Registration Dossier

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
other information
Study period:
1997
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
secondary literature

Data source

Reference
Reference Type:
secondary source
Title:
no data
Author:
MHW, Japan
Year:
1997
Bibliographic source:
cited in OECD SIDS Final 08/02

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- purity: 99.9%

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on mating procedure:
- premating exposure period: 14 days
Duration of treatment / exposure:
male: from 14 days before mating to 14 days after mating; males were exposed for 42 days
female: from 14 days before mating to day 3 of lactation
Frequency of treatment:
daily
Doses / concentrationsopen allclose all
Dose / conc.:
2 mg/kg bw/day (nominal)
Dose / conc.:
10 mg/kg bw/day (nominal)
Dose / conc.:
50 mg/kg bw/day (nominal)
No. of animals per sex per dose:
13
Control animals:
yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:
BODY WEIGHT: Yes
- Time schedule for examinations: twice a week the first week; weekly afterwards, including during the pregnancy of females. Additionally for females, days 0 and 4 of lactation.

FOOD CONSUMPTION AND COMPOUND INTAKE
- Food consumption determined in four day blocks during the first week and weekly afterwards.
- Compound intake calculated in mean diet as g food/rat/average over the period of calcul.
Sperm parameters (parental animals):
Parameters examined in all male parental generations: testis weight, epididymis weight
Litter observations:
STANDARDISATION OF LITTERS
no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, live births, postnatal mortality (on day 4), body weight recorded on day 0 and day 4.

GROSS EXAMINATION OF DEAD PUPS
yes, morphological
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals on day 43.
- Maternal animals: All surviving animals on day 4 post partum.

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues were prepared for microscopic examination: Heart, thymus, spleen and liver, kidney, adrenal gland, testis, epididymis. These ones were also weighed.
Postmortem examinations (offspring):
SACRIFICE
- Supposed on day 4. Not accurately indicated.

GROSS NECROPSY
- Gross necropsy consisted of external examinations
Reproductive indices:
Copulation index, fertility index, gestation index, duration of pregnancy
Offspring viability indices:
viability index

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Males: Temporary salivation after each administration was observed in the animals exposed at 10 mg/kg and more.
Females: One female in the 50 mg/kg group died on post-partum day 3.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Males: Suppression of body weight gain and food consumption during the early administration period were noted in the 50 mg/kg group.
Females: Slight decrease in body weight gain and food consumption during the early administration period were observed in the animals exposed at 10 mg/kg and more.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Males: Increased eosinophilic bodies and basophilic changes of the renal tubular epithelial cells in the kidneys of all the animals exposed were noted, as well as granular casts in the lower nephrons. Centrilobular hypertrophy of hepatocytes was also detected in the groups of animals exposed at 10 mg/kg and more.
Females: Centrilobular hypertrophy of hepatocytes was observed in the groups of animals exposed at 10 mg/kg and more.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
The compound showed no adverse effects on copulation and fertility, duration of pregnancy, gestation index and parturition at any of the dose levels tested. Three dams in the 50 mg/kg group showed abnormal lactation.

Details on results (P0)

There were no adverse effects of 2,2'-azobis(2- methylpropanitrile) on copulation and fertility, duration of pregnancy, gestation index and parturition at all treated group. Three of 12 dams at 50 mg/kg showed the difficulty of nursling and two of them let all their offsprings die within the first 4 days after birth.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day
Based on:
test mat.
Sex:
male
Basis for effect level:
other: No effect on reproductive toxicity in males
Dose descriptor:
NOAEL
Effect level:
10 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: At 50 mg/kg bw/d, abnormal lactation in 3 dams

Target system / organ toxicity (P0)

Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Gross pathological findings:
no effects observed

Details on results (F1)

This compound showed no adverse effects on viability, sex ratio and body weight gain of pups. However, viability of newborns at birth and body weight of nurslings on postnatal day 4 was lower than the control levels at 50 mg/kg/day. These changes were considered to be caused by maternal toxicity. There were no morphological abnormalities in pups at all treated groups.
For reproductive and developmental toxicity the NOEL was 10 mg/kg in pups according to the authors.
It was considered that the effects on pups were caused by maternal toxicity since a difficulty in nursing (lactation) was reported. Then the NOAEL for reproduction in offspring was considered to be 50 mg/kg.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
10 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: decreased viability index and body weight on day 4 at 50 mg/kg/day

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

General parenteral toxicity:

There were no adverse effects of 2,2'-azobis(2- methylpropanitrile) on copulation and fertility, duration of pregnancy, gestation index and parturition at all treated group. Three of 12 dams at 50 mg/kg showed the difficulty of nursling and two of them let all their offsprings die within the first 4 days after birth.

Toxicity to offspring:

This compound showed no adverse effects on viability, sex ratio and body weight gain of pups. However, viability of newborns at birth and body weight of nurslings on postnatal day 4 was lower than the control levels at 50 mg/kg/day. These changes were considered to be caused by maternal toxicity. There were no morphological abnormalities in pups at all treated groups.

Table 1: Summary of development of pups

Dose (mg/kg)

0

2

10

50

Number of pregnant females

12

12

11

12

Number of pregnant females with pup alive

12

12

11

12

Gestation Index

100

100

100

100

Gestation length in days

22.3 +/-0.5 (12)

22.1 +/-0.3 (12)

22.2 +/-0.4 (11)

22.4 +/-0.5 (12)

Number of corpora lutea

18.8 +/-2.5 (12)

17.8 +/-1.5 (12)

18.1 +/-2.0 (11)

16.9 +/-2.0 (12)

Number of implantation sites

17.3 +/-25 (12)

16.8 +/-1.5 (12)

17.1 +/-2.1 (11)

15.6 +/-1.4 (12)

Implantation index

92.9 +/-10.6 (12)

95.0 +/-5.7 (12)

94.7 +/-7.7 (11)

92.8 +/-9.3 (12)

Day 0 of lactation

 

 

 

 

Number of pups born

15.4 +/-2.5 (12)

15.8 +/-1.9 (12)

15.5 +/- 1.8 (11)

14.8 +/-1.5 (12)

Delivery index

88.8 +/-6.2 (12)

94.0 +/-6.1 (12)

91.6 +/-10.3 (11)

94.6 +/-4.4 (12)

Number of pups alive

15.0 +/-2.5 (12)

15.8 +/-1.9 (12)

15.5 +/-1.8 (11)

14.8 +/-1.5 (12)

Birth index

86.6 +/- 8.3 (12)

93.5 +/- 5.8 (12)

91.6 +/-10.3 (11)

94.6 +/-4.4 (12)

Live birth index

97.4 +/-4.9 (12)

99.4 +/- 1.9 (12)

 100.0 +/-0.0 (11)

100.0 +/- 0.0 (12)

Pup weight in grams

 

 

 

 

Male

6.2 +/-0.6 (12)

6.3 +/-0.5 (12)

6.3 +/-0.4 (11)

6.1 +/-0.2 (12)

Female

5.9 +/-0.5 (12)

6.0 +/-0.5 (12)

6.0 +/-0.3 (11)

5.9 +/-0.2 (12)

Sex ratio

50.5 +/-9.9 (12)

43.9 +/-12.8(12)

54.4 +/-10.9 (11)

53.6 +/-11.7 (12)

Day 4 of lactation

 

 

 

 

Number of pups alive

14.6 +/-2.3 (12)

15.6 +/-1.9 (12)

15.5 +/-1.8 (11)

11.6 +/-5.9 (12)

Viability index

 97.5 +/-4.1 (12)

99.0 +/-2.4 (12)

100.0 +/-0.0 (11)

77.9 +/-38 (12)

Pup weight in grams

 

 

 

 

Male

9.8 +/-1.4 (12)

9.8 +/-1.1 (12)

9.8 +/-1.0 (11)

9.0 +/-1.2 (12)

Female

9.5 +/-1.2 (12)

9.5 +/-1.1 (12)

9.4 +/-1.0 (11)

8.5 +/-1.1 (12)

Gestation Index: Number of pregnant female with pups alive/Number of pregnant females

Implantation Index: Number of implantation sites/Number of corpora lutea *100 (%)

Delivery Index: Number of pups born/Number of implantation sites *100 (%)

Birth Index: Number of pups alive on day 0/ Number of implantation sites *100 (%)

Live Birth Index: Number of pups alive on day 0/ Number of pups born *100 (%)

Sex ratio: Number of male pups alive on day 0/ Number of pups alive on day 0 *100 (%)

Viability index: Number of pups alive on day 4/ Number of pups alive on day 0 *100 (%)

Applicant's summary and conclusion

Conclusions:
In conclusion, the NOEL for reproductive and developmental toxicity was 50 mg/kg/day in males and 10 mg/kg/day in females and in pups. According to CLP criteria, the test substance is not classified.
Executive summary:

The reproductive / developmental toxicity of the test substance was evaluated in male and female rats after oral administration (gavage) at doses of 0, 2, 10 and 50 mg/kg/day until day 43 for males and day 4 of post partum for females.

The test substance had no effects on the copulation index or fertility index at 50 mg/kg or lower doses. The test substance did not affect the length of gestation period or delivery index in maternal animals, either. No abnormal parturition was observed. Abnormal nursing behavior was noted in 3 animals from the 50 mg/kg group.

The findings in offspring showed no effects on the parturition index, live pup delivery index, overall delivery index, or the sex ratio and body weight on day 0 at all dose-levels. In the 50 mg/kg group, the offspring viability index and body weight on day 4 of lactation showed a tendency to decrease. No offspring from the test substance treated groups showed any morphological anomaly.

Based on these results, the NOEL for reproductive and developmental toxicity was 50 mg/kg/day in males and 10 mg/kg/day in females and in pups.


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