Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The OECD Guideline for the Testing of Chemicals, Proposal for Updating Guideline 414, "Prenatal Developmental Toxicity Study", Adopted on 22nd January 2001, GLP compliant

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
Name of test material (as cited in study report): GRO 266
- Sponsor’s internal code: SAT 090019
- Physical state: White to beige powder
- Analytical purity: >99 (area % by HPLC); 100.3 wt % (by 1H-NMR)
- Lot/batch No.: GRO-RN-9963-191

Test animals

Species:
rat
Strain:
other: RccHan:WIST
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories, Netherland
- Age at study initiation: 10 to 11 weeks old at the time of initiation of acclimatisation
- Weight at study initiation: between 167.00 – 251.00 gm
- Fasting period before study: No
- Housing: Individually in clean sterilised solid floor polypropylene rat cages
- Diet (e.g. ad libitum): Rat Pellet Feed
- Water (e.g. ad libitum):Clean drinking water
- Acclimation period: At list 6 days

ENVIRONMENTAL CONDITIONS
- Standard international environmental condition

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Solutions were prepared fresh everyday prior to dosing.
Dose volume : 10 mg/ml
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The content of active ingredient and the homogenicity in the used vehicle were analytically determined
Details on mating procedure:
-Impregnation procedure: Post acclimatisation, male and female rats (1:1)
-Length of cohabitaion: till 100 sperm positive females were obtained
-Proof of pregnancy: by examination of the vaginal smear, collected after lavage. Females showing presence of spermatozoa in the smear were separ ated. The day of recording of spermatozoa in the vaginal smear was termed as Day " 0" of gestation.
-Further mating of females after unsuccessful attenots : no
Duration of treatment / exposure:
Starting from gestaion day 5th to 19th
Frequency of treatment:
Once Daily
Doses / concentrations
Remarks:
Doses / Concentrations:
200, 400 and 800 mg/kg body weight/day
Basis:
actual ingested
No. of animals per sex per dose:
To obtain minimum 20 gravid females per group at time of sacrifice required 25 sperm positive females per group
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale:
During range finding study, pregnant female rats (5/group) were exposed daily by oral gavage to GRO 266 in distilled water at dose levels of 250, 500 and 1000 mg /kg body weight/day during gestational days (GD) 5th to 19th. The range finding study was conducted in an identical manner to main prenatal developmental toxicity study including all parameter for assessing the effects of prenatal exposure on the pregnant test rats and on the developing foetus.

Result of the Dose Range finding Study:
• Mortality was not observed up to the dose level of 1000mg/kg body weight/day.
• Gestational body weight, percent body weight change and feed consumption data did not reveal any significant changes during the gestation period up to the dose level of 1000 mg/kg body weight/day.
• Prenatal and foetal data were not affected up to the dose level of 1000 mg/kg body weight/day.
• Foetal external, visceral, head razor section and skeletal evaluation reveal no treatment related lesion/alteration up to the dose level of 1000 mg/kg body weight/day

Based on the above results of the dose range finding study, the dose levels selected for main study were 0, 200, 400 and 800 mg/kg body weight/day.


Examinations

Maternal examinations:
MORTALITY AND MORBIDITY: Yes
Time Schedule: Twice daily
CLINICAL OBSERVATION: Yes
Time Schedule: once daily after treatment
GESTATIONAL BODY WEIGHT: Yes
Time Schedule: On day 0, 3, 5, 8, 11, 14, 17 and 20 of gestation
FEED CONSUMPTION: Yes
Time Schedule: During 0-3, 3-5, 5-8, 8-11, 11-14, 14-17, and 17-20 gestation period.
An amount of 200 g of rat pellet feed was given to each pregnant female on each occasion and the feed left over was recorded.
POST-MORTEM EXAMINATION (NECROPSY): Yes
Tme Schedule: On day 20th of gestation
Gross Pathology Observation: The maternal viscera including uteri were examined macroscopically
Ovaries and uterine content:
OVARIES AND UTERINE CONTENT WAS EXAMINATION AFTER TERMINATION: Yes
Examination inclluded:
Number of corpora lutea
Gravid uterus weight (g)
Number of implantations
Number of resorptions (early/late)
Number of dead foetuses
Number of live foetuses
Fetal examinations:
Number of male foetuses
Number of female foetuses
The sex of the foetus
Individual foetal weight (g)
Gross external examination : all foetuses per litter
Visceral (Soft tissue) and Head razor examination : half foetuses per litter
Skeletal Examination: half foetuses per litter
Statistics:
Mean Values were compared using Bartlett's test for their variation. If the result is insignificant then mean values were compared by one-way analysis of variance (ANOVA) and the Dunnett test (mean values being considered as normally distributed and variances being considered as homogeneous). Other wise mean values of treated groups are compared with treated groups using Student't test. Parcentage values were compared by Chi-Square test
Indices:
Per cent Maternal Mortality: (Number of female dead during the test/Number of positive spermatozoan female at the start) X 100
Pregnancy Rate: (Number of female with implantation site at term/Number of spermatozoan positive female sacrificed at term) X 100
20th Day Maternal Corrected Body Weight: 20th day maternal gestational body weight - uterine weight
Body Weight Change (%) at each interval:
((Body weight on a particular interval - Body weight on previous interval)/ Body weight on previous interval) X 100
Relative Uterine Weight (%): (Gravid uterine weight/20th day maternal body weight)X 100
Total resorptions: Early resorptios + Late resorptions
Pre-implantation Loss (%): ((Number of corpora lutea - Number of implants)/Number of corpora lutea) X 100
Post-implantation Loss (%): (Total number of resorption/ Number of implants) X 100
Live Foetus (%): (Number of live foetus/ Number of implants) X 100
Dead Foetus (%): (Number of dead foetus/ Number of implants) X 100
Average foetus Weight: Sum of Individual foetus weight/number of foetus
Sex Ratio : Number of male foetus/Number of female foetus

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Details on maternal toxic effects:
The mean maternal body weights of pregnant rats were significantly reduced on 20th day of gestation at the dose levels of 400 and 800 mg/kg body weight/day.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Basis for effect level:
other: maternal toxicity
Basis for effect level:
other: effect type not specified

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Statistically significant reduction was recorded in the mean male, female and total (male + female) foetus weight at the dose level of 800 mg/kg body weight/day as compared to the control group.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
400 mg/kg bw/day (actual dose received)
Basis for effect level:
other: fetotoxicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Executive summary:

This study was performed to evaluate the possible prenatal developmental toxicity of GRO 266 by oral gavage in pregnant Wistar rats. The method followed was as per the guideline of OECD N° 414 (January 2001).

Pregnant Wistar rats were exposed during gestational days (GD) 5 to 19, to GRO 266 dissolved in distilled water once daily, by oral gavage at the dose levels of 200, 400 and 800 mg/kg body weight/day, based on test results of a dose-range finder study. Pregnant rats from the control group were administered distilled water only. Maternal body weights, clinical symptoms and feed consumption were recorded throughout the gestation period. The treated and control rats were sacrificed on GD 20, the uteri were removed, weighed and examined for the number of implantation, resorption and for live and dead foetus. The ovaries were checked for corpora lutea. The foetus were weighed and examined for external, visceral, head razor section and skeletal abnormalities.

Mortality was not observed upto the dose level of 800 mg/kg body weight/day dose group. Pregnancy data remained comparable between rats from various treated groups and concurrent control group. All the rats were normal during the treatment period.

The mean maternal body weights of pregnant rats were significantly reduced on 20th day of gestation at the dose levels of 400 and 800 mg/kg body weight/day. The percent body weight change data remained comparable between the rats from various treated groups and control group. The feed consumption of the E+ rats was significantly reduced in all test groups during 5th and 8th day of gestation, just starting with the oral application of the test substance. This temporary reduction of feed consumption was compensated in low dosed animals after the 8th day of gestation and in the mid and high dosed groups after the 14th day of gestation to be equivalent to the control group at the end of the treatment phase. Based on pathological examinations, the test substance up to the highest tested dose of 800 mg/kg body weight/day was found to be non-toxic in maternal rats.

No treatment related effects were observed in the mean prenatal data upto the dose level of 800 mg/kg body weight/day. Statistically significant reduction was recorded in the mean male, female and total (male + female) foetus weight at the dose level of 800 mg/kg body weight/day as compared to the control group.

No significant difference in the incidences of malformation or birth defects was recorded during external, visceral, head razor and skeletal examination of foetus from the control and various treated groups.

From the present study, it is concluded that the "No Observed Adverse Effect Level (NOAEL)" for maternal toxicity of GRO 266 is conservatively considered to be 200 mg/kg body weight/day due to reduced absolute body weight in the 400 and 800 mg/kg body weight/day dose groups. It should be mentioned that body weight of treated animals was not different to the control group, when corrected by the individual uteri weights. The NOAEL for fetal toxicity of GRO 266 is 400 mg/kg body weight/day due to reduced body weight seen in 800 mg/kg body weight/day dose group.

The results of the present study indicate that GRO 266 is non-teratogenic in rats upto the dose level of 800 mg/kg body weight/day.