Name: OECD_SIDS / SUBSTANCE : 2,2&#39;-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane /2224-15-9 Tue, 29 Nov 2022, 15:19:07+0900 /Legal entity owner: National Institute of Health Sciences
Printing date: 2022-11-29T15:19:07.849+09:00

iTable of Contents
0/0 ............................................................................................................................. 1
National Institute of Health Science ..................................................................... 2
2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane ................................................... 3
1 General information .................................................................................. 3
1.1 Identification ................................................................................. 3
Identification ............................................................................... 3
Identification ...................................................................... 3
7 Toxicological information ......................................................................... 4
7.5 Repeated dose toxicity ................................................................... 4
7.5.1 Repeated dose toxicity: oral ................................................. 4
Repeated dose toxicity: oral. 001 .......................................... 4
7.6 Genetic toxicity ............................................................................ 15
7.6.1 Genetic toxicity in vitro ...................................................... 15
Genetic toxicity in vitro.001 ................................................ 15
Genetic toxicity in vitro.002 ................................................ 22
7.8 Toxicity to reproduction ............................................................... 26
7.8.1 Toxicity to reproduction ..................................................... 26
Toxicity to reproduction. 001 .............................................. 26
References ................................................................................................................ 36
Reference Substances ...................................................................................... 36
2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane ......................................... 36
Test Materials .................................................................................................. 37
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane ......................................... 37
Literatures ....................................................................................................... 38
Combined repeated dose toxicity study with thereproductive/developmental toxicity screening test of 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane by oral administration in rats .......... 38
In Vitro Chromosomal Aberration Test of on 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane Cultured Chinese Hamster Cells. ..... 39
Reverse Mutation Test of 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxiraneon Bacteria. ............................................................................................... 40
Legal Entities ................................................................................................... 41
National Institute of Health Sciences .......................................................... 41ii

DOSSIER:UUID: 0
Dossier UUID:
Author:Date: 2022-11-29T15:19:07.707+09:00
Remarks:

Dossier header
Dossier submission type
Name
OECD SIDS
Versioncore 7.0
Name (given by user)

Dossier subject
Dossier subject
2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane / 2224-15-9
Public nameSubmitting legal entity
National Institute of Health Science
Dossier creation date/time
Tue, 29 Nov 2022, 15:19:07+0900
Used in category

1

LEGAL_ENTITY: National Institute of Health
Science
UUID: f51e7b54-9211-4863-90ce-fcf8a155d647
Dossier UUID:
Author:Date: 2022-11-07T16:24:02.822+09:00
Remarks:

General information
Legal entity nameNational Institute of Health Science22,2'-[1,2-Ethanediylbis(oxymethylene)]bisoxirane
General information
Identification
Identification
SUBSTANCE: 2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane

UUID: a9230384-e00d-455b-9f00-e6a378a6b446
Dossier UUID:
Author:Date: 2022-11-29T15:13:13.559+09:00
Remarks:
Substance name2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane
Legal entity
National Institute of Health Sciences / Kawasaki / Japan

Identification of substance
Reference substance
2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane / 2224-15-9
EC number

EC name

CAS number

CAS name

2224-15-9
IUPAC nameRole in the supply chain
Manufacturer
false
Importer
false
Only representativefalse
Downstream user
false

3

Toxicological information
Repeated dose toxicity
Repeated dose toxicity: oral
ENDPOINT_STUDY_RECORD: Repeated dose toxicity: oral. 001

UUID: c1195a3f-e3c2-49bc-93e1-55e2155c846d
Dossier UUID:
Author:Date: 2022-11-29T15:13:13.559+09:00
Remarks:

Administrative data
Endpoint
short-term repeated dose toxicity: oral
Type of information
experimental study
Adequacy of study
key study
Robust study summary
false
Used for classification
false
Used for SDS
false
Reliability
1 (reliable without restriction)
Rationale for reliability incl. deficiencies
guideline study OECD Test Guideline study under GLP condition
Reliability 1
Cross-reference
Reason / purpose for cross-reference
reference to same study 7.8.1 Toxicity to reproduction: Toxicity to reproduction. 001
Related information
OECD / Toxicity to reproduction / Toxicity
Ethanediylbis(oxymethylene)]bis-oxirane / 2224-15-9

toreproduction.

001/

2,2&#39;-[1,2-Data source
Reference
Combined repeated dose toxicity study with the reproductive/developmental toxicity screening test of /Ministry of Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published4Materials and methods
Test guideline
Qualifieraccording to guideline
Guideline
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction /Developmental Toxicity Screening Test)
Deviationsno
GLP compliance
yes
Limit test
no

Test material
Test material information
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
Specific details on test material used for the study- Name of test material (as cited in study report): 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane- Analytical purity: contain 0.1% water- Storage condition of test material: Room temperature, shading, airtightness- Stability under test conditions: The stability of test material was identified by analysis of theremainder.

Test animals
Speciesratcommon rodent species
Strainother: Crl:CD(SD)Sexmale/female
Details on test animals or test system and environmental conditions
TEST ANIMALS- Source: Charles River Japan, Inc., Hino Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation:
Males (main study groups): 365-433 g, females (main study groups): 236-266 g, females in (mating
study groups): 219-265 g- Housing: Animals were individually housed in stainless steel suspension cage (240W × 380D × 2
00H mm), from gestation day 18 to lactation day 4, Dams were bred individually or with individual littermates in plastic cages (310W x 360D x 175H mm) and bedding.
- Diet: Solid feed (CRF-1: Oriental Yeast Co., ltd.) was given ad libitum.
- Water: Tap water was given ad libitum.
- Acclimation period: Males (main study groups): 21 days, females (main study groups): 22 days, females (mating study groups): 21 days
5

ENVIRONMENTAL CONDITIONS- Temperature (°C): 20-26°C (actual temperature: 22.5-24.7°C)- Humidity (%): 40.0-70.0% (actual humidity: 41.3-58.4%)- Air changes (per hr): 12- Photoperiod (hrs dark / hrs light): 12 hr dark/12 hr light (light: 6:00~18:00)

Administration / exposure
Route of administration
oral: gavage
Vehiclewater for injection
Details on oral exposure- Amount of vehicle (if gavage): 5 mL/kg- Dosing volume: 5 mL/kg
Analytical verification of doses or concentrations
yes
Details on analytical verification of doses or concentrations
Concentrations of the test solutions using administration on day 1 were analyzed with GC. Analyticalconcentrations of the test solutions were all within the range of 92.2-106.4% of the nominal concen
trations and both values were within the acceptable range (concentration: percentage of nominal
concentration, 100±10%).
Duration of treatment / exposure
Males: 28 days including 14 days pre-mating
Females (main study groups): 28 days
Females (mating groups): 42-46 days including 14 days pre-mating, mating and gestation periods andthe days until day 4 of lactation
Frequency of treatment
Once/day, 7 days/week
Doses / concentrations
Dose / conc.
0mg/kg bw/day (actual dose received)

Dose / conc.
12.5

mg/kg bw/day (actual dose received)

Dose / conc.
50

mg/kg bw/day (actual dose received)

Dose / conc.
200

mg/kg bw/day (actual dose received)

No. of animals per sex per dose
- Main study groups:
6

Control- and high-dose groups: 12 males and 10 females per group (half of both sexes assigned as t
he treatment groups, and the remaining half assigned as the recovery groups)
Low- and middle-dose groups: 12 males and 5 females per group (half of males assigned as thetreatment groups, and the remaining half assigned as the recovery groups)- Mating groups:
12 females per dose
Control animals
yes, concurrent vehicle
Details on study design- Dose selection rationale: Based on the results of a 14-day preliminary study, the high dose was set to
200 mg/kg bw/day, and the intermediate dose and low dose were set to 50 mg/kg bw/day and 12.5
mg/kg bw/day, respectively.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 100, 200 or 500 mg/kg bw/day).
At 500 mg/kg bw/day, all males died, and 4 females died or were moribund. At 200 mg/kg bw/
day, transient salivation, decreased body weight and food consumption, and decreased eosinophil
percentage were observed in males and females, and increased platelets, shortened PT and APTT,
reddened gastric mucosa, and decreased absolute relative thymus weight were observed in males,
and increased neutrophil percentage, and decreased ALT and total protein were observed in females.
At 100 mg/kg bw/day, decreased body weight, increased platelets, shortened PT, and decreased
absolute relative thymus weight were observed in males, and decreased eosinophil percentage, anddecreased ALT and total protein were observed in females.
- Rationale for animal assignment (if not random): Body weight-balanced randomization

Examinations
Observations and examinations performed and frequency
CAGE SIDE OBSERVATIONS: Yes- Time schedule:
Males and females (main study groups): 2 times/day (before administration, 1-183 minutes afteradministration) during the administration period. Once a day during the recovery period.
Females (mating groups): 2 times/day (before administration, 3-150 minutes after administration) dur
ing the administration period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Males and females (main study groups): on day of grouping, on days 7, 14, 21 and 27 of adminis
tration period.
Females (mating groups): on day of grouping, on days 7 and 14 of administration period, on days 1, 8
and 15 of gestation period, on day 4 of lactation period.
BODY WEIGHT: Yes- Time schedule for examinations:
Males and females (main study groups):
Twice a week (On days 1, 4, 8, 11, 15, 18, 22, 25, 28 and 29 of administration period, on days 1, 4, 8, 11,
14 and 15 of recovery period).
Females (mating groups): Twice a week (On days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32, 36, 39, 43, 46, 50
and 53 of administration period, on days 0, 7, 14 and 20 of gestation period, on days 0, 4and 5 of lac
tation period).
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):- Food consumption: Yes
7

- Time schedule for examinations:
Males and females (main study groups):
Twice a week (Males: On days 2, 5, 9 and 12 of administration period, on days 2, 5, 9 and 12 of recovery period; Females: On days 2, 5, 9, 12, 16, 19, 23 and 26 of administration period, on days 2, 5, 9
and 12 of recovery period).
Females (mating groups): Twice a week (On days 2, 5, 9 and 12 of administration period, on days 2, 9,
16 and 20 of gestation period, on days 2 of lactation period).
WATER INTAKE- Time schedule for examinations:
Males and females (main study groups):
Twice a week (Males: On days 2, 5, 9 and 12 of administration period, on days 2, 5, 9 and 12 ofrecovery period; Females: On days 2, 5, 9, 12, 16, 19, 23 and 26 of administration period, on days 2, 5, 9
and 12 of recovery period).
Females (mating groups): Twice a week (On days 2, 5, 9 and 12 of administration period, on days 2, 9,
16 and 20 of gestation period, on days 2 of lactation period).
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes- Time schedule for collection of blood:
Males and females (main study groups): At the end of administration period, or at the end of recovery
period in both sexes- Anaesthetic used for blood collection: Pentobarbital sodium- Animals fasted: Yes- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and 2
00 mg/kg bw/day)- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume,
mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte perc
entage, platelet count, white blood cell count, differential white blood cell count, prothrombin time,
activated partial thromboplastin time, fibrinogen.
CLINICAL BIOCHEMISTRY: Yes- Time schedule for collection of blood:
Males and females (main study groups): At the end of administration period, or at the end of recovery
period in both sexes- Animals fasted: Yes- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0,12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and
200 mg/kg bw/day)- Parameters checked: ALP, total cholesterol, triglyceride, total bilirubin, glucose, urea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein, albumin, A/G ratio,
AST, ALT, γ-GT
BLOOD HORMONE: Yes- Time schedule for collection of serum:
Males and females (main study groups): At the end of administration period in both sexes- Animals fasted: Yes- How many animals:
6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)- Parameters checked: Triiodothyronine (T3), Thyroxin (T4), and thyroid stimulating hormone (TSH)
URINALYSIS: Yes- Time schedule for collection of urine:8Males and females (main study groups): Before the end of the administration period (day 23 of admi
nistration period) and before the end of recovery (day 12 of recovery period).
- Metabolism cages used for collection of urine: Yes
A urine collector to collect fresh urine samples under fasting but ad libitum drinking conditions,followed by collection of 24-hour urine samples under ad libitum feeding and drinking conditions.
- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and 2
00 mg/kg bw/day)- Parameters checked:
Fresh urine: Color, pH, protein, glucose, ketones, bilirubin, occult blood, urobilinogen, sediment24-urine: Specific gravity, urine volume (24-hour volume),
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males and females (main study groups): Final week of administration (Manipulative test and me
asurement of grip strength: Day 27 of administration, measurement of motor activity: Day 26 of admin
istration)- Dose groups that were examined: Autopsy animals after the end of the administration period- Battery of functions tested:
1) Manipulative Test. Pupillary reflex, approaching behavior, response to touch, auditory reflex, pain r
eflex2) Measurement of Grip Strength. Following manipulative test, grip strength of forelimb and hind limb
were measured by CPU gauge (San Diego Instruments Inc.).
3) Measurement of Spontaneous Motor Activity. Spontaneous motor activity (Ambulatory and verticalcounts) was measured by Activity Monitor (MED Associates Inc.).
The measurements were collected at 10-minute intervals from 1 hour to 2 hours after administration.
Sacrifice and pathology
GROSS PATHOLOGY: Yes
ORGAN WEIGHT: Yes [main study groups: brain, pituitary, salivary glands, thyroids, adrenal gland, thymus, spleen, heart, liver, kidney, testes, epididymides, ventral prostate, seminal vesicles, ovaries,
uterus; females in mating group: ovary, uterus]
HISTOPATHOLOGY: Yes, [main study groups: heart, lung, trachea, liver, pancreas, sublingual gland,
submandibular gland, esophagus, stomach, duodenum, jejunum, ileum (including Peyer’s patch),cecum, colon, rectum, thymus, spleen, mandibular lymph nodes, mesenteric lymph nodes, kidney,urinary bladder, testis, epididymis, ventral prostate, seminal vesicles (including coagulating gland),ovaries, uterus, vagina, pituitary, adrenal glands, thyroid (including parathyroid), cerebrum, cereb
ellum, pons, spinal cord, sciatic nerve, eye ball, Harderian gland, sternum and femur (including bonemarrows), muscle (rectus femoris), mammary gland; females in mating group: ovaries, uterus, and va
gina]
Statistics
For quantitative data, homogeneity of variance was tested using Bartlett method first. If the variance
was homogenous, statistical difference between each treatment group and the control group was
analyzed using Dunnett method. If not homogenous, statistical difference between each treatmentgroup and the control group was tested using Steel method. For comparison of quantitative data be
tween two groups in the recovery test, homogeneity of variance was analyzed by F-test. Then, if homo
genous, student's t-test was applied. If not homogenous, Aspin-Welch's t-test was used.
For histopathological findings, statistical analysis was carried out in combination with Steel-test andCochran-Armitage trend test.
Regarding clinical observation (except for frequency of urination, defecation, rearing and grooming)and sensory reactivity, Steel test was applied.

9

Results and discussion
Results of examinations
Clinical signs
effects observed, non-treatment-related
Description (incidence and severity)
CLINICAL SIGNS:[At the administration period]:
Transient salivation was observed in males and females at 200 mg/kg bw/day. This was considered
to be due to the irritant properties of the test substance.
[At the recovery period]:
There were no changes related to the test substance in any groups.
DETAILED CLINICAL OBSERVATIONS:[At the administration period]:
Transient salivation was observed in males and females at 200 mg/kg bw/day. This was considered to be due to the irritant properties of the test substance.
Mortality
no mortality observed
Body weight and weight changes
effects observed, treatment-related
Description (incidence and severity)[At the administration period]:
Decreases in body weights were observed throughout the administration period in males at 50 mg/kg/day and above, in late the administration period in females (main study and mating groups) at 200 m
g/kg bw/day.
[At the recovery period]:
Decrease in body weight was observed throughout the recovery period in males at 200 mg/kg/day.
Food consumption and compound intake (if feeding study)
effects observed, treatment-related
Description (incidence and severity)[At the administration period]:
Decreases in food consumption were observed throughout the administration period in males at 200
mg/kg bw/day and early in the administration period in females (main study and mating groups) at
200 mg/kg bw/day.
[At the recovery period]:
There were no changes related to the test substance in any groups.
Food efficiency
not examined
Water consumption and compound intake (if drinking water study)
effects observed, treatment-related
Description (incidence and severity)[At the administration period]:
Increase in water consumption was observed late in the administration period in females (main study
groups) at 200 mg/kg bw/day.
10

[At the recovery period]:
Increase in water consumption was observed early in the recovery period in females (main study
groups) at 200 mg/kg bw/day.
Ophthalmological findings
not examined
Haematological findings
effects observed, treatment-related
Description (incidence and severity)[At the end of administration period]:
Increases in platelet count and lymphocyte percentage, and decrease in neutrophil percentage wereobserved in females (main study groups) at 50 mg/kg bw/day and above. Increases in reticulocyte
reticulocyte percentage and PT were observed in females (main study groups) at 200 mg/kg bw/day.
[At the end of recovery period]:
There were no changes related to the test substance in any groups.
Clinical biochemistry findings
effects observed, treatment-related
Description (incidence and severity)
Including blood hormones (T3, T4, TSH)
CLINICAL BIOCHEMISTRY:[At the end of administration period]:
Decrease in triglyceride was observed in males at 50 mg/kg bw/day and above.
[At the end of recovery period]:
There were no changes related to the test substance in any groups.
BLOOD HOLMONES:
There were no changes related to the test substance in any groups at the end of administration andrecovery periods.
Urinalysis findings
no effects observed
Behaviour (functional findings)
no effects observed
Immunological findings
not examined
Organ weight findings including organ / body weight ratios
effects observed, treatment-related
Description (incidence and severity)[At the end of administration period]:
Decreases in absolute and relative ovaries weights were observed in females (mating groups) at 12.5
mg/kg bw/day and 50 mg/kg bw/day. Decreases in absolute and relative uterus weights observed infemales (mating groups) at 50 mg/kg bw/day.
[At the end of recovery period]:
There were no changes related to the test substance in any groups.
Gross pathological findings
no effects observed
Neuropathological findings
not examined
11

Histopathological findings: non-neoplastic
effects observed, treatment-related
Description (incidence and severity)[At the end of administration period]:
Stomach: Chronic ulcer of the glandular stomach was observed in males and females (main study
groups) at 50 mg/kg bw/day and above. Squamous epithelium hyperplasia of the forestomach wasobserved in males and females (main study groups) at 200 mg/kg bw/day.
[At the end of recovery period]:
Stomach: Scar of the glandular stomach was observed in males at 50 mg/kg bw/day and above.
Squamous epithelium hyperplasia of the forestomach was observed in males and females (mainstudy groups) at 200 mg/kg bw/day.
Histopathological findings: neoplastic
not examined

Effect levels
Key result
false
Dose descriptor
NOAEL
Effect level
12.5

mg/kg bw/day (actual dose received)

Based ontest mat.
Sex
male
Basis for effect level
body weight and weight gain
At 50 mg/kg bw/day, decrease in body weight was observed in males.
clinical biochemistry
At 50 mg/kg bw/day, decrease in triglyceride was observed in males.
histopathology: non-neoplastic
At 50 mg/kg bw/day, chronic ulcer of the glandular stomach was observed in males.
Key result
false
Dose descriptor
NOAEL (non-mating females)
Effect level
12.5

mg/kg bw/day (actual dose received)

Based ontest mat.
Sex
female12Basis for effect level
haematology
At 50 mg/kg bw/day, Increases in platelet count and lymphocyte percentage, and decrease inneutrophil percentage were observed in females (main study groups).
histopathology: non-neoplastic
At 50 mg/kg bw/day, chronic ulcer of the glandular stomach was observed in females (main study
groups).
Key result
false
Dose descriptor
NOAEL (maternal toxicity)
Effect level<

12.5

mg/kg bw/day (actual dose received)

Based ontest mat.
Sex
femaleBasis for effect level
organ weights and organ / body weight ratios
At 12.5 mg/kg bw/day, decreases in absolute and relative ovaries weights were observed infemales (mating study groups).

Any other information on results incl. tables
Figures and Tables (in English) are available in the following full report of the study.
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9d.pdf

Applicant's summary and conclusion
Conclusions
The NOAELs for repeated-dose toxicity were determined to be 12.5 mg/kg bw/day for males andfemales (non-mating groups), and the NOAEL for maternal toxicity was determined to be less than 12.5 mg/kg bw/day.
Executive summary
In the combined repeated dose and reproductive/developmental screening test (OECD TG422), SD rats
were treated orally with the test substance at the doses of 0, 12.5, 50 and 200 mg/kg bw/day. Males (12animals/dose: 6 animals were treated as a recovery group) were dosed for 28 days including a 14 day
pre-mating period. Females (12 animals/dose) were dosed for 42-46 days including 14 day pre-mating,mating, and gestation periods and days until day 4 of lactation. In addition, as the main study group of
females, 5 or 10 females/group was dosed for 28 days without mating (5 females at 0 and 200 mg/kg
bw/day were treated as recovery groups).
Decreases in body weights were observed during the administration period in males at 50 mg/kg bw/
day and above and in females (main study and mating groups) at 200 mg/kg bw/day. Decreases infood consumption were observed during the administration period in males and females (main study
and mating groups) at 200 mg/kg bw/day. An increase in water consumption was observed during the13administration period and early recovery periods in females (main study group) at 200 mg/kg bw/day.
The following findings were observed in the examination at the end of the administration period. In thehaematological examination, increases in platelet count and lymphocyte percentage, and decreases inneutrophil percentage were observed in females (main study groups) at 50 mg/kg bw/day and above.
Increases in reticulocyte reticulocyte percentage and PT were observed in females (main study groups)
at 200 mg/kg bw/day. In the clinical chemistry, decreases in triglyceride were observed in males at
50 mg/kg bw/day and above. In the organ weights, decreases in absolute and relative ovaries weights
were observed in females (mating groups) at 12.5 mg/kg bw/day and 50 mg/kg bw/day. Decreases inabsolute and relative uterus weights were observed in females (mating groups) at 50 mg/kg bw/day. In
the histopathological examination, chronic ulcer of the glandular stomach was observed in males andfemales (main study groups) at 50 mg/kg bw/day and above. Squamous epithelium hyperplasia of theforestomach was observed in males and females (main study groups) at 200 mg/kg bw/day. At the endof the recovery period, all changes observed in haematological examination, clinical chemistry, organ
weight and histopathological examination were disappeared or reduced
Based on the above results, the NOAELs for repeated dose toxicity of 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane were determined to be 12.5 mg/kg bw/day in male and female(non-mating groups), and NOAEL for maternal toxicity was determined to be less than 12.5 mg/kg bw/
day.

14

Genetic toxicity
Genetic toxicity in vitro
ENDPOINT_STUDY_RECORD: Genetic toxicity in vitro.001

UUID: 1066351b-57a7-49e8-9ed4-e7477de31a7f
Dossier UUID:
Author:Date: 2022-11-07T15:03:48.000+09:00
Remarks:

Administrative data
Endpoint
in vitro gene mutation study in bacteria
Type of information
experimental study
Adequacy of study
key study
Robust study summary
false
Used for classification
false
Used for SDS
false
Reliability
1 (reliable without restriction)
Rationale for reliability incl. deficiencies
guideline study under GLP condition
Reliability 1

Data source
Reference
Reverse Mutation Test of 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane on Bacteria. / Ministry of
Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published

Materials and methods
Test guideline
Qualifieraccording to guideline
15

Guideline
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
in vitro gene mutation study in bacteria
Deviations
no
Qualifieraccording to guideline
Guideline
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
genetic toxicity in vitro, otherDeviationsno
GLP compliance
yesType of assay
bacterial reverse mutation assay
in vitro gene mutation study in bacteria

Test material
Test material information
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
Specific details on test material used for the study
Purity: contain 0.1% water

MethodSpecies / strain
Species / strain / cell typeS. typhimurium TA 1535, TA 1537, TA 98 and TA 100
bacteria
Species / strain / cell type
E. coli WP2 uvr A
bacteria
Metabolic activation
with and without
Metabolic activation system
S9 mix: SD male rat liver, induced by phenobarbital and 5,6-benzoflavone
Test concentrations with justification for top dose
-S9 mix:
9.77, 19.5, 39.1, 78.1, 156.3, 312.5, 625, 1250, 2500, 5000 μg/plate (TA 1535 and TA 100 strains)
78.1, 156.3, 312.5, 625, 1250, 2500, 5000 μg/plate (WP2uvrA strain)
312.5, 625, 1250, 2500, 5000 μg/plate (TA 98 and TA 1537 strains)+S9 mix:
16

39.1, 78.1, 156.3, 312.5, 625, 1250, 2500, 5000 μg/plate (TA 1535 and TA 100 strains)
78.1, 156.3, 312.5, 625, 1250, 2500, 5000 μg/plate (WP2uvrA strain)
312.5, 625, 1250, 2500, 5000 μg/plate (TA 98 and TA 1537 strains)
Maximum concentration was established based on the result of the preliminary test at concentration up to 5000 ug/plate. In this test, the growth inhibition was observed at 5000 μg/plate for WP2uvrAstrain with or without S9 mix.

Vehicle / solvent- Vehicle(s)/solvent(s) used: water for injection
Controls
Untreated negative controlsnoNegative solvent / vehicle controls
yes
True negative controlsnoPositive controls
yes
Positive control substance
9-aminoacridine
9-amimoacridine hydrochloride (9AA): -S9 mix: (TA1537)
sodium azide
NaN3: -S9 mix: (TA1535)
furylfuramide2-(2-Furyl)-3-(5-nitro-2-furyl) acrylamide (AF-2): -S9 mix: (TA100, TA98, WP2 uvrA)
other: 2-aminoanthracene (2AA)+S9 mix: (TA1535, TA100, TA98, TA1537 and WP2 uvrA)
Details on test system and experimental conditions
METHOD OF APPLICATION: Preincubation
DURATION- Preincubation period: 20 min at 37°C
- Exposure duration: ca.48 hrs
NUMBER OF PLATES: 3
NUMBER OF REPLICATIONS: 2
DETERMINATION OF CYTOTOXICITY- Method: other: growth inhibition
Evaluation criteria
A chemical was judged to be mutagenic when the mean number of revertant colonies per plateincreased more than twice that of the negative control and when the dose-related and reproducible increase was observed.
StatisticsnoResults and discussion
17

Test results
Key result
trueSpecies / strain
S. typhimurium TA 1535
bacteria
Metabolic activation
with and without
Genotoxicity
positive
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
false
Species / strain
S. typhimurium TA 1537
bacteria
Metabolic activation
with
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
true
18Species / strain
S. typhimurium TA 1537
bacteria
Metabolic activation
without
Genotoxicity
positive
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
S. typhimurium TA 98
bacteria
Metabolic activation
with and without
Genotoxicity
positive
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
S. typhimurium TA 100
bacteria
19

Metabolic activation
with and without
Genotoxicity
positive
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
E. coli WP2 uvr A
bacteria
Metabolic activation
with
Genotoxicity
positive
Cytotoxicity / choice of top concentrations
cytotoxicity 5000 μg/plate
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Key result
false
Species / strain
E. coli WP2 uvr A
bacteria
Metabolic activation
without
Genotoxicity
negative20

Cytotoxicity / choice of top concentrations
cytotoxicity 2500 μg/plate or more
Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Additional information on results
The maximum specific activity of mutation was 2151.1 revertants/mg/plate, which was observedin plates of Salmonella typhimurium TA100 treated with the test article at 78.1 μg/plate without
metabolic activation.

Any other information on results incl. tables
Figures and Tables (in English) are available in the following full report of the study.
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9e.pdf

Applicant's summary and conclusion
Conclusions
Positive with or without metabolic activation
Executive summary
In a bacterial reverse mutation assay using Salmonella typhimurium TA100, TA1535, TA98, and TA
1537, and Escherichia coli WP2uvrA (OECD TG 471), 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
was positive for TA100, TA1535 and TA98 with or without metabolic activation, positive for Escherichia
coli WP2uvrA/pKM101 with metabolic activation, positive for TA1537 without metabolic activation.
The maximum specific activity of mutation was 2151.1 revertants/mg/plate, which was observed inplates of Salmonella typhimurium TA100 treated with the test article at 78.1 μg/plate without metabolic
activation.

21

ENDPOINT_STUDY_RECORD: Genetic toxicity in vitro.002

UUID: 193b622d-2af8-4946-818b-3a1f18c337b8
Dossier UUID:
Author:Date: 2022-11-07T15:04:52.000+09:00
Remarks:

Administrative data
Endpoint
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information
experimental study
Adequacy of study
key study
Robust study summary
false
Used for classification
false
Used for SDS
false
Reliability
1 (reliable without restriction)
Rationale for reliability incl. deficiencies
guideline study under GLP condition
Reliability 1

Data source
Reference
In Vitro Chromosomal Aberration Test of on 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane Culture /Ministry of Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published

Materials and methods
Test guideline
Qualifieraccording to guideline
Guideline
OECD Guideline 473 (In Vitro Mammalian Chromosomal Aberration Test)in vitro cytogenicity / chromosomal aberration study in mammalian cells (from 26 September 2014)22

Deviations
no
Qualifieraccording to guideline
Guideline
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
genetic toxicity in vitro, otherDeviationsno
GLP compliance
yesType of assay
in vitro mammalian chromosome aberration testin vitro cytogenicity / chromosome aberration study in mammalian cells

Test material
Test material information
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
Specific details on test material used for the study
Purity: contain 0.1% water

MethodSpecies / strain
Species / strain / cell type
Chinese hamster lung (CHL/IU)
mammalian cell line
Cytokinesis block (if used)colcemid
Metabolic activation
with and without
Metabolic activation system
S9 mix: SD male rat liver, induced by phenobarbital and 5,6-benzoflavone
Test concentrations with justification for top dose
Cell growth inhibition study-S9 mix (short-term treatment): 14.1, 28.1, 56.3, 112.5, 225, 450, 900, 1800 ug/mL+S9 mix (short-term treatment): 14.1, 28.1, 56.3, 112.5, 225, 450, 900, 1800 ug/mL-S9 mix (continuous treatment, 24hr): 14.1, 28.1, 56.3, 112.5, 225, 450, 900, 1800 ug/mL
Main study-S9 (short-term treatment): 62.5, 125, 250, 500 ug/mL+S9 (short-term treatment): 125, 250, 500, 1000 ug/mL-S9 mix (continuous treatment, 24hr): 31.3, 62.5, 125, 250 ug/mL
Vehicle / solvent- Vehicle(s)/solvent(s) used: water for injection
23

Controls
Untreated negative controlsnoNegative solvent / vehicle controls
yes
True negative controlsnoPositive controls
yes
Positive control substanceN-dimethylnitrosamine+S9
mitomycin C-S9Details on test system and experimental conditions
METHOD OF APPLICATION: Exposure duration: [short-term treatment]:6 hrs + 18 hr, [continuous
treatment]: 24 hrs
SPINDLE INHIBITOR: Colcemid
STAIN: Giemsa stain (2 v/v%) for 15 min.
NUMBER OF REPLICATIONS: 2
NUMBER OF CELLS EVALUATED: 100 + 100 cells /concentration
DETERMINATION OF CYTOTOXICITY- Method: relative total growth
Evaluation criteria
For the evaluation of the frequencies of structural aberrations and of polyploidy induced, the following
criteria were employed. Appearance incidence of cell with chromosomal aberrations: Negative (-):
less than 5%, Equivocal(±): 5% or more and less than 10%, Positive(+): 10% or more
StatisticsnoResults and discussion
Test results
Key result
trueSpecies / strain
Chinese hamster lung (CHL/IU)
mammalian cell line
Metabolic activation
with and without
Genotoxicity
positive structural aberration, D20: 0.051 mg/mL, TR: 1300
Cytotoxicity / choice of top concentrations
cytotoxicity
24

Vehicle controls validity
valid
Untreated negative controls validity
not examined
True negative controls validity
not examined
Positive controls validity
valid
Additional information on results
RANGE-FINDING/SCREENING STUDIES (if applicable):
50% cell growth inhibition (IC50): 650 ug/mL (short-term treatment, +S9 mix), 270 ug/mL (short-term treatment, -S9 mix), 150 ug/mL (continuous treatment)

Any other information on results incl. tables
Figures and Tables (in English) are available in the following full report of the study.
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9f.pdf

Applicant's summary and conclusion
Conclusions
Positive with or without metabolic activation
Executive summary
In an in vitro chromosomal aberration test using CHL/IU cells (OECD TG 473), 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane induced structural chromosomal aberrations but did not
induce chromosome numerical aberrations under the conditions of this study.

25

Toxicity to reproduction
Toxicity to reproduction
ENDPOINT_STUDY_RECORD: Toxicity to reproduction. 001

UUID: ff758fd1-dff2-4058-87b4-6afdc80229cf
Dossier UUID:
Author:Date: 2022-11-29T14:38:14.704+09:00
Remarks:

Administrative data
Endpoint
reproductive toxicity, other A combined repeated dose/reproductive developmental toxicity study
Type of information
experimental study
Robust study summary
false
Used for classification
false
Used for SDS
false
Reliability
1 (reliable without restriction)
Rationale for reliability incl. deficiencies
guideline study OECD Test Guideline study under GLP condition
Reliability 1
Cross-reference
Reason / purpose for cross-reference
reference to same study 7.5.1 Repeated dose toxicity: oral: Repeated dose toxicity: oral. 001
Related information
OECD / Repeated dose toxicity: oral / Repeated dose toxicity: oral. 001 / 2,2&#39;-[1,2Ethanediylbis(oxymethylene)]bis-oxirane / 2224-15-9

Data source
Reference
Combined repeated dose toxicity study with the reproductive/developmental toxicity screening test of /Ministry of Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9d.pdf

Materials and methods
26

Test guideline
Qualifieraccording to guideline
Guideline
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction /Developmental Toxicity Screening Test)
Deviationsno
GLP compliance
yes
Limit test
no

Test material
Test material information
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
Specific details on test material used for the study- Name of test material (as cited in study report): 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane- Analytical purity: contain 0.1% water- Storage condition of test material: Room temperature, shading, airtightness- Stability under test conditions: The stability of test material was identified by analysis of theremainder.

Test animals
Speciesrat
Strainother: Crl:CD(SD)Sexmale/female
Details on test animals or test system and environmental conditions
TEST ANIMALS- Source: Charles River Japan, Inc., Hino Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation:
Males (main study groups): 365-433 g, females (main study groups): 236-266 g, females in (mating
study groups): 219-265 g- Housing: Animals were individually housed in stainless steel suspension cage (240W × 380D × 2
00H mm), from gestation day 18 to lactation day 4, Dams were bred individually or with individual littermates in plastic cages (310W x 360D x 175H mm) and bedding.
- Diet: Solid feed (CRF-1: Oriental Yeast Co., ltd.) was given ad libitum.
- Water: Tap water was given ad libitum.
- Acclimation period: Males (main study groups): 21 days, females (main study groups): 22 days, females (mating study groups): 21 days
ENVIRONMENTAL CONDITIONS- Temperature (°C): 20-26°C (actual temperature: 22.5-24.7°C)- Humidity (%): 40.0-70.0% (actual humidity: 41.3-58.4%)- Air changes (per hr): 1227

- Photoperiod (hrs dark / hrs light): 12 hr dark/12 hr light (light: 6:00~18:00)

Administration / exposure
Route of administration
oral: gavage
Vehiclewater for injection
Details on exposure- Amount of vehicle (if gavage): 5 mL/kg- Dosing volume: 5 mL/kg
Details on mating procedure- M/F ratio per cage: 1/1- Length of cohabitation: up to 14 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
Analytical verification of doses or concentrations
yes
Details on analytical verification of doses or concentrations
Concentrations of the test solutions using administration on day 1 were analyzed with GC. Analyticalconcentrations of the test solutions were all within the range of 92.2-106.4% of the nominal concen
trations and both values were within the acceptable range (concentration: percentage of nominal
concentration, 100±10%).
Duration of treatment / exposure
Males: 28 days including 14 days pre-mating
Females (main study groups): 28 days
Females (mating groups): 42-46 days including 14 days pre-mating, mating and gestation periods andthe days until day 4 of lactation
Frequency of treatment
Once/day, 7 days/week
Doses / concentrations
Dose / conc.
0mg/kg bw/day (actual dose received)

Dose / conc.
12.5

mg/kg bw/day (actual dose received)

Dose / conc.
50

mg/kg bw/day (actual dose received)

Dose / conc.
200

mg/kg bw/day (actual dose received)

No. of animals per sex per dose
- Main study groups:
28

Control- and high-dose groups: 12 males and 10 females per group (half of both sexes assigned as t
he treatment groups, and the remaining half assigned as the recovery groups)
Low- and middle-dose groups: 12 males and 5 females per group (half of males assigned as thetreatment groups, and the remaining half assigned as the recovery groups)- Mating groups:
12 females per dose
Control animals
yes, concurrent vehicle
Details on study design- Dose selection rationale: Based on the results of a 14-day preliminary study, the high dose was set to
200 mg/kg bw/day, and the intermediate dose and low dose were set to 50 mg/kg bw/day and 12.5
mg/kg bw/day, respectively.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 100, 200 or 500 mg/kg bw/day).
At 500 mg/kg bw/day, all males died, and 4 females died or were moribund. At 200 mg/kg bw/day,transient salivation, decreased body weight and food consumption, and decreased eosinophil percent
age were observed in males and females, and increased platelets, shortened PT and APTT, reddened
gastric mucosa, and decreased absolute relative thymus weight were observed in males, and increased neutrophil percentage, and decreased ALT and total protein were observed in females. At 100
mg/kg bw/day, decreased body weight, increased platelets, shortened PT, and decreased absolute
relative thymus weight were observed in males, and decreased eosinophil percentage, and decreased
ALT and total protein were observed in females.
- Rationale for animal assignment (if not random): Body weight-balanced randomization

Examinations
Parental animals: Observations and examinations
CAGE SIDE OBSERVATIONS: Yes- Time schedule:
Males and females (main study groups): 2 times/day (before administration, 1-183 minutes afteradministration) during the administration period. Once a day during the recovery period.
Females (mating groups): 2 times/day (before administration, 3-150 minutes after administration) dur
ing the administration period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Males and females (main study groups): on day of grouping, on days 7, 14, 21 and 27 of adminis
tration period.
Females (mating groups): on day of grouping, on days 7 and 14 of administration period, on days 1, 8
and 15 of gestation period, on day 4 of lactation period.
BODY WEIGHT: Yes- Time schedule for examinations:
Males and females (main study groups):
Twice a week (On days 1, 4, 8, 11, 15, 18, 22, 25, 28 and 29 of administration period, on days 1, 4, 8, 11,
14 and 15 of recovery period).
Females (mating groups): Twice a week (On days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32, 36, 39, 43, 46, 50
and 53 of administration period, on days 0, 7, 14 and 20 of gestation period, on days 0, 4and 5 of lac
tation period).
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):- Food consumption: Yes- Time schedule for examinations:29Males and females (main study groups):
Twice a week (Males: On days 2, 5, 9 and 12 of administration period, on days 2, 5, 9 and 12 of recovery period; Females: On days 2, 5, 9, 12, 16, 19, 23 and 26 of administration period, on days 2, 5, 9
and 12 of recovery period).
Females (mating groups): Twice a week (On days 2, 5, 9 and 12 of administration period, on days 2, 9,
16 and 20 of gestation period, on days 2 of lactation period).
WATER INTAKE- Time schedule for examinations:
Males and females (main study groups):
Twice a week (Males: On days 2, 5, 9 and 12 of administration period, on days 2, 5, 9 and 12 ofrecovery period; Females: On days 2, 5, 9, 12, 16, 19, 23 and 26 of administration period, on days 2, 5, 9
and 12 of recovery period).
Females (mating groups): Twice a week (On days 2, 5, 9 and 12 of administration period, on days 2, 9,
16 and 20 of gestation period, on days 2 of lactation period).
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes- Time schedule for collection of blood:
Males and females (main study groups): At the end of administration period, or at the end of recovery
period in both sexes- Anaesthetic used for blood collection: Pentobarbital sodium- Animals fasted: Yes- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and 2
00 mg/kg bw/day)- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume,
mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte perc
entage, platelet count, white blood cell count, differential white blood cell count, prothrombin time,
activated partial thromboplastin time, fibrinogen.
CLINICAL BIOCHEMISTRY: Yes- Time schedule for collection of blood:
Males and females (main study groups): At the end of administration period, or at the end of recovery
period in both sexes- Animals fasted: Yes- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0,12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and
200 mg/kg bw/day)- Parameters checked: ALP, total cholesterol, triglyceride, total bilirubin, glucose, urea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein, albumin, A/G ratio,
AST, ALT, γ-GT
BLOOD HORMONE: Yes- Time schedule for collection of serum:
Males and females (main study groups): At the end of administration period in both sexes- Animals fasted: Yes- How many animals:
6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)- Parameters checked: Triiodothyronine (T3), Thyroxin (T4), and thyroid stimulating hormone (TSH)
URINALYSIS: Yes- Time schedule for collection of urine:30

Males and females (main study groups): Before the end of the administration period (day 23 of admi
nistration period) and before the end of recovery (day 12 of recovery period).
- Metabolism cages used for collection of urine: Yes
A urine collector to collect fresh urine samples under fasting but ad libitum drinking conditions,followed by collection of 24-hour urine samples under ad libitum feeding and drinking conditions.
- How many animals:
At the end of administration period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0, 12.5, 50, 200 mg/kg bw/day)
At the end of recovery period: 6 males/dose (0, 12.5, 50, 200 mg/kg bw/day), 5 females/dose (0 and 2
00 mg/kg bw/day)- Parameters checked:
Fresh urine: Color, pH, protein, glucose, ketones, bilirubin, occult blood, urobilinogen, sediment24-urine: Specific gravity, urine volume (24-hour volume),
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males and females (main study groups): Final week of administration (Manipulative test and me
asurement of grip strength: Day 27 of administration, measurement of motor activity: Day 26 of admin
istration)- Dose groups that were examined: Autopsy animals after the end of the administration period- Battery of functions tested:
1) Manipulative Test. Pupillary reflex, approaching behavior, response to touch, auditory reflex, pain r
eflex2) Measurement of Grip Strength. Following manipulative test, grip strength of forelimb and hind limb
were measured by CPU gauge (San Diego Instruments Inc.).
3) Measurement of Spontaneous Motor Activity. Spontaneous motor activity (Ambulatory and verticalcounts) was measured by Activity Monitor (MED Associates Inc.).
The measurements were collected at 10-minute intervals from 1 hour to 2 hours after administration.
Oestrous cyclicity (parental animals)
Vaginal smears were collected from all females in the mating groups and microscopically examined
every day from the day after the start of administration until the day copulation was confirmed.
Sperm parameters (parental animals)
Parameters examined in all P male parental generations: testis, epididymis and seminal vesicle weight, histopathological examinations for testes, epididymides, seminal vesicle and ventral prostate.
Litter observations
PARAMETERS EXAMINED: The following parameters were examined in F1 offspring: Number and sexof pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, and weight gain.
GROSS EXAMINATION OF DEAD PUPS: Yes, for external and internal abnormalities.
Postmortem examinations (parental animals)
METHOD OF SACRIFICED: All animals were sacrificed by exsanguination under pentobarbital sodium
anesthesia.
SACRIFICE: Males and females (main study groups): On next day after the last administration,
Maternal animals: on Day 5 of lactation, and males and females recovery group: on Day 14 of recovery.
GROSS PATHOLOGY: Yes
ORGAN WEIGHT: Yes [main study groups: brain, pituitary, salivary glands, thyroids, adrenal gland,
thymus, spleen, heart, liver, kidney, testes, epididymides, ventral prostate, seminal vesicles, ovaries, ut
erus; females in mating group: ovary, uterus]
HISTOPATHOLOGY: Yes, [main study groups: heart, lung, trachea, liver, pancreas, sublingual gland,
submandibular gland, esophagus, stomach, duodenum, jejunum, ileum (including Peyer’s patch),31

cecum, colon, rectum, thymus, spleen, mandibular lymph nodes, mesenteric lymph nodes, kidney,urinary bladder, testis, epididymis, ventral prostate, seminal vesicles (including coagulating gland),ovaries, uterus, vagina, pituitary, adrenal glands, thyroid (including parathyroid), cerebrum, cerebellum, pons, spinal cord, sciatic nerve, eye ball, Harderian gland, sternum and femur (including bone marrows), muscle (rectus femoris), mammary gland; females in mating group: ovaries, uterus, vagina ]Postmortem examinations (offspring)
SACRIFICE- The F1 offsprings were euthanized on PND4 by exsanguination under 20%Isoflurane anesthesia.
GROSS NECROPSY: Yes- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, andabdominal viscera.
HISTOPATHOLOGY / ORGAN WEIGTHS- Not examined.
Statistics
For quantitative data, homogeneity of variance was tested using Bartlett method first. If the variance
was homogenous, statistical difference between each treatment group and the control group was
analyzed using Dunnett method. If not homogenous, statistical difference between each treatmentgroup and the control group was tested using Steel method. For comparison of quantitative data be
tween two groups in the recovery test, homogeneity of variance was analyzed by F-test. Then, if homo
genous, student's t-test was applied. If not homogenous, Aspin-Welch's t-test was used.
For histopathological findings, statistical analysis was carried out in combination with Steel-test a
nd Cochran-Armitage trend test. Regarding clinical observation (except for frequency of urination, defecation, rearing and grooming) and sensory reactivity, Steel test was applied. Regarding implantat
ion index, delivery index, birth index, live birth index, viability index, sex ratio and external abnormalities, Steel test was performed between administration groups and control groups. Regarding copulation,
fertility index, and gestation index, Fisher's test was applied.
Reproductive indices
Each parameter was determined by the following equations:
Copulation index (%) = (No. of pairs with successful copulation / No. of pairs) × 100
Fertility index (%) = (No. of pregnant females / No. of pairs with successful copulation) × 100
Gestation index (%) = (No. of dams having live pups / No. of pregnant dams) × 100
Length of gestation (days)
Implantation index (%) = (No. of implantation scars / No. of corpora lutea) × 100
Delivery index (%) = (No. of pups born / No. of implantation scars) × 100
Birth index (%) = (No. of live pups born / No. of implantation scars) × 100
Live birth index (%) = (No. of live pups born / No. of pups born) × 100
Sex ratio on Day 4 of lactation = No. of male pups / No. of female pups
External abnormalities (%) = (No. of pups with external abnormalities / No. of live pups) × 100
Offspring viability indices
Viability index (%) = (No. of live pups on Day 4 of lactation/ No. of live pups born) × 100

Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
Clinical signs
effects observed, non-treatment-related
32

Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Mortality
no mortality observed
Body weight and weight changes
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Food consumption and compound intake (if feeding study)
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Food efficiency
not examined
Water consumption and compound intake (if drinking water study)
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Ophthalmological findings
not examined
Haematological findings
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Clinical biochemistry findings
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Urinalysis findings
no effects observed
Behaviour (functional findings)
no effects observed
Immunological findings
not examined
Organ weight findings including organ / body weight ratios
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Gross pathological findings
no effects observed
Neuropathological findings
not examined
33

Histopathological findings: non-neoplastic
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Histopathological findings: neoplastic
not examined

Reproductive function / performance (P0)
Reproductive function: oestrous cycle
no effects observed
Reproductive function: sperm measures
no effects observed
Reproductive performance
effects observed, treatment-related

Details on results (P0)General toxicity: See 7.5.1 Repeated dose toxicity.001
Reproductive performance: An increased non-pregnant females, decreased fertility index were ob
served at 50 mg/kg bw/day and above. Decreases in number of corpora lutea, number of impla
ntations, implantation index, and birth index, prolonged gestation length were observed at 50 mg/kg
bw/day.

Effect levels (P0)
Key result
false
Dose descriptor
NOAEL
Effect level
12.5

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
reproductive performance
At 50 mg/kg bw/day, decreases in number of corpora lutea, number of implantations, implantationindex, and birth index, prolonged gestation length were observed.

Results: F1 generation
General toxicity (F1)Clinical signs
no effects observed
34Mortality / viability
mortality observed, treatment-related
Body weight and weight changes
effects observed, treatment-related
Gross pathological findings
no effects observed

Effect levels (F1)
Key result
false
Dose descriptor
NOAEL
Generation
F1
Effect level
12.5

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
other: At 50 mg/kg bw/day, decreases in pups born, live pups born, live pups on Day 4 of lactation,
viability index, increases in body weights at Day0 and Day4 of lactation were observed.

Any other information on results incl. tables
Figures and Tables (in English) are available in the following full report of the study.
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9d.pdf

Applicant's summary and conclusion
Conclusions
In the combined repeated oral dose toxicity study with the reproduction/developmental toxicity screening test (OECD TG 422) described above, reproductive effects in parent animals and effects on offsp
ring were observed at 50 mg/kg bw/day. The NOAEL for the rat reproductive/developmental toxicity
of 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane was regarded as 12.5 mg/kg bw/day.

35

References
Reference Substances
REFERENCE_SUBSTANCE: 2,2'-[1,2Ethanediylbis(oxymethylene)]bis-oxirane
UUID: e03f3fed-a3ac-4bd4-b972-e670071cb252
Dossier UUID:
Author:Date: 2022-11-07T15:00:24.000+09:00
Remarks:
Reference substance name2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane

Inventory
CAS number
2224-15-9

Synonyms
Synonyms
Identifier
other: Ethylene glycol diglycidyl ether

Molecular and structural information
Molecular formula
C8H14O4
Molecular weight174.1936

Test Materials
TEST_MATERIAL_INFORMATION: 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane
UUID: 931e20ce-f068-4902-9747-41c723b5f5e0
Dossier UUID:
Author:Date: 2022-11-07T15:02:41.000+09:00
Remarks:Name
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane

Composition
Composition
Reference substance
2,2'-[1,2-Ethanediylbis(oxymethylene)]bis-oxirane / 2224-15-9
EC number

EC name

CAS number

CAS name

2224-15-9
IUPAC name37

Literatures
LITERATURE: Combined repeated dose toxicity study withthe reproductive/developmental toxicity screening test of
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane by oral
administration in rats
UUID: bd9f6f04-067d-4005-9e92-e1af12f2c22b
Dossier UUID:
Author:Date: 2022-11-07T15:00:55.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Combined repeated dose toxicity study with the reproductive/developmental toxicity screening test of
2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane by oral administration in rats
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Year
2013
Bibliographic source
available in the web of Japan Existing Chemical Data Base (JECDB) https://dra4.nihs.go.jp/
mhlw_data/home/pdf/PDF2224-15-9d.pdf
Testing facility
Nihon Bioresearch Inc.
Report number
10013038

LITERATURE: In Vitro Chromosomal Aberration Test of
on 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane
Cultured Chinese Hamster Cells.
UUID: a091c3ef-9a83-4112-8765-688872d9dd16
Dossier UUID:
Author:Date: 2022-11-07T15:04:18.000+09:00
Remarks:

General information
Reference Typestudy report
Title
In Vitro Chromosomal Aberration Test of on 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane Cultured
Chinese Hamster Cells.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Year
2011
Bibliographic source
Japan Existing Chemical Data Base (JECDB)
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9f.pdf
Testing facility
Nihon Bioresearch Inc.
Report number
97053039

LITERATURE: Reverse Mutation Test of 2,2'-[1,2ethanediylbis(oxymethylene)]bis-oxirane on Bacteria.
UUID: 475e1779-7bd5-4491-96f7-4ef097fb790f
Dossier UUID:
Author:Date: 2022-11-07T15:02:24.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Reverse Mutation Test of 2,2'-[1,2-ethanediylbis(oxymethylene)]bis-oxirane on Bacteria.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Year
2011
Bibliographic source
Japan Existing Chemical Data Base (JECDB)
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2224-15-9e.pdf
Testing facility
Nihon Bioresearch Inc.
Report number
90083040Legal Entities
LEGAL_ENTITY: National Institute of Health Sciences
UUID: IUC4-b036ff75-0f3c-323b-b200-ed5f46cf5101
Dossier UUID:
Author:Date: 2022-11-07T15:49:29.000+09:00
Remarks:

General information
Legal entity nameNational Institute of Health Sciences
Remarks
Disclaimer: The contents in this document were created based on the MHLW (Ministry of Health,Labour and Welfare) peer reviewed study reports (in Japanese) in JECDB (Japan Existing Chemical
Database) at http://dra4.nihs.go.jp/mhlw_data/jsp/SearchPageENG.jsp. Authorship is in the Divisionof Risk Assessment, the National Institute of Health Sciences, and the contents do not reflect any official MHLW opinions or any other regulatory policies.

Address
Address 1
Tonomachi 3-25-26
Address 2
Kawasaki-ku
Postal code
210-9501
Town
KawasakiRegion / State
Kanagawa
CountryJapan
JP

Identifiers
Other IT system identifiers
IT system
LEO
ID
10767
IT systemIUCLID441

ID
16558402024DIV750

42