levels (125, 62.5 and 31.3 mg/kg).

As a result of the additional dose range finding study, neither death nor severe clinical sign was observed. Therefore, the high dose level of the main study was set at 250 mg/kg and it was sequentially diluted by applying a geometric ratio of 2 to produce 2 lower dose levels (125 and 62.5 mg/kg) in males. In females, the high dose level of the main study was set at 125 mg/kg and it was sequentially diluted by applying a geometric ratio of 2 to produce 2 lower dose levels (62.5 and 31.3 mg/kg). In addition, positive and negative control groups were set. The main study was conducted with males and females, since differences in clinical signs and mortality were observed between the sexes.

In the main study, there was no statistically significant difference in the body weight and the ratio of polychromatic erythrocytes (PCE) among total erythrocytes in any test substance group of males and females compared to the negative control group. However, there were statistically significant differences and dose-related effects on the incidence of micronucleated polychromatic erythrocytes (MNPCE) in PCE in all test substance groups of males and in the 125 mg/kg group of females compared to the negative control group.

The incidence of MNPCE in PCE in the positive control group was statistically significantly increased in males and females compared to the negative control group.

Based on these results, the test substance, 2,2’-[ethylenebis(oxymethylene)]bisoxirane, had potential to induce micronuclei formation in bone marrow cells of mice under the conditions of this study.