Name: COMPLETE / SUBSTANCE : 2,4-Di-tert-pentylphenol / 120-95-6 Tue, 5 Sep 2023,
13:10:56+0900 /Legal entity owner: National Institute of Health Sciences / Kawasaki / Japan
Printing date: 2023-09-05T13:10:57.157+09:00

iTable of Contents
0/0 ............................................................................................................................. 1
National Institute of Health Sciences .................................................................... 2
2,4-Di-tert-pentylphenol ........................................................................................ 4
OECD .......................................................................................................... 4
D Health Effects .................................................................................. 4
67 Repeated dose toxicity: oral .................................................... 4
RepeatedDoseToxicityOral. 001 ............................................ 470 Genetic toxicity in vitro ......................................................... 14
Genetic toxicity in vitro.001 ................................................ 14
Genetic toxicity in vitro.002 ................................................ 19
73 Toxicity to reproduction ........................................................ 23
ToxicityReproduction. 001 .................................................. 23
DOMAIN .................................................................................................... 33
2,4-Di-tert-pentylphenol ...................................................................... 33
References ................................................................................................................ 34
Reference Substances ...................................................................................... 34
2,4-Di-tert-amylphenol ................................................................................ 34
Test Materials .................................................................................................. 35
2,4-Di-tert-pentylphenol .............................................................................. 35
Literatures ....................................................................................................... 36
Combined repeat dose and reproductive/developmental toxicity screening
test of 2,4-di-tert-pentylphenol by oral administration in rats ......................... 36
In Vitro Chromosomal Aberration Test of Phenol, 2,4-bis (1,1dimethylpropyl) on Cultured Chinese Hamster Cells. .................................... 37
Reverse Mutation Test of Phenol, 2,4-bis (1,1-dimethylpropyl) on
Bacteria. ................................................................................................... 38ii

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Dossier subject
2,4-Di-tert-pentylphenol / 120-95-6
Public nameSubmitting legal entity
National Institute of Health Sciences / Kawasaki / Japan
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Tue, 5 Sep 2023, 13:10:56+0900
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LEGAL_ENTITY: National Institute of Health
Sciences
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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 systemIUCLID42

ID
16558402024DIV750

3

2,4-Di-tert-pentylphenol
OECD
Health Effects
Repeated dose toxicity: oral
ENDPOINT_STUDY_RECORD: RepeatedDoseToxicityOral. 001

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Author:Date: 2022-03-25T11:50:41.000+09:00
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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
guideline study OECD Test Guideline study under GLP condition
Reliability 1
Cross-reference
Reason / purpose for cross-reference
reference to same study
Related information
OECD / Toxicity to reproduction / ToxicityReproduction. 001 / 2,4-Di-tert-pentylphenol / 120-95-6

Data source
Reference
Combined repeat dose and reproductive/developmental toxicity screening test of 2,4-di-tert-pentylphe /Ministry of Health, Labour and Welfare (MHLW), Japan / study report4Data accessdata published https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF120-95-6d.pdf

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
2,4-Di-tert-pentylphenol
Specific details on test material used for the study- Name of test material (as cited in study report): Phenol, 2,4-bis (1,1-dimethylpropyl)- Analytical purity: 99.7%- Storage condition of test material: Cold and dark place (3-6°C)- 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., Atsugi Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation: Male: 330.1-413.5 g), Female: 228.9-300.4 g- Housing: Animals were individually housed in bracket-type metallic wire-mesh cages (220W × 270D ×
190H mm), from gestation day 18 to lactation day 4, Dams were bred individually or with individual lit
termates in plastic cages (350W x 400D x 180H mm) and bedding.
- Diet: Solid feed (CE-2: CLEA Japan Inc.) was given ad libitum.
- Water: Tap water was given ad libitum.
5

- Acclimation period: 18 days
ENVIRONMENTAL CONDITIONS- Temperature (°C): 21.0-25.0 (actual temperature: 23.0-25.5°C)- Humidity (%): 40.0-75.0% (actual humidity: 42.5-70.5%)- Air changes (per hr): 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
Details on analytical verification of doses or concentrations
The concentrations of each test suspensions used in day1 of administration were analyzed by HPLC.
Results showed that the concentrations of each test suspensions were 93.8 to 97.7% of the nominal
concentration.
Duration of treatment / exposure(P) Males: 42 days including 14 days pre-mating(P)Females: 41-44 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.
10

mg/kg bw/day (actual dose received)

Dose / conc.
50

mg/kg bw/day (actual dose received)

Dose / conc.
100

mg/kg bw/day (actual dose received)

Remarks
At the start of administration: 250 mg/kg bw/day
No. of animals per sex per dose
Mating group: 13 animals/sex/dose (0, 10, 50, and 100 mg/kg bw/day)6

Non-mating group: 10 females/dose (0 and 100 mg/kg bw/day)Recovery group: 5 males/dose in the mating group (0 and 50 mg/kg bw/day) and 5 females/dose in
the non-mating groups (0 and 100 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 250
mg/kg bw/day was selected as an expected obvious toxic dose, and the lowest dose of 10 mg/kg
bw/day was selected as an expected no toxic dose. The middle dose levels of 50 mg/kg bw/day wereselected.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 250, 500 or 1,000 mg/kg bw/day.
At 500 mg/kg bw/day and above, all animals died or were moribund, and necropsy showed lesions in the stomach, lung and kidney. At 250 mg/kg bew/day, soft stool, salivation, decrease in body weight, decreased body weight, pale spots on the kidneys, increased liver and kidney weights, decreases in
RBC, Ht, Hb and total cholesterol, increases in triglyceride, γ-GTP, ALP, BUN, and LDH 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: 2 times/day (before administration, after administration) during the administration p
eriod. Once a day during the recovery period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Males in the main groups and females in the non-mating groups: the end of acclimation period, day 7,
14, 21, 28, 35, and 42 during the administration.
Females in the mating groups: the end of acclimation period, day 7, 14, 21, 28, 35, and 42 during theadministration period, and once during lactation period (lactation day 0from day 4)
Males and females in the recovery groups: the end of acclimation period, day 7, 14, 21, 28, 35, and 42
during the administration period and day 7 and 14 during recovery period.
BODY WEIGHT: Yes- Time schedule for examinations:
Males in the main groups and females in the non-mating groups: days 1, 7, 10, 12, 14, 17, 21, 24, 28,
31, 35, 38 and 42 of administration period and on the day of necropsy.
Males and females in the recovery groups: days1, 7, 10, 12, 14, 17, 21, 24, 28, 31, 35, 38 and 42 ofadministration period, and days 1, 7 and 14 of recovery period and on the day of necropsy.
Females in the mating groups: days 1, 7, 10, 12, 14 and 17 of administration period (uncopulated animals : day 26 ), days 0, 3, 7, 10, 14, 17 and 20 of gestation, days 0, 4 of lactation, and on the day of
necropsy.
Food consumption: Yes
Measurement of food consumption was conducted on all animals at the following frequencies:
Males in the main groups and females in the non-mating groups: days 1-2, 7-8, 14-15, 29-30, 35-36 and41-42 of administration period.
Males and females in the recovery groups: days 1-2, 7-8, 14-15, 29-30, 35-36 and 41-42 of admin
istration, period and days 6-7 and 12-13 of recovery period.
Females in the mating groups: days 1-2, 7-8 and 14-15 of administration period, days 0-1, 7-8, 14-15,
and 20-21 of gestation period, days 3-4 of lactation period.
OPHTHALMOSCOPIC EXAMINATION: No
7

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: Pentbarbital sodium- Animals fasted: Yes- How many animals:
5 animals/sex/group- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume, mea
n corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte percentage,platelet count, white blood cell count, differential white blood cell count, prothrombin time, activated p
artial thromboplastin time.
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:
5 animals/sex/group- Parameters checked: ALP, total cholesterol, triglyceride, phospholipids, total bilirubin, glucose, bloodurea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein,
albumin, A/G ratio, AST, ALT, LDH, γ-GTP, bile acid.
URINALYSIS OF MALES: Yes- Time schedule for collection of urine: final week of administration (days 37 of administration) and in
the final week of recovery (days 13 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, osmotic pressure, sodium, potassium, chloride, urine volume (24-hour volume)
BLOOD HORMONE: No
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males in the main groups and females in the non-mating groups: final week of administration(Manipulative Test: day 42, Measurement of Grip Strength and Motor Activity: day 39 for males, day
41 for females)
Females in the mating groups: lactation day 5
Males and females in the recovery groups: final week of administration (Manipulative Test: day 42,
Measurement of Grip Strength and Motor Activity: day 39 for males, day 41 for females) and in the
final week of recovery (day 14 of recovery).
- Dose groups that were examined: All dose groups (5 animals/sex/group)- Battery of functions tested:
1) Manipulative Test. Prayer's reaction, pupillary reflex, visual placing, startle reaction, withdrawal
reflex, eyelid reflex, and righting reflex2) Measurement of Grip Strength. Grip strength of forelimb and hind limb were measured by grip
strength meter.
3) Measurement of Motor Activity. Motor activity was measured by a motor activity sensor for e
xperimental animals SUPER-MEX (Muromachi Kikai. C0., Ltd.). The measurement was conducted for
20 min.
Sacrifice and pathology
GROSS PATHOLOGY: Yes
ORGAN WEIGHT: Yes [brain, thyroids (including parathyroids), adrenal gland, thymus, spleen, heart,
liver, kidney, testis, epididymis, prostate, seminal vesicles, ovary, uterus]
8HISTOPATHOLOGY: Yes, [brain, spinal cord, pituitary, eye ball (Harderian gland), submandibular
gland, sublingual gland, trachea, thyroid, parathyroid, thymus, heart, lung (including bronchial), liver,kidney, spleen, pancreas, adrenal gland, stomach, duodenum, jejunum, ileum, cecum, colon, rectum,
submandibular lymph node, mesenteric lymph node, testis, epididymis, prostate, seminal vesicles, ova
ry, uterus, vagina, bladder, femur (including bone marrows), skeletal muscle, sciatic nerve, mammary
gland, and gross abnormalities site.
Statistics
Changes in estrous cyclicity and conception rate were analyzed by Fisher’s test. Graded pathologica
l data was analyzed by Mann-Whitney’s U test (significance level = 0.05) and pathological data withnumber of positive and negative animals was analyzed by one-sided Fisher’s test. Other data, obtaine
d values in each animal or mean of a litter was one data, and these data were compared among thesatellite groups and other among the groups. These data were analyzed using F-test for homogeneityof distribution. The Student’s t-test and the Aspin-Welch’s t-test were conducted for homogenous
and non-homogenous distribution, respectively to compare the control and individual treatment gro
ups. Three or more groups setting, these data were analyzed using Bartlett’s test for homogeneity of
distribution. The Dunnett’s multiple comparison test after the ANOVA and the Dunnett’s-type meanrank sum test after Kruskal-Wallis’s H test were conducted for homogenous and non-homogenous
distribution, respectively to compare the control and individual treatment groups. Significance level
was set at 0.05 compared with the control group and among the groups.

Results and discussion
Results of examinations
Clinical signs
effects observed, treatment-related
Mortality
mortality observed, treatment-related
Body weight and weight changes
effects observed, treatment-related
Food consumption and compound intake (if feeding study)
effects observed, treatment-related
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
effects observed, treatment-related
Urinalysis findings
effects observed, treatment-related
Behaviour (functional findings)
effects observed, treatment-related
9

Immunological findings
not examined
Organ weight findings including organ / body weight ratios
effects observed, treatment-related
Gross pathological findings
effects observed, treatment-related
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:[At the dosing period]: In the 100 (250) mg/kg bw/day group, death or moribund animals were obser
ved in 9 of 13 males from day 6 to day 13 of administration and in 12 of 23 females from day 6 to day
9 of administration.
In the 50 mg/kg bw/day group, maternal death or moribund death occurred in 1 animal each on the day of parturition.
[At the recovery period]: No mortality observed.
Clinical signs:[At the dosing period]: Salivation, soiled perineal region, emaciation, hypothermia, piloerection, paleskin were observed in males and females at 50 mg/kg bw/day and above. Loose stool, decrease inamount feces, decrease in locomotor activity, mucous feces, lacrimation, reddish tear, smudge ofperinasal area, reddish urine, moribundity were observed in males and females, and blackish feces,no-feces, loss of locomotor activity, bradypnea, lateral position were observed in males at 100 (250)mg/kg bw/day.
[At the recovery period]: There were no changes related to the test substance in any groups.
DETAILED CLINICAL OBSERVATIONS:[At the dosing period]: Decrease in locomotor activity, and soiled fur were observed in males andfemales at 100 mg/kg bw/day. Crouching position, bradycardia, decrease in temperature, pale skin,anemic skin, hypopnea, reddish tear, tip toe gait, no resistance, piloerection were observed in femal
es at 100 (250) mg/kg bw/day. loss of locomotor activity, soiled fur, reddish tear, crouching position,
bradycardia, decrease in temperature, hypopnea, anemic skin, tip toe gait were observed in satellite
females at 100 mg/kg bw/day.
[At the recovery period]: There were no changes related to the test substance in any groups.
BODY WEIGHT:[At the dosing period]:A decrease in body weight gain was observed in males at 50 mg/kg bw/day and above.
[At the recovery period]:
There were no changes related to the test substance in any groups.
FOOD CONSUMPTION:[At the dosing period]:A decrease in food consumption was observed in males and females at 100 mg/kg bw/day.
[At the recovery period]:
There were no changes related to the test substance in any groups.
URINALYSIS:[At the dosing period]: An increase tendency in urine volume was observed in males, an increase in ur
ine volume was observed in females at 100 mg/kg bw/day.
10

[At the recovery period]: There were no changes related to the test substance in any groups.
HAEMATOLOGY:[At the end of dosing period]: Decreases in hemoglobin and hematocrit, an increase tendency in RBC,
an increase in prothrombin time, and an increase tendency in activated partial thromboplastin time
were observed in males at 100 mg/kg bw/day. Decreases in RBC, hemoglobin and hematocrit wereobserved in non -mating females at 100 mg/kg bw/day. A decrease in mean corpuscular hemoglobin
concentration (MCHC), an increase in reticulocyte (%) were observed in mating-females at 50 mg/kg
bw/day.
[At the end of recovery period]: There were no changes related to the test substance in any groups.
CLINICAL CHEMISTRY:[At the end of dosing period]: An increase tendency in ALP was observed in males at 100 mg/kg bw/
day.
[At the end of recovery period]: There were no changes related to the test substance in any groups.
NEUROBEHAVIOURAL EXAMINATION:
1) MANIPULATIVE TEST:
There were no changes related to the test substance in any groups at the dosing and recovery periods.
2) GRIP STRENGTH TEST:
There were no changes related to the test substance in any groups at the dosing and recovery periods.
3) LOCOMOTOR ACTIVITY MEASUREMENT:[At the dosing period]: A decrease in locomotor activity was observed in non-mating females at 100
mg/kg bw/day.
[At the recovery period]: There were no changes related to the test substance in any groups.
ORGAN WEIGHTS:[At the end of dosing period]:
An increase in relative liver weight was observed in males at 50 mg/kg bw/day. Increases in absoluteand relative liver weights were observed in males and non-mating females at 100 mg/kg bw/day andin mating-females at 50 mg/kg bw/day. An increase in spleen weight was observed in non-mating
females at 100 mg/kg bw/day[At the end of recovery period]: An increase in spleen weight was observed in non-mating females at
100 mg/kg bw/day
GROSS PATHOLOGY:[At the end of dosing period]:
Enlargement of liver was observed in males at 50 mg/kg bw/day and above. Enlargement of kidney
was observed in males at 100 mg/kg bw/day.
[At the end of recovery period]:
There were no changes related to the test substance in any groups.
HISTOPATHOLOGY: NON-NEOPLASTIC:[At the end of dosing period]:
Kidney: Basophilic tuble, hyalin cast in cortex/medulla were observed in males at 100 mg/kg bw/day.
Basophilic tuble in cortex were observed in non-mating females at 100 mg/kg bw/day.
[At the end of recovery period]:
Kidney: Basophilic tuble in cortex were observed in non-mating females at 100 mg/kg bw/day.

Effect levels
Key result
false
Dose descriptor
NOAEL
11

Effect level10

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
clinical signs
Salivation, soiled perineal region, emaciation, hypothermia, piloerection, pale skin were observed inmales and females at 50 mg/kg bw/day and above.
gross pathology
Enlargement of liver was observed in males at 50 mg/kg bw/day and above.
haematology
A decrease in mean corpuscular hemoglobin concentration (MCHC), an increase in reticulocyte (%)
were observed in mating-females at 50 mg/kg bw/day.
mortality
In the 50 mg/kg bw/day group, maternal death or moribund death occurred in 1 animal each on the
day of parturition.
organ weights and organ / body weight ratios
An increase in relative liver weight was observed in males at 50 mg/kg bw/day. Increases inabsolute and relative liver weights were observed in males and non-mating females at 100 mg/kg
bw/day and in mating-females at 50 mg/kg bw/day.

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/PDF120-95-6d.pdf

Applicant's summary and conclusion
Conclusions
The NOAEL for repeated dose toxicity in this study was determined to be 10 mg/kg bw/day both formales and females.
Executive summary
In the combined repeated dose and reproductive/developmental screening test (OECD TG422), SD rats
were treated orally with 2,4-di-tert-pentylphenol at the doses of 0, 10, 50 and 100 (250) mg/kg bw/
day. After the start of treatment, the dose was changed from 250 mg/kg bw/day to 100 mg/kg bw/
day after day 12 of treatment because about half of the animals in the high dose group died or were
moribund. No fertility study was performed in this group.
The following findings were observed in the examination during the administration period or at theend of administration period. In the 100 (250) mg/kg bw/day group, died or moribund animals wereobserved in 9 of 13 males from day 6 to day 13 of the administration and in 12 of 23 females fromday 6 to day 9 of the administration. In the 50 mg/kg bw/day group, maternal death or moribund deathoccurred in 1 animal each on the day of parturition. In the clinical signs, salivation, soiled perineal region,
emaciation, hypothermia, piloerection, pale skin were observed in males and females at 50 mg/kg bw/
day and above. Loose stool, a decrease in amount of feces, a decrease in locomotor activity, mucous
feces, lacrimation, reddish tear, smudge of perinasal area, reddish urine, moribundity were observed inmales and females, and blackish feces, no-feces, loss of locomotor activity, bradypnea, lateral position12

were observed in males at 100 (250) mg/kg bw/day. In the locomotor activity measurement, a decreasein locomotor activity was observed in non-mating females at 100 mg/kg bw/day. In the urinalysis, anincrease tendency in urine volume was observed in males and an increase in urine volume was observedin females at 100 mg/kg bw/day. In the hematology, decreases in hemoglobin and hematocrit, an
increase tendency in RBC, an increase in prothrombin time, and an increase tendency in activated partial
thromboplastin time were observed in males at 100 mg/kg bw/day. Decreases in RBC, hemoglobin
and hematocrit were observed in non -mating females at 100 mg/kg bw/day. A decrease in mean
corpuscular hemoglobin concentration (MCHC), increase in reticulocyte (%) were observed in matingfemales at 50 mg/kg bw/day. In the clinical chemistry, increase tendency in ALP was observed in males
at 100 mg/kg bw/day. In the organ weights, an increase in relative liver weight was observed in males
at 50 mg/kg bw/day. Increases in absolute and relative liver weights were observed in males and nonmating females at 100 mg/kg bw/day and in mating-females at 50 mg/kg bw/day. Increased spleen
weight was observed in non-mating females at 100 mg/kg bw/day. In the gross pathology, enlargementof liver was observed in males at 50 mg/kg bw/day and above. In the histopathology, basophilic tuble,
hyalin cast in cortex/medulla of kidney were observed in males at 100 mg/kg bw/day and basophilic
tuble in cortex of kidney were observed in non-mating females at 100 mg/kg bw/day. Based on theabove results, NOAEL for the repeated dose toxicity of 2,4-di-tert-pentylphenol was determined to be 10
mg/kg bw/day for both males and female rats.

13

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

UUID: 00b89fa3-84f9-4dad-a3b7-49a2272d3b64
Dossier UUID:
Author:Date: 2022-03-25T10:19:38.000+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
other: OECD Test Guideline study under GLP condition

Data source
Reference
Reverse Mutation Test of Phenol, 2,4-bis (1,1-dimethylpropyl) on Bacteria. / Ministry of Health, Labourand Welfare (MHLW), Japan / study report
Data accessdata published

Materials and methods
Test guideline
Qualifieraccording to guideline
Guideline
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
genetic toxicity in vitro, other14

Deviations
not specified
GLP compliance
yesType of assay
bacterial reverse mutation assay
in vitro gene mutation study in bacteria

Test material
Test material information
2,4-Di-tert-pentylphenol
Specific details on test material used for the study- Name of test material (as cited in study report): Phenol, 2,4-bis (1,1-dimethylpropyl)- Analytical purity: 99.7%

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
Justification for deviation from the high dose level-S9 mix:
19.5, 39.1, 78.1, 156, 313, 625 μg/plate (TA100, TA98 strains)
4.88, 9.77, 19.5, 39.1, 78.1, 156 μg/plate (TA1535, WP2uvrA strains)
0.610, 1.22, 2.44, 4.88, 9.77, 19.5 μg/plate (TA1537 strain)+S9 mix:
39.1, 78.1, 156, 313, 625, 1250, 2500 μg/plate (TA100, TA1535 strains)
156, 313, 625, 1250, 2500, 5000 μg/plate (WP2uvrA, TA98 strains)
19.5, 39.1, 78.1, 156, 313, 625 μg/plate (TA1537 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 500 μg/plate and above forS. typhimurium TA 100 and TA98, at 150 μg/plate and above for S. typhimurium TA1535 and E. coli
WP2uvrA, at 15.0 μg/plate and above for S. typhimurium TA1537 strain without S9 mix and at 1500 μg/plate and above for S. typhimurium TA 100 and TA1535, at 5000 μg/plate for S. typhimurium
TA98 and E. coli WP2uvrA, at 500 μg/plate and above for S. typhimurium TA 1537 with S9 mix.
Vehicle / solvent- Vehicle(s)/solvent(s) used: DMSO
15

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 (AF-2), Sodium azide (SAZ) and 9-Amino
acridine (９AA)+S9 mix, 2-Aminoanthracene (2AA) and Benzo[a]pyrene (B[a]P)

Details on test system and experimental conditions
METHOD OF APPLICATION: Preincubation
DURATION- Preincubation period: 20 min at 37°C
- Exposure duration:48 hrs
NUMBER OF PLATES: 2
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 and above, +S9 mix: 2500 μg/plate
16

Vehicle controls validity
valid
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: 9.77 μg/plate and above, +S9 mix: 625 μg/plate
Vehicle controls validity
valid
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: 625 μg/plate, +S9 mix: 5000 μg/plate
Vehicle controls validity
valid
Positive controls validity
valid
Key result
trueSpecies / strain
S. typhimurium TA 100
bacteria
Metabolic activation
with and without
17

Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 313 μg/plate and above, +S9 mix: 2500 μg/plate
Vehicle controls validity
valid
Positive controls validity
valid
Key result
trueSpecies / strain
E. coli WP2 uvr A
bacteria
Metabolic activation
with and without
Genotoxicity
negative
Cytotoxicity / choice of top concentrations
cytotoxicity -S9 mix: 156 μg/plate, +S9 mix: 5000 μg/plate
Vehicle controls validity
valid
Positive controls validityvalid

Any other information on results incl. tables
Figures and Tables (in Japanease) are available in the following full report of the study.
https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF120-95-6e.pdf
Please also see the attached files (Tables in English)

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, phenol, 2,4-bis (1,1-dimethylpropyl) was negative with or without
metabolic activation.

18

ENDPOINT_STUDY_RECORD: Genetic toxicity in vitro.002

UUID: d818e117-30a7-4c59-ac6f-e7fc8a0b6e4bDossier UUID:
Author:Date: 2023-01-10T14:34:00.000+09:00
Remarks:

Administrative data
Endpoint
in vitro 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 Phenol, 2,4-bis (1,1-dimethylpropyl) on Cultured Chinese Ham /Ministry of Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published

Materials and methods
Test guideline
Qualifieraccording to guideline
Guideline
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
genetic toxicity in vitro, otherDeviations
no19

GLP compliance
yesType of assay
other: in vitro mammalian chromosome aberration test

Test material
Test material information
2,4-Di-tert-pentylphenol
Specific details on test material used for the study- Name of test material (as cited in study report): Phenol, 2,4-bis (1,1-dimethylpropyl)- Analytical purity: 99.7%

MethodSpecies / strain
Species / strain / cell type
Chinese hamster lung (CHL/IU)
mammalian cell line
Metabolic activation
with and without
Metabolic activation system
S9 mix; SD male rat liver, induced by phenobarbital and 5,6-benzoflavone
Justification for deviation from the high dose level
Cell growth inhibition study-S9 mix (short-term treatment): 0.0091, 0.018, 0.037, 0.073, 0.15, 0.29, 0.59, 1.2, 2.3 mg/mL+S9 mix (short-term treatment): 0.0091, 0.018, 0.037, 0.073, 0.15, 0.29, 0.59, 1.2, 2.3 mg/mL-S9 mix (continuous treatment, 24hr): 0.0091, 0.018, 0.037, 0.073, 0.15, 0.29, 0.59, 1.2, 2.3 mg/mL
Main study-S9 (short-term treatment): 0.0031, 0.0063, 0.013, 0.025, 0.050, 0.10 mg/mL+S9 (short-term treatment): 0.00094, 0.0019, 0.0038, 0.0075, 0.015, 0.030 mg/mL-S9 (continuous treatment, 24hr): 0.0019, 0.0038, 0.0075, 0.015, 0.030, 0.060 mg/mL
Vehicle / solvent- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controlsnoNegative solvent / vehicle controls
yes
True negative controlsnoPositive controls
yes
Positive control substanceother: [-S9]: mitomycin C; [+S9]: cyclophosphamide20

Details on test system and experimental conditions
METHOD OF APPLICATION:
Exposure duration:- [short-term treatment]: 6 hr + 18 hr- [continuous treatment]: 24 hr
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
The frequency of cells with structural chromosomal aberrations and polyploid cells was tested for si
gnificance by Fisher's exact test (one-sided test, P = 2.5%) between the negative control and test s
ubstance treated groups. If a significant difference was observed, a Chochran-Armitage trend tests (one-sided test, P = 2.5%) was performed for dose dependency. The results of these tests were used
as a reference for a comprehensive evaluation, taking into account biological considerations.
Statistics
yes

Results and discussion
Test results
Key result
trueSpecies / strainother: Chinese hamster lung (CHL/IU) cells
Metabolic activation
with and without
Genotoxicity
positive +S9 mix (short-term treatment): Significant increases in cells with structural chromosomal
abnormalities and polyploid cells were observed in the high concentration group (frequencies: 12%
and 4.8%, respectively).
Cytotoxicity / choice of top concentrations
cytotoxicity
Vehicle controls validity
valid
Positive controls validity
valid
Additional information on results
RANGE-FINDING/SCREENING STUDIES (if applicable):
50% cell growth inhibition (IC50): 0.015 mg/mL (short-term treatment, +S9 mix), 0.049 m g/mL (shortterm treatment, -S9 mix), 0.029 mg/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.
21https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF120-95-6f.pdf

Applicant's summary and conclusion
Conclusions
Interpretation of results (migrated information): Positive with metabolic activation
In an in vitro chromosomal aberration test using CHL/IU cells, phenol, 2,4-bis (1,1-dimethylpropyl)
was positive with metabolic activation.

22

Toxicity to reproduction
ENDPOINT_STUDY_RECORD: ToxicityReproduction. 001

UUID: c7b88315-6407-4869-b0f8-48f79ac4e1a3
Dossier UUID:
Author:Date: 2022-03-25T11:51:46.000+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
guideline study OECD Test Guideline study under GLP condition
Reliability 1
Cross-reference
Reason / purpose for cross-reference
reference to same study
Related information
OECD / Repeated dose toxicity: oral / RepeatedDoseToxicityOral. 001 / 2,4-Di-tert-pentylphenol /120-95-6

Data source
Reference
Combined repeat dose and reproductive/developmental toxicity screening test of 2,4-di-tert-pentylphe /Ministry of Health, Labour and Welfare (MHLW), Japan / study report
Data accessdata published https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF120-95-6d.pdf

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
2,4-Di-tert-pentylphenol
Specific details on test material used for the study- Name of test material (as cited in study report): Phenol, 2,4-bis (1,1-dimethylpropyl)- Analytical purity: 99.7%- Storage condition of test material: Cold and dark place (3-6°C)- 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., Atsugi Breeding Center.
- Age at study initiation: 10 weeks old- Weight at study initiation: Male: 330.1-413.5 g), Female: 228.9-300.4 g- Housing: Animals were individually housed in bracket-type metallic wire-mesh cages (220W × 270D ×
190H mm), from gestation day 18 to lactation day 4, Dams were bred individually or with individual lit
termates in plastic cages (350W x 400D x 180H mm) and bedding.
- Diet: Solid feed (CE-2: CLEA Japan Inc.) was given ad libitum.
- Water: Tap water was given ad libitum.
- Acclimation period: 18 days
ENVIRONMENTAL CONDITIONS- Temperature (°C): 21.0-25.0 (actual temperature: 23.0-25.5°C)- Humidity (%): 40.0-75.0% (actual humidity: 42.5-70.5%)- Air changes (per hr): 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
Vehiclecorn oil
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
The concentrations of each test suspensions used in day1 of administration were analyzed by HPLC.
Results showed that the concentrations of each test suspensions were 93.8 to 97.7% of the nominal
concentration.
Duration of treatment / exposure(P) Males: 42 days including 14 days pre-mating(P)Females: 41-44 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.
10

mg/kg bw/day (actual dose received)

Dose / conc.
50

mg/kg bw/day (actual dose received)

Dose / conc.
100

mg/kg bw/day (actual dose received)

Remarks
At the start of administration: 250 mg/kg bw/day
No. of animals per sex per dose
Mating group: 13 animals/sex/dose (0, 10, 50, and 100 mg/kg bw/day)
Non-mating group: 10 females/dose (0 and 100 mg/kg bw/day)25

Recovery group: 5 males/dose in the mating group (0 and 50 mg/kg bw/day) and 5 females/dose in
the non-mating groups (0 and 100 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 250
mg/kg bw/day was selected as an expected obvious toxic dose, and the lowest dose of 10 mg/kg
bw/day was selected as an expected no toxic dose. The middle dose levels of 50 mg/kg bw/day wereselected.
[14-day preliminary study]
A 14-day repeated dose oral toxicity test (Crl:CD(SD) rats, doses: 0, 250, 500 or 1,000 mg/kg bw/day.
At 500 mg/kg bw/day and above, all animals died or were moribund, and necropsy showed lesions in the stomach, lung and kidney. At 250 mg/kg bew/day, soft stool, salivation, decrease in body weight, decreased body weight, pale spots on the kidneys, increased liver and kidney weights, decreases in
RBC, Ht, Hb and total cholesterol, increases in triglyceride, γ-GTP, ALP, BUN, and LDH were observed.
- Rationale for animal assignment (if not random): Body weight-balanced randomization

Examinations
Parental animals: Observations and examinations
CAGE SIDE OBSERVATIONS: Yes- Time schedule: 2 times/day (before administration, after administration) during the administration
period. Once a day during the recovery period.
DETAILED CLINICAL OBSERVATIONS: Yes- Time schedule:
Males in the main groups and females in the non-mating groups: the end of acclimation period, day 7,
14, 21, 28, 35, and 42 during the administration.
Females in the mating groups: the end of acclimation period, day 7, 14, 21, 28, 35, and 42 during theadministration period, and once during lactation period (lactation day 0from day 4)
Males and females in the recovery groups: the end of acclimation period, day 7, 14, 21, 28, 35, and 42
during the administration period and day 7 and 14 during recovery period.
BODY WEIGHT: Yes- Time schedule for examinations:
Males in the main groups and females in the non-mating groups: days 1, 7, 10, 12, 14, 17, 21, 24, 28, 3
1, 35, 38 and 42 of administration period and on the day of necropsy.
Males and females in the recovery groups: days1, 7, 10, 12, 14, 17, 21, 24, 28, 31, 35, 38 and 42 ofadministration period, and days 1, 7 and 14 of recovery period and on the day of necropsy.
Females in the mating groups: days 1, 7, 10, 12, 14 and 17 of administration period (uncopulated animals : day 26 ), days 0, 3, 7, 10, 14, 17 and 20 of gestation, days 0, 4 of lactation, and on the day of
necropsy.
Food consumption: Yes
Measurement of food consumption was conducted on all animals at the following frequencies:
Males in the main groups and females in the non-mating groups: days 1-2, 7-8, 14-15, 29-30, 35-36 and41-42 of administration period.
Males and females in the recovery groups: days 1-2, 7-8, 14-15, 29-30, 35-36 and 41-42 of admi
nistration, period and days 6-7 and 12-13 of recovery period.
Females in the mating groups: days 1-2, 7-8 and 14-15 of administration period, days 0-1, 7-8, 14-15,
and 20-21 of gestation period, days 3-4 of lactation period.
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes
26

- Time schedule for collection of blood: At the end of administration period, or at the end of recovery p
eriod in both sexes- Anaesthetic used for blood collection: Pentbarbital sodium- Animals fasted: Yes- How many animals:
5 animals/sex/group- Parameters examined: red blood cell count, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, reticulocyte percentage,platelet count, white blood cell count, differential white blood cell count, prothrombin time, activatedpartial thromboplastin time.
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:
5 animals/sex/group- Parameters checked: ALP, total cholesterol, triglyceride, phospholipids, total bilirubin, glucose, bloodurea nitrogen, creatinine, sodium, potassium, chloride, calcium, inorganic phosphorus, total protein,
albumin, A/G ratio, AST, ALT, LDH, γ-GTP, bile acid.
URINALYSIS OF MALES: Yes- Time schedule for collection of urine: final week of administration (days 37 of administration) and in
the final week of recovery (days 13 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, osmotic pressure, sodium, potassium, chloride, urine volume (24-hour volume)
BLOOD HORMONE: No
NEUROBEHAVIOURAL EXAMINATION: Yes- Time schedule for examinations:
Males in the main groups and females in the non-mating groups: final week of administration(Manipulative Test: day 42, Measurement of Grip Strength and Motor Activity: day 39 for males, day
41 for females)
Females in the mating groups: lactation day 5
Males and females in the recovery groups: final week of administration (Manipulative Test: day 42,
Measurement of Grip Strength and Motor Activity: day 39 for males, day 41 for females) and in the
final week of recovery (day 14 of recovery).
- Dose groups that were examined: All dose groups (5 animals/sex/group)- Battery of functions tested:
1) Manipulative Test. Prayer's reaction, pupillary reflex, visual placing, startle reaction, withdrawal
reflex, eyelid reflex, and righting reflex2) Measurement of Grip Strength. Grip strength of forelimb and hind limb were measured by grip
strength meter.
3) Measurement of Motor Activity. Motor activity was measured by a motor activity sensor forexperimental animals SUPER-MEX (Muromachi Kikai. C0., Ltd.). The measurement was conducted for
20 min.
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.
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.

27

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 pentbarbital sodium
anesthesia.
SACRIFICE: Males in main groups and females in non-mating groups: On next day after the last administration (Day 42), Maternal animals: on Day 4 of lactation, and Male and females recovery anima
ls: on Day 14 of recovery.
ORGAN WEIGHT: Yes [brain, thyroids (including parathyroids), adrenal gland, thymus, spleen, heart, liv
er, kidney, testis, epididymis, prostate, seminal vesicles, ovary, uterus]
HISTOPATHOLOGY: Yes, [brain, spinal cord, pituitary, eye ball (Harderian gland), submandibular
gland, sublingual gland, trachea, thyroid, parathyroid, thymus, heart, lung (including bronchial), liver,kidney, spleen, pancreas, adrenal gland, stomach, duodenum, jejunum, ileum, cecum, colon, rectum,
submandibular lymph node, mesenteric lymph node, testis, epididymis, prostate, seminal vesicl
es, ovary, uterus, vagina, bladder, femur (including bone marrows), skeletal muscle, sciatic nerve,
mammary gland, and gross abnormalities site.
Postmortem examinations (offspring)
SACRIFICE- The F1 offsprings were euthanized on PND4 by exsanguination under sevoflurane anesthesia.
GROSS NECROPSY：Yes- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, andabdominal viscera.
HISTOPATHOLOGY / ORGAN WEIGTHS- Not examined.
Statistics
Changes in estrous cyclicity and conception rate were analyzed by Fisher’s test. Graded pathological
data was analyzed by Mann-Whitney’s U test (significance level = 0.05) and pathological data with number of positive and negative animals was analyzed by one-sided Fisher’s test. Other data, obtained
values in each animal or mean of a litter was one data, and these data were compared among thesatellite groups and other among the groups. These data were analyzed using F-test for homogeneity of distribution. The Student’s t-test and the Aspin-Welch’s t-test were conducted for homogenous
and non-homogenous distribution, respectively to compare the control and individual treatment grou
ps. Three or more groups setting, these data were analyzed using Bartlett’s test for homogeneity of
distribution. The Dunnett’s multiple comparison test after the ANOVA and the Dunnett’s-type meanrank sum test after Kruskal-Wallis’s H test were conducted for homogenous and non-homogenous
distribution, respectively to compare the control and individual treatment groups. Significance level
was set at 0.05 compared with the control group and among the groups.
Reproductive indices
Each parameter was determined by the following equations:
Copulation index (%) = (No. of copulated pares / No. of mated pares) × 100
Fertility index (%) = (No. of fertile males / No. of copulated pares) × 100
Delivery index (dams, %) = (No. of dams with live offspring / No. of pregnant dams) × 100
Implantation index (%) = (No. of implantation scars / No. of corpora lutea) × 100
Sex ratio = No. of male offspring / (No. of male offspring + No. of female offspring)
Delivery index (offspring) = (No. of offspring at birth/ No. of implantation scars) × 100
Birth index = (No. of live offspring at birth/No. of implantation scars) × 100
28

Live birth index = (No. of live offspring at birth/No. of offspring at birth) × 100
Offspring viability indices
Viability index = (No. of live offspring 4days after birth / No. of live offspring at birth) × 100

Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
Clinical signs
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Mortality
mortality observed, treatment-related
Description (incidence)See 7.5.1 Repeated dose toxicity.001
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)
not examined
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
effects observed, treatment-related29

Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Behaviour (functional findings)
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
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
effects observed, treatment-related
Description (incidence and severity)
See 7.5.1 Repeated dose toxicity.001
Neuropathological findings
not examined
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 performanceno effects observed

Details on results (P0)General toxicity: See 7.5.1 Repeated dose toxicity.001
Reproductive function / performance: no effects observed

Effect levels (P0)
Key result
false
Dose descriptor
NOAEL
30

Effect level10

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
clinical signs
Salivation, soiled perineal region, emaciation, hypothermia, piloerection, pale skin were observed inmales and females at 50 mg/kg bw/day and above.
mortality
In the 50 mg/kg bw/day group, maternal death or moribund death occurred in 1 animal each on the
day of parturition.
haematology
A decrease in mean corpuscular hemoglobin concentration (MCHC), an increase in reticulocyte (%)
were observed in mating-females at 50 mg/kg bw/day.
organ weights and organ / body weight ratios
An increase in relative liver weight was observed in males at 50 mg/kg bw/day. Increases inabsolute and relative liver weights were observed in males and non-mating females at 100 mg/kg
bw/day and in mating-females at 50 mg/kg bw/day.
gross pathology
Enlargement of liver was observed in males at 50 mg/kg bw/day and above.
Key result
false
Dose descriptor
NOAEL
Effect level50

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
reproductive performance
No reproductive effects were observed in both males and females up to 50 mg/kg bw/day.

Results: F1 generation
General toxicity (F1)Clinical signs
no effects observed
Mortality / viability
no mortality observed
Body weight and weight changes
no effects observed31Gross pathological findings
no effects observed

Details on results (F1)
No effects observed.

Effect levels (F1)
Key result
false
Dose descriptor
NOAEL
Generation
F1
Effect level50

mg/kg bw/day (actual dose received)

Based ontest mat.
Sexmale/female
Basis for effect level
other:
There were no effects on developmental parameters up to 50 mg/kg bw/day.

Overall reproductive toxicity
Key result
false
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/PDF120-95-6d.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, there were no effects on the reproductive and dev
elopmental parameters up to 50 mg/kg bw/day. The NOAEL for the rat reproductive/developmental
toxicity of 2,4-di-tert-pentylphenol was regarded as 50 mg/kg bw/day, the highest dose tested.
32

DOMAIN
SUBSTANCE: 2,4-Di-tert-pentylphenol
UUID: 75638419-5081-47af-a448-15d10d68a280
Dossier UUID:
Author:Date: 2023-09-05T13:10:50.154+09:00
Remarks:
Substance name
2,4-Di-tert-pentylphenol
Legal entity
National Institute of Health Sciences / Kawasaki / Japan

Identification of substance
Reference substance
2,4-Di-tert-amylphenol / 120-95-6
EC number

EC name

CAS number

CAS name

120-95-6
IUPAC nameRole in the supply chain
Manufacturer
false
Importer
false
Only representativefalse
Downstream user
false

33

References
Reference Substances
REFERENCE_SUBSTANCE: 2,4-Di-tert-amylphenol
UUID: 6c2e6826-0a81-4119-b2a4-3af2da131516
Dossier UUID:
Author:Date: 2021-10-29T14:20:21.000+09:00
Remarks:
Reference substance name
2,4-Di-tert-amylphenol

Inventory
CAS number
120-95-6

Molecular and structural information
Molecular weight234.3834

Test Materials
TEST_MATERIAL_INFORMATION: 2,4-Di-tertpentylphenol
UUID: e27c5e05-4e22-475d-b4ec-ac7dc5708bf2
Dossier UUID:
Author:Date: 2021-10-29T14:32:39.000+09:00
Remarks:
Name
2,4-Di-tert-pentylphenol

Composition
Composition
Reference substance
2,4-Di-tert-amylphenol / 120-95-6
EC number

EC name

CAS number

CAS name

120-95-6
IUPAC name

Other characteristics
Test material form
solid: crystalline

35

Literatures
LITERATURE: Combined repeat dose and reproductive/
developmental toxicity screening test of 2,4-di-tertpentylphenol by oral administration in rats
UUID: ab7756b4-037b-4784-b5cd-379d3d2326a7
Dossier UUID:
Author:Date: 2021-11-01T14:51:20.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Combined repeat dose and reproductive/developmental toxicity screening test of 2,4-di-tert-pentylphe
nol 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) at https://dra4.nihs.go.jp/
mhlw_data/home/pdf/PDF120-95-6d.pdf
Testing facility
Food and drug safety center
Report number
R-11-003

36

LITERATURE: In Vitro Chromosomal Aberration Test of
Phenol, 2,4-bis (1,1-dimethylpropyl) on Cultured Chinese
Hamster Cells.
UUID: 56577b83-5a18-456d-a9fa-6d79becbe16f
Dossier UUID:
Author:Date: 2022-03-03T17:30:33.000+09:00
Remarks:

General information
Reference Typestudy report
Title
In Vitro Chromosomal Aberration Test of Phenol, 2,4-bis (1,1-dimethylpropyl) on Cultured Chinese
Hamster Cells.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Year
2012
Bibliographic source
Japan Existing Chemical Data Base (JECDB) https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF12
0-95-6f.pdf
Testing facility
the Hatano Research Institute, Food and Drug Safety Center
Report date2012-03-15
Report number
G-11-031

37

LITERATURE: Reverse Mutation Test of Phenol, 2,4-bis(1,1-dimethylpropyl) on Bacteria.
UUID: 76cf0669-ff0a-487e-a31c-90ee8a106701
Dossier UUID:
Author:Date: 2022-03-08T10:55:52.000+09:00
Remarks:

General information
Reference Typestudy report
Title
Reverse Mutation Test of Phenol, 2,4-bis (1,1-dimethylpropyl) on Bacteria.
Author
Ministry of Health, Labour and Welfare (MHLW), Japan
Year
2012
Bibliographic source
Japan Existing Chemical Data Base (JECDB) https://dra4.nihs.go.jp/mhlw_data/home/pdf/PDF12
0-95-6e.pdf
Testing facility
the Hatano Research Institute, Food and Drug Safety Center
Report date2012-03-09
Report number
M-11-042

38