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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (according to OECD 422 and GLP)
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2001
Report Date:
2001

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: particulates

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Japan, Inc. (Atsugi Breeding center)
- Age at study initiation: 8 weeks
- Weight at study initiation: males 309-355 g, females: 206 to 232 g
- Housing: individually; mating period: 1 male, 1 female; 1 litter during lactation
- Diet: autoclaved pellet die for laboratory animals (CRF-1, Oriental Yeast Industry Co., LTD), ad libitum
- Water: sterile tap water, ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/-2
- Humidity (%): 55 +/- 15
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 0.5 % sodium-carboxymethylcellulose (CMC-NA) aqueous solution containing 0.1 % Tween 80
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of test substance : once or twice a week
- Storage temperature of suspension: 4°C

VEHICLE
- Justification for use and choice of vehicle (if other than water): no data
- Amount of vehicle (if gavage): 10 mL/kg bw
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: 14 days
- Proof of pregnancy: vaginal plug or sperm in vaginal smear referred to as day 0 of pregnancy
- Further matings after two unsuccessful attempts: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
At the first preparation, the dosing formulations were analyzed by HPLC and it was confirmed that the concentrations were within the target value +/-10%.
Duration of treatment / exposure:
males: from 14 days before mating and through the mating period until the day before necroscopy for a total of 37 days
females: from 14 days beformating, through the matingand gestation periods and delivery until day 4 of lactation for a total of 42 to 47 days. Non-delivering females, however, were treated until the day before necropsy.
Frequency of treatment:
daily
Details on study schedule:
no data
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 300, 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: highest dose is the upper limit dose level stipulated in the OECD guideline
- Rationale for animal assignment (if not random): stratified-by-weight randomization methodon the basis of the body weights to get almost the same mean body weights for each group.
Positive control:
none

Examinations

Parental animals: Observations and examinations:
For deatails on parental animals see section 7.5.1 (repeated dose toxicity oral).
Oestrous cyclicity (parental animals):
From the first day of dosing to the start of mating, the estrous cycle was examined by collecting vaginal smears every day in the morning for all females of each group, and the mean estrous cycle length of each female was calculated. Females that had an estrous cycle length other than 4 to 6 days were judged as having irregular estrous cycles.
Sperm parameters (parental animals):
no data
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies

GROSS EXAMINATION OF DEAD PUPS: yes
Postmortem examinations (parental animals):
For deatials on parental animals see section 7.5.1 (repeated dose toxicity oral).

Examination of the ovary and uteri of the dams for the numbers of corpora lutea and implantations.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring were sacrificed at 4 days of age.
- These animals were subjected to postmortem examinations (macroscopic and microscopic examination) as follows:

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

Statistics:
Multiple comparison test:
Body weight, body weight gain, food consumption, hematology, blood chemistry, organ weight, number of corpora lutea, number of implantations, number of offspring

Kruskal-Wallis test and Dunnett-type multiple comparison test:
Days until copulation, number of estrus stages without copulation, mean estrous cycle length, gestation length, implantation index, delivery index, live birth index, incidence of offspring with external anomaly, viability index an Day 4

Chi-square test: Histopathological findings

Fisher's exact probability test:
Incidence of females with irregular estrous cycles, copulation index, fertility index, gestation index, sex ratio (rnale/fernale), incidence of dams with externally abnormal offspring
Reproductive indices:
(1) Days until copulation: Number of days from the start of mating to detection of copulation
(2) Number of estrus stages without copulation
(3) Copulation index (%): (Number of animals with successful copulation/Number of animals paired) x 100
(4) Fertility index (%): (Number of pregnant animals/Number of animals with successful copulation) x 100
(5) Gestation length: Days until completion of delivery from Day 0 of gestation
(6) Gestation index (%): (Number of females with live offspring/Number of pregnant females) x 100
(7) Implantation index (%): (Number of implantations/Number of corpora lutea) x 100
(8) Delivery index (%): (Total number of offspring born/Number of implantations) x 100
Offspring viability indices:
(1) Live birth index (%): (Number of offspring born alive/ Total number of offspring born) x 100
(2) Viability index on Day 4 (%): (Number of live offspring on Day 4/ Number of offspring born alive) x 100

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
details see section 7.5.1
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
details see section 7.5.1
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not examined

Reproductive function / performance (P0)

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

Details on results (P0)

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
In the estrous cycle examination, there was no significant difference in the mean estrous cycle length between the control and treated groups. Moreover, there were no females with irregular estrous cycles in any group including the control group.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
As a results of mating, all the mating pairs of each group copulated including the control group, and there was no significant difference in the copulation index, nor were there any in the number of estrus stages without copulation or the days until copulation between the control and treated groups. Moreover, there were only 1 and 2 non-pregnant females in the 100 and 300 mg/kg groups, respectively, and no significant difference was found in the fertility index between the control and treated groups.

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There were no effects on reproduction or development up to the highest dose tested.
Remarks on result:
other: Generation: P and F1 (migrated information)

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Details on results (F1)

1) Observation of offspring
There were no significant differences in the number of offspring born or born alive, sex ratio, live birth index or viability index on Day 4 between the control and each treated group. In the clinical observation, subcutaneous hemorrhage and abdominal hemorrhage were observed in 1 or 2 pups each in the control and 100 mg/kg groups, respectively; however, they were judged not treatment-related because of the lack of dose-dependency in their incidences. Although loss of suckling was observed in some neonates on the birthday in the control, 300 and 1000 mg/kg groups, it was judged not treatment-related because all pups were confirmed to have been suckled by the dams on the next day. Moreover, no abnormal external features were found in neonates in any group including the control group.

2) Body weight
There were no significant differences in body weight or body weight gain of either sex between the control and treated groups.

3) Necropsy findings
Hemorrhage in the abdominal cavity, and white and yellow patches on the liver were observed only in 1 male in the 100 mg/kg group, as was dilatation of the renal pelvis in 2 males in the 1000 mg/kg group. However, they were judged not treatrnent-related, because the same findings did not occur frequently in the 1000 mg/kg group. Moreover, there were no abnormal findings in pups that died until postnatal Day 4.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Under the conditions of the test, there were no effects on reproduction or development; therefore, the NOAEL for the reproductive / developmental toxicity was considered to be 1000 mg/kg bw/day in both parental animals and offspring.
Executive summary:

A combined repeated dose toxicity study with the reproduction/developmental toxicity screening test with the test item administered by oral gavage in Sprague Dawley rats (12/sex/dose) was performed according to OECD TG 422. The dose levels for this study were 0, 100, 300 and 1000 mg/kg/day for a total of 37 days (males) and through mating, gestation and delivery until day 4 of lactation (females).

There were no effects on reproduction or development; therefore, the NOAEL for the reproductive/developmental toxicity was considered to be 1000 mg/kg/day in both parental animals and offspring.