Name: COMPLETE / SUBSTANCE : Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate /2915-49-3 Fri, 16 Dec 2022, 11:34:45+0900 /Legal entity owner: National Institute of Health Sciences
Printing date: 2022-12-16T11:34:45.191+09:00

iTable of Contents
0/0 ............................................................................................................................. 1
National Institute of Health Science ..................................................................... 2
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate ......................................... 3
CORE .......................................................................................................... 3
1 General information .......................................................................... 3
1.10 Assessment approach (assessment entities) ......................... 3
Assessment approach (assessment entities) ........................ 3
OECD .......................................................................................................... 4
D Health Effects .................................................................................. 4
67 Repeated dose toxicity: oral .................................................... 4
Repeated dose toxicity: oral.001 ........................................... 470 Genetic toxicity in vitro ......................................................... 13
Genetic toxicity in vitro.001 ................................................ 13
Genetic toxicity in vitro.002 ................................................ 19
73 Toxicity to reproduction ........................................................ 23
Toxicity to reproduction.001 ............................................... 23
DOMAIN .................................................................................................... 33
Substance ......................................................................................... 33
Substance ................................................................................ 33
References ................................................................................................................ 34
Reference Substances ...................................................................................... 34
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate ............................... 34
Test Materials .................................................................................................. 35
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate ............................... 35
Literatures ....................................................................................................... 36
Combined repeated dose toxicity study with the reproductive/
developmental toxicity screening test of Bis(2-ethylhexan-1-yl) cyclohex-4ene-1,2-dicarboxylate by oral administration in rats ..................................... 36
In Vitro Chromosomal Aberration Test of Bis (2-ethylhexan-1-yl)
cyclohex-4-ene-1,2-dicarboxylate on Cultured Chinese Hamster Cells. ........... 37
Reverse Mutation Test of Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2dicarboxylate on Bacteria. .......................................................................... 38
Legal Entities ................................................................................................... 39
National Institute of Health Sciences, Japan .............................................. 39ii

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Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate / 2915-49-3
Public nameBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
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National Institute of Health Science
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LEGAL_ENTITY: National Institute of Health
Science
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General information
Legal entity nameNational Institute of Health Science2Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2dicarboxylate
CORE
General information
Assessment approach (assessment entities)
FIXED_RECORD: Assessment approach

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OECD
Health Effects
Repeated dose toxicity: oral
ENDPOINT_STUDY_RECORD: Repeated dose toxicity: oral.001

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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
other: The study was conducted in accordance with Test Guidelines and under GLP
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 to reproduction.001 / Bis (2-ethylhexan-1-yl) cyclohex-4ene-1,2-dicarboxylate / 2915-49-3

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 report4Data accessdata published

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
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Specific details on test material used for the study- Name of test material (as cited in study report): Bis(2-ethylhexyl) 4-cyclohexene-1,2-dicarboxylate- Analytical purity: 99.0%- Storage condition of test material: Cold and dark place (3 - 7°C)
- Stability under test conditions: The stability of test material was identified by analysis of the remainder.

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., Atsugi Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation:Male: 382 g (356-419 g), Female: 235 g (219-268 g)- Housing: Animals were individually housed in bracket-type metallic wire-mesh cages (254W × 350D
× 170H mm), from gestation day 17 to lactation day 4, Dams were bred individually or with individual
littermates in plastic cages (340W x 400D x 185H mm) and bedding.
- Diet: Solid feed (NMF: Oriental Yeast Co., ltd.) was given ad libitum.
5

- Water: Tap water was given ad libitum.
- Acclimation period: 19 days
ENVIRONMENTAL CONDITIONS- Temperature (°C): 23±3 (actual temperature: 22-25°C)- Humidity (%): 50±20% (actual humidity: 38-68%)- Air changes (per hr): 10-15- Photoperiod (hrs dark / hrs light): 12 hr dark/12 hr light (light: 7:00~19:00)

Administration / exposure
Route of administration
oral: gavage
Vehiclecorn oil
Details on oral exposure- Amount of vehicle (if gavage): 5 mL/kg- Dosing volume: 5 mL/kg
Analytical verification of doses or concentrations
yes
Duration of treatment / exposure(P) Males: 42 days including 14 days pre-mating(P)Females: 41-46 days including 14 days pre-mating, mating and gestation periods and the days until
day 4 of lactation
Female (no mating, satellite group): 42 days
Frequency of treatment
Once/day, 7 days/week
Doses / concentrations
Dose / conc.
0mg/kg bw/day (actual dose received)

Dose / conc.
30

mg/kg bw/day (actual dose received)

Dose / conc.
100

mg/kg bw/day (actual dose received)

Dose / conc.
300

mg/kg bw/day (actual dose received)

No. of animals per sex per dose
Main group: 12 females/dose (0, 30, 100, and 300 mg/kg bw/day), 7, 12, 12, and 7 males/dose (0, 30,
100, and 300 mg/kg bw/day)Satellite group: 5 females/dose (0 and 300 mg/kg bw/day)6

Recovery group: 5 males/dose and 5 females (satellite group)/dose (0 and 300 mg/kg bw/day)Control animals
yes, concurrent vehicle
Details on study design- Dose selection rationale: Based on the results of a 14-day preliminary study, the highest dose of 300
mg/kg bw/day was selected as an expected obvious toxic dose, and the lowest dose of 30 mg/kg
bw/day was selected as an expected no toxic dose. The middle dose levels of 100 mg/kg bw/day
were selected.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 100, 300 or 1000 mg/kg bw/day).
In the 300 mg/kg bw/day or more, high liver weight were observed.
- Rationale for animal assignment (if not random): Body weight-balanced randomization

Examinations
Observations and examinations performed and frequency
CAGE SIDE OBSERVATIONS: Yes- Time schedule: 3 times/day (before administration, immediately after administration and 2 hours
after administration) during the administration period. Once a day during the recovery period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Male main and female satellite groups: once before the start of administration, once every weekly
during the administration.
Female main group: once before the start of administration, days 1, 7, 14 and 20 of gestation, and day
4 of lactation.
Male and female recovery groups: once before the start of administration, once every weekly during the administration and recovery periods.
BODY WEIGHT: Yes- Time schedule for examinations:
Males in the main and females satellite groups were weighed on days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32,
36, 39 and 42 of administration and on the day of necropsy, and males and females in the recovery
groups were weighed on days 1, 4, 8, 11 and 14 of recovery and on the day of necropsy in addition to
the measurement days for males in the main groups.
Females in the main groups were weighed on days 1, 4, 8, 11 and 15 of administration (uncopulated
animals were weighed on days 18 and 22 of administration as well), days 0, 4, 7, 11, 14, 17 and 20 of
gestation, days 0 and 4 of lactation and the day of necropsy.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):- Food consumption: Yes
Measurement of food consumption was conducted on all animals at the following frequencies:
males in the main and females satellite groups on days 1, 4, 8, 11, 15, 32, 36, 39 and 42 of admini
stration; males and females in the recovery groups on days 1, 4, 8, 11 and 14 of recovery in addition to
the measurement days for males in the main groups; and females in the main groups on days 1, 4, 8,
11 and 15 of administration, days 1, 4, 7, 11, 14, 17 and 20 of gestation and days 2 and 4 of lactation.
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes- Time schedule for collection of blood: At the end of administration period, or at the end of recovery
period in both sexes- Anaesthetic used for blood collection: ether- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),7

5 animals/sex/group (30 and 100 mg/kg/day)- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte pe
rcentage, platelet count, white blood cell count, differential white blood cell count, absolute number of
each white blood cell, prothrombin time, activated partial thromboplastin time, fibrinogen.
CLINICAL CHEMISTRY: Yes- Time schedule for collection of blood: At the end of administration period, or at the end of recovery
period in both sexes- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),5 animals/sex/group (30 and 100 mg/kg/day)- Parameters checked: ALP, total cholesterol, triglyceride, phospholipids, total bilirubin, glucose, blood urea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein,
albumin, A/G ratio, AST (GOT), ALT (GPT), LDH, γ-GTP
URINALYSIS: Yes- Time schedule for collection of urine: final week of administration (days 37 to 38 of administration)and in the final week of recovery (days 9 to 10 of recovery)- Metabolism cages used for collection of urine: Yes
A urine collector to collect four-hour urine samples under fasting but ad libitum drinking conditions,followed by collection of 20-hour urine samples under ad libitum feeding and drinking conditions.
- How many animals: 5 animals/group- Parameters checked: pH, protein, ketones, glucose, occult blood, bilirubin, urobilinogen, color, sediment, urine volume (4-hour volume), osmotic pressure, urine volume (20-hour volume), water intake(24-hour volume)
BLOOD HORMONE: Yes- Time schedule for collection of serum: Same as clinical chemistry- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),5 animals/sex/group (30 and 100 mg/kg/day)- Parameters checked: Triiodothyronine (T3), Thyroxin (T4), and thyroid stimulating hormone (TSH)
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males in the main groups: final week of administration (day 40 of administration)
Females in the main groups: lactation day 4 (day 41 to day 44 of administration) after necropsy of F1
pups
Males and females in the recovery groups: final week of administration (day 40 of administration) andin the final week of recovery (day 12 of recovery).
- Dose groups that were examined: All dose groups (5 animals/sex/group)- Battery of functions tested:
1) Manipulative Test. Auditory response, approach response, touch response, tail pinch response,
pupillary reflex, aerial righting reflex, landing foot splay
2) Measurement of Grip Strength. Following manipulative test, grip strength of forelimb and hind limb
was measured by CPU gauge MODEL-RX-5 (AIKOH Engineering Co., Ltd.).
3) Measurement of Motor Activity. Following measurement of grip strength, motor activity wasmeasured by a motor activity sensor for experimental animals NS-AS01 (Neuro Science, Inc). The
measurement was conducted for 1 hour, and measured values at 10-minute intervals and from 0 to
60 minutes were collected.
Sacrifice and pathology
GROSS PATHOLOGY: Yes
ORGAN WEIBHT: Yes [brain, pituitary, thyroids (including parathyroids), adrenal gland, thymus, spleen,
heart, liver, kidney, testis, epididymis, ovary, uterus]
8HISTOPATHOLOGY: Yes, [cerebrum, cerebellum (including pons) , pituitary, spinal cord (thoracic), sc
iatic nerve, eye ball, thyroid, parathyroid, adrenal glands, thymus, spleen, submandibular lymph nod
es, mesenteric lymph nodes, heart, trachea, lung (including bronchial), stomach, duodenum, jejunum,
ileum, cecum, colon, rectum, liver, kidney, bladder, testis, epididymis, ovary, uterus, vagina, prostate,seminal vesicles, sternum and femur (including bone marrows),and macroscopic lesions]
Statistics
The data were analyzed for homogeneity of variance by the Bartlett test. If variances were
homogeneous, data was analyzed by the Dunnett test, whereas heterogeneous data was analyzed by
the Dunnett type mean rank test (p<0.05, two-sided).
In the recovery test, these values of two groups were analyzed by F test. If variances were homogeneo
us, data was analyzed by the Student t-test, whereas heterogeneous data was analyzed by the AspinWelch t-test (p<0.05, two-sided).

Results and discussion
Results of examinations
Clinical signs
no effects observed
Mortality
no mortality observed
Body weight and weight changes
no effects observed
Food consumption and compound intake (if feeding study)no effects observed
Food efficiency
not examined
Water consumption and compound intake (if drinking water study)
not examined
Ophthalmological findings
not examined
Haematological findings
effects observed, treatment-related
Clinical biochemistry findings
no effects observed
Description (incidence and severity)
Including blood hormones (T3, T4, TSH)
Urinalysis findings
no effects observed
Behaviour (functional findings)
no effects observed
Immunological findings
not examined
9Organ weight findings including organ / body weight ratios
effects observed, treatment-related
Gross pathological findings
no effects observed
Neuropathological findings
not examined
Histopathological findings: non-neoplastic
effects observed, treatment-related
Histopathological findings: neoplastic
not examined
Details on results
CLINICAL SIGNS AND MORTALITY:
Mortality: There was no death.
Clinical signs: There were no effects related to the test substance in any groups at the dosing andrecovery periods.
DETAILED CLINICAL OBSERVATIONS: There were no changes related to the test substance in anygroups at the dosing and recovery periods.
BODY WEIGHT:
There were no changes related to the test substance in any groups at the dosing and recovery periods.
FOOD CONSUMPTION:
There were no changes related to the test substance in any groups at the dosing and recovery peri
ods.
URINALYSIS:There were no changes related to the test substance in any groups at the dosing and recovery
periods.
NEUROBEHAVOURAL EXAMINATION: There were no changes related to the test substance in anygroups at the dosing and recovery periods.
HAEMATOLOGY:[At the end of dosing period]: Decreases in white blood cell count, lymphocyte count, neutrophil countand large unstained cell count were observed in males receiving 300 mg/kg bw/day.
[At the end of recovery period]: There were no changes related to the test substance in any groups.
CLINICAL CHEMISTRY (Including blood hormones (T3, T4, TSH)):
There were no changes related to the test substance in any groups at the end of dosing and recovery
periods.
ORGAN WEIGHTS:[At the end of dosing period]: Increases in absolute and relative weights of liver were observed inmating and non-mating females receiving 300 mg/kg bw/day.
Increases in relative weights of liver and absolute and relative weights of kidney were observed inmales receiving 300 mg/kg bw/day.
[At the end of recovery period]: Increases in absolute and relative weights of liver were observed inmales receiving 300 mg/kg bw/day.
GROSS PATHOLOGY:
There were no changes related to the test substance in any groups at the end of dosing and recovery
periods.
10

HISTOPATHOLOGY: NON-NEOPLASTIC:[At the end of dosing period]:
Liver: Minimal centrilobular hypertrophy of hepatocytes were observed in females receiving 100 mg/
kg bw/day or more, and males receiving 300 mg/kg bw/day.
[At the end of recovery period]: There were no changes related to the test substance in any groups.

Effect levels
Key result
true
Dose descriptor
NOAEL
Effect level30

mg/kg bw/day (actual dose received)

Based ontest mat.
Sex
femaleBasis for effect level
histopathology: non-neoplastic
At 100 mg/kg bw/day and above, centrilobular hypertrophy of hepatocytes were observed in females.

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/PDF2915-49-3d.pdf

Applicant's summary and conclusion
Conclusions
Based on the effects of the liver at 100 mg/kg bw/day in females, the NOAEL for repeated-dose toxici
ty of bis(2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate was 30 mg/kg bw/day in rats.
Executive summary
A combined repeated-dose toxicity study with the reproduction/developmental toxicity screening test
was done according to OECD TG 422. Male and female rats (12 animals/sex/dose) were administeredwith bis(2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate via oral gavage at doses of 0 [vehicle: cornoil], 30, 100, and 300 mg/kg bw/day. Males (12/dose) were administered with bis(2-ethylhexan-1yl) cyclohex-4-ene-1,2-dicarboxylate for 42 days, which include a 14-day premating period and asubsequent mating period, while females (12/dose) were treated for 41–46 days, which include 14-day
premating, mating, and gestation periods until lactation day 4. Among the 12 males administered with 0
and 300 mg/kg bw/day, 5 were assigned as the recovery group. Additionally, 10 females administeredwith 0 and 300 mg/kg bw/day were assigned as a satellite group and treated with bis(2-ethylhexan-111

yl) cyclohex-4-ene-1,2-dicarboxylate for 42 days, without mating, and assessed after a 14-day recovery
period.
No deaths were recorded, and there are no changes in clinical signs, manipulative test, grip strength,
motor activity, body weight, food consumption, urinalysis, blood chemistry, blood hormone (T3, T4,
and TSH), and gross pathological findings resulting from the treatment in any of the dose groups forboth sexes at the end of the treatment and recovery periods. The level of white blood cell, lymphocyte,
neutrophil, and large unstained cells was significantly decreased in males receiving 300 mg/kg bw/day
at the end of the administration period. The absolute and relative weights of liver were significantly
increased as well in both sexes receiving 300 mg/kg bw/day at the end of the administration period.
There was also significant increase in absolute and relative weights of kidney in the males receiving 300
mg/kg bw/day at the end of the administration period. Histopathological analysis has indicated a slight
hypertrophy of centrilobular hepatocytes in males receiving 300 mg/kg bw/day and in females 100 and
300 mg/kg bw/day at the end of the administration period. Since these changes lessen or disappear at
the end of the recovery period, they are thought to be reversible. Based on the effects of the liver at 100
mg/kg bw/day in females, the NOAEL for repeated-dose toxicity of bis(2-ethylhexan-1-yl) cyclohex-4ene-1,2-dicarboxylatewas 30 mg/kg bw/day in rats.

12

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

UUID: 9ffd22b5-6134-4e88-bdef-7bc740e4aec4
Dossier UUID:
Author:Date: 2022-12-14T16:25:23.602+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
other: OECD Test Guideline study under GLP condition

Data source
Reference
Reverse Mutation Test of Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate on Bacteria. / Ministryof Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published

Materials and methods
Test guideline
Qualifieraccording to guideline
Guideline
OECD Guideline 471 (Bacterial Reverse Mutation Assay)13

in vitro gene mutation study in bacteria
Deviationsno
GLP compliance
yesType of assay
bacterial reverse mutation assay
in vitro gene mutation study in bacteria

Test material
Test material information
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Specific details on test material used for the study- Name of test material (cited in study report): Bis(2-ethylhexyl) 4-cyclohexene-1,2-dicarboxylate- Purity 99.0%

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 pKM 101
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, 313 μg/plate (TA100 strain)
2.44, 4.88, 9.77, 19.5, 39.1, 78.1μg/plate (TA1535, TA 98, TA 1537 strains),
156, 313, 625, 1250, 2500, 5000 μg/plate (WP2uvrA/pKM101 strain)+S9 mix:
39.1, 78.1, 156, 313, 625, 1250 μg/plate (TA100, TA1535, TA98, TA1537 strains),
156, 313, 625, 1250, 2500, 5000 μg/plate (WP2uvrA/pKM101 strain)
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 78.1 μg/plate and above forS. typhimurium 1535, TA98 and 1537 without S9 mix, at 313 μg/plate and above for S. typhimurium
TA100 without S9 mix, at 1250 μg/plate and above for S. typhimurium TA strains with S9 mix and at
5000 μg/plate for E. coli WP2 uvrA with or without S9 mix.
Vehicle / solvent- Vehicle(s)/solvent(s) used: Acetone
14

Controls
Untreated negative controlsnoNegative solvent / vehicle controls
yes
True negative controlsnoPositive controls
yes
Positive control substanceother: -S9 mix: 2-(2-Furyl)-3-(5-nitro-2-furyl) acrylamide, sodium azide and 2-Methoxy-6-chloro-9-[3-(2chloroethyl)-aminopropylamino]acridine 2HCl; +S9 mix: 2-aminoanthracene, benzo(a)pyrene
Details on test system and experimental conditions
METHOD OF APPLICATION: Preincubation
DURATION- Preincubation period: 20 min at 37°C
- Exposure duration:48 or 49 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
Test results
Key result
trueSpecies / strain
S. typhimurium TA 1535
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 78.1 μg/plate; +S9 mix: 1250 μg/plate
Vehicle controls validity
valid
15

Untreated negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
S. typhimurium TA 1537
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 39.1 μg/plate or more (test 1: 78.1 μg/plate) ; +S9 mix: 625 μg/plate or more (test 1: 1250 μg/plate)
Vehicle controls validity
valid
Untreated negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
S. typhimurium TA 98
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 78.1 μg/plate; +S9 mix: 1250 μg/plate
Vehicle controls validity
valid
Untreated negative controls validity
not examined
Positive controls validity
valid
Key result
true
16Species / strain
S. typhimurium TA 100
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 313 μg/plate or more; +S9 mix: 1250 μg/plate or more
Vehicle controls validity
valid
Untreated negative controls validity
not examined
Positive controls validity
valid
Key result
trueSpecies / strain
E. coli WP2 uvr A pKM 101
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 5000 μg/plate; +S9 mix: 2500 μg/plate or more (test 2: 5000 μg/plate)
Vehicle controls validity
valid
Untreated negative controls validity
not examined
Positive controls validityvalid

Any other information on results incl. tables
Figures and Tables (in Japanese) are available in the following full report of the study.
http://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2915 -49 -3e.pdf
Please also see the attached files (Tables in English)

Overall remarks, attachments
17

Attachments
Attached (sanitised) documents for publication
35948-25-5_Ames.xlsx / 34.77 KB (application/vnd.openxmlformatsofficedocument.spreadsheetml.sheet)

Applicant's summary and conclusion
Conclusions
Interpretation of results (migrated information): negative
In a bacterial reverse mutation assay using Salmonella typhimurium TA100, TA1535, TA98, and TA
1537, and Escherichia coli WP2uvrA/pKM101 (OECD TG 471), Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate was negative with or without metabolic activation.

18

ENDPOINT_STUDY_RECORD: Genetic toxicity in vitro.002

UUID: f14b50e2-3b33-4b45-b578-61865da78280
Dossier UUID:
Author:Date: 2022-12-14T16:26:43.880+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
other: OECD Test Guideline study under GLP condition

Data source
Reference
In Vitro Chromosomal Aberration Test of Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate on /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 Chromosome Aberration Test)in vitro cytogenicity / chromosome aberration study in mammalian cells
19

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

Test material
Test material information
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Specific details on test material used for the study- Name of test material (cited in study report): Bis(2-ethylhexyl) 4-cyclohexene-1,2-dicarboxylate- Purity 99.0%

MethodSpecies / strain
Species / strain / cell typeother: Chinese hamster lung(CHL/IU) cells
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): 31.3, 62.5, 125, 250, 500, 1000, 2000, 4000 ug/mL+S9 mix (short-term treatment): 31.3, 62.5, 125, 250, 500, 1000, 2000, 4000 ug/mL-S9 mix (continuous treatment, 24hr): 31.3, 62.5, 125, 250, 500, 1000, 2000, 4000 ug/mL-S9 mix (continuous treatment, 48hr): 31.3, 62.5, 125, 250, 500, 1000, 2000, 4000 ug/mL (IC50=52.9
ug/mL)
Main study-S9 (short-term treatment): 1000, 2000, 4000 ug/mL+S9 (short-term treatment): 1000, 2000, 4000 ug/mL-S9 (continuous treatment, 24hr):1000, 2000, 4000 ug/mL-S9 (continuous treatment, 48hr): 7.81, 15.6, 31.3, 62.5, 125 ug/mL
Vehicle / solvent- Vehicle(s)/solvent(s) used: Acetone
20

Controls
Untreated negative controlsnoNegative solvent / vehicle controls
yes
True negative controlsnoPositive controls
yes
Positive control substanceother: [-S9]: mitomycin C; [+S9]: cyclophosphamide
Details on test system and experimental conditions
METHOD OF APPLICATION: Exposure duration: [short-term treatment]:6 hrs + 18 hr, [continuous
treatment]: 24, 48 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 / strainother: Chinese hamster lung (CHL/IU) cells
Metabolic activation
without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
21

Positive controls validity
valid
Key result
trueSpecies / strainother: Chinese hamster lung (CHL/IU) cells
Metabolic activation
with
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
no cytotoxicity
Vehicle controls validity
valid
Untreated negative controls validity
not examined
Positive controls validityvalid

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/PDF2915-49-3f.pdf

Applicant's summary and conclusion
Conclusions
Interpretation of results (migrated information): negative with or without metabolic activation
The in vitro chromosomal aberration test using CHL/IU cells (OECD TG 473) was negative with orwithout metabolic activation.

22

Toxicity to reproduction
ENDPOINT_STUDY_RECORD: Toxicity to reproduction.001

UUID: 23453dda-38c1-4d2b-bbfe-d40fb559c6da
Dossier UUID:
Author:Date: 2022-12-16T11:34:08.444+09:00
Remarks:

Administrative data
Endpoint
reproductive toxicity, other A combined repeated dose/reproductive developmental toxicity study
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
other: OECD Test Guideline study under GLP condition
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 / Bis (2-ethylhexan-1-yl)
cyclohex-4-ene-1,2-dicarboxylate / 2915-49-3

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

Materials and methods
23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
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Specific details on test material used for the study- Name of test material (as cited in study report): Bis(2-ethylhexyl) 4-Cyclohexene-1,2-dicarboxylate- Analytical purity: 99.0%- Storage condition of test material: Cold and dark place (3 - 7°C)
- Stability under test conditions: The stability of test material was identified by analysis of the remainder.

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., Atsugi Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation: Male: 382 g (356-419 g), Female: 235 g (219-268 g)- Housing: Animals were individually housed in bracket-type metallic wire-mesh cages (254W × 350D
× 170H mm), from gestation day 17 to lactation day 4, Dams were bred individually or with individual
littermates in plastic cages (340W x 400D x 185H mm) and bedding.
- Diet: Solid feed (NMF: Oriental Yeast Co., ltd.) was given ad libitum.
- Water: Tap water was given ad libitum.
- Acclimation period: 19 days
ENVIRONMENTAL CONDITIONS- Temperature (°C): 23±3 (actual temperature: 22-25°C)- Humidity (%): 50±20% (actual humidity: 38-68%)- Air changes (per hr): 10-15- Photoperiod (hrs dark / hrs light): 12 hr dark/12 hr light (light: 7:00~19:00)

24

Administration / exposure
Route of administration
oral: gavage
Vehicleother: Water 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 6 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
Test suspensions at each concentration to be used for males in week 1 and six week of admi
nistration were analyzed by HPLC method at BoZo Research Center Inc. Results showed that theconcentration of test article in each concentration was 102.2 to 106.0% of the nominal concentration
and both values were within the acceptable range (concentration: percentage of nominal concentration, 100±10%)Duration of treatment / exposure(P) Males: 42 days including 14 days pre-mating(P)Females: 41-46 days including 14 days pre-mating, mating and gestation periods and the days until
day 4 of lactation
Female (no mating, satellite group): 42 days

Frequency of treatment
Once/day, 7 days/week
Doses / concentrations
Dose / conc.
0mg/kg bw/day (actual dose received)

Dose / conc.
30

mg/kg bw/day (actual dose received)

Dose / conc.
100

mg/kg bw/day (actual dose received)

Dose / conc.
300

mg/kg bw/day (actual dose received)

25

No. of animals per sex per dose
Main group:12 females/dose (0, 30, 100, and 300 mg/kg bw/day), 7, 12, 12, and 7 males/dose (0, 30, 100, and 300 mg/kg bw/day)Satellite group: 5 females/dose (0 and 300 mg/kg bw/day)Recovery group: 5 males/dose and 5 females (satellite group)/dose (0 and 300 mg/kg bw/day)Control animals
yes, concurrent no treatment
Details on study design- Dose selection rationale: Based on the results of a 14-day preliminary study, the highest dose of 300
mg/kg bw/day was selected as an expected obvious toxic dose, and the lowest dose of 30 mg/kg
bw/day was selected as an expected no toxic dose. The middle dose levels of 100 mg/kg bw/day
were selected.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 100, 300 or 1000 mg/kg bw/day).
In the 300 mg/kg bw/day or more, high liver weight was observed.
- Rationale for animal assignment (if not random): Body weight-balanced randomization

Examinations
Parental animals: Observations and examinations
CAGE SIDE OBSERVATIONS: Yes- Time schedule: 3 times/day (before administration, immediately after administration and 2 hours
after administration) during the administration period. Once a day during the recovery period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Male main and female satellite groups: once before the start of administration, once every weekly
during the administration.
Female main group: once before the start of administration, days 1, 7, 14 and 20 of gestation, and day
4 of lactation.
Male and female recovery groups: once before the start of administration, once every weekly during the administration and recovery periods.
BODY WEIGHT: Yes- Time schedule for examinations:
Males in the main and females satellite groups were weighed on days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32,
36, 39 and 42 of administration and on the day of necropsy, and males and females in the recovery
groups were weighed on days 1, 4, 8, 11 and 14 of recovery and on the day of necropsy in addition to
the measurement days for males in the main groups.
Females in the main groups were weighed on days 1, 4, 8, 11 and 15 of administration (uncopulated
animals were weighed on days 18 and 22 of administration as well), days 0, 4, 7, 11, 14, 17 and 20 of
gestation, days 0 and 4 of lactation and the day of necropsy.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):- Food consumption: Yes
Measurement of food consumption was conducted on all animals at the following frequencies:
males in the main and females satellite groups on days 1, 4, 8, 11, 15, 32, 36, 39 and 42 of admini
stration; males and females in the recovery groups on days 1, 4, 8, 11 and 14 of recovery in addition to
the measurement days for males in the main groups; and females in the main groups on days 1, 4, 8,
11 and 15 of administration, days 1, 4, 7, 11, 14, 17 and 20 of gestation and days 2 and 4 of lactation.
OPHTHALMOSCOPIC EXAMINATION: No

26

HAEMATOLOGY: Yes- Time schedule for collection of blood: At the end of administration period, or at the end of recovery
period in both sexes- Anaesthetic used for blood collection: ether- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),5 animals/sex/group (30 and 100 mg/kg/day)- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte pe
rcentage, platelet count, white blood cell count, differential white blood cell count, absolute number of
each white blood cell, prothrombin time, activated partial thromboplastin time, fibrinogen.
CLINICAL CHEMISTRY: Yes- Time schedule for collection of blood: At the end of administration period, or at the end of recovery
period in both sexes- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),5 animals/sex/group (30 and 100 mg/kg/day)- Parameters checked: ALP, total cholesterol, triglyceride, phospholipids, total bilirubin, glucose, blood urea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein,
albumin, A/G ratio, AST (GOT), ALT (GPT), LDH, γ-GTP
URINALYSIS: Yes- Time schedule for collection of urine: final week of administration (days 37 to 38 of administration)and in the final week of recovery (days 9 to 10 of recovery)- Metabolism cages used for collection of urine: Yes
A urine collector to collect four-hour urine samples under fasting but ad libitum drinking conditions,followed by collection of 20-hour urine samples under ad libitum feeding and drinking conditions.
- How many animals: 5 animals/group- Parameters checked: pH, protein, ketones, glucose, occult blood, bilirubin, urobilinogen, color, sediment, urine volume (4-hour volume), osmotic pressure, urine volume (20-hour volume), water intake(24-hour volume)
BLOOD HORMONE: Yes- Time schedule for collection of serum: Same as clinical chemistry- Animals fasted: Yes- How many animals:
All animals/sex/group (Control and 300 mg/kg/day),5 animals/sex/group (30 and 100 mg/kg/day)- Parameters checked: Triiodothyronine (T3), Thyroxin (T4), and thyroid stimulating hormone (TSH)
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males in the main groups: final week of administration (day 40 of administration)
Females in the main groups: lactation day 4 (day 41 to day 44 of administration) after necropsy of F1
pups
Males and females in the recovery groups: final week of administration (day 40 of administration) andin the final week of recovery (day 12 of recovery).
- Dose groups that were examined: All dose groups (5 animals/sex/group)- Battery of functions tested:
1) Manipulative Test. Auditory response, approach response, touch response, tail pinch response,
pupillary reflex, aerial righting reflex, landing foot splay
2) Measurement of Grip Strength. Following manipulative test, grip strength of forelimb and hind limb
was measured by CPU gauge MODEL-RX-5 (AIKOH Engineering Co., Ltd.).
3) Measurement of Motor Activity. Following measurement of grip strength, motor activity wasmeasured by a motor activity sensor for experimental animals NS-AS01 (Neuro Science, Inc). The

27

measurement was conducted for 1 hour, and measured values at 10-minute intervals and from 0 to
60 minutes were collected.
Oestrous cyclicity (parental animals)
Vaginal smears were collected from all females in the main groups and microscopically examined
every day from the day after the start of administration until the day copulation was confirmed.
During the pre-mating administration period, vaginal smear pictures were classified as proestrus,
estrus, metestrus or diestrus and examined for the frequency of estrus and interval between estruses(estrous cycle). During the mating period, vaginal smears were examined for the presence of sperm.
Sperm parameters (parental animals)
Parameters examined in all P male parental generations: testis weight, epididymis weight, histopatho
logical 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 ether anesthesia.
SACRIFICE: Male main and female satellite animals: On next day after the last administration (Day43), Maternal animals: on Day 4 of lactation, and male and females recovery animals: on Day 14 of recovery.
GROSS PATHOLOGY: Yes
ORGAN WEIBHT: Yes [brain, pituitary, thyroids (including parathyroids), adrenal gland, thymus, spleen,
heart, liver, kidney, testis, epididymis, ovary, uterus]
HISTOPATHOLOGY: Yes, [cerebrum, cerebellum (including pons) , pituitary, spinal cord (thoracic), scia
tic nerve, eye ball, thyroid, parathyroid, adrenal glands, thymus, spleen, submandibular lymph nodes, mesenteric lymph nodes, heart, trachea, lung (including bronchial), stomach, duodenum, jejunum,
ileum, cecum, colon, rectum, liver, kidney, bladder, testis, epididymis, ovary, uterus, vagina, prostate,seminal vesicles, sternum and femur (including bone marrows),and macroscopic lesions]
Postmortem examinations (offspring)
SACRIFICE- The F1 offsprings were euthanized on PND4 by exsanguination under ether anesthesia.
GROSS NECROPSY: Yes- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, andabdominal viscera.
HISTOPATHOLOGY / ORGAN WEIGTHS- Not examined.
Statistics
The data were analyzed for homogeneity of variance by the Bartlett test. If variances were
homogeneous, data was analyzed by the Dunnett test, whereas heterogeneous data was analyzed by
the Dunnett type mean rank test (p<0.05, two-sided).
In the recovery test, these values of two groups were analyzed by F test. If variances were homogeneo
us, data was analyzed by the Student t-test, whereas heterogeneous data was analyzed by the AspinWelch t-test (p<0.05, two-sided).
Reproductive indices
Each parameter was determined by the following equations:
Copulation index (%) = (No. of copulated animals / No. of mated animals) × 100
Fertility index (%) = (No. of pregnant females / No. of copulated females) × 100
Insemination index (%) = (No. of impregnated males / No. of copulated males) × 100
28

Gestation length (days) = No. of days from pregnancy 0 to delivery 0
Delivery index (%) = (No. of females which delivered liveborns / No. of pregnant females) × 100
Implantation index (%) = (No. of implantation sites / No. of corpora lutea) × 100
Stillborn index (%) = (No. of stillborn / No of liveborns and stillborns) × 100
Live birth index (%) = (No. of liveborn / No. of implantation sites) × 100
External abnormalities (%) = (No. of pups with external abnormalities / No. of liveborns) × 100
Sex ratio = No. of liveborns males / No. of liveborns
Offspring viability indices
Viability index on postnatal day 4 (%) = (No. of live pups on day 4 / No. of liveborns on day 0) × 100

Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
Clinical signs
no effects observed
Mortality
no mortality observed
Body weight and weight changes
no effects observed
Food consumption and compound intake (if feeding study)no effects observed
Food efficiency
not examined
Water consumption and compound intake (if drinking water study)
not examined
Ophthalmological findings
not examined
Haematological findings
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1
Clinical biochemistry findings
no effects observed
Description (incidence and severity)
Including blood hormones (T3, T4, TSH)
Urinalysis findings
no effects observed
Behaviour (functional findings)
no effects observed29Immunological findings
not examined
Organ weight findings including organ / body weight ratios
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1
Gross pathological findings
no effects observed
Neuropathological findings
not examined
Histopathological findings: non-neoplastic
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1
Histopathological findings: neoplastic
not examined

Reproductive function / performance (P0)
Reproductive function: oestrous cycle
no effects observed
Reproductive function: sperm measures
not examined
Reproductive performanceno effects observed

Details on results (P0)1) Estrous Cycle
There were no animals showing abnormal estrous cycles, and there were no significant differences in
the average length of the estrous cycle between the control group and any treatment groups.
2) Results of Mating
There were no significant differences in the incidence of females with irregular estrus cycle, mating
period with the number of estrus and day of conceiving, copulation index, and fertility index between
the control group and any treatment groups.
3) Delivery Data and Delivery
There were no significant differences in the gestation length, number of corpora lutea, number of imp
lantation sites, implantation index, and delivery index between the control group and any treatmentgroups.
Haematological findings
See 7.5.1
Organ weight findings
See 7.5.1
HISTOPATHOLOGY
See 7.5.1

Effect levels (P0)30Key result
true
Dose descriptor
NOAEL
Effect level300

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
other: No effects on reproduction

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

Effect levels (F1)
Key result
true
Dose descriptor
NOAEL
Generation
F1
Effect level300

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
other: No effects on development

Overall reproductive toxicity
31

Key result
true
Reproductive effects observednoAny 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/PDF2915-49-3d.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) , there were no effects on reproductive and developmental parameters up to
300 mg/kg bw/day. The NOAEL for the rat reproductive/developmental toxicity of bis(2-ethylhexan-1yl) cyclohex-4-ene-1,2-dicarboxylate was regarded as 300 mg/kg bw/day, the highest dose tested.
Executive summary
A combined repeated-dose toxicity study with the reproduction/developmental toxicity screening test
was done according to OECD TG 422. Male and female rats (12 animals/sex/dose) were administeredwith bis(2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate via oral gavage at doses of 0 [vehicle: cornoil], 30, 100, and 300 mg/kg bw/day. Males (12/dose) were administered with bis(2-ethylhexan-1yl) cyclohex-4-ene-1,2-dicarboxylate for 42 days, which include a 14-day premating period and asubsequent mating period, while females (12/dose) were treated for 41–46 days, which include 14-day
premating, mating, and gestation periods until lactation day 4. Among the 12 males administered with 0
and 300 mg/kg bw/day, 5 were assigned as the recovery group. Additionally, 10 females administeredwith 0 and 300 mg/kg bw/day were assigned as a satellite group and treated with bis(2-ethylhexan-1yl) cyclohex-4-ene-1,2-dicarboxylate for 42 days, without mating, and assessed after a 14-day recovery
period.
No mortalities were recorded with any dose during the treatment period. There were also no effects
on reproductive toxicity (fertility and reproductive organs) and developmental toxicity up to the highest
dose. Because there was no effect observed at 300 mg/kg bw/day administration, the NOAEL for the
reproduction and development toxicity was 300 mg/kg bw/day in rats.

32

DOMAIN
Substance
SUBSTANCE: Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
UUID: e7c03a78-48a6-4292-9759-dac5a84ae551
Dossier UUID:
Author:Date: 2022-12-16T11:34:31.593+09:00
Remarks:
Substance nameBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Public nameBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
Legal entity
National Institute of Health Sciences, Japan

Identification of substance
Reference substanceBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate / 2915-49-3
EC number

EC name

CAS number

CAS name

2915-49-3
IUPAC nameRole in the supply chain
Manufacturer
false
Importer
false
Only representativefalse
Downstream user
false

33

References
Reference Substances
REFERENCE_SUBSTANCE: Bis (2-ethylhexan-1-yl)
cyclohex-4-ene-1,2-dicarboxylate
UUID: 2d5c054f-75a7-4fd4-9f3e-4b68ed1f75a1
Dossier UUID:
Author:Date: 2019-12-19T09:31:18.000+09:00
Remarks:
Reference substance nameBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate

Inventory
CAS number
2915-49-3

34

Test Materials
TEST_MATERIAL_INFORMATION: Bis (2-ethylhexan-1-yl)
cyclohex-4-ene-1,2-dicarboxylate
UUID: 492974ed-339a-4115-8a33-fe2da576efe4
Dossier UUID:
Author:Date: 2019-12-19T09:39:02.000+09:00
Remarks:Name
Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate

35

Literatures
LITERATURE: Combined repeated dose toxicity study withthe reproductive/developmental toxicity screening test ofBis(2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
by oral administration in rats
UUID: fd162373-8e99-476c-8cd6-83e09587026b
Dossier UUID:
Author:Date: 2020-03-24T08:44:51.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Combined repeated dose toxicity study with the reproductive/developmental toxicity screening test ofBis(2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate by oral administration in rats
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Bibliographic source
available in the web of Japan Existing Chemical Data Base (JECDB) https://dra4.nihs.go.jp/
mhlw_data/home/pdf/PDF2915-49-3d.pdf
Testing facility
BoZo Research Center
Report number
R-1051

36

LITERATURE: In Vitro Chromosomal Aberration Test ofBis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate
on Cultured Chinese Hamster Cells.
UUID: 8c0542f6-e903-4fd8-a962-902ea68b154d
Dossier UUID:
Author:Date: 2020-03-17T15:32:41.000+09:00
Remarks:

General information
Reference Typestudy report
Title
In Vitro Chromosomal Aberration Test of Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate on
Cultured Chinese Hamster Cells.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Bibliographic source
Japan Existing Chemical Data Base (JECDB) https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF2
915-49-3f.pdf
Testing facility
Bozo Research Center Inc.
Report number
M-140837

LITERATURE: Reverse Mutation Test of Bis (2ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate on
Bacteria.
UUID: 2b1d855f-1987-4bac-a8e1-a21083261583
Dossier UUID:
Author:Date: 2020-03-17T15:11:21.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Reverse Mutation Test of Bis (2-ethylhexan-1-yl) cyclohex-4-ene-1,2-dicarboxylate on Bacteria.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Bibliographic source
Japan Existing Chemical Data Base (JECDB) http://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF291
5-49-3e.pdf
Testing facility
Bozo Research Center Inc.
Report number
T-046738

Legal Entities
LEGAL_ENTITY: National Institute of Health Sciences,Japan
UUID: 0952b3b9-2d0c-4bc8-925e-b069be7789b7
Dossier UUID:
Author:Date: 2020-02-19T14:42:16.000+09:00
Remarks:

General information
Legal entity nameNational Institute of Health Sciences, Japan

39