Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
December 22, 2015 to March 20, 2017
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: crystalline
Specific details on test material used for the study:
dentification Acido oxalico - STD
Appearance Fine White Crystal
Chemical Name Oxalic Acid Dihydrated
Batch Number 15000900
CAS No. 6153-56-6
Manufactured by OXAQUIM S.A.
FÁBRICA Y DEPARTAMENTO TÉCNICO
Polígono Industrial "Las Horcas" s/n
44600 Alcañiz (Teruel)
España
Teléfono: +34 978 83 31 13
Fax: +34 978 83 38 61
Supplied by OXAQUIM S.A.
C/ Gregal, 3 Urbanización "Parc Llevant"
43764 EL CATLLAR (Tarragona)
España
Teléfono: +34 977 65 38 98
Fax: +34 977 65 39 30
Purity (%) 99.6%
Molecular Weight 126.07 g/mol
Molecular Formula C2O4H2.2H2O
Manufacture Date May 21, 2015
Expiry Date May 20, 2017
Stability of the test item dilution Seven Days
Stability of the test item 1 year
Storage Conditions Room Temperature (20 to 30 oC)

Test animals

Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Sainath Agencies,
- Age at study initiation: 5- 7 Months
- Weight at study initiation: Minimum 2710.3 and Maximum 3792.6 gm
- Housing: Individually in EN130S Noryl (Tecniplast) cages (approximately 653 mm x 653 mm). After randomization, male and female were individually. During the mating phase, animals were housed on one male: one female basis within each dose group. After successful mating, the females were housed individually during gestation.
- Diet (e.g. ad libitum): Teklad Certified Global High Fiber Rabbit Diet (Lot No. 2031C-062816MA) from ENVIGO was provided ad libitum
- Water (e.g. ad libitum):Aquaguard filtered tap water was provided ad libitum
- Acclimation period:10 -19 days under laboratory conditions, after veterinary examination.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.9 to 20.5 0C
- Humidity (%): 55.5 to 65.1 %,
- Air changes (per hr): air-conditioned with adequate (above 10) air changes per hour.
- Photoperiod (hrs dark / hrs light):light cycle of 12 hours light and 12 hours dark.

IN-LIFE DATES: From: November 29, 2016 To:January 22, 2017

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): Twice a Weekly
- Diet Preparation: Dietary admixtures were prepared using the powdered feed.
Test item was triturated and weighed into a tarred glass beaker using a suitable weighing balance, mixed with powdered feed to prepare the pre-mix at higher concentration. Pre-mix was used to prepare the low, intermediate and high dose concentrations. Required amount of pre-mix and powdered feed was mixed to achieve different dose concentrations in feed. Feed for the control animals were prepared similarly without addition of test item. The feed was prepared and homogenized with the help of Homogenizer before providing to the animals.
- Storage temperature of food: Room Temperature
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical method was validated and the linearity response was found to be linear (r2 = 1.000) in the range of 10.12 to 500.16 mg. The applied concentration was found homogeneously distributed in the feed at concentrations 1,000 mg/L, 5,000 mg/L and 10,000 mg/L for the test item respectively. (i.e. % coefficient of variation (RSD) ranged in between 1.18 to 2.62). The dose concentration obtained in all the dose groups was in agreement with the target concentration (i.e. ranged between 92.072 % to 97.949 %)
Details on mating procedure:
- Impregnation procedure: [cohoused]
- If cohoused:
- M/F ratio per cage: 1: 1
- Length of cohabitation: Animals were paired on a one male: two female, within each dose group, for a period of fourteen days
Duration of treatment / exposure:
The dosing was commenced on day 6 of gestation (post mating) and continued till day 29 of gestation.
Frequency of treatment:
Daily
Duration of test:
30 days
Doses / concentrationsopen allclose all
Dose / conc.:
1 000 ppm
Remarks:
Low dose
Dose / conc.:
5 000 ppm
Remarks:
Intermediate dose
Dose / conc.:
10 000 ppm
Remarks:
High dose: For Pre-natal developmental toxicity study in Rabbit, the doses for main study (10000PPM) was selected based up on dose range finding study. In Dose Range finding study, up-to 10000 PPM there were no test item related changes in reproductive indices observed.
No. of animals per sex per dose:
24 females.
Control animals:
yes
yes, plain diet
Details on study design:
TEST SYSTEM

Test System Young Adult New Zealand White Rabbits
Scientific Name Oryctolagus cuniculus
Justification Recommended by the guideline
Source Sainath Agencies,
1-6-197/45/D, Bapujinagar, Musheerabad,
Hyderabad-48, India
Registration No. 282/PO/Bt/S/2000/CPCSEA
Total number of Groups 4
Age at start of mating 5-7 months
Total number of animals 60 Male* and 120 Female
Total number of animals used 10 Male and 20 Female (Dose Range Finding Study)
48* Male and 96 Female (Main Study)
(Females were nulliparous and non-pregnant)
*Note: Male animals were sent for sacrifice after confirmation of last mating.

Total number of animals per group 5 Mated female (Dose Range Finding Study)
24 Mated female (Main Study)
Body weight when treated Minimum 2710.3 and Maximum 3792.6 gm
Identification By unique cage number and individual animal numbers marked with an indelible marker p en on the ear. The animals were marked with permanent animal numbers before the start of test item administration and refreshed weekly thereafter. The animals were marked with the temporary animal numbers at start acclimatization. Fetuses were identified with tag.
Randomization Female were assigned to groups on confirmation of amting i.e. on ‘0’ day of gestation in se quence beginning with the group following the last group assigned on the previous day.
Acclimatization 10-19 days under laboratory conditions, after veterinary examination. Only animals without any visible signs of illness were used for the study.
RATIONALE FOR DOSE LEVEL SELECTION

A GLP Dose Range finding (DRF) study was carried out in New Zealand White Rabbit before commencement of the main study to confirm the high dose level for test item. A total of three groups (Low, Intermediate and High) consisting of 5 mated female were treated at dose levels of 1000, 5000 and 10000 PPM. Control group animals were treated with powdered feed alone. Dose levels for Main Study were selected based on Dose Range Finding Study observation and results.
Dose levels for main study selected for Low, Intermediate and High at dose levels were 1000, 5000 and 10000 PPM, respectively. Control group animals were treated with powdered feed alone.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes / No / No data
- Time schedule:
No clinical signs were observed in any of the female animals treated at control (0 PPM) Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups

DETAILED CLINICAL OBSERVATIONS: No
- Time schedule: NA

BODY WEIGHT: Yes
No significant difference in the body weight and body weight gain (%) was observed in pregnant female animals treated at control (0 PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
Feed consumption of pregnant female animals at (0PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups was not significantly different throughout the treatment period


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 30
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes / No / No data
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes: all per litter

- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes:all per litter
- Head examinations: Yes: half per litter
Statistics:
• Data are summarized in tabular form. Statistical analysis was performed using statplus program. Values are given as mean ± standard deviation (SD).
• All the data were checked for Normality with Shapiro-Wilk W test.
• All the data were checked for Homogeneity with Bartlett Chi-Square test.
• Data were subjected to perform Analysis of Variance (ANOVA) for continuous data.
• Discontinuous data were subjected to nonparametric test (Mann-Whitney U-Test).
P ≤ 0.05 (5% level of significance) was considered to represent significance in the respective parameters, P > 0.05 was considered not Significant
Indices:
Pre–implantation loss (%)
(Number of Corpora Lutea - number of implantation sites) / Number of Corpora Lutea x 100
Post–implantation loss (%)
(Number of implantation sites - Total number of live foetuses) / Number of implantation sites x 100
Sex Ratio (% males)
Number of male foetuses (Day 0) / Total number of foetuses (Day 0) x 100
Variation Incidence (%)
Number of foetuses with variation/ Total Number of foetuses examined x 100

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
No significant difference in the body weight and body weight gain (%) was observed in pregnant female animals treated at control (0 PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups in the DRF and main study. Howerever in main study marginal decrease in body weight gain percentage was observed on day 3 to day 30 gestation day in Low (1000 PPM), Intermediate (5000 PPM) and High dose (10000 PPM) groups compared to cotrol group.
The body weight changes observed are marginal and could not be attributed to the test item administartion and does not represent a change of any biological significance
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Feed consumption of pregnant female animals at (0PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups was not significantly different throughout the treatment period in dose range finding study except significant decrease in feed consumption was observed in intermediate (5000 PPM) and high dose (10000 PPM) group on day 27 og gestation period when compared with control (0 PPM).
In main study in females during gestation day 0 marginal increase in feed consumption was observed in low (1000 PPM) and intermediate dose (5000 PPM) compareed to control (0 PPM) animals. On day 6 marginal decrease in feed consumption was noticed in Low (1000 PPM) dose compared to control, whereas on day 9,12, 15 and day 24 of gestation significant decrease in feed was noticed low and intermediate dose. On day 18 and 21 decrease in feed consumption was observed in low dose group whereas on day 27 significant decrease in feed consumption was recorded in intermediate dose while compared to control group of animals. These changes were within the normal range with respect to age and sex of the animals. Hence, the changes in feed consumption were considered as biological variation
Food efficiency:
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
No significant difference was observed in gravid uterine weight
Gross pathological findings:
no effects observed
Description (incidence and severity):
No abnormality was observed in any of the female animals from control, low, intermediate and high dose groups in both dose range finding and main study.
No test item related a change in organ weights of the animals was observed in dose range finding

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
no significant difference was observed in the extent of pre-implantation loss and post-implantation loss in all treated groups
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
No significant difference was observed in early and late resorption and number of live foetuses in all tretment groups
Early or late resorptions:
no effects observed
Description (incidence and severity):

No significant difference was observed in early and late resorption and number of live foetuses in all tretment groups
Parameter Group G1 G2 G3 G4
Dose (PPM) 0 1000 5000 10000
No. of Dams 21 20 20 21
No. of CL Mean 7.38 7.55 7.35 6.57
SD 2.20 2.37 2.76 1.72
No. of Implantation Mean 6.43 6.00 6.30 5.43
SD 2.01 2.18 2.81 1.54
No. of Foetuses Mean 5.00 4.83 4.96 4.38
SD 2.77 3.13 3.17 2.41
Early Resorption Mean 0.38 0.20 0.20 0.33
SD 0.67 0.52 0.52 0.73
Late Resorption Mean 0.05 0.00 0.10 0.10
SD 0.22 0.00 0.31 0.30
Pre implantation loss Mean 0.95 1.55 1.05 1.14
SD 1.20 1.19 1.23 1.06
% 12.72 20.79 16.36 16.61
Post implantation loss Mean 0.67 0.20 0.40 0.43
SD 0.97 0.52 0.68 0.75
% 11.13 6.96 3.83 10.87
Dam with resorption/s Number 8 3 5 7
Number of dams with abortions Number
0 0 0 0
Number of dams with early deliveries Number
0 0 0 0
Number of dams with stillbirths Number
0 0 0 0
Number of dams with dead foetuses Number
1 0 1 0
Dead fetuses:
effects observed, non-treatment-related
Description (incidence and severity):
Parameter Group G1 G2 G3 G4
Dose (PPM) 0 1000 5000 10000
No. of Dams 21 20 20 21
Total No. of Foetuses 122 116 120 105
Mean Litter Size 5.81 5.80 5.95 5.00
No. of Live Foetuses Number 120 116 119 105
Mean 5.71 5.80 5.67 5.00
SD 2.12 2.44 2.71 1.84
% 98.36 100.00 99.17 100.00
No. of Dead Foetuses Number 2 0 1 0
Mean 0.10 0.00 0.05 0.00
SD 0.44 0.00 0.22 0.00
% 1.64 0.00 0.83 0.00
No. of Live Male Foetuses Number 59 60 53 48
Mean 2.46 2.50 2.21 2.00
SD 1.74 2.15 1.74 1.35
% 49.17 51.72 44.54 45.71
No. of Live Female Foetuses Number 61 56 66 57
Mean 2.54 2.33 2.75 2.38
SD 1.86 1.66 2.09 1.69
% 50.83 48.28 55.46 54.29
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
ca. 10 000 ppm
Based on:
test mat.
Basis for effect level:
other: observed effects were not treatment related hence highest tested dose considered as NOAEL

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
No significant difference was observed in litter and foetus weight in low, intermediate and high dose groups in dose range finding study. However, in main study combined foetus body weight was decrease in high dose (10000 PPM) groups as compared to control (0 PPM). Whereas in female foetuses of High dose revelead decrese in body weight compare to control group of animals
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): effects observed, non-treatment-related
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): in main study combined foetus body weight was decrease in high dose (10000 PPM) groups as compared to control (0 PPM). Whereas in female foetuses of High dose revelead decrese in body weight compare to control group of animals
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
No significant difference was observed in the sex ratio of foetuses in low, intermediate and high dose groups
Changes in litter size and weights:
no effects observed
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
External examination of foetuses revealed two small sized foetus in control, four in low dose, three in intermediate dose and two in high dose group.
No significant difference was observed in external findings in all treatment groups

Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Skeletal examination of foetuses revealed incompletely ossified, not ossified, bipartite, bipartite thoracic vertebrae, not ossified lumbar vertebrae; not ossified or incompletely ossified sacral vertebrae; not ossified caudal vertebrae; not ossified, incompletely ossified and/or misshapen/irregular sternal centers; not ossified or incompletely ossified manubrium and/or xyphoid; waviness in control, low, intermediate and high dose groups. These variations were isolated and when expressed on a foetus/litter basis, it was found to be not statistically significant.
Type and distribution of variations noted during skeletal examination at the dose levels of 1000, 5000 an 10000 PPM did not indicate any test item-related effects
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Visceral examination of foetuses revealed Lungs: Absent Azygos lobe in two foetuses of control and one in low and high dose group foetuses, respectively. Microphthalmia was observed in four foetuses of control and high dose each. Whereas two in intermediate and three foetuses in low dose group was observed. Brain: Dilated ventricles were observed in two foetuses of control and one each in intermediate and high dose groups.
No significant difference was observed in visceral findings in all treatment groups
Details on embryotoxic / teratogenic effects:
The oral (dietary) administration of oxalic acid to pregnant New Zealand white Rabbits from day 6 to day 29 of gestation at dose levels of 1000, 5000 and 10000 PPM resulted in no treatment-related maternal and embryofoetal toxicity

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
ca. 10 000 ppm
Based on:
test mat.
Sex:
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: observed effects were not treatment related hence highest tested dose considered as NOAEL

Fetal abnormalities

Key result
Abnormalities:
effects observed, non-treatment-related

Overall developmental toxicity

Developmental effects observed:
no

Any other information on results incl. tables

Study No.: 5671

 

Study Title: Prenatal Developmental Toxicity Study of OXALIC ACID in Rabbits (Dietary)

1.        Number of pregnant and non-pregnant dams

GROUP

G1

G2

G3

G4

DOSE (PPM)

0

1000

5000

10000

No. of Pregnant females

21

20

20

21

No. of Non-Pregnant females

03

04

04

03

 

2.        Number of dams with abortions, early deliveries, stillbirths, resorptions, and/or dead foetuses

GROUP

G1

G2

G3

G4

DOSE (PPM)

0

1000

5000

10000

Number of dams with abortions

0

0

0

0

Number of dams with early deliveries

0

0

0

0

Number of dams with stillbirths

0

0

0

0

Number of dams with resorptions

8

3

5

7

Number of dams with dead foetuses

1

0

1

0

 

3.        Pre-and post-implantation loss, number and percent  Body weight, body weight change and gravid uterine weight, including optionally, body weight change corrected for gravid uterine weight

GROUP

G1

G2

G3

G4

DOSE (PPM)

0

1000

5000

10000

Pre implantation loss (Number)

0.95

1.55

1.05

1.14

Pre implantation loss (%)

12.72

20.79

16.36

16.61

Post implantation loss (Number)

0.67

0.20

0.40

0.43

Post implantation loss (%)

11.13

6.96

3.83

10.87

Gravid Uterine Weight (G)

376.0784

378.2618

361.4792

324.3361

 

 

 

 

 

 

Group

Body weights (G)

Gestation Day

0

3

6

9

12

15

18

21

24

27

29

30

1

2904.8

2937.8

2974.0

3012.0

3055.2

3102.2

3156.8

3214.2

3274.6

3342.2

3413.8

3431.1

2

2981.3

3005.4

3032.7

3063.4

3099.1

3138.9

3183.2

3230.4

3282.0

3337.7

3399.5

3415.0

3

2972.2

2996.0

3023.7

3056.3

3090.1

3129.8

3186.1

3223.6

3276.5

3333.2

3395.4

3411.1

4

2981.2

3004.6

3032.5

3063.6

3099.1

3138.5

3183.5

3232.0

3284.5

3338.0

3406.2

3422.3

 

 

Group

Body weight Gain (%)

Gestation Day

3

6

9

12

15

18

21

24

27

29

30

1

1.14

2.40

3.71

5.20

6.83

8.71

10.69

12.78

15.11

17.58

18.18

2

0.81

1.72

2.76

3.96

5.30

6.79

8.37

10.11

11.98

14.05

14.57

3

0.80

1.73

2.82

3.96

5.29

7.20

8.45

10.23

12.14

14.23

14.76

4

0.79

1.72

2.77

3.96

5.29

6.80

8.43

10.20

11.99

14.29

14.84

4.        Mean number and percent of live offspring  Mean foetal/pup body weight by sex and sexes combined

Parameter

Group

G1

G2

G3

G4

Dose (PPM)

0

1000

5000

10000

No. of Dams

21

20

20

21

Total No. of Foetuses

122

116

120

105

No. of Live Foetuses

Mean

5.71

5.80

5.95

5.00

Number

120

116

119

105

%

98.36

100.00

99.17

100.00

Foetus weight by sexes combined (G): Mean

44.693

45.678

43.687

41.132

Foetus Weight (G): Mean

Male

45.195

48.682

43.403

41.719

Female

44.200

42.458

43.907

40.618

 

 

 

 

 

5.        Number and percent of foetuses and litters with malformation (including runts) and/or variation as well as description and incidences of malformations and main variations (and/or retardation)

EXTERNAL FINDINGS

Group

G1

G2

G3

G4

Dose (PPM)

0

1000

5000

10000

No. of Litters Examined

21

20

20

21

No. of Litters affected

2

4

3

2

% Litters affected

9.52

20.00

15.00

9.52

No. of Foetuses Examined

120

116

119

105

Variation Incidence – Number (%)

No. of Foetus with Variations

Total Variations

2 (1.67)

4 (3.45)

3 (2.52)

2 (1.90)

Body small in size

2 (1.67)

4 (3.45)

3 (2.52)

2 (1.90)

 

 

 

VISCERAL FINDINGS

Group

G1

G2

G3

G4

Dose (PPM)

0

1000

5000

10000

No. of Litters Examined

21

20

20

21

No. of Litters affected

5

5

3

6

% Litters affected

23.81

25.00

15.00

28.57

No. of Foetuses Examined

120

116

119

105

Variation Incidence – Number (%)

No. of Foetus with Variations

Total Variations

8 (6.67)

5 (4.31)

3 (2.52)

6 (5.71)

Liver: Pale

0

1 (0.86)

0

0

Kidneys: Pale

0

1 (0.86)

0

0

Lungs: Absent Azygos lobe

2 (1.67)

1 (0.86)

0

1 (0.95)

Eyes: Microphthalmia

4 (3.33)

3 (2.59)

2 (1.68)

4 (3.81)

Brain: Dilated ventricles

2 (1.67)

0

1 (0.84)

1 (0.95)

 

 

 

 

 

 

 

 

 

 

 

 

 

SKELETAL FINDINGS

Group

G1

G2

G3

G4

Dose (PPM)

0

1000

5000

10000

No. of Litters Examined

21

20

20

21

No. of Litters affected

9

7

11

6

% Litters affected

42.86

35.00

55.00

28.57

No. of Foetuses Examined

120

116

119

105

Variation Incidence – Number (%)

Total Variations

13 (10.83)

7 (6.03)

16 (13.45)

12 (11.43)

Thoracic Vertebra

Hemicentric

 1 (0.83)

0

0

0

Caudal Vertebra

Bipartite

1 (0.83)

0

0

0

Sternal centers (2)

Misshapen/Irregular shaped

0

0

1 (0.84)

0

Ribs: Left

Small Size

0

0

1 (0.84)

0

Detached

0

0

1 (0.84)

1 (0.95)

Rudimentary

4 (3.33)

3 (2.59)

8 (6.72)

8 (7.62)

Incompletely ossified

1 (0.83)

0

0

0

Ribs: Right

Small Size

0

0

1 (0.84)

0

Not ossified

1 (0.83)

0

0

0

Incompletely ossified

1 (0.83)

0

0

0

Rudimentary

8 (6.67)

3 (2.59)

4 (3.36)

9 (8.57)

Bifid

0

0

1 (0.84)

0

Detached

0

1 (0.86)

1 (0.84)

0

Sternal centers (2)

Misshapen/Irregular shaped

0

0

1 (0.84)

0

Xyphoid

Bent

0

1 (0.86)

0

0

 

 

 

Applicant's summary and conclusion

Conclusions:
The test item, OXALIC ACID was administered daily by dietary route to three treatment groups of twenty four New Zealand white Rabbits per group from day 6 to 29 day of gestation at dose levels of 1000, 5000 and 10000 PPM,. A control group of twenty four females were treated with dietary powder feed alone.
No mortality was observed in any of the treated animal throughout the scheduled treatment period.
No clinical signs were observed in any of the female animals treated at control (0 PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups.
No significant difference was observed in body weight, body weight gain (%) and feed consumption attributable to treatment in all treatment groups when compared with control group.
No treatment-related effects were observed in reproduction parameters such as corpora lutea count, early and late resorption and number of live and dead foetuses, pre-implantation loss, post-implantation loss as well as foetal sex ratio at any dose level treated at 1000, 5000 and 10000 PPM.
Litter weight and foetus body weight in both sexes were significantly not different all treatment groups.
The external and visceral variations observed were randomly distributed across the groups. Therefore these findings were considered as incidental findings and not test item-related.
Type and distribution of variations noted during skeletal examination at the dose levels of 1000, 5000 and 10000 PPM did not indicate any test item-related effects.
The oral (dietary) administration of OXALIC ACID to pregnant New Zealand white Rabbits from day 6 to day 29 of gestation at dose levels of 1000, 5000 and 10000 PPM resulted in no treatment-related maternal and embryo foetal toxicity.
Hence, the No Observed Adverse Effect Level (NOAEL) of test item OXALIC ACID for maternal and embryo foetal toxicity in New Zealand white Rabbits via oral (dietary) route was concluded to be the highest dose level employed i.e., 10000 PPM


Executive summary:

Study Title:Prenatal Developmental Toxicity Study of OXALIC ACID in Rabbits (Dietary)

This study was designed to investigate theeffects of OXALIC ACID on the pregnant female and on embryonic and foetal development when administered orally (dietary) daily to mated female Rabbits from day of implantation to one day prior to scheduled caesarean section (i.e. day 6 through day 29 post mating) including assessment of maternal effects as well as death, structural abnormalities (soft tissue and skeletal), or altered growth in the foetus.

The studyis compatible with the requirements of theOECD Guidelines for the Testing of Chemicals, Number 414 “PrenatalDevelopmental Toxicity Study”, Adopted by the Council on 22ndJanuary 2001.

In dose Range Finding (DRF) study was carried out before the main experiment to confirm the dose level of test item for main study. Three groups (Low, Intermediate and high) consisting of 5 pregnant Female Rabbits each were treated dailyfrom day 6 to day 29 of gestation at dose level of1000, 3000 and 10000 PPM. All animals were observed for mortality/viability and clinical signs of toxicity. Feed consumption and body weight were recorded during treatment period. On day 30thof gestation period all the animals were sent for necropsy and gross pathological examinations.

Dose Range Finding (DRF), no effects were observed in mortality/viability, clinical signs, feed consumption, body weight, gross pathological andvariations/malformations in foetuses subjected for external examination.Dose levels for main study were selected based on dose range finding study results. The doses selected for main study were 1000, 5000 and 10000 PPM.

The test item (formulated in powder feed) was administered daily by oral (dietary) to threetreatmentgroups oftwenty four mated female New Zealand White Rabbits per group from day 6 to day 29 of gestation at dose level of1000, 5000 and 10000 PPM. A control group comprised of 24 female Rabbits was administered powder feed alone.

The analytical method was validated and the linearity response was found to be linear(r2= 1.000)in the range of 10.12 to 500.16 mg. The applied concentration was found homogeneously distributed in the feed at concentrations 1,000 mg/L, 5,000 mg/L and 10,000 mg/L for the test item respectively. (i.e. % coefficient of variation (RSD) ranged in between 1.18 to 2.62). The dose concentration obtained in all the dose groups was in agreement with the target concentration (i.e. ranged between 92.072 % to 97.949 %).Details of the analysis of dose formulation are given inAPPENDIX I.

The mated females were observed twice daily for viability/mortality. Clinical signs were observed twice daily for first three days and once daily thereafter until sacrifice.

Animals were paired on a one male: two female, within each dose group, for a period of fourteen days. Each female was examined for the presence of a copulation plug in the vagina, vaginal mucus membrane and mating behavior. The presence vaginal plug (Presence of sperm), vaginal mucus membrane (swollen valva and congested mucus membrane and mating behavior were considered as positive evidence of mating (Day 0 of gestation).

Body weights were recorded ongestation day 0, 3, 6, 9, 12, 15, 18, 21, 24, 27 and day 30. Feed consumption was recorded on gestation day 3, 6, 9, 12, 15, 18, 21, 24, 27 and 29.All surviving animals were sacrificed by intravenous injection of thiopentone sodium. On cesarean section uteri were removed, weighed and examined for the number of implantation sites, resorption and number of live and dead foetuses. The number of corpora lutea was counted on ovaries. The foetuses were removed from uterus, identified, weighed, sexed and evaluated for external malformation/variation. All the fetuses (alternating foetuses within the litter) were processed for skeletal alterations and for visceral (soft tissue) alterations.

Results

Analytics:

Analysis of stability, homogeneity and dose concentration of the prepared dose formulations was within acceptable levels.

Mortality:

No mortality was observed in any of the treated animal through out the scheduled treatment periodin main study.

Clinical signs:                                                                                                                        

No clinical signs were observed in any of the female animals treated at control (0PPM) Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups.

Body weight:

No significant difference in the body weight and body weight gain (%) was observed in pregnant female animals treated at control (0PPM), Low (1000 PPM), Intermediate (5000 PPM and High dose (10000 PPM) groups.

Feed Consumption:

No significant difference was observed in the feed consumption attributable to treatment in low, intermediate and high dose group compared with control group.

  

Mating Record:

All female animals of control, low, intermediate and high dose groups showed positive evidence of mating. At necropsy, three females each of control, high and and four females of low and intermediate dose group were found non-pregnant.

Necropsy:

Necropsy performed on 30thday of gestation, no test item-related macroscopic findings were observed in any of the animal in all treatment groups in dose range finding and main study.

No test item related microscopic findings were observed in treated animals ofdose range finding study.

All other histopathological findings observed in various tissues during evaluation of test item group animals were comparable with control group and were considered incidental. These changes can usually be considered to be species, age, gender, congenital, physiological or mode of death related and are covered in background historical data of pathology.

Examinationof Uterine Contents:

No significant difference was observed in the gravid uterine weight, corpora lutea count, in early and late resorption and number of live foetuses as well as in pre and post implantation losses between control and treated groups.

Foetus weight:

No significant difference was observed in foetus weight between control and treated groups.

Sex ratio:

Sex ratio was calculated as percent (%) male and there was no significant difference was observed in sex ratio between control and treated groups.

Examination of Foetuses:

ExternalandVisceral examination

No test-item related variations/malformations were observed in foetuses subjected for external and visceral (soft tissue) examination.

Skeletalvariations/malformations

The number of foetuses showing skeletal variation in treatment groups were comparable with control group. In general, incidences of skeletal variations were unaffected by test item treatment.

No test item related external, visceral and skeletal variations/malformations were observed in any of the treated group. Most of the observations recorded were minor abnormalities and considered as routine variables.