Registration Dossier

Toxicological information

Toxicity to reproduction

Currently viewing:

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Data is from study report

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Principles of method if other than guideline:
According to OECD 421 (Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Chemical Name: 5-Decanolide
Molecular Formula: C10H18O2
Molecular Weight: 170.25 g/mol
Consistency: Colorless to light yellow oily liquid
Activity (Clinical Indication):Industrial Chemical
Storage Condition: At room temperature (RT)in ventilated cabinets.
Safety Precautions: Safety precautions included use of protective clothing, gloves, masks and eye protection (glasses).

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source:In-house bred animals
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) x wks; (F1) x wks: 9 to 11 weeks
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g: Males: 240.91 g to 288.02 g
Females: 200.02 g to 232.05 g
- Fasting period before study:
- Housing: Animals were housed in a standard polypropylene cage (size: L 430 x B 285 x H 150 mm) with stainless steel mesh top grill having facilities for holding pelleted food and drinking water in water bottle fitted with stainless steel sipper tube. Clean sterilized paddy husk was provided as bedding material.
i. Pre mating
Two animals of same sex and group per cage were housed.
ii. Mating
During mating, two animals (one male and one female) of same group were housed.
iii. Post mating
After confirming presence of sperm in the vaginal smear (Day 0 of pregnancy), the mated pairs were separated. Males were housed with their former cage mates while females were housed individually. Sterilized paper shreds were provided as a nesting material from gestation day 20 onwards.
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): Altromin Maintenance diet for rats and mice 1324 manufactured by Altromin Spezialfutter GmbH & Co. KG was provided ad libitum to the animals throughout the experimental period.
- Water (e.g. ad libitum): Water was provided ad libitum throughout the acclimatization and experimental period. Deep bore-well water passed through reverse osmosis unit was provided in plastic water bottles with stainless steel sipper tubes.
- Acclimation period:

ENVIRONMENTAL CONDITIONS
- Temperature (°C):19.1 to 23.6oC
- Humidity (%):40 to 69%
- Air changes (per hr): 12 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light and 12 hours dark cycle

IN-LIFE DATES: From: 29 November 2017
To: 09 May 2018

Administration / exposure

Route of administration:
oral: gavage
Type of inhalation exposure (if applicable):
not specified
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:The test item formulations were freshly prepared before dose administration on each treatment day. The required quantity of test item was weighed into a clean beaker and there by adding little volume of the vehicle into the beaker,mixed well using glass rod and transferred into measuring cylinder. This rinsing procedure was repeated until complete transfer of test item formulation into the measuring cylinder. Finally, the volume was made up to the required quantity with vehicle to get a desired concentration of 25, 50 and 100 mg/mL of test item for low, mid and high dose groups respectively.

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

VEHICLE
- Justification for use and choice of vehicle (if other than water): The test item is not miscible with distilled water at a concentration of 100 mg/mL (high dose concentration selected for the study) and forms uniform suspension with corn oil. Hence, corn oil was selected as a vehicle for test item formulation.
- Concentration in vehicle: 0, 250, 500 and 1000 mg/kg bw
- Amount of vehicle (if gavage): 10 mL/kg
- Lot/batch no. (if required): Batch No: A1708001
- Purity:
Details on mating procedure:
- M/F ratio per cage:1:1 ratio
- Length of cohabitation: The female was placed with the same male until pregnancy occurs by evidence of sperm in vaginal smear until two weeks have elapsed.
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: Day ‘0’ pregnancy was confirmed by the presence of sperm in the vaginal smear.
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility: Not specified
- Further matings after two unsuccessful attempts: [no / yes (explain)]Not specified
- After successful mating each pregnant female was caged (how): females were housed individually. Sterilized paper shreds were provided as a nesting material from gestation day 20 onwards.
- Any other deviations from standard protocol:
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Stability and Homogeneity were analysis.
Duration of treatment / exposure:
Male: 37 days
Female: Approx 62 days
Frequency of treatment:
Daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
Total: 96
0 mg/kg bw: 12 male, 12 female
250 mg/kg bw: 12 male, 12 female
500 mg/kg bw: 12 male, 12 female
1000 mg/kg bw: 12 male, 12 female
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The doses of 0, 250, 500 and 1000 mg/kg body weight for vehicle control, low dose, mid dose and high dose respectively were selected based on the NOAEL of test item is >1000 mg/kg body weight in a 28 days repeated dose study (oral) in rats
- Rationale for animal assignment (if not random): The animals were weighed and arranged in ascending order of their body weights. These body weight stratified animals were distributed to all the groups using Microsoft Excel Spreadsheet, such that body weight variation of animals selected for the study did not exceed ± 20% (+2.99% and -11.62% for males and +4.34% and -7.91% for females) of the mean body weight of each sex. The grouping was done one day prior to the initiation of treatment. Body weight of the animals was analyzed statistically for mean body weight to rule out the statistical significant difference between groups within each sex.
- Other:

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily
- Cage side observations checked in table [No.?] were included. : mortality and morbidity were examined.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: The animals were weighed at receipt, on the first day of dosing, weekly thereafter (varied by -1 day) and at termination. The females were weighed on gestation days 0, 7, 14 and 20 during pregnancy and on days 1, 4, 7 and 13 during lactation period.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

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

OTHER:
Oestrous cyclicity (parental animals):
Oestrus cycles were monitored for two weeks after five days of acclimatization to evaluate its normal oestrus cyclicity (4 to 5 days). Only females with normal oestrus cyclicity were selected for the treatment. Vaginal smears were monitored daily from the beginning of the treatment period until evidence of mating. When obtaining vaginal/cervical cells, care was taken to avoid disturbance of mucosa. Oestrus cyclicity was also monitored on the day of sacrifice for females.
Litter observations:
The number of pups born (dead and live) in a litter, sex, live births and external observations were recorded at birth. Individual body weight of live pups on lactation day 1 (within 24 hours of parturition), 4, 7 and 13 were recorded. The anogenital distance of each pup was measured on postnatal day 4 (lactation day 4) and the ratio of AGD to the cube root of pup body weight was calculated. All survived male pups were examined for appearance of nipples/areolae on postnatal day 13 (lactation day 13). The litter was observed daily in order to note the number of alive, dead and cannibalized pups.
Postmortem examinations (parental animals):
Hormone analysis, organ weight and external and internal gross pathological and histopathological examination.
Postmortem examinations (offspring):
organ weight and The pups were sacrificed on lactation day 13 and the sacrificed pups and dead pups were examined for gross abnormalities
Statistics:
The raw data was subjected to computer statistical processing. The computer printout of the data (in the form of appendix) was verified with the raw data. After verification, the data was subjected to various statistical analyses using SPSS software version 22.
All analysis and comparisons were evaluated at the 95% level of confidence (P<0.05), indicated by the aforementioned tests designated by the superscripts throughout the report as stated below:
* Statistically significant (P<0.05) change than the vehicle control group.
Reproductive indices:
Male Mating Index, Male Fertility Index, Female Mating Index, Female Fertility Index, Gestation Index, Live Birth Index (%) per dam and Pup Survival index (%) on lactation day 4 were examined.
Offspring viability indices:
yes, on daye 0 and 13

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
There were no clinical signs of toxicity observed at any of the tested dose group animals of either sex during the experimental period.
Dermal irritation (if dermal study):
not specified
Mortality:
no mortality observed
Description (incidence):
no mortality/morbidity observed at any of the tested dose group animals of either sex during the experimental period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
There were no changes observed in mean body weight and percent change in body weight with respect to day 1 at all the tested group animals of either sex during gestation, lactation and the experimental period.

Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There were no changes observed in feed consumption at any of the tested dose group animals of either sex during during gestation, lactation and the experimental period.
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:
no effects observed
Description (incidence and severity):
There were no treatment related changes observed in serum T4 levels in adult males at any of the tested dose groups when compared with vehicle control group.
However, statistically significant increase in T4 levels at the high dose group males when compared with vehicle control group males was noted.
This significant variation could be due to two males in 1000 mg/kg bw with a serum T4 level of 142.851 and 140.210 respectively which are significantly more than other animals of same group. Hence, the variations observed in the T4 levels are considered to be incidental.
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no effects observed in absolute and relative organ weights at any of the tested dose group animals of either sex when compared with vehicle control group
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no gross pathological changes (both external and internal) observed at all the tested dose group adult animals and pups of either sex at all the tested dose groups examined at termination.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No treatment related histopathological findings noticed in the present study.
Lesions considered spontaneous and incidental were observed in treated and control rats. These lesions consisted of unilateral interstitial mononuclear cell (MNC) infiltration in epididymides.
A detailed qualitative examination of the testes was made, taking into account the tubular stages of the spermatogenic cycle. The examination was conducted in order to identify treatment related effects such as missing germ cell layers or types, retained spermatids, multinucleate or apoptotic germ cells and sloughing of spermatogenic cells into the lumen or any cell or stage specificity of testicular findings.
Testes and ovaries did not show any pathological findings/lesions.
Histopathological findings: neoplastic:
not specified
Other effects:
not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
There were no changes observed in the oestrus cyclicity at any of the tested dose group females during pre-mating treatment, mating treatment and on lactation day 14.
Reproductive function: sperm measures:
not specified
Reproductive performance:
no effects observed
Description (incidence and severity):
There were no changes observed in number of corpora lutea, number of implantations and no changes were noted in pre and post-implantation loss, pre-natal loss, post-natal loss at any of the tested dose group animals when compared with vehicle control group animals. No resorptions were noted at all the dose group animals observed during necropsy.

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:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
clinical biochemistry
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic
reproductive function (oestrous cycle)
reproductive performance
Remarks on result:
other: No effect observed

Target system / organ toxicity (P0)

Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
There were no clinical signs observed in any of the pups of tested dose group animals during lactation period.
Dermal irritation (if dermal study):
not specified
Mortality / viability:
no mortality observed
Description (incidence and severity):
There were no treatment related changes observed in the number of pups and pup survival index of each litter at any of the tested dose group animals during lactation period when compared with vehicle control group animals.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
There were no changes observed in mean pup (male and female) weight on lactation day 1, 4, 7 and 13 at any of the tested dose groups when compared with vehicle control group dams.
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:
no effects observed
Description (incidence and severity):
There were no treatment related changes observed in serum T4 levels in lactation day 13 pups at any of the tested dose groups when compared with vehicle control group.
Urinalysis findings:
not specified
Sexual maturation:
no effects observed
Description (incidence and severity):
There were no changes observed in ano-genital distance ratio on lactation day 4 and no occurrences of nipples in male pups of dams at any of the tested dose group litters and vehicle control group litters observed on lactation day 13.
Anogenital distance (AGD):
not specified
Nipple retention in male pups:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no gross pathological changes (both external and internal) observed at all the tested dose group pups of either sex at all the tested dose groups examined at termination.
Histopathological findings:
not specified
Other effects:
not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):
not specified

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not specified

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
sexual maturation
clinical signs
mortality
body weight and weight gain
clinical biochemistry
gross pathology
Remarks on result:
other: No effect observed

Target system / organ toxicity (F1)

Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified

Overall reproductive toxicity

Reproductive effects observed:
not specified
Treatment related:
not specified

Any other information on results incl. tables

SUMMARY OF THE STUDY


Parameters ↓

Group & Dose (mg/kg body weight/day)

G1 & 0

G2 & 250

G3 & 500

G4 & 1000

Reproductive Indices

Mating Indices 

Pairs started (No.)

12

12

12

12

Males showing evidence of mating (No.) within 14 days*

12

12

10

11

Females showing evidence of copulation (No.)

12

12

12

12

Male Mating Index (%) within 14 days*

100.00

100.00

83.33

91.67

Female Mating Index (%)

100.00

100.00

100.00

100.00

 

Fertility Indices

Females achieving pregnancy (No.)

12

12

12

12

Male Fertility Index (%)*

100.00

100.00

83.33

91.67

Female Fertility Index (%)

100.00

100.00

100.00

100.00

 

Copulatory Indices

Conceiving days 1 to 5 (No.)

7

6

6

5

Conceiving days >6 (No.)

5

6

6

7

Mean Precoital Interval (Days)

5.83

6.75

7.83

7.92

 

Gestation Indices

Pregnancy≤21 days (No.)

1

1

0

0

Pregnancy = 22 days (No.)

4

6

6

4

Pregnancy≥23 days (No.)

7

5

6

8

Mean Gestation length (Days)

22.50

22.33

22.67

22.75

Gestation Index (%)

100.00

100.00

100.00

100.00

 

Dams with live young born (No.)

12

12

12

12

Dams with live young at day 4 post-partum (No.)

12

12

12

12

Dams with live young at day 13 post-partum (No.)

12

12

12

12

 

Implantation Index and Pre and Post implantation Losses 

Implants/dam (Mean)

10.58

11.75

11.17

11.00

Corpora luetea/dam (Mean)

10.67

11.75

11.25

11.00

Implantation Index (%)

99.17

100.00

99.07

100.00

Pre-Implantation Loss (%)

0.83

0.00

0.93

0.00

Post-Implantation Loss (%)

1.52

0.00

1.39

0.00

  

Offspring Viability Indices

Live Birth Indices and Sex Ratio at Birth 

Live pups/dam at birth (mean)

10.42

11.75

11.00

11.00

Litter Size (Total No. of pups born/dam) at birth (mean)

10.58

11.75

11.00

11.00

Mean Live Birth Index/dam (%)

98.48

100.00

100.00

100.00

Male Live pups/dam at birth (mean)

4.92

5.92

4.83

4.92

Female Live pups/dam at birth (mean)

5.50

5.83

6.17

6.08

Sex Ratio (male/female)

1.23

1.22

0.90

1.05

*: The females were placed with proven male of same test group after 14 days of cohabitation period.

Parameters ↓

Group & Dose (mg/kg body weight/day)

G1 & 0

G2 & 100

G3 & 300

G4 & 1000

Pup Survival Indices and Sex Ratio during lactation  

Mean No. of Pups survived per dam ( LD1 to 4)

10.42

11.75

11.00

11.00

Mean No. of Pups dead per dam ( LD1 to 4)

0.00

0.00

0.00

0.00

Mean Pup Survival Index (%) per dam (LD1 to 4) 

100.00

100.00

100.00

100.00

Sex Ratio (male/female) per dam at LD 4

1.23

1.22

0.90

1.05

Mean No. of Pups Sacrificed for Blood Collection on LD4

0.58

1.25

1.08

0.92

Mean No. of Pups survived per dam (LD4 to 7)

9.83

10.50

9.92

10.08

Mean No. of Pups dead per dam (LD4 to 7)

0.00

0.00

0.00

0.00

Mean Pup Survival Index (%) per dam (LD4 to 7)

100.00

100.00

100.00

100.00

Sex Ratio (male/female) per dam at LD 7

1.22

1.55

1.14

1.21

Mean No. of Pups survived per dam (LD7 to 13)

9.83

10.50

9.92

10.08

Mean No. of Pups dead per dam (LD7 to 13)

0.00

0.00

0.00

0.00

Mean Pup Survival Index (%) per dam ( LD7 to 13)

100.00

100.00

100.00

100.00

Sex Ratio (male/female) per dam at LD 13

1.22

1.55

1.14

1.21

 

Pre and Postnatal loss 

Mean Pre-natal (implantations minus live births) (No.)

0.17

0.00

0.17

0.00

Females with 0 (No.)

11

12

11

12

Females with 1 (No.)

1

0

1

0

Post-natal (live births minus alive at post natal day 13)

Females with 0 (No.)

12

12

12

12

Females with ≥ 1 (No.)

0

0

0

0

 

Litter Observations 

Male Pup weight at birth (mean) in gram

7.10

7.00

7.40

7.20

Female Pup weight at birth (mean) in gram

6.68

6.38

6.83

6.39

Male Pup weight on LD4 (mean) in gram

12.07

11.87

11.90

12.28

Female Pup weight on LD4 (mean) in gram

11.20

10.89

10.60

10.61

Male Pup weight on LD7 (mean) in gram

17.10

17.02

17.52

17.43

Female Pup weight on LD7 (mean) in gram

15.88

15.59

15.58

15.22

Male Pup weight on LD13 (mean) in gram

28.06

27.91

28.67

28.17

Female Pup weight on LD13 (mean) in gram

25.98

25.95

25.89

25.60

 SUMMARYOF CLINICAL SIGNSOF TOXICITY, DETAILED CLINICAL EXAMINATIONAND MORTALITY RECORD         

Group, Sex & Dose

(mg/kg body weight/day)

No. of Animals

Clinical Signs of Toxicity/

Detailed Clinical Examination

Mortality

(No. of Mortality /

No. of Animals dosed)

G1, M & 0

12

N

0/12

G2, M & 250

12

N

0/12

G3, M & 500

12

N

0/12

G4, M & 1000

12

N

0/12

M: Male; N: Normal

Group, Sex & Dose

(mg/kg body weight/day)

No. of Animals

Clinical Signs of Toxicity/

Detailed Clinical Examination)

Mortality

(No. of Mortality /

No. of Animals dosed)

G1, F & 0

12

N

0/12

G2, F & 250

12

N

0/12

G3, F & 500

12

N

0/12

G4, F & 1000

12

N

0/12

  SUMMARY OF ABSOLUTE ORGAN WEIGHT (g) RECORD

Group, Sex & Dose
(mg/kg body weight/day)

Epididymes

Testes

Prostate+Seminal vesicles with coagulating glands (PSC)

Thyroid along with parathyroid#

G1, M & 0

Mean

1.5304

3.4862

3.0974

0.2577

±SD

0.1461

0.2643

0.3581

0.0253

n

12

12

12

12

G2, M & 250

Mean

1.5855

3.3497

3.2505

0.2581

±SD

0.1608

0.2477

0.3081

0.0202

n

12

12

12

12

G3, M & 500

Mean

1.6304

3.5343

3.1775

0.2575

±SD

0.1677

0.2791

0.2398

0.0169

n

12

12

12

12

G4, M & 1000

Mean

1.5877

3.2692

3.1080

0.2476

±SD

0.1734

0.1988

0.3843

0.0207

n

12

12

12

12

M: Male;SD: Standard Deviation; n: Number of animals; #: Weighed post fixation

Group, Sex & Dose

(mg/kg body weight/day)

 

Thyroid along with parathyroid#

G1, F & 0

Mean

0.2341

±SD

0.0105

n

11

G2, F & 100

Mean

0.2414

±SD

0.0113

n

10

G3, F & 300

Mean

0.2457

±SD

0.0126

n

11

G4, F & 1000

Mean

0.2354

±SD

0.0162

n

11

SUMMARY OF FASTING BODY WEIGHT (g) AND ORGAN WEIGHT (%) RELATIVE TO FASTING BODY WEIGHT RECORD

Group, Sex & Dose

(mg/kg body weight/day)

Fasting Body Weight (g)

Epididymes

Testes

Prostate+Seminal vesicles with coagulating glands (PSC)

Thyroid along with parathyroid

G1, M & 0

Mean

379.21

0.4050

0.9226

0.8197

0.0678

±SD

24.03

0.0477

0.0933

0.1089

0.0033

n

12

12

12

12

12

G2, M & 250

Mean

389.42

0.4070

0.8614

0.8382

0.0663

±SD

23.79

0.0305

0.0584

0.1005

0.0031

n

12

12

12

12

12

G3, M & 500

Mean

386.93

0.4216

0.9129

0.8228

0.0665

±SD

16.51

0.0410

0.0506

0.0725

0.0017

n

12

12

12

12

12

G4, M & 1000

Mean

376.98

0.4211

0.8679

0.8237

0.0656

±SD

21.53

0.0387

0.0384

0.0824

0.0020

n

12

12

12

12

12

M: Male;SD: Standard Deviation; n: Number of animals

Group, Sex & Dose

(mg/kg body weight/day)

 

Fasting Body Weight (g)

Thyroid along with parathyroid

G1, F & 0

Mean

264.08

0.0886

±SD

6.47

0.0024

n

11

11

G2, F & 250

Mean

272.77

0.0885

±SD

13.04

0.0020

n

10

10

G3, F & 500

Mean

274.96

0.0894

±SD

14.12

0.0014

n

11

11

G4, F & 1000

Mean

264.70

0.0889

±SD

16.47

0.0015

n

11

11

SUMMARY RECORD OF VAGINAL SMEAR EXAMINATION FOR DETERMINATION OF OESTRUS CYCLICITY

Group & Dose
(mg/kg body weight/day)

No. of Females

No. of Females with Regular Oestrus Cyclicity during

Pre-mating, Mating and on Lactation day 14

No. of Females with Irregular Oestrus Cyclicity during

Pre-mating, Mating and on Lactation day 14

G1 & 0

12

12

0

G2 & 250

12

12

0

G3 & 500

12

12

0

G4 & 1000

12

12

0

SUMMARY RECORD OF GESTATION LENGTH (DAYS) AND DELIVERY DATA

Group & Dose
(mg/kg body weight/day)

 

Gestation Length (Days)

 

Delivery Data

Litter Size (No.)

Live Pups (No.)

Dead Pups (No.)

 

Sex Ratio (M/F)

at Birth

Live Birth Index
(%)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

G1 & 0

Mean

23.00

11.45

11.09

5.09

6.00

0.36

0.36

0.00

1.04

97.35

±SD

0.89

2.02

1.64

1.70

1.90

0.67

0.67

0.00

0.76

5.05

n

11

11

11

11

11

11

11

11

11

11

G2 & 250

Mean

23.30

10.70

10.40

4.00

6.40

0.30

0.30

0.00

0.70

96.67

±SD

0.67

1.64

2.01

1.56

1.78

0.67

0.67

0.00

0.39

8.05

n

10

10

10

10

10

10

10

10

10

10

G3 & 500

Mean

23.00

12.09

12.00

6.09

5.91

0.09

0.00

0.09

1.23

99.24

±SD

0.45

1.14

1.18

2.30

1.70

0.30

0.00

0.30

0.87

2.51

n

11

11

11

11

11

11

11

11

11

11

G4 & 1000

Mean

23.64

11.18

10.82

4.45

6.36

0.36

0.09

0.27

0.76

96.74

±SD

0.50

2.18

2.27

1.51

1.86

0.67

0.30

0.65

0.34

6.11

n

11

11

11

11

11

11

11

11

11

11

  M: Male; F: Female;SD: Standard Deviation; n: Number of dams

SUMMARY OF LITTER OBSERVATION RECORD DURING LACTATION PERIOD

Group & Dose
(mg/kg body weight/day)

 

No. of Live Pups At Birth

During LD 1 to 4

Sex Ratio (M/F) at

LD 4

No. of Survived Pups during LD 1 to 4

Pup Survival Index
(%)

LD 1 to 4

Live Pups (No.)

 

 

 

 

 

Dead Pups (No.)

Total (No.)

Male

(No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

G1 & 0                  

Mean

11.09

11.09

5.09

6.00

0.00

0.00

0.00

1.04

11.09

100.00

±SD

1.64

1.64

1.70

1.90

0.00

0.00

0.00

0.76

1.64

0.00

n

11

11

11

11

11

11

11

11

11

11

G2 & 250                 

Mean

10.40

10.40

4.00

6.40

0.00

0.00

0.00

0.70

10.40

100.00

±SD

2.01

2.01

1.56

1.78

0.00

0.00

0.00

0.39

2.01

0.00

n

10

10

10

10

10

10

10

10

10

10

G3 & 500                  

Mean

12.00

12.00

6.09

5.91

0.00

0.00

0.00

1.23

12.00

100.00

±SD

1.18

1.18

2.30

1.70

0.00

0.00

0.00

0.87

1.18

0.00

n

11

11

11

11

11

11

11

11

11

11

G4 & 1000                  

Mean

10.82

10.82

4.45

6.36

0.00

0.00

0.00

0.76

10.82

100.00

±SD

2.27

2.27

1.51

1.86

0.00

0.00

0.00

0.34

2.27

0.00

n

11

11

11

11

11

11

11

11

11

11

M: Male: F: Female;SD: Standard Deviation; n: Number of dams; LD: Lactation Day

Group & Dose
(mg/kg body weight/day)

Animal No.

Live Pups (No.) on LD 4

Pups Sacrificed for Blood Collection on

LD 4 (No.)

Live Pups (No.) on LD 4 after Sacrificed for Blood Collection

During LD 4 to 7

Sex Ratio (M/F) at
LD 7

No. of Survived Pups during LD
4 to 7

Pup Survival Index
(%) during
          LD 4 to 7

Live Pups (No.)

 

Dead Pups (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

G1 & 0

Mean

5.09

6.00

11.09

0.00

1.00

1.00

5.09

5.00

10.09

5.09

5.00

10.09

0.00

0.00

0.00

1.30

10.09

100.00

±SD

1.70

1.90

1.64

0.00

0.89

0.89

1.70

1.67

0.94

1.70

1.67

0.94

0.00

0.00

0.00

1.05

0.94

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

G2 & 250

Mean

4.00

6.40

10.40

0.00

0.90

0.90

4.00

5.50

9.50

4.00

5.50

9.50

0.00

0.00

0.00

0.82

9.50

100.00

±SD

1.56

1.78

2.01

0.00

0.88

0.88

1.56

1.51

1.35

1.56

1.51

1.35

0.00

0.00

0.00

0.46

1.35

0.00

n

10

10

10

10

10

10

10

10

10

10

10

10

10

10

10

10

10

10

G3 & 500

Mean

6.09

5.91

12.00

0.00

1.64

1.64

6.09

4.27

10.36

6.09

4.27

10.36

0.00

0.00

0.00

2.27

10.36

100.00

±SD

2.30

1.70

1.18

0.00

0.50

0.50

2.30

1.74

0.92

2.30

1.74

0.92

0.00

0.00

0.00

2.62

0.92

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

G4 & 1000

Mean

4.45

6.36

10.82

0.00

0.73

0.73

4.45

5.64

10.09

4.45

5.64

10.09

0.00

0.00

0.00

0.87

10.09

100.00

±SD

1.51

1.86

2.27

0.00

1.01

1.01

1.51

1.36

1.38

1.51

1.36

1.38

0.00

0.00

0.00

0.44

1.38

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

11

Group & Dose
(mg/kg body weight/day)

Animal No.

Live Pups (No.) on LD 7

During LD 7 to 13

 

Sex Ratio (M/F) at

LD 13

No. of Survived Pups during

LD 7 to 13

Pup Survival Index
(%) during LD 7
to 13

Live Pups (No.)

 

Dead Pups (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

Male (No.)

Female (No.)

Total (No.)

G1 & 0

Mean

5.09

5.00

10.09

5.09

5.00

10.09

0.00

0.00

0.00

1.30

10.09

100.00

±SD

1.70

1.67

0.94

1.70

1.67

0.94

0.00

0.00

0.00

1.05

0.94

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

G2 & 250

Mean

4.00

5.50

9.50

4.00

5.50

9.50

0.00

0.00

0.00

0.82

9.50

100.00

±SD

1.56

1.51

1.35

1.56

1.51

1.35

0.00

0.00

0.00

0.46

1.35

0.00

n

10

10

10

10

10

10

10

10

10

10

10

10

G3 & 500

Mean

6.09

4.27

10.36

6.09

4.27

10.36

0.00

0.00

0.00

2.27

10.36

100.00

±SD

2.30

1.74

0.92

2.30

1.74

0.92

0.00

0.00

0.00

2.62

0.92

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

G4 & 1000

Mean

4.45

5.64

10.09

4.45

5.64

10.09

0.00

0.00

0.00

0.87

10.09

100.00

±SD

1.51

1.36

1.38

1.51

1.36

1.38

0.00

0.00

0.00

0.44

1.38

0.00

n

11

11

11

11

11

11

11

11

11

11

11

11

    SUMMARY OF UTERI OBSERVATIONS RECORD

Group & Dose
(mg/kg body weight/day)

No. of Corpora lutea

No. of Implantations

Implantation

Index

Pre-Implantation Loss (%)

Post-Implantation Loss (%)

Pre-natal Loss (No.)

Post-natal Loss (%)

Post-natal Loss (No.)

No. of Early Resorptions

No. of Late Resorptions

G1 & 0                  

Mean

11.73

11.45

98.07

1.93

2.65

0.36

0.00

0.00

0.00

0.00

±SD

2.33

2.02

3.35

3.35

5.05

0.67

0.00

0.00

0.00

0.00

n

11

11

11

11

11

11

11

11

11

11

G2 & 250  

Mean

10.70

10.70

100.00

0.00

3.33

0.30

0.00

0.00

0.00

0.00

±SD

1.64

1.64

0.00

0.00

8.05

0.67

0.00

0.00

0.00

0.00

n

10

10

10

10

10

10

10

10

10

10

G3 & 500           

Mean

12.27

12.09

98.59

1.41

0.76

0.09

0.00

0.00

0.00

0.00

±SD

1.19

1.14

3.14

3.14

2.51

0.30

0.00

0.00

0.00

0.00

n

11

11

11

11

11

11

11

11

11

11

G4 & 1000     

Mean

11.36

11.18

98.78

1.22

3.26

0.36

0.00

0.00

0.00

0.00

±SD

2.46

2.18

2.72

2.72

6.11

0.67

0.00

0.00

0.00

0.00

n

11

11

11

11

11

11

11

11

11

11

SD: Standard Deviation; n: Number of animals

 SUMMARY OF SERUM T4 LEVELS (ng/mL) RECORD - MALES

Group, Sex & Dose

(mg/kg body weight/day)

Serum T4 Levels

(ng/mL)

G1, M & 0

Mean

87.991

±SD

4.682

n

12

G2, M & 250

Mean

87.300

±SD

4.919

n

12

G3, M & 500

Mean

95.705

±SD

14.351

n

12

G4, M & 1000

Mean

104.278*

±SD

21.130

n

12

M: Male; SD: Standard Deviation; n: Number of animals

* Statistically significant (P<0.05) change than the vehicle control group.

SUMMARY OF SERUM T4 LEVELS (ng/mL) RECORD - LACTATION DAY 13 PUPS

Group & Dose

(mg/kg body weight/day)

Serum T4 Levels

(ng/mL)

G1 & 0

Mean

70.752

±SD

9.415

n

11

G2 & 250

Mean

62.065

±SD

8.076

n

10

G3 & 500

Mean

64.219

±SD

7.606

n

11

G4 & 1000

Mean

62.132

±SD

9.600

n

11

SD: Standard Deviation; n: Number of Dams (Serum collected from pups per dam pooled for analysis)

Applicant's summary and conclusion

Conclusions:
The No Observed Adverse Effect Level (NOAEL) of the test chemical was considered to be 1000 mg/kg body weight when administered to the males for two weeks pre-mating, during mating and up to the day before sacrifice during post-mating period (total of 37 days), to the females for two weeks pre-mating, during mating, pregnancy (gestation) and up to lactation day 13, under the experimental conditions
Executive summary:

In a Reproduction/Developmental Toxicity Screening Test, Sprague Dawley rats were treated with test chemical at dose levels of 250, 500 and 1000 mg/kg body weight orally by gavage. Males were treated for two weeks pre-mating, during mating and up to the day before sacrifice during post-mating period (total of 37 days of treatment). The females were treated for two weeks pre-mating period, during mating, pregnancy (gestation) and up to lactation day 13 after which the pups were sacrificed on lactation day 13 and females (dams) were sacrificed on lactation day 14 after overnight fasting (water allowed). All the tested dose group animals of either sex did not reveal any clinical signs of toxicity and no mortality/morbidity observed. There were no changes observed in mean body weight, percent change in body weight with respect to day 1 and feed consumption at all the tested dose group animals of either sex during the experimental period. No changes were observed in organ weights (both absolute and relative) at all the tested dose group animals. There were no treatment related changes observed in serum T4 levels of adult males and in serum T4 levels of lactation day 13 pups at any of the tested dose groups. Dams did not reveal any treatment related changes in oestrus cyclicity, copulatory interval, body weights and feed consumption during gestation and lactation periods, gestation length, live birth index, number of pups, sex ratio and pup survival index at all the tested dose groups throughout the lactation period. Similarly, all pups did not reveal any clinical signs or external anomalies throughout the lactation period. No treatment related changes in pup weights, ano-genital distance ratio were noted. No occurrences of nipples in male pups at any of the tested dose groups and vehicle control group. In addition, there were no gross pathological changes (both external and internal) observed at all the tested dose group adult animals and pups. There were no treatment related histopathological findings noticed during the microscopic examination. Testes were screened with special emphasis on stages of spermatogenesis and interstitial testicular cell structure, revealed normal progression of the spermatogenic cycle and presence of all germ layers (cells). In addition this, ovaries did not show any pathological findings/lesions. Therefore, The No Observed Adverse Effect Level (NOAEL) of the test chemical was considered to be 1000 mg/kg body weight when administered to the males for two weeks pre-mating, during mating and up to the day before sacrifice during post-mating period (total of 37 days), to the females for two weeks pre-mating, during mating, pregnancy (gestation) and up to lactation day 13, under the experimental conditions.