Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Weight of evidence based on the information about the test chemical.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

Data is from a HPVIS database.

Qualifier:

according to guideline

Guideline:

other: OECD 422 (Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test )

Principles of method if other than guideline:

According to OECD 422 (Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test )

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

No Data Available

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

The test material was administered by gavage to three groups each of ten male and ten female Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, for up to fifty-four consecutive days, at dose levels of 50, 250 and 600 mg/kg/day.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No Data Available

Details on mating procedure:

- M/F ratio per cage: 1:1
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy : The presence of sperm within the vaginal smear and/or vaginal plug in situ was taken as positive evidence of mating.
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. : N/A
- Further matings after two unsuccessful attempts: [no / yes (explain)] : no

Duration of treatment / exposure:

Males: 43 days;
Females: up to 54 days

Frequency of treatment:

Daily

Duration of test:

54 days prior to mating, throughout mating and gestation and continuing through lactation day 3

Remarks:

Doses / Concentrations:
0, 50, 250, and 600 mg/kg/day
Basis:

No. of animals per sex per dose:

20 animals in each group
0 mg/kg/day (control) - ten male and ten female Sprague-Dawley rats
50 mg/kg/day - ten male and ten female Sprague-Dawley rats
250 mg/kg/day - ten male and ten female Sprague-Dawley rats
600 mg/kg/day - ten male and ten female Sprague-Dawley rats

Control animals:

yes, concurrent vehicle

Details on study design:

No Data Available

Maternal examinations:

Survival, clinical sign, body weight, food consumption, water consumption, Duration of gestation, corpora luteaand, implantation site, fatality, hematology, clinical chemistry, oragn weight, gross patholology, histopathology and reperoductive performance were observed.
Pregnancy and parturition: The following was recorded for each female:
i) Date of mating
jj) Date and time of observed start of parturition
iii) Date and time of observed completion of parturition
iv) Duration of gestation

Ovaries and uterine content:

Duration of gestation, corpora lutea and, implantation site was observed.

Fetal examinations:

Survival, clinical sign, body weight and development were observed.

Statistics:

No Data Available

Indices:

Implantation Index, Gestational Index, Pup viability/survival index.

Historical control data:

No Data Available

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

not specified

Early or late resorptions:

not specified

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not specified

Changes in number of pregnant:

no effects observed

Other effects:

no effects observed

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
No effects on mating performance, fertility or gestation were observed.

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Remarks on result:

other: Not Specified

Abnormalities:

not specified

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Mean offspring weights for treated animals were comparable to controls.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

not specified

Changes in litter size and weights:

no effects observed

Description (incidence and severity):

No treatment-related macroscopic abnormalities were detected for the interim death offspring or for the remaining offspring at terminal kill.

Changes in postnatal survival:

not specified

External malformations:

effects observed, treatment-related

Description (incidence and severity):

Study 2: Offspring from the 600 mg/kg/day dose group showed less successful completion of surface righting assessments. There were no treatment-related differences in pinna unfolding.

Skeletal malformations:

not specified

Visceral malformations:

no effects observed

Description (incidence and severity):

Study 2: no effects observed

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Litter Observations: There were no clinical signs to suggest an effect of treatment.
Offspring Viability: Mean offspring weights for treated animals were comparable to controls.
Offspring Development: Offspring from the 600 mg/kg/day dose group showed less successful completion of surface righting assessments. There were no treatment-related differences in pinna unfolding.
Necropsy of Offspring: No treatment-related macroscopic abnormalities were detected for the interim death offspring or for the remaining offspring at terminal kill.

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: fetotoxicity

Remarks on result:

other: Not Specified

Abnormalities:

not specified

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

Based on all the observation and results, the NOAEL for maternal and fetotoxicity study was considered to be 250 mg/Kg bw/day.

Executive summary:

In the combined repeated-dose/reproductive/developmental toxicity screening study, the test material was administered by gavage to three groups each of ten male and ten female Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, for up to fifty-four consecutive days, at dose levels of 50, 250 and 600 mg/kg/day. The following was recorded for each female: Date of mating, Date and time of observed start of parturition, Date and time of observed completion of parturitionand Duration of gestation. After dosing, it was observed that,Females treated with 600 mg/kg/day showed a higher percentage of pre-implantation losses in comparison to controls, resulting in the birth of less offspring per litter and lower total litter weights at this dose level. Thus, no adverse effects (NOAEL) on mating performance, fertility or gestation were observed at 250 mg/kg-bw/day of concentartion. Also, the same NOAEL was considered for fetal parameters. Thus, based on all the observation and results, it was concluded that the NOAEL of the test chemical for the maternal and fetal parameters considered to be 250 mg/kg bw after administration to the test animals.

Cross-reference 2

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Data is from a publication.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

The study was designed to investigate the effect of administration by intragastric intubation of the the test chemical on pregnancy of the New Zealand White rabbit and the in utero development of its offspring over an estimation period of 29 days.

GLP compliance:

not specified

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:Cheshire Rabbit Farms (Oudden Lodge. Nr. Tarporley.Cheshire). Ranch Rabbits (Crawley Down.Sussex). and Buxted Rabbit Co. Ltd (Gt. Totease Farm, Buxted. Sussex).
- Age at study initiation for p: Twelve to twenty weeks of age
- Weight at study initiation:2.9 to 4.1 kg
- Housing:The rabbits were housed individually in metal cages equipped with sheet stainless steel sides, back and top, a stainless steel wire front and an aluminium perforated floor panel.Individual undercage plastic trays were lined with absorbent paper which was inspected and changed daily.
- Diet (e.g. ad libitum):SDS Rabbit Diet SQC,ad libitum
- Water (e.g. ad libitum):Tap water,ad libitum
- Acclimation period:10 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C):18°C ± 3°C
- Humidity (%): 55% ± 15%
- Photoperiod (hrs dark / hrs light): 14 hours

Route of administration:

oral: gavage

Vehicle:

other: 1% Methyl-cellulose

Details on exposure:

Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The dosages of diphenylamine administered by oral gavage in the present study were 33,100 and 300 mg/kg bodyweight/day. Treatment by intragastric intubation commenced on Day 7 of pregnancy and continued daily up to and including Day 19 of gestation. The highest required concentration of suspension was prepared by suspending the test material in the vehicle. The lower concentrations were prepared by serial dilution with the vehicle. The suspensions were prepared freshly each day. Treatment by intragastric intubation commenced on Day 7 of pregnancy and continued daily up to and including Day 19 of gestation. Dose volumes were calculated for individual animals on Day 7 of pregnancy and adjusted according to bodyweight on Days 9, 11 and 15.

VEHICLE : Methyl-cellulose

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No Data Available

Details on mating procedure:

Details of mating : Does were mated with males of proven fertility: once coitus had been observed, each female was allowed to remain with the male for at least one hour. Sixty-six of those dams which successfully completed coituswere each injected intravenously with 25 i.u. of Chorulon (luteinizing hormone) to promote ovulation. The day of mating was considered as Day 0 of pregnancy.

- Mating was phased over consecutive week days (Monday to Thursday) ensuring that no more than 20 animals were mated on anyone day. On each of these days, mated does were allocated to four groups equalising distribution as far as possible, as to number allocated,source of animals and male to which they were mated.

- Length of cohabitation:1 hour

- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: The day of mating was considered as Day 0 of pregnancy.

- Further matings after two unsuccessful attempts: [no / yes (explain)] : No

- After successful mating each pregnant female was caged (how): Females were re-assigned to cages in the experimental room.

- Any other deviations from standard protocol: Structural deviations were classified as:
Malformations : rare and/or probably lethal, e.g. amelia,exencephaly.
Anomalies : minor differences from 'normal' that are detected relatively frequently either at initial examination. e.g. variations of the gall bladder, or at skeletal examination. e.g. hemicentric vertebra.

-Variants : alternative structures occurring regularly in the control population are classified as variants. These may be permanent structures e.g. an extra pair of ribs, or they may be transient stages of development. e.g. unossified sternebra(e).

Duration of treatment / exposure:

Days 7 to 19 of gestation

Frequency of treatment:

daily

Duration of test:

29 days

Remarks:

Doses / Concentrations:
0, 33, 100 and 300 mg/kg/day
Basis:

No. of animals per sex per dose:

Control - 16 females
33 mg/kg/day - 16 females
100 mg/kg/day – 18 females
300 mg/kg/day – 16 females

Control animals:

yes, concurrent vehicle

Details on study design:

No Data Available

Maternal examinations:

Maternal examinations
a) Clinical observations: All animals were regularly handled and observed daily;obvious changes or signs of reaction to treatment were recorded.

b) Mortalities : Any animals that died or were killed for reasons of animal welfare were weighed and subjected to post mortem examination.

At 100 and 300 mg/kg bw dose levels : No increase in mortality

c) Body weight: All rabbits were weighed on Days 1, 7, 9, 11, 15, 20, 24 and 29 of gestation.
At 300 mg/kg bw/day : Reduction of mean body weight

d) Food consumption: The food intake of all rabbits was measured from weigh-day to weigh-day.

At 300 mg/kg bw/day : decrease in mean food consumption observed.

Post-mortem examinations: Yes
- Sacrifice on gestation day 29
- Organs examined: ovaries and uteri
the foetuses were removed by caesarean section and examined for visceral and skeletal anomalies.

Ovaries and uterine content:

The uterine content was examined after termination: Yes
Examinations included:
number and distribution of live young: Yes
Gravid uterus weight: No
Number of corpora lutea: Yes
Number of implantations: Yes

Fetal examinations:

Fetal examinations
Individual foetal weight: Yes
Sex: Yes
Number and distribution of embryonic/foetal deaths: Yes
Embryonic/foetal deaths were classified as:
Early: only placenta visible at termination
Late: both placenta and embryonic remnants visible at
termination
Foetal abnormalities : Yes
Visceral and skeletal anomalies : Yes

Statistics:

Statistical analyses were performed routinely on litter data and incidences of skeletal anomalies and skeletal variants using the litter as the basic sample unit. Non-parametric methods (Kruskal-Wallis and Jonckheere tests) , were employed since these values rarely follow a normal distribution.

Indices:

Group mean values for litter size. embryonic death. pre- and post implantation loss were calculated in two ways:

Mean A: includes all surviving animals that provide evidence of pregnancy including those showing abortion or total
resorption.

Mean B: includes all animals with live young at termination.

Mean B has more meaning when group size is low in which case the mean values would be unduly influenced by the presence of a single animal with total litter loss.

Mean A is a more accurate index when several animals show total litter loss.

For each litter:
Pre-implantation loss was calculated as a percentage, from the formula:
(No. of corpora lutea - No. of implantations)/No. of corpora Iutea x 100

Post implantation loss was similarly calculated from the formula:
(No. of implantations - No. of live young)/NO. of implantations x 100

For each group:
All values expressed as a percentage or ratio were first
calculated within each litter and the group values derived as a mean of individual litter percentages.

For sex ratios of pups, litter weights, mean foetal weights and abnormality values, only Mean B were calculated.

Historical control data:

No Data Available

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

At all dosages but in particular at 100 and 300 mg/kg/day, treatment was generally associated with green discolouration of the urine. There were no other signs of reaction considered attributable to treatment.

Dermal irritation (if dermal study):

not specified

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Occasional animals were killed, found dead or excluded from the study for reasons of animal welfare. No association with treatment was indicated.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

At 300 mg/kg/day animals showed a slight initial mean weight loss from the start of treatment. Subsequently mean weight gain remained slightly lower than among control animals. Differences in mean weight gains at 33 and 100 mg/kg/day were not dosage related; overall weight gain was lower than for control animals at 33 mg/kg/day but was slightly higher at 100 mg/kg/day.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

At 300 mg/kg/day mean food consumption was reduced relative to control values. Differences from control values at 33 and 100 mg/kg/day showed no apparent relationship to dosage.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

No significant effects were observed by the administration of the test chemical in the animals.

Number of abortions:

not specified

Pre- and post-implantation loss:

not specified

Total litter losses by resorption:

not specified

Early or late resorptions:

not specified

Dead fetuses:

not specified

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

no effects observed

Other effects:

not specified

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
(a)Signs and mortalities :
At all dosages but in particular at 100 and 300 mg/kg/day, treatment was generally associated with green discolouration of the urine.
There were no other signs of reaction considered attributable to treatment.
Occasional animals were killed, found dead or excluded from the study for reasons of animal welfare. No association with treatment was indicated.

(b)Food consumption :
At 300 mg/kg/day mean food consumption was reduced relative to control values. Differences from control values at 33 and 100 mg/kg/day showed no apparent relationship to dosage.

(c) Bodyweight change :
At 300 mg/kg/day animals showed a slight initial mean weight loss from the start of treatment. Subsequently mean weight gain remained slightly lower than among control animals.

Differences in mean weight gains at 33 and 100 mg/kg/day were not dosage related; overall weight gain was lower than for control animals at 33 mg/kg/day but was slightly higher at 100 mg/kg/day.

(d) Pregnancy rate: Pregnancy rate, as judged by the numbers of animals pregnant at termination, was high in all groups.

(e) Terminal autopsy : There were no macroscopic findings at terminal autopsy that were considered attributable to treatment.

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

changes in number of pregnant

changes in pregnancy duration

clinical signs

effects on pregnancy duration

food consumption and compound intake

gross pathology

histopathology: non-neoplastic

mortality

Remarks on result:

other: not specified

Abnormalities:

not specified

Localisation:

not specified

Description (incidence and severity):

not specified

Fetal body weight changes:

no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

Changes in postnatal survival:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

Incidences of foetuses with extra thoraco-lumbar ribs or with variant sternebrae showed no statistically significant differences from controls and no relationship to dosage.

Visceral malformations:

no effects observed

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Litter data
(a)Total litter loss – No effect

(b)Litter size, pre and post implantation loss - not affected by the test chemical

(c) Litter and mean foetal weight - not affected by dthe test chemical

(d) Malformations and anomalies - No compound-related fetal malformations or anomalies observable.

At all dosages incidences of visceral and skeletal anomalies appeared unaffected by treatment.

(e) Skeletal variants : Incidences of foetuses with extra thoraco-lumbar ribs or with variant sternebrae showed no statistically significant differences from controls and no relationship to dosage.

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

changes in sex ratio

fetal/pup body weight changes

changes in litter size and weights

changes in postnatal survival

external malformations

skeletal malformations

visceral malformations

other: embryotoxicity

Remarks on result:

other: not specified

Abnormalities:

not specified

Localisation:

other: not specified

Description (incidence and severity):

not specified

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

Based on all the observation and results, it was concluded that, the NOAEL for the maternal animals was 100 mg/kg bw while the NOAEL for the fetal parameters was found to be 300 mg/kg bw.

Executive summary:

The effects of the test chemical on pregnancy of the New Zealand White rabbit were assessed in this study. Daily dosages of 0, 33, 100 or 300 mg of the test chemical. were administered by intragastic intubation during Days 7 to 19 inclusive of pregnancy. The maternal animals were observed for Clinical observations, wherein, all animals were regularly handled and observed daily;obvious changes or signs of reaction to treatment were recorded. Mortalities: Any animals that died or were killed for reasons of animal welfare were weighed and subjected to post mortem examination. Body weight: All rabbits were weighed on Days 1, 7, 9, 11, 15, 20, 24 and 29 of gestation. Food consumption: The food intake of all rabbits was measured from weigh-day to weigh-day. Post-mortem examinations: Yes, Sacrifice on gestation day 29, Organs examined: ovaries and uteri the foetuses were removed by caesarean section and examined for visceral and skeletal anomalies. The fetal prameters were observed for Individual foetal weight, Sex ratio, Number and distribution of embryonic/foetal deaths (classified as) Early i.e. only placenta visible at termination, Late i.e.both placenta and embryonic remnants visible at termination, Foetal abnormalities, and Visceral and skeletal anomalies. On Day 29, animals were sacrficed, litter values determined and foetuses examined for visceral and skeletal abnormality. There was no significant effect on: litter values, as assessed by litter size, pre- and post implantation loss, litter or mean foetal weights and embryonic and foetal development as assessed by incidence of malformations,anomalies and skeletal variants.

Thus, Based on all the observation and results, it was concluded that, the NOAEL for the maternal animals was 100 mg/kg bw while the NOAEL for the fetal parameters was found to be 300 mg/kg bw.

Cross-reference 3

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

Data is from a study report.

Qualifier:

according to guideline

Guideline:

other: OECD 422 (Combined Repeated and Reproduction/Developmental Toxicity Screening Test).

Principles of method if other than guideline:

According to OECD 422 (Combined Repeated and Reproduction/Developmental Toxicity Screening Test).

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

Crj: CD(SD)

Details on test animals or test system and environmental conditions:

Details on test animal
TEST ANIMALS
- Source: No data available
- Age at study initiation: (P) x wks; (F1) x wks: P- 9 weeks old
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g: No data available

- Fasting period before study: No data available
- Housing: No data available
- Diet (e.g. ad libitum): No data available
- Water (e.g. ad libitum): No data available
- Acclimation period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data available
- Humidity (%):No data available
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): No data available

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on exposure:

No data available

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No Data Available

Details on mating procedure:

No Data Available

Duration of treatment / exposure:

Male: 42 days
Female: 41 - 55 days (from 14 days before mating to day 4 of lactation)

Frequency of treatment:

daily

Duration of test:

No Data Available

Remarks:

Doses / Concentrations:
0, 25, 75 or 250 mg/kg/day
Basis:
no data

No. of animals per sex per dose:

No data available

Control animals:

yes, concurrent vehicle

Details on study design:

No Data Available

Maternal examinations:

Clinical sign, Body weight, Food consumption, Hematology, clinical chemistry and FOB were examined

Ovaries and uterine content:

Implantation Index, Resorption Index, Gestational Index.

Fetal examinations:

Pup Viability, Sex Ratio, Birth Index, Body weight of pups, and Skeletal malformations were observed.

Statistics:

No Data Available

Indices:

Pup viability index, Implantation Index, Resorption Index, Gestational Index.

Historical control data:

No Data Available

Clinical signs:

no effects observed

Description (incidence and severity):

No effect was observed on clinical sign of treated female rats as compared to control.

Dermal irritation (if dermal study):

not specified

Mortality:

not specified

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weight: No effect was observed on body weight of treated female rats as compared to control.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food consumption: No effect was observed on food consumption of treated female rats as compared to control.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

In female rats, decrease in the RBC and MCHC and increase in the RET was observed at recovery(250 mg/kg/day) as compared to control

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No effect was observed on clinical chemistry of treated male and female rats as compared to control.

Urinalysis findings:

not specified

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

No effect was observed on functional observation battery (FOB) of treated male and female rats as compared to control.

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Description (incidence and severity):

No effect was observed on body weight of treated male and female rats as compared to control.

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No Histopathological changes were observed in treated male and female rats as compared to control.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

No significant effects of the test chemical was observed on the test animals after the treatment.

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

Changes in number of pregnant:

no effects observed

Other effects:

not specified

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

behaviour (functional findings)

body weight and weight gain

changes in number of pregnant

changes in pregnancy duration

clinical biochemistry

clinical signs

dead fetuses

early or late resorptions

effects on pregnancy duration

food consumption and compound intake

haematology

histopathology: non-neoplastic

maternal abnormalities

mortality

number of abortions

organ weights and organ / body weight ratios

pre and post implantation loss

total litter losses by resorption

Remarks on result:

other: not specified

Abnormalities:

not specified

Fetal body weight changes:

no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

not specified

Changes in postnatal survival:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Visceral malformations:

no effects observed

Other effects:

not specified

Description (incidence and severity):

No effects were observed on fetal parameters at any given dose levels.

Details on embryotoxic / teratogenic effects:

No effect of the test chemical was observed on live pup index, viability index, and external anomalies.

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

changes in sex ratio

fetal/pup body weight changes

changes in postnatal survival

external malformations

skeletal malformations

visceral malformations

Remarks on result:

other: not specified

Abnormalities:

not specified

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

Based on all the observations and results, it was concluded that the NOAEL was considered to be 250 mg/kg/day when Crl:CD (SD) male and female rat were treated with the test chemical.

Executive summary:

In a combined repeated dose and reproduction/developmental screening test, male and female Crl:CD (SD) rats were treated orally with the test chemical in concentration of 0, 25, 75 or 250 mg/kg/day with recovery dose group of 0 and 250 mg/kg/day.No effect were observe on clinical sign, body weight and food consumption of treated rats as compared to control. Similarly, no effects were observed on blood chemistry, organ weight and histopathology of treated rats as compared to control. In addition, changes were observed as tendency to prolongation in the PT and prolongation in the APTT in male rats at 75 and 250 mg/kg/day dose group and decrease in the RBC and MCHC. An increase in the RET at 250 mg/kg/day dose group during recovery in female rats as compared to control. No reproductive effect were observed in treated male or female rats and on offspring’s as compared to control. Therefore,NOAEL for the parental generation and the F1 generation were considered to be 250 mg/kg/day when male and female Crl:CD (SD) rats were treated orally by gavage with the test chemical.

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

2006

Reference 2

Reference Type:

review article or handbook

Title:

SCREENING-LEVEL HAZARD CHARACTERIZATION Substituted Diphenylamines Category

Author:

U.S. Environmental Protection Agency

Year:

2009

Bibliographic source:

High Production Volume (HPV) Challenge Program;September, 2009

Reference 3

Reference Type:

publication

Title:

Effect Of Diphenylamine On Pregnancy Of The New Zealand White Rabbit

Author:

Edwards, J.A. et al.

Year:

1984

Bibliographic source:

Unpublished Report, Huntingdon Research Centre, submitted by Pennwalt Co., USA, to WHO: cited in FAO Plant Production and Protection Paper 67, Pesticide Residues in Food 1984, The Monographs, Rome 1985, pp. 641-643

Reference 4

Reference Type:

study report

Title:

Unnamed

Year:

2007

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

Equivalent or similar to OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine

EC Number:

239-816-9

EC Name:

Bis(4-(1,1,3,3-tetramethylbutyl)phenyl)amine

Cas Number:

15721-78-5

Molecular formula:

C28-H43-N

IUPAC Name:

4-(2,4,4-trimethylpentan-2-yl)-N-[4-(2,4,4-trimethylpentan-2-yl)phenyl]aniline

Details on test material:

Details on test material
- Name of test material (as cited in study report): 4,4'-Bis(1,1,3,3-tetramethylbutyl)diphenylamine
- Molecular formula (if other than submission substance): C28-H43-N
- Molecular weight (if other than submission substance): 393.655 g/mol
- Substance type: Organic
- Physical state: No Data Available
- Impurities (identity and concentrations): No Data Available

Test animals

Species:

other: Study 2: rat ; Study 3: rabbit Study 4: rat

Strain:

other: Study 2: Sprague-Dawley ; Study 3: New Zealand White Study 4: Crj: CD(SD)

Details on test animals or test system and environmental conditions:

Study 2: No Data Available

Study 3: TEST ANIMALS
- Source:Cheshire Rabbit Farms (Oudden Lodge. Nr. Tarporley.Cheshire). Ranch Rabbits (Crawley Down.Sussex). and Buxted Rabbit Co. Ltd (Gt. Totease Farm, Buxted. Sussex).
- Age at study initiation for p: Twelve to twenty weeks of age
- Weight at study initiation:2.9 to 4.1 kg
- Housing:The rabbits were housed individually in metal cages equipped with sheet stainless steel sides, back and top, a stainless steel wire front and an aluminium perforated floor panel.Individual undercage plastic trays were lined with absorbent paper which was inspected and changed daily.
- Diet (e.g. ad libitum):SDS Rabbit Diet SQC,ad libitum
- Water (e.g. ad libitum):Tap water,ad libitum
- Acclimation period:10 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C):18°C ± 3°C
- Humidity (%): 55% ± 15%
- Photoperiod (hrs dark / hrs light): 14 hours


Study 4: Details on test animal
TEST ANIMALS
- Source: No data available
- Age at study initiation: (P) x wks; (F1) x wks: P- 9 weeks old
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g: No data available

- Fasting period before study: No data available
- Housing: No data available
- Diet (e.g. ad libitum): No data available
- Water (e.g. ad libitum): No data available
- Acclimation period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data available
- Humidity (%):No data available
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): No data available

Administration / exposure

Route of administration:

other: Study 2: oral: gavage; Study 3: oral: gavage; Study 4: oral: gavage

Vehicle:

other: Study 2: corn oil; Study 3: 1% Methyl-cellulose; Study 4: Not Specified

Details on exposure:

Study 2: The test material was administered by gavage to three groups each of ten male and ten female Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, for up to fifty-four consecutive days, at dose levels of 50, 250 and 600 mg/kg/day.

Study 3: Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The dosages of diphenylamine administered by oral gavage in the present study were 33,100 and 300 mg/kg bodyweight/day. Treatment by intragastric intubation commenced on Day 7 of pregnancy and continued daily up to and including Day 19 of gestation. The highest required concentration of suspension was prepared by suspending the test material in the vehicle. The lower concentrations were prepared by serial dilution with the vehicle. The suspensions were prepared freshly each day. Treatment by intragastric intubation commenced on Day 7 of pregnancy and continued daily up to and including Day 19 of gestation. Dose volumes were calculated for individual animals on Day 7 of pregnancy and adjusted according to bodyweight on Days 9, 11 and 15.
VEHICLE : Methyl-cellulose

Study 4: No data available

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

Study 2: No Data Available

Study 3: No Data Available

Study 4: No Data Available

Details on mating procedure:

Study 2:
- M/F ratio per cage: 1:1
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy : The presence of sperm within the vaginal smear and/or vaginal plug in situ was taken as positive evidence of mating.
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. : N/A
- Further matings after two unsuccessful attempts: [no / yes (explain)] : no

Study 3:
Details of mating : Does were mated with males of proven fertility: once coitus had been observed, each female was allowed to remain with the male for at least one hour. Sixty-six of those dams which successfully completed coituswere each injected intravenously with 25 i.u. of Chorulon (luteinizing hormone) to promote ovulation. The day of mating was considered as Day 0 of pregnancy.

- Mating was phased over consecutive week days (Monday to Thursday) ensuring that no more than 20 animals were mated on anyone day. On each of these days, mated does were allocated to four groups equalising distribution as far as possible, as to number allocated,source of animals and male to which they were mated.

- Length of cohabitation:1 hour

- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: The day of mating was considered as Day 0 of pregnancy.

- Further matings after two unsuccessful attempts: [no / yes (explain)] : No

- After successful mating each pregnant female was caged (how): Females were re-assigned to cages in the experimental room.

- Any other deviations from standard protocol: Structural deviations were classified as:
Malformations : rare and/or probably lethal, e.g. amelia,exencephaly.
Anomalies : minor differences from 'normal' that are detected relatively frequently either at initial examination. e.g. variations of the gall bladder, or at skeletal examination. e.g. hemicentric vertebra.

-Variants : alternative structures occurring regularly in the control population are classified as variants. These may be permanent structures e.g. an extra pair of ribs, or they may be transient stages of development. e.g. unossified sternebra(e).

Study 4: No Data Available

Duration of treatment / exposure:

Study 2:
Males: 43 days;
Females: up to 54 days

Study 3: Days 7 to 19 of gestation

Study 4: Male: 42 days
Female: 41 - 55 days (from 14 days before mating to day 4 of lactation)

Frequency of treatment:

Study 2: Daily
Study 3: daily
Study 4: daily

Duration of test:

Study 2: 54 days prior to mating, throughout mating and gestation and continuing through lactation day 3
Study 3: 29 days
Study 4: No Data Available

Doses / concentrations

Remarks:

Study 2: Doses / Concentrations:
0, 50, 250, and 600 mg/kg/day
Basis:

Study 3: Doses / Concentrations:
0, 33, 100 and 300 mg/kg/day
Basis:

Study 4: Doses / Concentrations:
0, 25, 75 or 250 mg/kg/day
Basis:
no data

No. of animals per sex per dose:

Study 2: 20 animals in each group
0 mg/kg/day (control) - ten male and ten female Sprague-Dawley rats
50 mg/kg/day - ten male and ten female Sprague-Dawley rats
250 mg/kg/day - ten male and ten female Sprague-Dawley rats
600 mg/kg/day - ten male and ten female Sprague-Dawley rats

Study 3: Control - 16 females
33 mg/kg/day - 16 females
100 mg/kg/day – 18 females
300 mg/kg/day – 16 females

Study 4: No data available

Control animals:

yes, concurrent vehicle

Details on study design:

Study 2: No Data Available

Study 3: No Data Available

Study 4: No Data Available

Examinations

Maternal examinations:

Study 2: Survival, clinical sign, body weight, food consumption, water consumption, Duration of gestation, corpora luteaand, implantation site, fatality, hematology, clinical chemistry, oragn weight, gross patholology, histopathology and reperoductive performance were observed.
Pregnancy and parturition: The following was recorded for each female:
i) Date of mating
jj) Date and time of observed start of parturition
iii) Date and time of observed completion of parturition
iv) Duration of gestation

Study 3: Maternal examinations
a) Clinical observations: All animals were regularly handled and observed daily;obvious changes or signs of reaction to treatment were recorded.

b) Mortalities : Any animals that died or were killed for reasons of animal welfare were weighed and subjected to post mortem examination.

At 100 and 300 mg/kg bw dose levels : No increase in mortality

c) Body weight: All rabbits were weighed on Days 1, 7, 9, 11, 15, 20, 24 and 29 of gestation.
At 300 mg/kg bw/day : Reduction of mean body weight

d) Food consumption: The food intake of all rabbits was measured from weigh-day to weigh-day.

At 300 mg/kg bw/day : decrease in mean food consumption observed.

Post-mortem examinations: Yes
- Sacrifice on gestation day 29
- Organs examined: ovaries and uteri
the foetuses were removed by caesarean section and examined for visceral and skeletal anomalies.

Study 4: Clinical sign, Body weight, Food consumption, Hematology, clinical chemistry and FOB were examined

Ovaries and uterine content:

Study 2: Duration of gestation, corpora lutea and, implantation site was observed.

Study 3:
The uterine content was examined after termination: Yes
Examinations included:
number and distribution of live young: Yes
Gravid uterus weight: No
Number of corpora lutea: Yes
Number of implantations: Yes

Study 4: Changes in the estros cyclicity were observed.

Fetal examinations:

Study 2: Survival, clinical sign, body weight and development were observed.

Study 3:
Fetal examinations
Individual foetal weight: Yes
Sex: Yes
Number and distribution of embryonic/foetal deaths: Yes
Embryonic/foetal deaths were classified as:
Early: only placenta visible at termination
Late: both placenta and embryonic remnants visible at
termination
Foetal abnormalities : Yes
Visceral and skeletal anomalies : Yes

Study 4: No data available

Statistics:

Study 2: No Data Available

Study 3: Statistical analyses were performed routinely on litter data and incidences of skeletal anomalies and skeletal variants using the litter as the basic sample unit. Non-parametric methods (Kruskal-Wallis and Jonckheere tests) , were employed since these values rarely follow a normal distribution.

Study 4: No data available

Indices:

Study 2: Implantation Index, Gestational Index, Pup viability/survival index.

Study 3: Group mean values for litter size. embryonic death. pre- and post implantation loss were calculated in two ways:
Mean A: includes all surviving animals that provide evidence of pregnancy including those showing abortion or total
resorption.
Mean B: includes all animals with live young at termination.
Mean B has more meaning when group size is low in which case the mean values would be unduly influenced by the presence of a single animal with total litter loss.
Mean A is a more accurate index when several animals show total litter loss.
For each litter:
Pre-implantation loss was calculated as a percentage, from the formula:
(No. of corpora lutea - No. of implantations)/No. of corpora Iutea x 100
Post implantation loss was similarly calculated from the formula:
(No. of implantations - No. of live young)/NO. of implantations x 100
For each group:
All values expressed as a percentage or ratio were first
calculated within each litter and the group values derived as a mean of individual litter percentages.

For sex ratios of pups, litter weights, mean foetal weights and abnormality values, only Mean B were calculated.

Study 4: Implantation Index, Resorption Index, Gestational Index, Pup viability index.

Historical control data:

No Data Available

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Description (incidence and severity):

Study 2: no effects observed

Study 3: At all dosages but in particular at 100 and 300 mg/kg/day, treatment was generally associated with green discolouration of the urine. There were no other signs of reaction considered attributable to treatment.

Study 4: No effect was observed on clinical sign of treated male and female rats as compared to control.

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

Study 2: no mortality observed

Study 3: Occasional animals were killed, found dead or excluded from the study for reasons of animal welfare. No association with treatment was indicated.

Study 4: No data Available

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Study 3: At 300 mg/kg/day animals showed a slight initial mean weight loss from the start of treatment. Subsequently mean weight gain remained slightly lower than among control animals. Differences in mean weight gains at 33 and 100 mg/kg/day were not dosage related; overall weight gain was lower than for control animals at 33 mg/kg/day but was slightly higher at 100 mg/kg/day.

Study 4: Body weight: No effect was observed on body weight of treated male and female rats as compared to control.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Study 2: At 300 mg/kg/day mean food consumption was reduced relative to control values. Differences from control values at 33 and 100 mg/kg/day showed no apparent relationship to dosage.

Study 4: Food consumption: No effect was observed on food consumption of treated male and female rats as compared to control.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Study 4: In female rats, decrease in the RBC and MCHC and increase in the RET was observed at recovery(250 mg/kg/day) as compared to control

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

Study 4: No effect was observed on clinical chemistry of treated male and female rats as compared to control.

Urinalysis findings:

not specified

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Study 4: No effect was observed on functional observation battery (FOB) of treated male and female rats as compared to control.

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

Study 4: No effect was observed on body weight of treated male and female rats as compared to control.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Study 3: no effects observed

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

Study 3: no effects observed
Study 4: No Histopathological changes were observed in treated male and female rats as compared to control.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

Study 3: No significant effects were observed by the administration of the test chemical in the animals.

Maternal developmental toxicity

Number of abortions:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 4: no effects observed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 4: no effects observed

Total litter losses by resorption:

not specified

Early or late resorptions:

not specified

Dead fetuses:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 4: no effects observed

Changes in pregnancy duration:

no effects observed

Description (incidence and severity):

Study 3: no effects observed
Study 4: no effects observed

Changes in number of pregnant:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 3: no effects observed
Study 4: no effects observed

Other effects:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 4: no effects observed

Details on maternal toxic effects:

Study 2: Maternal toxic effects:no effects
Details on maternal toxic effects:
No effects on mating performance, fertility or gestation were observed.

Study 3: Maternal toxic effects:no effects
Details on maternal toxic effects:
(a)Signs and mortalities :
At all dosages but in particular at 100 and 300 mg/kg/day, treatment was generally associated with green discolouration of the urine.
There were no other signs of reaction considered attributable to treatment.
Occasional animals were killed, found dead or excluded from the study for reasons of animal welfare. No association with treatment was indicated.
(b)Food consumption :
At 300 mg/kg/day mean food consumption was reduced relative to control values. Differences from control values at 33 and 100 mg/kg/day showed no apparent relationship to dosage.
(c) Bodyweight change :
At 300 mg/kg/day animals showed a slight initial mean weight loss from the start of treatment. Subsequently mean weight gain remained slightly lower than among control animals.
Differences in mean weight gains at 33 and 100 mg/kg/day were not dosage related; overall weight gain was lower than for control animals at 33 mg/kg/day but was slightly higher at 100 mg/kg/day.
(d) Pregnancy rate: Pregnancy rate, as judged by the numbers of animals pregnant at termination, was high in all groups.
(e) Terminal autopsy : There were no macroscopic findings at terminal autopsy that were considered attributable to treatment.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

changes in number of pregnant

clinical signs

dead fetuses

effects on pregnancy duration

maternal abnormalities

mortality

number of abortions

pre and post implantation loss

Remarks on result:

other: Not Specified

Maternal abnormalities

Abnormalities:

not specified

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Study 2: Mean offspring weights for treated animals were comparable to controls.
Study 3: no effects observed
Study 4: no effects observed

Reduction in number of live offspring:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 3: no effects observed
Study 4: no effects observed

Changes in sex ratio:

no effects observed

Description (incidence and severity):

Study 3: no effects observed
Study 4: no effects observed

Changes in litter size and weights:

no effects observed

Description (incidence and severity):

Study 2: No treatment-related macroscopic abnormalities were detected for the interim death offspring or for the remaining offspring at terminal kill.
Study 3: no effects observed
Study 4: no effects observed

Changes in postnatal survival:

no effects observed

Description (incidence and severity):

Study 3: no effects observed
Study 4: no effects observed
Study 4: no effects observed

External malformations:

effects observed, treatment-related

Description (incidence and severity):

Study 2: Offspring from the 600 mg/kg/day dose group showed less successful completion of surface righting assessments. There were no treatment-related differences in pinna unfolding.
Study 3: no effects observed
Study 4: no effects observed

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

Study 3: Incidences of foetuses with extra thoraco-lumbar ribs or with variant sternebrae showed no statistically significant differences from controls and no relationship to dosage.

Visceral malformations:

no effects observed

Description (incidence and severity):

Study 2: no effects observed
Study 3: no effects observed

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

Study 2: Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Litter Observations: There were no clinical signs to suggest an effect of treatment.
Offspring Viability: Mean offspring weights for treated animals were comparable to controls.
Offspring Development: Offspring from the 600 mg/kg/day dose group showed less successful completion of surface righting assessments. There were no treatment-related differences in pinna unfolding.
Necropsy of Offspring: No treatment-related macroscopic abnormalities were detected for the interim death offspring or for the remaining offspring at terminal kill.

Study 3: Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Litter data
(a)Total litter loss – No effect
(b)Litter size, pre and post implantation loss - not affected by the test chemical
(c) Litter and mean foetal weight - not affected by dthe test chemical
(d) Malformations and anomalies - No compound-related fetal malformations or anomalies observable.
At all dosages incidences of visceral and skeletal anomalies appeared unaffected by treatment.
(e) Skeletal variants : Incidences of foetuses with extra thoraco-lumbar ribs or with variant sternebrae showed no statistically significant differences from controls and no relationship to dosage.

Study 4:

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

fetal/pup body weight changes

changes in litter size and weights

external malformations

visceral malformations

Remarks on result:

other: Not Specified

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Treatment related:

not specified

Applicant's summary and conclusion

Conclusions:

Study 2: Based on all the observation and results, the NOAEL for maternal and fetotoxicity study was considered to be 250 mg/Kg bw/day.

Study 3: Based on all the observation and results, it was concluded that, the NOAEL for the maternal animals was 100 mg/kg bw while the NOAEL for the fetal parameters was found to be 300 mg/kg bw.

Study 4: Based on all the observations and results, it was concluded that the NOAEL was considered to be 250 mg/kg/day when Crl:CD (SD) male and female rat were treated with the test chemical.

Executive summary:

Developmental Toxicity Study:

The summaries for the developmental toxicity studies are as follows:

Developmental Toxicity Study 2:

In the combined repeated-dose/reproductive/developmental toxicity screening study, the test material was administered by gavage to three groups each of ten male and ten female Sprague-Dawley Crl:CD® (SD) IGS BR strain rats, for up to fifty-four consecutive days, at dose levels of 50, 250 and 600 mg/kg/day. The following was recorded for each female: Date of mating, Date and time of observed start of parturition, Date and time of observed completion of parturitionand Duration of gestation. After dosing, it was observed that,Females treated with 600 mg/kg/day showed a higher percentage of pre-implantation losses in comparison to controls, resulting in the birth of less offspring per litter and lower total litter weights at this dose level. Thus, no adverse effects (NOAEL) on mating performance, fertility or gestation were observed at 250 mg/kg-bw/day of concentartion. Also, the same NOAEL was considered for fetal parameters. Thus, based on all the observation and results, it was concluded that the NOAEL of the test chemical for the maternal and fetal parameters considered to be 250 mg/kg bw after administration to the test animals.

Developmental Toxicity Study 3:

The effects of the test chemical on pregnancy of theNew Zealand Whiterabbit were assessed in this study. Daily dosages of 0, 33, 100 or 300 mg of the test chemical. were administered by intragastic intubation during Days 7 to 19 inclusive of pregnancy. The maternal animals were observed forClinical observations, wherein, all animals were regularly handled and observed daily;obvious changes or signs of reaction to treatment were recorded. Mortalities: Any animals that died or were killed for reasons of animal welfare were weighed and subjected to post mortem examination. Body weight: All rabbits were weighed on Days 1, 7, 9, 11, 15, 20, 24 and 29 of gestation. Food consumption: The food intake of all rabbits was measured from weigh-day to weigh-day. Post-mortem examinations: Yes, Sacrifice on gestation day 29, Organs examined: ovaries and uteri the foetuses were removed by caesarean section and examined for visceral and skeletal anomalies. The fetal prameters were observed forIndividual foetal weight, Sex ratio, Number and distribution of embryonic/foetal deaths (classified as) Early i.e. only placenta visible at termination, Late i.e.both placenta and embryonic remnants visible at termination, Foetal abnormalities, and Visceral and skeletal anomalies.On Day 29,animals were sacrficed, litter values determined and foetuses examined for visceral and skeletal abnormality.There was no significant effect on:litter values, as assessed by litter size, pre- and post implantation loss, litter or mean foetal weights andembryonic and foetal development as assessed by incidence of malformations,anomalies and skeletal variants.

Thus, Based on all the observation and results, it was concluded that, the NOAEL for the maternal animals was 100 mg/kg bw while the NOAEL for the fetal parameters was found to be 300 mg/kg bw.

Developmental Toxicity Study 4:

In a combined repeated dose and reproduction/developmental screening test, male and female Crl:CD (SD) rats were treated orally with the test chemical in concentration of 0, 25, 75 or 250 mg/kg/day with recovery dose group of 0 and 250 mg/kg/day.No effect were observe on clinical sign, body weight and food consumption of treated rats as compared to control. Similarly, no effects were observed on blood chemistry, organ weight and histopathology of treated rats as compared to control. In addition, changes were observed as tendency to prolongation in the PT and prolongation in the APTT in male rats at 75 and 250 mg/kg/day dose group and decrease in the RBC and MCHC. An increase in the RET at 250 mg/kg/day dose group during recovery in female rats as compared to control. No reproductive effect were observed in treated male or female rats and on offspring’s as compared to control. Therefore,NOAEL for the parental generation and the F1 generation were considered to be 250 mg/kg/day when male and female Crl:CD (SD) rats were treated orally by gavage with the test chemical.