4,4'-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-03-19 to 2018-09-13

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

2016-07-29

GLP compliance:

yes

Limit test:

no

Justification for study design:

SPECIFICATION OF STUDY DESIGN FOR EXTENDED ONE-GENERATION REPRODUCTION TOXICITY STUDY WITH JUSTIFICATIONS:

- Premating exposure duration for parental (P0) animals : 2 weeks
- Basis for dose level selection: dose interval 2.5; highest dose level = dose level of negative control
- Inclusion/exclusion of extension of Cohort 1B : n.a.
- Termination time for F2 : 13 d
- Inclusion/exclusion of developmental neurotoxicity Cohorts 2A and 2B : n.a.
- Inclusion/exclusion of developmental immunotoxicity Cohort 3 : n.a.
- Route of administration : oral
- Other considerations, e.g. on choice of species, strain, vehicle and number of animals [if applicable]

Test material

Test material information

Constituent 1

Reference substance name:

4,4'-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco

EC Number:

500-303-2

EC Name:

4,4'-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco

Cas Number:

115340-85-7

IUPAC Name:

4,4'-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco

Test material form:

liquid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material:
R-127 / batch 180131
- Expiration date of the lot/batch:
2019-01-31

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material:
Dry and dark at ambient temperature (10 – 30 °C)

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: no

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: InVivos Pte Ltd, 9 Perahu Road, Lim Chu Kang, Singapore 718793
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P)male 9 wks, (P)female 11 wks; (F1) 9-11 x wks
- Weight at study initiation: (P) Males: 334-468 g; Females: 198-278 g
- Housing: Individual Ventilated Cage System
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 d
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 26 °C
- Humidity (%): 30 - 70 %
IN-LIFE DATES: From: 2018-03-14 To: 2018-08-21

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The test item was dissolved in 0.9% sodium chloride (NaCl) saline

DIET PREPARATION
- Rate of preparation of diet (frequency): Altromin Maintenance Diet #1324 was purchased as finished product
- Mixing appropriate amounts with (Type of food): The test item was not mixed with the food but administered by gavage

VEHICLE
No vehicle

Details on mating procedure:

- M/F ratio per cage: 1 : 1
- Length of cohabitation: 2 weeks
- Proof of pregnancy: vaginal plug / sperm referred to as day 0 of pregnancy
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): yes, individually in ventilated cage system

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

Dosing of both sexes began 2 weeks prior to mating, and kept during the 14-day mating period. Parental males were euthanized when 14-day mating period was completed (28 days in total), while dosing of dams continued throughout the study until Day 13 post-partum. Females shown no-evidence of copulation were euthanized 24-26 days after the last day of mating period.

Frequency of treatment:

The doses were given to the parental males and females daily, seven days per week continuously until their termination days, respectively.

Details on study schedule:

Litter size and offspring parameters

On Day 4 after birth, the size of each litter was adjusted by eliminating extra pups by random selection to yield 4 or 5 pups per sex per litter. Blood samples were collected from 2 of the surplus pups in each litter (female preferred as male pups were kept for nipple retention assessment, except no remaining females for termination assessment), pooled, and used for serum T4 assessment. No pups were eliminated when litter size dropped below 8 or 10 pups per litter. If there was only 1 pup available above the normal litter size, only 1 pup was eliminated and used for blood collection for possible T4 assessment.

The duration of gestation was recorded and was calculated from Day 0 of pregnancy. Each litter was examined as soon as possible after delivery to establish the number and sex of pups, stillbirths, live births, runts (pups that were significantly smaller than corresponding control pups) and the presence of gross abnormalities.

Live pups were counted and sexed and litters weighed within 24 hours of parturition (Day 0 or 1 post-partum) and at least on Day 4 and 13 post-partum.

The anogenital distance (AGD) of each pup was measured on the same postnatal day between postnatal Day 0 to 4. Pup body weight was collected on the same day that AGD was measured, AGD was normalized to a measurement of pup size, preferably the cube root of body weight. The number of nipples/areolae in the male pups was counted on Postnatal Day 12 or 13.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

400 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

160 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 males, 12 females

Control animals:

yes

Details on study design:

- Dose selection rationale: dose interval 2.5; highest dose level = dosse level of negtaive control
- Rationale for animal assignment (if not random): random

Positive control:

None

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least once a day
- Cage side observations were included.
Cage side observations included (but not limited to) changes in skin and fur, eyes and mucous membranes, and also respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behavior patterns.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: At least once a day

BODY WEIGHT: Yes
- Time schedule for examinations:
Parentals: Day 1, day 8, mating day 1, mating date 8, endpoint date
Parental females from post-mating: Pregnancy day 1, pregnancy day 7, pregnancy day 20, within 24 h of parturition, post partum day 4, post partum day 13
Pups: Within 24 h after parturition, day 4 post-partum, day 13 post-partum

Oestrous cyclicity (parental animals):

Estrous cycles of parental females were monitored before dosing to select the females with regular cyclicity. Vaginal smear of each female was monitored frequently during 2-week pre-exposure period. In each group, 10 females with relatively typical 4- 5-day cycles were selected for dosing and mating.

Sperm parameters (parental animals):

Parameters examined in male parental generations:
Detailed histological examination was performed on the ovaries, testes and epididymides (with special emphasis on stages of spermatogenesis and histopathology of interstitial testicular cell structure) of the animals of Test group 1 (highest dose group) and negative control group.
Assessment is performed with special emphasis on stages of spermatogenesis and interstitial testicular structures (with focus on Leydig/interstitial cells).

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 4-5] pups/litter, each sex.
No pups were eliminated when litter size dropped below 8 or 10 pups per litter. If there was only 1 pup available above the normal litter size, only 1 pup was eliminated and used for blood collection for possible T4 assessment.

PARAMETERS EXAMINED
The following parameters were examined in offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, anogenital distance (AGD), presence of nipples/areolae in male pups

GROSS EXAMINATION OF DEAD PUPS: yes, for gross abnormalities. Particular attention was paid to the external reproductive genitals which were examined for signs of altered development.

Postmortem examinations (parental animals):

No parental animal was found dead prior to endpoint day.

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring not selected as parental animals were sacrificed at 13 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:
Blood was examined for for serum Thyroxine (T4) assessment. Any abnormal behavior of the offspring was recorded.

GROSS NECROPSY
- Gross necropsy consisted of:
Gross abnormalities. Particular attention was paid to the external reproductive genitals which were examined for signs of altered development.

HISTOPATHOLOGY / ORGAN WEIGTHS
Detailed histological examination was performed on the ovaries, testes and epididymides (with special emphasis on stages of spermatogenesis and histopathology of interstitial testicular cell structure) of the animals of Test group 1 (highest dose group) and negative control group.

Microscopic examination: Ovaries, testicles epididymis, prostate, seminal vesicles and additional tissue.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, non-treatment-related

Mortality:

mortality observed, non-treatment-related

Body weight and weight changes:

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

Effect levels (P0)

Dose descriptor:

NOEL

Effect level:

> 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

mortality

dermal irritation

body weight and weight gain

clinical biochemistry

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

histopathology: neoplastic

reproductive function (oestrous cycle)

reproductive function (sperm measures)

reproductive performance

Results: F1 generation

General toxicity (F1)

Clinical signs:

effects observed, non-treatment-related

Mortality / viability:

mortality observed, non-treatment-related

Body weight and weight changes:

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings:

no effects observed

Other effects:

no effects observed

Effect levels (F1)

Key result

Dose descriptor:

NOEL

Generation:

F1

Effect level:

> 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

viability

sexual maturation

clinical signs

mortality

body weight and weight gain

haematology

clinical biochemistry

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

histopathology: neoplastic

Overall reproductive toxicity

Key result

Reproductive effects observed:

no

Applicant's summary and conclusion

Conclusions:

Based on the results of the study, administrating the test item repeatedly at 1000, 400 and 160 mg/kg body weight through oral route (28-day for male parents and up to Day 13 post-partum for female parents), the test item 4, 4’-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco, did not produce reproductive / developmental toxicity to Sprague Dawley (SD) rats.

Executive summary:

The substance 4, 4’-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco, was examined in a Reproduction / developmental toxicity screening study according to OECD 421.

Based on the results of the study, administrating the test item repeatedly at 1000, 400 and 160 mg/kg body weight through oral route (28-day for male parents and up to Day 13 post-partum for female parents), the test item 4, 4’-Isopropylidenediphenol, ethoxylated, esters with fatty acids, coco, did not produce reproductive / developmental toxicity to Sprague Dawley (SD) rats.