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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013-11-24 to 2013-12-19
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
adopted 2001-01-22
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Version / remarks:
2004
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
signed 2013-07-09
Limit test:
no

Test material

Constituent 1
Reference substance name:
barium(2+) dichloride dihydrate
EC Number:
600-412-6
Cas Number:
10326-27-9
Molecular formula:
BaCl2.2(H2O)
IUPAC Name:
barium(2+) dichloride dihydrate
Test material form:
solid: crystalline
Details on test material:
- Name of test material (as cited in study report): Barium chloride dihydrate
- Molecular formula: BaCl2 2H2O
- Physical state: white crystalline powder
- Storage condition of test material: ambient temperature (15-25°C)
Specific details on test material used for the study:
- Source: Solvay Bario e Derivati S.p.A., Massa, Italy
- Batch number: 9284
- Purity: ≥ 98.5%

Test animals

Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
TEST ANIMALS- RccHan: WIST strain
- Source: Harlan, Horst, the Netherlands
- Age at study initiation: approx. 12 weeks of age
- Weight at study initiation: control group: 197.9 - 230.9 g; low dose group: 198.1 - 230.2 g; mid dose group: 190.9 - 232.5 g; high dose group: 191.6 - 239.4 g
- Housing: animals were housed in Macrolon cages with a bedding of wood shavings (Lignocel) and strips of paper (Enviro-dri) and a wooden block as environmental enrichment. During the quarantine and acclimatization periods, the animals were housed in groups of 4 per sex. Mated females were housed individually in Macrolon cages.
- Diet (ad libitum): cereal-based (closed formula) rodent diet (Rat & Mouse No.3 Breeding Diet; RM3) (supplier: SDS Special Diets Services, Witham, England)
- Water (ad libitum): domestic mains tap-water
- Quarantine period: 9 days (upon arrival the rats were quarantined and checked for overt signs of ill health and abnormalities. During the quarantine period, serological examinations of the microbiological status of the rats were conducted in a random sample.)
- Acclimation period: 2 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 - 24°C
- Relative humidity: exceeded 65% for short times only during cleaning activities
- Air changes: about 10 air changes per hour
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: demineralized water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Solutions of the test item in the vehicle were prepared weekly, and stored in a refrigerator (2-10°C) in the dark in aliquots sufficient for one day. The vehicle for dosing the controls was similarly stored.
The solutions of the test item in the vehicle were prepared by stirring on a magnetic stirrer. Subsequently, 8 aliquots (7 days plus 1 extra) were taken per dose level according to the daily volume required for each dosing. These aliquots were taken under continuous stirring. On each subsequent day, one aliquot for each group was removed from the refrigerator and allowed to equilibrate to ambient temperature prior to dosing.

A dosing volume of 10 mL/kg was applied for all animals, which was adjusted based on the latest body weight. After gestation day 14 dose volumes were not adjusted.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
From all three batches of the test items prepared in the study, samples were taken immediately after preparation and stored in a refrigerator until analysis. The following analyses were conducted by Inductively coupled plasma atomic emission spectrometry (ICP-AES) analysis. The test item was quantified using barium as a marker component:
- Homogeneity: the homogeneity of the test substance in the experimental test items was demonstrated in the first batch prepared, by analysing three samples (taken at different locations in the gavage liquid container) of each level.
- Concentration: the concentration of test substance at each level was determined in all three batches of test items prepared in the study.
- Stability: samples of the low-dose, mid-dose and high-dose level were analysed in the first batch prepared in the study at t=0 and after storage in the refrigerator (2 – 10 °C) for twelve days.

Results:
- Homogeneity: the relative standard deviations between the mean content at three different locations was < 5% in the low, mid and high dose level. Therefore barium chloride dihydrate was considered to be homogeneously distributed in each test.
- Stability: upon storage at refrigerator temperature from 22 November 2013 till 4 December 2013, the relative difference in test substance concentration between t=0 and t=4 days was -3.6, +1.5 and +4.2% in the low, mid and high dose level, respectively. And all the dose levels met the criteria for stability (relative difference ≤10%). Therefore it was concluded that there was no loss of test substance from any tests items during storage for twelve days in the refrigerator.
- Content: the content of barium chloride dihydrate determined in the test items are compared with the intended content. The relative difference between the mean determined content and the intended content was between 1.5 and 2.5% at all nominal levels of 1, 3 and 10 mg/ml which was within the acceptance criteria (relative difference ≤10%). Therefore, the actual content was considered to meet the intended level in each test item.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 2 females : one male
- Length of cohabitation: until a sperm positive smear was detected
- Proof of pregnancy: sperm in vaginal smear referred to as gestation day 0 of pregnancy
Duration of treatment / exposure:
gestation day 0 up to and including gestation day 20
Frequency of treatment:
daily
Duration of test:
25 days
Doses / concentrationsopen allclose all
Dose / conc.:
10 mg/kg bw/day
Dose / conc.:
30 mg/kg bw/day
Dose / conc.:
100 mg/kg bw/day
No. of animals per sex per dose:
24 mated female rats
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose levels have been selected in consultation with the study monitor on the basis of a dose range finding study with the test item in pregnant rats.
During the dose range finding study groups of 5 mated females, were administered different dose levels of the test substance by gavage from gestation day 0 up to gestation day 21. A dose volume of 10 mL/kg body weight was applied and demineralized water was used as vehicle and control item. Dose levels of 0, 50, 175 and 250 mg/kg were administered.
Based on the preterm death of 3/5 females in the high dose group and 2/5 females in the mid dose group after a single dose, dosing was discontinued in both groups.
All surviving animals in the mid and high dose group were re-allocated to a new mid dose group and received 100 mg/kg body weight barium chloride from gestation day 2 onwards.
On gestation day 21 the animals were sacrificed and caesarean section was performed.
In-life parameters included clinical signs, morbidity, mortality, body weight and food consumption. At sacrifice uterus weight, number of corpora lutea, number of implantation sites, early and late resorptions, number of live and dead foetuses and foetus weight were recorded. In addition, foetuses were examined for external abnormalities/malformations and dams were observed for gross anatomical changes.

Results:
Oral administration of 0, 50, 100, 175 and 250 mg/kg barium chloride to mated females resulted in:
- the preterm death of 3/5 animals in the 250 mg/kg group and 2/5 animals in the 175 mg/kg group after a single oral dose.
- the spontaneous death of one animal in the 100 mg/kg group on gestation day 21. This animal was found dead before cesarean section and had 11 dead foetuses. This animal had received one dose of 250 mg/kg on gestation day 0 and daily doses of 100 mg/kg from gestation day 2 to 21.
- limited clinical observations in the 250 and 175 mg/kg group, including hunched posture an piloerection.
- no effect on body weight or body weight gain, food consumption, mean number of corpora lutea, implantation sites, early and late resorptions and the mean number of live foetuses.
- although based on a limited number of litters (four in the 50 mg/kg group and three in the 100 mg/kg group) an effect on foetus weight could not be ruled out.
- no foetuses showing external malformations

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: clinical signs and mortality

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: gestation days (GD) 0, 3, 6, 10, 14, 17 and 21

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes (gestation days 0-3, 3-6, 6-10, 10-14, 14-17 and 17-21)
- 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: No

WATER CONSUMPTION AND COMPOUND INTAKE: No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day #21
The females were killed by decapitation after CO2/O2 anaesthesia on gestation day 21 and examined for gross abnormalities.

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Weight of empty uterus: Yes
- Weight of ovaries: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Gross evaluation of placentas: Yes
For seemingly non pregnant females (part of) the uterus was stained with Na2SO3 in order to visualize possible implantation sites (Salewski E, 1964). Upon staining non pregnancy was confirmed for these females.
Fetal examinations:
- External examinations: Yes
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No

Further examinations:
- number of live and dead foetuses
- sex of the foetuses
- live foetuses (individually) and corresponding placentas
- foetal weight
Statistics:
Tests were generally performed as two-sided tests with results taken as significant where the probability of the results was p<0.05 or p<0.01.
Continuous data were subjected to the ‘Decision tree for continuous data’ and dichotomous data to the ‘Decision tree for dichotomous data’.
Indices:
For each group the following indices were calculated:
- female fertility index = (no. of pregnant females/no. of inseminated females) x 100
- pre-implantation loss = [(no. of corpora lutea – no. of implantation sites) / no. of corpora lutea] x 100
- post-implantation loss = [(no. of implantation sites – no. of live foetuses) / No. of implantation sites] x 100
- gestation index = (no. of females with live foetuses/no. of females pregnant) x 100
- sex ratio = (no. of live male foetuses/no. of live foetuses) x 100

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Description (incidence and severity):
No treatment related clinical signs were observed in the control group, low dose group, mid dose group and the other animals in the high dose group.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, treatment-related
Description (incidence):
Two animals in the high dose group were found dead on day 21 of gestation. Both animals were pregnant and all foetuses were dead. Although death was not preceded by clinical signs, growth retardation or gross anatomical observations at necropsy that could clarify the death of these animals, the death of these rats is ascribed to treatment.
- one animal in the high dose group felt cold and was weakened and showed piloerection on gestation day 21. Upon necropsy this animal showed hydrothorax, haemorrhages in the liver and haemorrhagic discharge in the vagina. Also the death of this high-dose rat is ascribed to treatment.
- the spontaneous death of two rats, and the conditional decline of one rat on day 21 of gestation were considered to be treatment-related and to represent severe maternal toxicity in the high dose group.
- all foetuses were dead in the above three rats. The foetal deaths observed in these animals are considered to be related to the severe maternal toxicity in the high-dose group.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
- No effects were observed on body weight. A slightly, but statistically significantly reduced body weight gain was observed in the high dose group as compared to the control group during the first three days of dosing. This was considered to be related to treatment and recovered thereafter.
- No effects on body weight or body weight gain were observed in the low dose group and the mid dose group as compared to the control group.
Detailed results are provided in Table 1 and 2, see 'Any other information on results'.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There were no effects on food consumption. Detailed results are provided in Table 3 see 'Any other information on results'.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Endocrine findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no treatment related macroscopic findings in the dams, no effects on mean uterus weight, mean ovary weight and mean carcass weight.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
FEMALE REPRODUCTIVE ORGANS
- mean ovary weight, mean full and empty uterus weight were comparable in all groups
- mean carcass weight and net body weight change were comparable in all groups
- mean placenta weight was comparable in all groups

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
Reproduction indices were comparable for the control, low dose, mid dose and high dose group; no effects were noted in mean number of corpora lutea (13.0, 13.1, 12.8 and 13.0, respectively), mean number of implantation sites (10.9, 11.4, 11.0 and 10.8, respectively) and pre-implantation loss (16.8%, 14.0%, 13.4% and 16.7%, respectively). Detailed results are provided in Table 4 see 'Any other information on results'.
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Description (incidence and severity):
Mean number of early resorptions (0.7, 0.5, 0.6 and 0.3, respectively) and late resorptions (0.1, 0.1, 0 and 0.1, respectively) were comparable for the control, low dose, mid dose and high dose group.
Detailed results are provided in Table 4 see 'Any other information on results'.
Dead fetuses:
no effects observed
Description (incidence and severity):
Mean number of live fetuses was comparable for the control, low dose, mid dose and high dose group (10.1, 10.7, 10.4 and 9.9, respectively). Two females that were found dead in the high dose group (147 and 165) had dead fetuses only. In addition, the female in the high dose group that showed a bad general condition also had dead fetuses only. This effect is considered to be related to the ill health status of the animals and is therefore related to severe maternal toxicity.
Detailed results are provided in Table 4 see 'Any other information on results'.
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
not examined

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Remarks:
(barium chloride dihydrate)
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Remarks:
(barium chloride)
Effect level:
25.6 mg/kg bw/day
Based on:
other: bariumdichloride
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
Mean fetus weight was comparable in all groups (4.7 grams in the control group and mid dose group and 4.6 grams in the low dose and high dose groups, respectively).
Detailed results are provided in Table 5 see 'Any other information on results'.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
The mean number of live fetuses was comparable in all groups.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The mean percentages male littermates was comparable in all groups (49.2% in the control group and 53.9%, 54.3% and 48.4% in the low dose, mid dose and high dose group, respectively).
Detailed results are provided in Table 5 see 'Any other information on results'.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
No effects were observed on mean fetus weight or mean placenta weight.
Anogenital distance of all rodent fetuses:
not examined
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Description (incidence and severity):
Malformations: No external malformations were observed.
Variations: One fetus in the control group and one fetus in the mid dose group were considered to be small. In the low dose, mid dose and high dose groups, respectively 5, 8 and 3 fetuses (from 4, 6 and three litters) showed subcutaneous hemorrhages.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Malformations: One fetus in the mid dose group showed an absent lumbar centrum, resulting in a malformation of the vertebral column. No other skeletal malformations were observed.
Variations: In the high dose group a statistically significant percentage of fetuses per litter showed unossified metacarpals in the forelimbs or digits in the hindlimbs. The ossification status of the other digits of the forelimbs and fetus weight do not support a retardation in ossification and therefore the effects on ossification status of the forelimb metacarpals and hindlimb digits are not related to treatment.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Malformations: One fetus in the low dose group showed situs inversus. No other visceral malformations were observed. This is not considered to be related to treatment.
Variations: Visceral variations observed included unilateral or bilateral folding of the retina, unilateral or bilateral bent or kinked ureters and malposition of the kidneys. A statistically significantly increase in the number of fetuses showing distended urinary bladder
was observed in the high dose group (8 fetuses from 5 litters), whereas no fetuses showing distended urinary bladder were observed in the control group. Historical background data for this observation in the past 4 studies ranges from 4 fetuses (in 3 litters) to 15 fetuses (in 8 litters) in the control group. Therefore, based on historical background data for this observation, distended urinary bladder is considered a chance finding and not a treatment related effect.
Details on embryotoxic / teratogenic effects:
Based on the incidence and distribution of the external, visceral and skeletal malformations and variations observed, no test substance related effects were observed. At the dose levels tested the test substance showed no teratogenic potential.

Effect levels (fetuses)

open allclose all
Dose descriptor:
NOAEL
Remarks:
(barium chloride dihydrate)
Effect level:
>= 100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: No effects observed at the highest dose level
Dose descriptor:
NOAEL
Remarks:
(barium chloride)
Effect level:
>= 85.3 mg/kg bw/day
Based on:
other: barium chloride
Basis for effect level:
other: No effects observed at the highest dose level
Remarks on result:
other: NOAEL calculated for barium chloride from the barium chloride dihydrate data.

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
no

Any other information on results incl. tables

Table 1: Body weight of pregnant rats (g)

 

Day(s) Relative to mating

Day 0

[k]

Day 3

 [a]

Day 6

 [a]

Day 10

 [a]

Day 14

 [k]

Day 17

 [a]

Day 21

 [k]

0 mg/kg d

Mean

SD

N

218.59

9.12

23

230.96

8.61

23

239.37

9.97

23

252.25

11.61

23

265.58

12.71

23

289.87

13.64

23

309.03

21.17

23

10 mg/kg d

Mean

SD

N

213.48

8.14

22

225.31

8.75

22

232.85

9.54

22

246.61

9.62

22

261.33

10.47

22

286.60

12.78

22

299.87

17.70

22

30 mg/kg d

Mean

SD

N

215.78

10.62

23

227.60

10.93

23

235.34

11.68

23

247.91

12.86

23

261.65

13.17

23

285.34

13.41

23

299.57

17.81

23

100 mg/kg d

Mean

SD

N

216.41

11.27

22

225.60

12.64

22

233.77

14.62

22

246.67

16.44

22

262.11

17.95

22

286.88

21.60

22

312.97

26.13

22

k=KRUSKAL-WALLIS; a=ANOVA

Table 2: Body weight changes of animals (g/day)

 

Day(s) Relative to mating

0 → 3

[d]

3 → 6

[a]

6 → 10

[k]

10→ 14

[k]

14→ 17

[k]

17→ 21

[u]

0→ 21

[a]

0 mg/kg d

Mean

SD

N

12.37

2.96

23

8.41

3.06

23

12.87

3.31

23

13.33

4.09

23

24.29

5.26

23

19.16

14.04

23

90.44

17.93

23

10 mg/kg d

Mean

SD

N

11.83

3.27

22

7.54

3.49

22

13.76

4.30

22

14.72

3.58

22

25.27

5.49

22

13.27

13.89

22

86.40

15.55

22

30 mg/kg d

Mean

SD

N

11.83

3.06

23

7.74

3.28

23

12.57

5.38

23

13.74

4.10

23

23.69

6.34

23

14.23

13.40

23

83.80

16.49

23

100 mg/kg d

Mean

SD

N

9.20**

3.40

22

8.17

3.92

22

12.90

4.42

22

15.45

3.02

22

24.76

6.02

22

26.09

11.41

22

96.56

19.18

22

* = p < 0.05, ** = p < 0.01

d=ANOVA-DUNNETT; a=ANOVA; k=KRUSKAL-WALLIS; u=KRUSKAL-WALLIS-DUNN

Table 3: Food consumption of animals (g/day)

 

Day(s) Relative to mating

0 → 3

[a]

3 → 6

[k]

6 → 10

[a]

10→ 14

[a]

14→ 17

[k]

17→ 21

[d]

0 mg/kg d

Mean

SD

N

16.34

1.87

23

18.12

2.11

23

18.17

2.30

23

19.91

2.60

23

19.62

2.66

23

15.34

5.11

23

10 mg/kg d

Mean

SD

N

15.57

1.68

22

16.97

2.59

22

18.08

1.70

22

19.04

2.04

22

18.50

2.64

22

13.68

4.95

22

30 mg/kg d

Mean

SD

N

15.73

1.85

23

17.04

1.62

23

17.71

1.90

23

20.50

2.47

23

19.41

1.76

23

13.84

4.95

23

100 mg/kg d

Mean

SD

N

14.91

2.64

22

16.70

2.51

22

18.10

1.99

22

20.45

3.58

22

20.29

1.81

22

17.34

3.96

22

a=ANOVA; k=KRUSKAL-WALLIS; d=ANOVA-DUNNETT

Table 4: Developmental endpoint results

 

0 mg/kg d

10 mg/kg d

30 mg/kg d

100 mg/kg d

Mated female

(n)

24

24

24

24

Pregnant female

(n)

23

22

23

20

Corpora Lutea

(No. per animal)[k]

Mean

SD

Total

13.0

1.7

299

13.1

1.6

250

12.8

3.1

254

13.0

1.6

217

Implantation sites (No. per animal) [k]

Mean

SD

Total

10.9

3.0

250

11.4

3.3

250

11.0

3.7

254

10.8

3.0

217

Pre-implantation loss

(No. per animal) [k]

Mean

SD

Total

2.1

2.5

49

1.8

2.7

39

1.8

2.5

41

2.1

2.6

43

Pre-implantation loss (% per animal)

%

16.8

14.0

13.4

16.7

Fetuses (No. per animal) [k]

Mean

SD

Total

10.1

2.9

232

10.7

3.2

236

10.4

3.7

239

10.4

3.2

209

Alive

%

100.0

100.0

100.0

94.7

Dead

%

0.0

0.0

0.0

5.3

Live fetuses

(No. per animal) [k]

Mean

SD

Total

10.1

2.9

232

10.7

3.2

236

10.4

3.7

239

9.9

4.0

198

Malformed fetuses, external observations

(No. per animal)

Mean

SD

Total

0.0

0.0

0

0.0

0.0

0

0.0

0.0

0

0.0

0.0

0

Fertility index

Mean

96%

92%

96%

91%1

Gestation index

Mean

100%

100%

100%

95%1

Early resorption

(No. per animal) [k]

Mean

SD

Total

0.7

0.9

16

0.5

0.7

12

0.6

1.1

14

0.3

0.5

6

Early resorption (% per animal)

%

6.2

5.4

5.7

3.3

Late resorption

(No. per animal) [k]

Mean

SD

Total

0.1

0.3

2

0.1

0.3

2

0.0

0.2

1

0.1

0.3

2

Late resorption (% per animal)

%

0.7

0.7

0.3

1.3

Abortion sites

(No. per animal)

Mean

SD

Total

0.0

0.0

0

0.0

0.0

0

0.0

0.0

0

0.0

0.0

0

Postimplantation loss

No. per animal) [k]

Mean

SD

Total

0.8

0.9

18

0.6

0.8

14

0.7

1.2

15

0.9

2.4

19

Post-implantation loss (% per animal)

%

6.9

6.1

6.0

9.6

Placenta weight (g)

Mean

SD

0.51

0.13

0.49

0.08

0.48

0.10

0.50

0.09

Uterine weight (g)

day 21

Mean

SD

62.716

15.253

66.400

18.324

62.991

18.621

63.072

17.293

1Two females in the high dose group were found dead on day 21 of gestation and were excluded from the mean data table: Female 147 had 12 dead fetuses and female 165 had 11 dead fetuses.

k = KRUSKAL-WALLIS

Postimplantation Loss = Early/Late resorptions + Aborted Fetuses + Dead Fetuses

Table 5: Fetal examination results

 

0 mg/kg d

10 mg/kg d

30 mg/kg d

100 mg/kg d

Body weight of fetus [k] (g)

Mean

SD

4.71

0.44

4.62

0.46

4.67

0.51

4.63

0.29

Number of male fetuses

111

126

124

95

Number of female fetuses

121

110

115

103

Sex distribution (males/total) [c] (%)

49.2

53.9

54.3

48.4

k=KRUSKAL-WALLIS

 

Applicant's summary and conclusion

Conclusions:
The No-Observed-Adverse-Effect-Level (NOAEL) of barium chloride dihydrate for maternal and embryo-fetal development toxicity in rats was considered to be 30 and ≥ 100 mg/kg body weight, respectively.
Executive summary:

A GLP compliant prenatal developmental toxicity study with barium chloride dihydrate was conducted in Wistar rats according to OECD Guideline 414. Barium Chloride Dihydrate was administered daily via gavage at dose levels of 0, 10, 30 or 100 mg/kg body weight to groups of 24 mated rats from gestation day 0 up to and including gestation day 20. A dosing volume of 10 mL/kg bodyweight was applied for all animals.

The study animals were observed daily for mortality and clinical signs. Body weight and food consumption of the dams were also recorded. At day 21 of gestation dams were sacrificed and caesarean section was performed. Necropsy parameters included gross examination of the dam, uterus weight (full and empty), ovary weight and number of corpora lutea. The number of implantation sites, early and late resorptions, live and dead fetuses, fetus weight, placenta weight, fetus sex and external abnormalities were recorded. Pre-implantation loss and post-implantation loss were calculated. Approximately half of the fetuses were examined for soft tissue abnormalities. The other half was examined for skeletal abnormalities and ossification state.

Daily administration of barium chloride dihydrate at dose levels of 0, 10, 30 or 100 mg/kg body weight to pregnant rats from gestation day 1 up to and including gestation day 20, resulted in maternal toxicity as evidenced by the spontaneous deaths of two animals on gestation day 21 and the conditional decline of another animal on gestation day 21 in the high dose group. No developmental toxicity was observed.

The NOAEL for maternal toxicity was therefore 30 mg/kg body weight (recalculated for barium chloride: 25.6 mg/kg bw/day). In absence of developmental effects the NOAEL for prenatal developmental toxicity in the rat was ≥ 100 mg/kg body weight (recalculated for barium chloride: ≥ 85.3 mg/kg bw/day).