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Toxicity to reproduction

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Administrative data

Endpoint:
one-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Study period:
10 Aug 1987 - 12 Jan 1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
GLP - Guideline study. According to the ECHA guidance document "Practical guide 6: How to report read-across and categories (March 2010)", the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1988
Report Date:
1988
Reference Type:
secondary source
Title:
Diesters Category of the Aliphatic Esters Chemicals (Test Plan and Robust Summaries for Substances in the HPV Test Plan)
Author:
US-EPA (American Chemistry Council's Aliphatic Esters Panel)
Year:
2010
Bibliographic source:
High Production Volume (HPV) Chemical Challenge Program (201-16837A and 201-16837B)
Reference Type:
secondary source
Title:
Bis(2-ethylhexyl)adipate (DEHA) CAS N°: 103-23-1
Author:
OECD
Year:
2000
Bibliographic source:
SIDS Initial Assessment Report for SIAM 10; Tokyo, Japan, 15-17 March 2000

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 415 [One-Generation Reproduction Toxicity Study (before 9 October 2017)]
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Di-(2-ethylhexyl) Adipate (DEHA)
- Physical state: clear liquid
- Analytical purity: 99.2% w/w
- Batch No: Y02259/003/001
- Storage: in a ventilated cupboard at 20 ºC
- Stability: Chemical stability of DEHA in diet was determined on three batches of diet at nominally 300 and 12000 ppm. Satisfactory chemical stability was established for diet stored at room temperature for up to 34 days.

Test animals

Species:
rat
Strain:
other: Alpk:APfSD (WIstar-derived)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Barriered SPF area at ICI Central Toxicology Laboratory, Alderley Park, Cheshire, UK.
- Age at study initiation: (P) 3 wks;
- Housing: two female or one male per cage, individually during gestation and lactation
- Diet: Pasteurised CTI diet, Special Diets Services Ltd., Essex, UK, ad libitum
- Water: Filtered tap water, ad libitum
- Acclimation period: 6-7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-21
- Humidity (%): 40-60
- Air changes (per hr): 15-25
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: Treatment of parental animals started on 10 Aug 1987.

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
An appropriate quantity of test compound was added to 1 kg of CTI diet and mixed in a pestle and mortar before being made up into a premix.
The amount of DEHA to be added to the 30 kg diet to obtain the required concentrations were 9.07 g for the 300 ppm group, 54.44 g for the 1800 ppm group and 362.9 g for the 12000 ppm group.

DIET PREPARATION
- Rate of preparation of diet (frequency): Diets were fed for no longer than 21 days after preparation.

Details on mating procedure:
- M/F ratio per cage: 1/2
- Length of cohabitation: 10 days
- Proof of pregnancy: sperm in vaginal smear referred to as day 1 of pregnancy
- After 10 days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): individually
In all pairings brother sister mating was avoided.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of the first batches of all dietary levels were taken from the food jars, prior to feeding, for quantitative analysis.
Further samples were taken at regular intervals for full quantitative analysis by Vortex extration and Soxhlet extraction.
Mean concentrations of all diets analysed were within 2% of target concentration for all dose groups.
Duration of treatment / exposure:
(P) Males: 10 weeks before mating.
(P) Females: 10 weeks before mating
After 10 weeks, the animals were mated to produce a single litter (F1A) which were reared to day 36 post partum.
Test diets were fed continuously throughout the study.
Frequency of treatment:
continuously
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
300, 1800 and 12000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
304, 1038 and 11840 ppm
Basis:
other: analytical
Remarks:
Doses / Concentrations:
approx. 52, 178 and 2102 mg/kg bw/day for males and 61, 203 and 2399 mg/kg bw/day for females
Basis:
other: The values were calculated by the mean body weights and the mean food consumption with the analytical compound values
No. of animals per sex per dose:
15 males
30 females
Control animals:
yes, plain diet
Details on study design:
- Strain selection rationale: Historical data on the used rat strain were available in the laboratory.
- Dose selection rationale: The dose levels for this study were based on data from the literature (NTP, 1982) and included an anticipated no effect level and a level at which toxic effects of the test substance were expected at some stage during the course of the study.

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly throughout the premating period (10 weeks). Female rats were weighed on days 1, 8, 15 and 22 of pregnancy and at termination.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes
Food consumption for each cage of rats was recorded throughout the premating periods and calculated on a weekly basis, with the exception of several femal cages where a set of residue and top up values were not recorded during the first week of the study.
Oestrous cyclicity (parental animals):
Not measured
Sperm parameters (parental animals):
Not measured
Litter observations:
Litters were examined for dead or moribund pups at least once daily and any such pups were subjected to a gross post mortem examination.
A count of all live and dead pups was made within 24 hours of parturition (day 1) and thereafter at days 5, 11, 22, 29 and 36 post partum. The sexes of the pups were also recorded at these times. Any clinical abnormalities seen in the pups were recorded.
Individual pup bodyweights were recorded within 24 h of birth (day1) and at days 5, 11, 22, 29 and 36 post partum.
The litters were weaned at day 22 post partum.
Since pups were not individually identified, data were recorded by sex and litter.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals after completion of mating.
- Maternal animals: All surviving animals after weaning their litters.

GROSS NECROPSY
- Gross necropsy consisted of full necropsy.

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues were prepared for microscopic examination:
Cervix, Prostate, Epididymis, Seminal vesicle, Liver, Testis, Mammary Gland, Uterus, Ovary, Abnormal tissues.
The number of implantation sites in each uterine horn was recorded for each mated female.
Liver weights were recorded from F0 animals with exception of those killed intercurrently.
Postmortem examinations (offspring):
SACRIFICE
All surviving pups were killed as soon as possible after day 36 post partum.

GROSS NECROPSY
Any pups up to and including 18 days of age found dead or with behavioural, functional, or morphological abnormalities were examined by free hand dissection; abnormalities were recorded and the pups were discarded. Pups over 18 days of age found either dead or requiring euthanasia were subjected to a post mortem examination.
All clinical abnormal pups and further normal pups received a gross necropsy so that a minimum of 2 pups of each sex were examined from each litter where possible. All remaining normal pups were killed and discarded after clinical examination.
Statistics:
The following parameters were assessed using appropriate statistical tests:
mean bodyweight gain, food consumption and food utilisation during the premating period, female bodyweight gain during pregnancy, parental liver weights and pup (litter) bodyweight gain until day 36 post partum. Male and female fertility indices, mean length of gestation, mean pre-coital interval, mean live born index, mean survival index, mean litter size, total litter weight and whole litter losses.
Reproductive indices:
- The reproductive performance of the parents was assessed from the records of mating and parturition.
- The fertility of the parents was established by the success of each mating (production of a viable litter).
- The length of gestation was measured in days from date of positive smear to date of birth (but only in fertile females fulfilling the criterion above)
- The pre-coital interval, the time in days between the date of pairing and the date of positive vaginal smear, was measured, and mean value per group was estimated from all pairings with a positive smear.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
marginal reduction in body weight gain in females of the highest dose group during premating phase; reduction in body weight in week 3 and 4 of pregnancy in females at highest dose; slight increase in food consumption in males at highest dose
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
marginal reduction in body weight gain in females of the highest dose group during premating phase; reduction in body weight in week 3 and 4 of pregnancy in females at highest dose; slight increase in food consumption in males at highest dose
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Test substance intake: slightly less efficient food utilisation in males of highest dose group

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
There were no clinical abnormalities which could be attributed to treatment.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Bodyweight gain was marginally reduced for females receiving 12000 ppm DEHA during the premating phase.
At the start of pregnancy, the body weight of the females treated with 12000 ppm DEHA was marginally lower than controls and there was reduced body weight gain in this treatment group throughout pregnancy, the effect being most marked at weeks 3 and 4 (6% lower on day 22 compared to controls). There was no effect on bodyweight or body weight gain in any other treatment group during the premating period and during pregnancy, respectively.
A slight increase in food consumption in males treated with 12000 ppm DEHA was observed at week 6 to 10, the effects being statisticaly significant at weeks 6-9. The food consumption was unaffected for males receiving 1800 ppm and 300 ppm and for females of all dose groups.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS) Food utilisation was slightly less efficient overall for males receiving 12000 ppm.
REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
There was no effect on either male or female fertility, gestation length or pre-coital interval in any dose group which could be attributed to treatment with DEHA.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Absolute liver weights were increased in both males and females receiving 12000 ppm, by approximately 18%.
Relative to the bodyweights these liver weight increases were 18.9% in males and 19.7% in females.
No effects were seen in any other dose group.

GROSS PATHOLOGY (PARENTAL ANIMALS)
There were no gross changes detected in adults which could be attributed to treatment with exception of accentuated lobular pattern in the liver of two rats fed 12000 ppm DEHA.

HISTOPATHOLOGY (PARENTAL ANIMALS)
No microscopic changes were detected in reproductive organs of those animals which failed to breed and were thus supected infertile.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Remarks:
fertility
Effect level:
>= 2 102 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: see 'Remark'
Dose descriptor:
NOAEL
Remarks:
fertility
Effect level:
>= 2 399 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: see 'Remark'
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
178 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: The nominal dose was 1800 ppm in diet.
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
203 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: The nominal dose was 1800 ppm in diet.
Dose descriptor:
LOAEL
Remarks:
systemic
Effect level:
2 102 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: The nominal dose was 12000 ppm in diet. An increase in liver weight was observed in both male and female parents receiving 12000 ppm DEHA.
Dose descriptor:
LOAEL
Remarks:
systemic
Effect level:
2 399 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: The nominal dose was 12000 ppm in diet. An increase in liver weight was observed in both male and female parents receiving 12000 ppm DEHA. Reduction in bodyweight gain during gestation in the 12000 ppm DEHA group compared with controls.

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
mean litter size was slightly reduced at highest dose group (statistically not significant)
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
reduction in mean pup weight for male and female offspring in the highest dose group
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Details on results (F1)

VIABILITY (OFFSPRING)
The incidence of whole litter losses was not attributed to the treatment with DEHA, although there were four whole litter losses. None in the control group, one in the 300 ppm group, two in the 1800 ppm group and one in the 12000 ppm group. The low incidence and no clear dose-relation was considered as non treatment-related effects.
Mean litter size was slightly (without statistically significance) reduced at 12000 ppm throughout the post partum period. The number of live born pups was unaffected by treatment.
The pup survival rate was not different from control group, when the four whole litter losses were excluded from calculation.

CLINICAL SIGNS (OFFSPRING)
The pups were in good clinical condition. The observed clinical signs were of low incidence with no relationship to DEHA-treatment.

BODY WEIGHT (OFFSPRING)
Mean pup weight gain and total litter weight for both male and female offspring receiving 12000 ppm of the test substance were reduced throughout the whole of the post partum phase in days 1 - 36. No effects were observed in any other dose group.

GROSS PATHOLOGY (OFFSPRING)
There were no findings, which could be considered to be treatment-related.

Effect levels (F1)

open allclose all
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
178 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Mean pup weight gain and total litter weight were reduced at the highest dose. Litter size was slightly reduced throughout the post-partum period at the highes dose.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
203 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: Mean pup weight gain and total litter weight were reduced at the highest dose. Litter size was slightly reduced throughout the post-partum period at the highest dose.
Dose descriptor:
LOAEL
Generation:
F1
Effect level:
2 102 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Mean pup weight gain and total litter weight were reduced. Slighly reduction in litter size throughout the post-partum period.
Dose descriptor:
LOAEL
Generation:
F1
Effect level:
2 399 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: Mean pup weight gain and total litter weight were reduced. Slighly reduction in litter size throughout the post-partum period.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion