Registration Dossier

Diss Factsheets

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
21 January 2014 to 31 March 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose for cross-reference:
other: read-across target
Reference
Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study conducted on read-across material
Justification for type of information:
Read across to structurally similar substance monobutyltin trichloride (MBTC, CAS No.: 1118-46-3), see attached justification.
Reason / purpose for cross-reference:
read-across source
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake
Abnormalities:
not specified
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
Abnormalities:
not specified
Developmental effects observed:
not specified

Data source

Reference
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)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
N-butyltin trichloride
EC Number:
214-263-6
EC Name:
N-butyltin trichloride
Cas Number:
1118-46-3
Molecular formula:
C4H9Cl3Sn
IUPAC Name:
n-butyltin trichloride
Test material form:
liquid
Details on test material:
- Physical state: Clear, beige liquid
- Storage conditions of test material: Room temperature, protected from light

Test animals

Species:
rat
Strain:
other: Crl:CD (SD)
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at study initiation: 8 to 10 weeks
- Weight at study initiation: 177 to 247 g
- Housing: individually housed in solid bottom cages with nonaromatic woodchip bedding
- Diet: ad libitum
- Water: tap water available ad libitum from an automatic watering system

ENVIRONMENTAL CONDITIONS
- Temperature: 68 to 79 °F
- Humidity: 30 to 70 %
- Photoperiod: 12 hour light/dark cycle

IN-LIFE DATES: From: 21 and 22 January 2014 To: 11 February 2014

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Formulations of the test material in the vehicle were prepared weekly and stored refrigerated at 2 to 8 °C. Formulations were prepared at nominal concentrations of 20, 60 and 200 mg/mL.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The vehicle was selected based on information from a pilot study.
- Concentration in vehicle: 20, 60 and 200 mg/mL.
- Amount of vehicle (if gavage): 5 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dosing samples were evaluated for homogeneity and concentration. 1 mL samples were collected for homogeneity during week 1 from the top, middle and bottom of the 100 and 1000 mg/kg formulations. Samples from all concentrations were taken during weeks 1 and 2 from the middle stratum for analysis of concentration. The dose formulations were analysed using HPLC/UV.

HPLC/UV CONDITIONS
Column: Agilent Poroshell 120 SB-C8 column, 3.0 x 50 mm
Gradient flow: 0.05 % trifluoroacetic acid in water (mobile phase A) and acetonitrile (mobile phase B) at a flow rate of 1.0 mL/minute.
Wavelength: 230 nm
Derivatisation: Prior to analysis, duplicate samples were derivatised in hexane and phenylmagnesium bromide then diluted with isopropyl alcohol to within the range of the calibration curve. Vehicle samples were also derivitised and then diluted using a dilution factor of 100.

RESULTS
- Homogeneity
Analysis of the low- and high-dose formulations (20 and 200 mg/mL, respectively) used for dosing the first week of study confirmed they were homogeneous as prepared as each formulation met the laboratory’s acceptance criteria of 100 ± 15 % of nominal and % RSD (Relative Standard Deviation) ≤ 10.
- Concentration
The test formulations used for dosing the first two weeks of study analysed between 97.8 and 101.4 % of nominal with the % RSDs ranging between 0.127 to 3.344, confirming that animals were receiving the appropriate dose levels when treated at 5 mL/kg/dose. The laboratory’s acceptance criteria for concentration were similar to that for homogeneity (100 ± 15 % of nominal and %RSD ≤ 10). No test material was found in the control vehicle samples.
Details on mating procedure:
- Impregnation procedure: purchased timed pregnant
Duration of treatment / exposure:
Gestation day 0 to 19
Frequency of treatment:
Once daily
Duration of test:
19 days
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
25 females per dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Doses were selected based on information from a pilot study. In the pilot study, dose levels of 75, 150, 300, and 600 mg/kg/day were evaluated. Animals were treated orally from GD 0 to 19. No maternal or developmental toxicity was observed at these dose levels in the pilot study. Therefore, in the absence of maternal toxicity at the 600 mg/kg/day dose level in the pilot study, the high dose level in the definitive study was increased to 1000 mg/kg/day, a limit dose.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
- Cage side observations: Morbidity, mortality, injury and availability of food and water.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily
- Examinations: Animals were removed from the cage and given a detailed clinical examination. The observations included, but were not limited to, evaluation of the skin, fur, eyes, ears, nose, oral cavity, thorax, abdomen, external genitalia, limbs and feet, respiratory and circulatory effects, autonomic effects such as salivation, nervous system effects including tremors, convulsions, reactivity to handling and unusual behaviour.

BODY WEIGHT: Yes
- Time schedule for examinations: Gestation days 0, 3, 6, 9, 12, 15, 18 and 20. Body weight change was calculated for gestation days 0-3, 3-6, 6-9, 9-12, 12-15, 15-18, 18-20 and 0-20. Adjusted body weight (minus gravid uterus) and adjusted bodyweight change were also calculated.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Time schedule for examinations: Food consumption was measured and recorded on the corresponding body weight days and calculated for the same intervals.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20. Each surviving female was euthanised by carbon dioxide inhalation, followed by exsanguination of the abdominal vena cava and immediately subjected to a caesarean section.
- Organs examined: Dams were subjected to a complete necropsy. Special emphasis was placed on structural abnormalities or pathologic changes that may have influenced pregnancy.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
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: half per litter
- Head examinations: Yes: all per litter
Statistics:
All parental in-life data, gravid uterine weights, corpora lutea/dam, total implantations/dam, litter size/dam, viable foetuses/dam, total number of resorptions/dam, number of early resorptions/dam and number of late resorptions/dam were assessed using group pair-wise comparisons. Levene's test was used to assess homogeneity of group variances for each specified endpoint and for all collection intervals. If Levene's test was not significant (p≥0.01), a pooled estimate of variance (Mean Square Error) was computed from a one-way analysis of variance (ANOVA) and utilised by a Dunnett's comparison of each treatment group with the control. If the Levene's test was significant (p<0.01), then Welch's t-test was used with a Bonferroni correction for comparisons with the control group. All endpoints were analysed using two-tailed tests.
Foetal sex ratio (% males/litter), % preimplantation loss and % postimplantation loss were analysed using Arcsin-Square-Root Transformation. Data presented as percentages were transformed using the arcsin of the square root and analysed in accordance with the group pair-wise comparisons.
Pregnancy index, malformations and variations were all evaluated using Fisher’s exact test. An overall test for association between response and treatment was performed. If this test was significant (p<0.05) and there were more than two groups, then each treatment group was compared to the control group. All endpoints were analysed sing two-tailed tests. Overall testing and follow-up pair-wise testing was conducted for the control and all treatment groups that had sufficient sample size (n≥3).
Mean foetal bodyweights were analysed by covariate analysis by comparing treatment groups for each endpoint (mean of foetal bodyweights per female). The covariate was litter size. Each treatment satisfying the sample size assumption was compared to the control using Dunnett's test under the analysis of covariance model. Endpoints were analysed using two tailed tests.
Indices:
- Viable foetuses
- Postimplantation loss = ((No. implantations - No. viable foetuses) / No. implantations) x 100
- Preimplantation loss = ((No. corpora lutea - No. implantations) / No. corpora lutea) x 100
- Pregnancy index = (No. pregnant females - No. females with evidence of mating) x 100
- Foetal sex ratio
Historical control data:
A large database of historical control data for the rat was available for comparison.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
- Salivation was observed at least once in 1, 1 and 16 animals in the 100, 300 and 1000 mg/kg bw/day groups, respectively. Audible breathing, likely related to the salivation, was observed at least once in two, three, and ten animals in the low, mid and high dose groups respectively. Audible breathing was not heard post administration in the control group. This was considered to be a pharmacologic response to the test material.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
- Administration of the test material had no effect on maternal survival. All animals in the control group and 100 mg/kg bw/day group survived until necropsy.
- One death occurred in each of the 300 and 1000 mg/kg bw/day groups on gestation days 17 and 20, respectively. These were not attributed to test material administration.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
- No effect on bodyweight or body weight change was observed in the 100 and 300 mg/kg bw/day groups.
- In the 1000 mg/kg bw/day group, mean body weights on gestation day 18 and 20 were 6 and 7 % lower, respectively, than mean control values. These differences were statistically significant and attributed to test material administration. Mean body weight gain for this group between gestation days 9 to 12 was 14 % higher than the control group and 62, 32 and 18 % lower between gestation days 12 to 15, 15 to 18 and 18 to 20, respectively. Between gestation days 0 to 20 the bodyweight gain for this group overall was 15 % lower than controls. Several animals in the 1000 mg/kg bw/day group were found to have experienced weight loss during the study. With the exception of the animal found dead on GD 20 which was found at necropsy to have a white paste-like foreign material extending the length of the oesophagus and into the stomach (which was determined to be ingested bedding material), the cause of the weight loss for the animals in this group was attributed to lower food consumption.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
- No effect on maternal food consumption was noted in the 300 and 100 mg/kg bw/day dose groups and values were comparable to mean control values.
- In the high dose group, food consumption was statistically lower than controls over GD 0 to 3 (- 12 %), GD 15 to 18 (-22 %), GD 18 to 20 (-14 %) and GD 0 to 20 (-9 %). This was attributed to test material administration and correlated, particularly in late in gestation, with lower body weights and body weight gains.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
- At necropsy, no effect of treatment with the test material was noted in the 100 mg/kg bw/day dose level. The few findings in this group occurred at a low incidence and were not attributed to test material administration.
- Foreign material generally identified as white in colour, which appeared to be bedding material, was observed in seven and twenty three females in the 300 and 1000 mg/kg bw/day groups, respectively. This was not observed in the control or low dose group animals. The toxicological significance of this finding is unclear. Other macroscopic findings in the 300 mg/kg bw/day group were of low incidence and not considered to be related to test material administration.
- In the 1000 mg/kg bw/day dose group, all 25 animals were found to have white foci in the nonglandular portion of the stomach. The occurrence of this finding was considered to be test material related.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not examined

Maternal developmental toxicity

Number of abortions:
not specified
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
There were no treatment related effects on uterine implantation data.
Total litter losses by resorption:
not specified
Early or late resorptions:
not specified
Dead fetuses:
no effects observed
Description (incidence and severity):
- 25, 24, 24 and 24 litters, in the control, low, mid and high dose groups respectively, were produced with viable foetuses to examine on GD 20.
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
Pregnancy indices were 100, 96, 100 and 100 % in the control, low, mid and high dose groups, respectively.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
- There were no treatment related effects on uterine implantation data. The mean number of corpora lutea in the 300 mg/kg bw/day group was statistically lower than the control group. This was considered to be spurious as ovulation occurred prior to the start of treatment. Additionally the reduction in number of corpora lutea in this group had no impact on uterine implantation data.
- Mean gravid uterine weights, adjusted GD 20 body weights and adjusted GD 0 to 20 body weight change in the 100 and 300 mg/kg bw/day groups were comparable to the mean control values. In the 1000 mg/kg bw/day group, the mean gravid uterine weight was statistically lower than the mean control value. This was attributed to lower foetal body weights. The mean adjusted GD 20 body weight and GD 0 to 20 change were also statistically lower, which correlated with lower bodyweights and bodyweight change at GD 20 in this group.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake

Maternal abnormalities

Abnormalities:
not specified

Results (fetuses)

Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
- In the 100 and 300 mg/kg bw/day group, no effect on foetal body weight was observed and body weights were comparable to the control group. In the 1000 mg/kg bw/day group, foetal body weights were 8.6 % lower for combined sexes and 8.1 and 8.6 % lower for males and females, respectively. This was found to be statistically significant and considered an effect of test material administration.
Reduction in number of live offspring:
not examined
Changes in sex ratio:
no effects observed
Description (incidence and severity):
Foetal sex ratios were not affected by the administration of the test material. Mean sex ratios ranged from 51.5 to 58.3 % and were comparable to the mean control ratio of 52.0 %.
Changes in litter size and weights:
not specified
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No external malformations or variations were attributed to test material administration at all dose levels tested. Two foetuses in one litter of the 300 mg/kg bw/day group were found to have oedema and a meningoencephalocele defect and entire body discolouration and protruding tongue. These observations were only noted in two foetuses from the same litter. In the absence of any findings in the 1000 mg/kg bw/day group, these were therefore not considered to be test material related.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
- No skeletal malformations or variations were attributed to test material administration. In the 100 mg/kg bw/day group, five foetuses from a single litter had bent long bones of the forelimbs (humerus, radius and/or ulna) and bent scapulae. With the exception of bent scapulae, these malformations are known to have low historical incidence in the performing laboratory and in the absence of similar malformations at higher dose groups, this was considered to be incidental. The dam with the affected litter was noted to have intervals of weight loss and low food consumption during the study and was found to have adhesions within the thoracic cavity and clear fluid at necropsy. The observations in the foetuses for this dam were attributed to the poor health of the dam during the study. In the 300 mg/kg bw/day group two foetuses from separate litters were noted to have dissimilar skeletal malformations. In the 1000 mg/kg bw/day group, two foetuses from the same litter had shortened bones of the fore and hind limbs, short or misshapen exoccipital bones of the skull and misshapen scapula. These were not attributed to test material administration. No other skeletal malformations were noted in foetuses from this group.
- Skeletal developmental variations in the treated groups were found to be comparable to the control group. Their occurrence within the treated groups were generally within the historical control data range.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
In the 1000 mg/kg bw/day group, the only visceral malformation observed was a folded retina in a single foetus. This was considered to be spontaneous and unrelated to test material administration.
Other effects:
not examined

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Table 1: Mean gestation bodyweights, bodyweight change, food consumption and gravid uterine weights

Day / Interval

Dose Group (mg/kg bw/day)

0

100

300

1000

Bodyweight (g)

Body weight Change (g)

Food Consumption (g/animal/day)

Bodyweight (g)

Body weight Change (g)

Food Consumption (g/animal/day)

Bodyweight (g)

Body weight Change (g)

Food Consumption (g/animal/day)

Bodyweight (g)

Body weight Change (g)

Food Consumption (g/animal/day)

0 / -

213.4

-

-

211.2

-

-

212.7

-

-

212.7

-

-

3 / 0-3

228.3

14.8

12.8

229.3

18.1

13.4

233.3

20.6**

13.2

230.2

17.5

11.3*

6 / 3-6

243.9

15.6

17.3

243.5

14.2

16.6

249.1

15.8

17.0

243.8

13.6

16.6

9 / 6-9

262.0

18.1

19.2

261.0

17.5

18.1

266.4

17.3

19.2

260.8

17.0

19.1

12 / 9-12

284.1

22.1

20.2

285.1

24.1

19.6

293.3

26.9**

20.3

286.2

25.3*

19.8

15 / 12-15

305.6

21.5

21.7

303.0

17.9

21.4

309.5

16.2

22.6

294.2

8.1**

20.1

18 / 15-18

337.6

32.0

24.3

335.6

32.7

23.4

343.4

34.0

23.7

316.0*

21.7**

19.0**

20 / 18-20

370.9

33.2

21.4

364.3

28.7

20.4

377.9

34.5

22.8

345.5*

27.2*

18.4*

- / 0-20

 

Overall change: 157.4

Overall mean: 19.5

 

Overall change: 153.1

Overall mean: 18.9

 

Overall: 165.0

Overall mean: 19.7

 

Overall change: 133.3**

Overall mean: 17.7**

Adjusted Final Bodyweight (g)

20

296.8

294.9

305.7

278.6*

Adjusted weight Change from Day 0 (g)

20

83.4

83.7

92.8

66.4**

Gravid Uterine Weight (g)

20

74.1

69.4

72.2

66.9*

* p < 0.05

** p < 0.01

 

Table 2: Mean foetal bodyweights

Dose Group (mg/kg bw/day)

Mean Bodyweight of Males

(g)

Mean Bodyweight of Females

(g)

Mean Bodyweight of Males & Females

(g)

0

4.29 (4.30)

4.07 (4.08)

4.19 (4.20)

100

4.19 (4.18)

3.96 (3.94)

4.09 (4.07)

300

4.24 (4.24)

4.05 (4.05)

4.15 (4.15)

1000

3.94 (3.95)**

3.73 (3.74)**

3.83 (3.84)**

** p < 0.01

( ) Least square mean

Applicant's summary and conclusion

Conclusions:
Under the conditions of the test, the NOAEL for developmental toxicity and maternal toxicity were 300 mg/kg bw/day based on lower foetal body weight and lower bodyweight, bodyweight gain and food consumption observed in dams treated with 1000 mg/kg bw/day. There were no teratogenic effects recorded in the study.
Executive summary:

The developmental toxicity potential of the test material was investigated in the rat in a study performed in accordance with the standardised guidelines OECD 414 and US EPA OPPTS 870.3700 under GLP conditions.

The test material was administered to timed pregnant Crl:CD (SD) rats via oral gavage in corn oil from gestation day 0 to gestation day 19 at doses of 0, 100, 300 or 1000 mg/kg bw/day. Observations of the animals included clinical signs, body weights and food consumption. Animals were sacrificed on gestation day 20. The dams underwent ovarian and uterine examinations and necropsy. The foetuses were examined for external, skeletal and visceral malformations and variations.

No treatment-related effect was observed on survival. The death of one animal in each of the 300 and 1000 mg/kg/day dose groups was not considered test material related. Salivation and audible breathing were observed in the treated groups and considered a pharmacologic response to the test material. No effect of treatment at the 100 and 300 mg/kg/day dose levels was observed on gestation body weights, body weight change, or food consumption. At the 1000 mg/kg/day dose level, maternal toxicity was observed due to statistically significant changes from mean control values of the following: lower body weights on GD 18 (-6 %) and GD 20 (-7 %); lower body weight gain GD 12 to 15 (-62 %), GD 15 to 18 (-32 %), GD 18 to 20 (-18 %), and GD 0 to 20 (-15 %); and lower food consumption GD 0 to 3 (-12 %), GD 15 to 18 (-22 %), GD 18 to 20 (-14 %), and GD 0 to 20 (-9 %). A common finding in all the 1000 mg/kg/day animals only at macroscopic examination was white foci in the nonglandular portion of the stomach. Another finding included foreign material generally identified as white in colour and having the appearance of bedding was observed in 7/25 and 23/25 females in the 300 and 1000 mg/kg/day dose groups, respectively, but not observed among the control or 100 mg/kg/day animals. The reason for these animals ingesting the bedding and its toxicological significance in relation to the test material are unclear.

No effect of the test material at dose levels ≤1000 mg/kg/day was observed on pregnancy indices, uterine implantation data, foetal sex ratios, or foetal external, visceral, or skeletal examinations. Lower foetal body weights (8 % lower than controls) were observed at 1000 mg/kg/day. Thus, the No-Observed-Adverse-Effect Level (NOAEL) for maternal and developmental toxicity was 300 mg/kg/day. There were no teratogenic effects recorded in the study.