Diallyl isophthalate

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

one-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 April to 07 May 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

ANALOGUE APPROACH JUSTIFICATION
Please refer to attached document.

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Sprague-Dawley Crl:CD (SD) IGS BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Commercial laboratory animal supplier.
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) x wks; (F1) x wks
- Weight at study initiation: (P) Males: 308 - 366g; Females: 196 - 234g;
- Fasting period before study: No
- Housing: Polypropylene cages with stell grid floors and tops, suspended over paper-lined polypropylene trays.
- Diet: Rodent pellets ad libitum
- Water: ad libitum
- Acclimation period: no data
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 55 ± 15%
- Air changes (per hr): 15
- Photoperiod: 12 hrs dark / 12 hrs light

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): once

VEHICLE
- Justification for use and choice of vehicle (if other than water): No data
- Concentration in vehicle: No data
- Amount of vehicle (if gavage): No data
- Lot/batch no. (if required):
- Purity:

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation:
- Proof of pregnancy: vaginal plug and/or sperm in vaginal smear referred to as day 0 / day 1 no data of pregnancy

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The standard and sample solutions were analysed by HPLC using the following conditions;
HPLC; Agilent technologies 1050 or 1100, incorporating autosampler and workstation
Column; Prodigy ODS (250 x 4.6 mm id)
Mobile phase; acetonitrile:water (75:25 v/v)
Flow-rate; 1.0 ml/min
UV detector wavelength; 223 nm
Injection volume; 25 µl
Retention time; ~5.6 min

Duration of treatment / exposure:

14 days pre coitus

Frequency of treatment:

Daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

16.7 mg/kg bw/day (nominal)

Dose / conc.:

50 mg/kg bw/day (nominal)

Dose / conc.:

150 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily in the week and once daily during weekends and public holidays.


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily, immediately before dosing, immediately after dosing and one hour after dosing for clinical signs of toxicity.

BODY WEIGHT: Yes
- Time schedule for examinations: During maturation and mating weighed weekly. Following mating weighed weekly until termination. Parental females showing evidence of mating weighed on days 0, 7, 14 and 20 post coitum. Parental generation females with a live litter were weighed on days 1 and 4 post partum.

Oestrous cyclicity (parental animals):

No data

Sperm parameters (parental animals):

No data

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities,

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities;

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals killed on day 5 post partum
- Maternal animals: All surviving animals killed on day 5 post partum

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
All main organs were prepared for microscopic examination and weighed

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed at 5 days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
All main tissues were prepared for microscopic examination and weighed, respectively.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

All animals showed increased salivation immediately post dose at 150 mg/kg/day

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, treatment-related

Description (incidence):

At 150 mg/kg/day there were 3 mortalities

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

At 150 mg/kg/day there was a slight reduction in bodyweight gain.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Immunological findings:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

not examined

Reproductive performance:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

50 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

gross pathology

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

150 mg/kg bw/day (nominal)

System:

hepatobiliary

Organ:

liver

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

not examined

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

Behaviour (functional findings):

no effects observed

Developmental immunotoxicity:

not examined

There were no treatment related effects on litter sizes at birth or on subsequent offspring survival throughout lactation. There were no effects on offspring growth or development. There were no effects on offspring reflexological responses and no effects on the intra-litter sex ratios.

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

> 150 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No effects seen clinically or histopathologically.

Key result

Critical effects observed:

no

Key result

Reproductive effects observed:

no

Conclusions:

The F1 generation displayed no treatment-related effects on growth or development. No macroscopic abnormalities were seen at terminal necropsy.

The P0 parent showed dose-related effects to their livers; periportal hepatocyte necrosis, enlargement and basophilia, bile duct proliferation and periportal fibrosis.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Based on two oral studies in rats no treatment-related effects were recorded on fertility, offspring viability, growth or development . The oraldoses across the two studies ranged from 16.7 to 250 mg/kg/day. In one study (Knox and Brooks) the dose of 150 mg/kg/day induced severe effects in females probably due to dystocia. However, this was not repeated at the same dose in the other study (Saillenfait et al) and only recorded for two females exposed at 200 mg/kg/day. Maternal weight gain and food consumption were significantly reduced at and above 200 mg/kg/day.

There was no significant increase in the incidence of resorptions, or malformations, at any dose in either of the two studies. In the

Saillenfait et al study fetal body weight was significantly reduced at and above 200 mg/kg/day but significant effects on skeletal development were only recorded at 250 mg/kg/day. Thus, DAP caused fetal toxicity at doses which also produced maternal effects (> 150 mg DAP/kg/day), but which did not affect fertility or induce significant embryo-lethality or teratogenicity.

The Safepharm study showed that the F1 generation displayed no treatment-related effects on growth or development. No macroscopic abnormalities were seen at terminal necropsy. The P0 parent showed dose-related effects to their livers; periportal hepatocyte necrosis, enlargement and basophilia, bile duct proliferation and periportal fibrosis.

Short description of key information:
The key information is based on the results of a guideline study performed under GLP in 2003 and a study published in 2008 (see the section of Developmental Toxicity). The study method used concerns a reproduction/developmental toxicity screening test [OECD TG 421] with parental exposure prior to mating until 1 to 4 days post partum. Endpoints related parameters examined include histopathological changes in the reproductive organs of parental animals as well as offspring viability, growth and development.

Effects on developmental toxicity

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

ANALOGUE APPROACH JUSTIFICATION
Please refer to attached document.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Commercial laboratory animal supplier
- Age at study initiation:
- Weight at study initiation:
- Fasting period before study:
- Housing: clear polycarbonate cages with stainless steel wire lids and corn cob granules for bedding.
- Diet; food pellets ad libitum
- Water; ad libitum
- Acclimation period: 1-2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 50 ± 5%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12h light-dark photocycle

IN-LIFE DATES: From: To:

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food: ambient

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: no data
- Length of cohabitation:
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- Any other deviations from standard protocol:

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

Daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

150 mg/kg bw/day (nominal)

Dose / conc.:

200 mg/kg bw/day (nominal)

Dose / conc.:

250 mg/kg bw/day (nominal)

No. of animals per sex per dose:

12

Control animals:

yes

Details on study design:

- Dose selection rationale: Based on study in 2004 giving LD50 of 891 mg/kg in males and 656 mg/kg in females
- Rationale for animal assignment (if not random):
- Other:

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Maternal bodyweights, first weight after GD=0 was GD = 6. After that every 3 days until GD = 21.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
- Time schedule for examinations:

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: uterus, uterine contents, ovaries, fetuses,

OTHER:

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other:

Fetal examinations:

- External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes:
- Head examinations: No data

Statistics:

Whenever possible, the data were presented as mean +/- SD. The number of corpora lutea, implantaion sites and live foetuses, and various body weights were analysed by one-way analysis of variance, followed by Dunnett's test if differences were found. The frequency of post-implantation loss, dead foetuses, resorptions and alterations among litterswas evaluated by using Kruskal-Wallis test followed by the Mann-Whitney test where appropriate. Rates of pregnancy and of litters with dead foetuses or resorptions and incidences of foetal alterations per dose were analysed by using Fishers's test. Where applicable, least-squares analysis was caried out. The reported level of statitsical significance was p < 0.05. The litter was used as the basis for the analysis of foetal variables.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Macroscopic changes in the liver. This mainly consisted of a pale and mottled organ.

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Description (incidence):

one dam from the 250 mg/kg dose group was found dead on GD 21. No obvious cause for death was established.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Reduced body weight gain.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

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:

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Foetal skeletal variations and delayed ossification in fore and hinf limbs and caudal vertebra centra.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed

Changes in number of pregnant:

no effects observed

Details on maternal toxic effects:

Macroscopic changes were noted in most of the dams at 150 mg/kg dose and higher.

Key result

Dose descriptor:

dose level:

Effect level:

150 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

necropsy findings

Remarks on result:

other: Liver pale and mottled.

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

other:

Description (incidence and severity):

Liver pale and mottled.

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in postnatal survival:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

effects observed, treatment-related

Visceral malformations:

effects observed, treatment-related

Key result

Dose descriptor:

dose level:

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Reduced foetal body weight.

Remarks on result:

other: Dose related

Key result

Abnormalities:

effects observed, treatment-related

Localisation:

skeletal: skull

skeletal: forelimb

skeletal: sternum

skeletal: rib

skeletal: vertebra

skeletal: hindlimb

Description (incidence and severity):

Significantly delayed ossification at 250 mg/kg/day as evidenced by the reduced numbers of osified phalanged in fore and hind limbs, metatarsals in hindlimbs and caudal vertebral centra.

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

200 mg/kg bw/day (nominal)

Treatment related:

yes

Relation to maternal toxicity:

not specified

Dose response relationship:

yes

Relevant for humans:

yes

Conclusions:

At 150 mg/kg/day, maternal response to treatment was indicated by macroscopic changes in the liver.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

50 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

Klimisch 1

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

The studies were carried out on DAP, a closely related structural analogue to DAIP. The classification system for reproductive toxicity is subdivided in adverse effects on sexual function and fertility and adverse effects on development of the offspring. The key information provided here indicates that sexual function and fertility are not affected by oral DAP and therefore DAIP, exposure at doses that induce parental and foetal toxicity. Although, there are indications that the gestation period might be delayed at lower dose levels, these proved not to be consistent or reproducible. Further, these were recorded at a dose that also induced histopathological changes in the liver. Developmental toxicity concerned mainly fetal body weight and only minimal differences in skeletal variations and malformations. Also these were only recorded at dose levels, which also affected maternal body weight

Classification was not considered, because only minor effects were recorded in skeletal examinations, and foetal weights were affected at dose levels that also affected maternal body weights.