Registration Dossier

Administrative data

Endpoint:
toxicity to reproduction: other studies
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Guideline study conducted under GLP. Preferred study for this SIDS endpoint. The method of analysis involved derivatization. This method only measures the amount of the alkyltin moiety, MMT, present and does not identify the other ligands attached to the tin. Currently there is no analytical method available that can quantify the actual named substance, i.e., the entire organotin compound with its associated chloride ligand.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD 421
GLP compliance:
yes
Type of method:
in vivo

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Trichloromethylstannane
- Molecular formula (if other than submission substance): CH3Cl3Sn
- Molecular weight (if other than submission substance): 240.8 g/mol
- Smiles notation (if other than submission substance): Cl[Sn](Cl)(Cl)C
- InChl (if other than submission substance): IUPAC : Stannane, trichloromethyl
- Structural formula attached as image file (if other than submission substance): see Fig.
- Physical state: liquid
- Analytical purity: 82.85%
- Impurities (identity and concentrations):
- Composition of test material, percentage of components:
Alkyl group distribution % (mass/mass)
Monomethyltin trichloride 82.85
Dimethyltin dichloride 9.29
Trimethyltin chloride 0.02
Tin tetrachloride 4.68
Me2ClSnCH2SnCl3 0.59
MeCl2SnCH2SnCl3 1.78
Cl3SnCH2SnCl3 0.80

- Lot/batch No.: 82420
- Expiration date of the lot/batch: December 1, 2003
- Storage condition of test material: <-18 deg C, in the dark

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: (P) x 15-16 weeks; (F1) x wks
- Weight at study initiation: (P) Males: 113.9 - 13.8 g (mean 126.3g); Females: 178.8 - 213.5 g (mean 199.7 g); (F1) Males: x-x g; Females: x-x g
- Fasting period before study: not reported
- Housing: 3 or 4 per group per cage; during gestation and lactation they w ere housed individually
- Use of restrainers for preventing ingestion (if dermal): not applicable
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 13 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3 deg C
- Humidity (%): 30% not exceeding 70%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 hours light/ 12 hours dark


IN-LIFE DATES: From: May 20, 2003 To: June 26- July 14, 2003

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test substance was incorporated into the basal diet by mixing in a mechanial blender


DIET PREPARATION
- Rate of preparation of diet (frequency): shortly before the start of the studies and every 6 weeks thereafter
- Mixing appropriate amounts with (Type of food): Rat and Mouse No. 3 Breeding Diet, RM3
- Storage temperature of food: stored in a freezer until use


VEHICLE- no vehicle
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
ANALYTICAL METHODS: GC-MS was used to determine the achieved  concentration, homogeneous distribution and stability of the test  substance in diet samples.  The method of analysis involved  derivatization.  This method only measures the amount of the alkyltin  moiety, MMT, present and does not identify the other ligands attached to  the tin.  Currently there is no analytical method available that can  quantify the actual named substance, i.e., the entire organotin compound  with its associated chloride ligand.
From each diet sample, 2.0 g was transferred into a 50 mL Greiner tube. An aliquot of the internal standard solution was added. Subsequently, methanol, acetate buffer solution, 20% aqueous NaBEt4 solution and hexane were added to each sample and this mixture was shaken and heated to 60 deg C. Prior to GC/MS analysis, the hexane layer was washed with 2 mol/l HCl in order to remove the ethylboron compounds. The concentration of each test substance in feed was determined by GC-MS analysis of the hexane extracts.
Duration of treatment / exposure:
2 consecutive weeks during the premating period,  daily during gestation (up to 26 days) and up to euthanasia at or shortly after postnatal day (PN 4)
Frequency of treatment:
daily
Duration of test:
13-week
Doses / concentrations
Remarks:
Doses / Concentrations:
30, 150, 750 mg trichloromethylstannane/kg diet
Basis:
nominal in diet
No. of animals per sex per dose:
10 females per dose level for satellite group
Control animals:
yes, concurrent no treatment
Details on study design:
The test substance was administered at constant concentrations in the diet for 13 consecutive weeks. The test substance was administered via the diet two weeks premating, during gestation and up to euthanasia at or shortly after postnatal day 4.
Statistics:
- Test substance analysis: Homogeneity: one way analysis of variance (Anova) using the sample  location (1-5) as grouping factor.  The test substance was considered to  be homogeneously distributed in the diets if p > 0.01 and/or if the  relative standard deviation (RSD) between the sample means was less than  or equal to 15%. Stability: one way analysis of variance (Anova) using time as grouping  factor.  The test substance was considered to be stable in the diets if p  > 0.01 and/or if the mean concentration on the last day was between 80%  and 120% of the mean concentration on the first day (t =0). Achieved concentration: for each concentration level, the mean of the  concentrations, as measured in the diet samples used for the assessment  of the homogeneity, was considered to represent the achieved  concentration.  The content of the test substance in the diet was  considered to be 'close to intended' if the mean measured concentration  was between 80% and 120% of the intended concentration. - Body weight: one way analysis of covariance (covariate: body weight on  day 0) followed by Dunnett's multiple comparison tests. - Food consumption and food efficiency: one way analysis of variance  (Anova) followed by L.S.D. tests. - Fisher's exact probability test was used to evaluate the number of  mated and pregnant females and females with live pups.  Number of  implantation sites, live and dead pups were evaluated by Kruskal-Wallis  non-parametric analysis of variance followed by the Mann-Whitney U-test. - The litter was used as the statistical unit for calculation of fetal  values. - Histopathological changes: Fisher's exact probability test. All tests were two-sided.  Probability values of p<0.05 were considered  significant.

Results and discussion

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
750 mg/kg diet
Sex:
male/female
Basis for effect level:
other: Based on the absence of  statistically significant reproductive or developmental effects observed  after mating of females of the 750 mg/kg satellite group with male  animals of the main study,(= 49.7 mg/kg bw/day in males and 26.5 - 45.8 mg/kg females)
Dose descriptor:
NOAEL
Effect level:
150 mg/kg diet
Sex:
male/female
Basis for effect level:
other: Based on the incidences of missing pups in  the 750 mg/kg group
Dose descriptor:
NOAEL
Effect level:
150 mg/kg diet
Sex:
female
Basis for effect level:
other: Based on the effects observed on body weight  and food consumption in the 750 mg/kg group, 150 mg/kg diet (equivalent  to 6.2 - 11.7 mg MMTC/kg bw/day in females) 

Observed effects

NOAEL (fertility and developmental effects): 750 mg test substance/kg diet
NOAEL (postnatal toxicity): 150 mg test substance/kg diet
NOAEL (maternal toxicity): 150 mg test substance/kg diet

Any other information on results incl. tables

TEST SUBSTANCE INTAKE:
The test substance intake of the female animals of the 30, 150 and 750  mg/kg dose groups was respectively:
Premating period
days 0-7: 1.8, 9.0 and 44.5 mg/kg bw/day
days 7-14: 1.8, 8.8 and 43.9 mg/kg bw/day
Gestation period
GD 0-7: 1.9, 9.6 and 44.5 mg/kg bw/day
GD 7-14: 2.0, 9.6 and 45.8 mg/kg bw/day
GD 14-21: 1.2, 6.2 and 35.9 mg/kg bw/day
Lactation period
PN 1-4: 1.7, 11.7 and 26.5 mg/kg bw/day
Analysis of the test substance in diet samples revealed that the test  substance dose was close to the nominal level for all diets.  Mean  measured concentrations ranged from 96 to 113% of nominal concentrations.
Homogeneity: The test substance was considered to be homogenously  distributed in all diets.
Stability: The test substance was considered to be stable in the diets  upon storage at room temperature for 7 days, and upon storage at < -18°C  for 6 weeks. 
MATERNAL TOXIC EFFECTS: 
- Mortality and day of death: One animal of the 750 mg/kg group was found  dead on GD 22 (i.e. 37 days after the start of exposure).
- Pre-coital time: comparable among the control and the treated groups.
- Mating index: 90 -100%
- Number of pregnant females per dose level: 9, 8, 9 and 8 for the control, 30, 150 and 750 mg/kg groups, respectively.
- Female fecundity index: comparable among the control and the treated  groups.
- Female fertility index: comparable among the control and the treated  groups.
- Male fertility index: comparable among the control and the treated  groups.
- Number aborting: No data available.
- Number of resorptions: No data available.
- Mean number of implantations: 11.2 (control group), 10.8 (30 mg/kg),  11.6 (150 mg/kg), 10.5 (750 mg/kg).
- Gestation index: 89, 100, 100 and 88% in the control, 30, 150 and 750  mg/kg groups, respectively.
- Live birth index: 98, 97, 100 and 96% in the control, 30, 150 and 750  mg/kg groups, respectively.
- Number of pups born (number of litters): 90(8), 86(8), 99(9) and 50(7)  for the control, 30, 150 and 750 mg/kg groups, respectively.
- Number of stillborn pups (number of litters): 2(1), 3(2), 0 and 2(2)  for the control, 30, 150 and 750 mg/kg groups, respectively.
- Post implantation losses [total implantation sites minus total live births at the first observation]: 13(18.6%), 16(15.3%), 5(4.7%) and  36(42.9%) for the control, 30, 150 and 750 mg/kg groups, respectively.
Interpretation of these data was complicated by the incidence of missing  pups across groups.  A variable incidence of pups "missing" after birth  was recorded.  The incidence of missing pups was statistically significantly higher than controls in the high-dose group, statistically significantly lower than controls in the mid-dose group, and not  statistically different than controls in the low-dose group.  The missing  pups were presumed to have been cannibalized by the dams, but it is not  known if the missing pups were alive or dead.  It is also not known if  some pups were cannabilized prior to being counted for litter size.  This  could account for the slightly lower number of recorded live births and  the slightly higher post-implantation loss in the high-dose versus  controls.

The reason for missing pups can not be determined on the basis of the data within the study.  Missing pups could be due soley to a toxic behavioral effect on dams which caused a lack of, or abnormal, nurturing.   It is clear that no malformations were noted at any observation point  for any of the missing pups and no overt behavioral effects were noted,  however some other effect could have caused the dam to eat them.
- Number of corpora lutea: No data available.
- Duration of Pregnancy: 21 -23 days
- Body weight: Increased body weight change from GD 7-14 of the females of the 30 mg/kg group, which was considered a chance finding.  Mean body  weights (change) of the females were similar among the control, 30 and 150 mg/kg group during the entire study.
Mean body weight (changes) between PN 1-4 of the 750 mg/kg group was decreased; however, no statistical significance was reached for these  findings.
- Food consumption: Food consumption of the female animals of the 750  mg/kg group was decreased (not statistically significantly) during the  lactation period.  During the premating and gestation periods food consumption of the females was similar in the control, 30, 150 and 750  mg/kg groups.
- Description, severity, time of onset and duration of clinical signs: No data available.
- Hematological findings incidence and severity: No data available.
- Clinical biochemistry findings incidence and severity: No data  available.
- Gross pathology incidence and severity: No treatment related changes were observed.  The animal found dead on day 37 was necropsied. Findings included yellow patches on the liver, yellow appearance of the small  intestines, haemorrhagic discharge from the vagina and a haemothorax.   The haemothorax was considered to be the probable cause of death.  Most probably the haemothorax was caused by severe dystocia, since at necropsy the uterus contained 12 dead fetuses.
- Organ weight changes: Mean absolute and relative uterus, ovary and thymus weights were similar in all groups.
- Histopathology incidence and severity: No treatment related histopathological changes were observed in the uterus, ovary and thymus of the female animals of the control and 750 mg/kg groups.
FETAL DATA:
- Litter size:  The mean number of pups delivered per litter amounted to 11.2, 10.8, 11.0 and 7.1 for the control, 30, 150 and 750 mg/kg groups, respectively.
- Litter weight: The mean pup weights and pup weight changes were similar in the treated groups when compared to the control group.
- Pup mortality: 2.2, 3.5, 0 and 4% for the control, 30, 150 and 750 mg/kg groups, respectively (PN 1); 16, 25, 3 and 65% for the control, 30, 150 and 750 mg/kg groups, respectively (PN 4).
- Number viable: The viability index (PN 1-4) was 84, 75, 97 and 35% in the control, 30, 150 and 750 mg/kg groups, respectively.
- Number live pups per litter: 11.0, 10.4, 11.0 and 6.9 for the control, 30, 150 and 750 mg/kg groups, respectively (PN 1); 10.6, 7.8, 10.7 and  4.2 for the control, 30, 150 and 750 mg/kg groups, respectively (PN 4).
- Sex ratio: No difference was observed in the sex ratio between the groups.
- Postnatal growth: No data available; screening study.
- Postnatal survival: No data available; screening study.
- Grossly visible abnormalities: In the 30 mg/kg group, the number of cold  pups was statistically significantly increased on PN 1 and 4.  In  addition on PN 4, the number of runts, pale pups and pups with no milk in  the stomach was statistically significantly increased in the 30 mg/kg  group.  In the 150 mg/kg group the number of pale pups on PN 4 was statistically significantly increased.  In the 750 mg/kg group the number of cold pups was statistically significantly increased on PN 1.  As most effects were observed in the 30 mg/kg group only, these effects were not  considered to be treatment related.  Macroscopic observation of the stillborn pups revealed no abnormalities of the pups.
- External abnormalities: No data available; screening study.
- Soft tissue abnormalities: No data available; screening study.
- Skeletal abnormalities:  No data available; screening study.

Applicant's summary and conclusion

Executive summary:

The toxicity of trichloromethylstannane [CAS # 993-16-8] in Wistar rats was examined using continuous administration via the diet for 13 consecutive weeks (OECD Test Guideline 408). In satellite groups of female rats a reproduction/developmental screening test (OECD Test Guideline 421) was performed to provide initial data on possible reproductive and developmental effects of trichloromethylstannane. The main study used four groups of 10 rats/sex (13 -week study) and the satellite study used four groups of 10 female rats (reproduction/developmental screening study). For both studies the control group was kept on control diet and three test groups received experimental diets containing 30, 150 and 750 mg/kg [ppm] of the test substance. The dose levels used in both studies were based on the results of a preceding dose range finding study.

 

In the satellite study female rats were fed their respective test diets beginning 2 weeks prior to the mating period, and continued on test diets through mating, gestation, and up to PN 4 or shortly thereafter. Male rats from the main study were mated after a premating period of 10 weeks with female rats of the satellite groups which were fed the same dose of test diets.

Clinical observations, growth, food consumption, food conversion efficiency, neurobehavioural testing, ophthalmoscopy, haematology, clinical chemistry, renal concentration test, urinalysis, organ weights and gross examination at necropsy, microscopic examination of various organs and tissues and assessment of various reproductive and developmental parameters were used as criteria for detecting the effects of treatment.

 

Results

Satellite study – Reproduction/developmental screening study OECD Test Guideline421

 

The calculated doses during the pre-mating, gestation and lactation periods for the females receiving 30, 150, or 750 mg/kg trichloromethylstannane in the diet ranged from 1.2-2.0, 6.2-11.7 and 26.5-45.8 mg/kg body weight, respectively.

 

During the study one animal of the 750 mg/kg group was found dead on GD22. Most probably haemothorax caused by dystocia was the cause of death. No other mortalities or treatment-related clinical signs were observed in the female animals of the satellite groups.

 

Mean body weight, body weight change and food consumption was similar in the control and the 30 and 150 mg/kg groups. Mean body weight on PN 4 and mean body weight change PN 1-4 of the 750 mg/kg group was decreased, although not statistically significantly.

 

During the premating and gestation periods, mean food consumption of the female animals was similar in the control, 30 and 150 mg/kg groups. During the lactation period food consumption of the 750 mg/kg group was decreased (not statistically significantly).

 

No treatment-related effects on reproduction and development of the pups were observed in the 30 and 150 mg/kg groups.

In the 750 mg/kg group, a number of changes were seen, indicative of a reproductive and developmental effect. One animal was found dead on GD22 (death was most probably caused by haemothorax caused by dystocia), post-implantation loss was 43%, the number of pups delivered was 7.1 versus 11.2 in the control group, 3 females lost all pups between PN 1-4, and pup mortality between PN 1-4 in the 750 mg/kg group was 65% versus 16% in the control group.

 

No treatment-related effects on pup weight and pup abnormalities were observed.

 

Absolute and relative organ weights (uterus, ovary and thymus) were similar among the groups. At microscopic examination of these organs no treatment-related histopathological changes were observed.

 

Conclusions

Satellite study - Reproduction/developmental screening study (OECD Test Guideline 421)

Based on reproductive and developmental effects (decreased number of pups delivered and increased post implantation loss and pup mortality) observed after mating of female animals of the 750 mg/kg satellite group with male animals of the main study, the mid-dose level 150 mg Trichloromethylstannane/kg diet (equivalent to 9.8 mg/ kg body weight/day in males and 6.2-11.7

mg/kg body weight for females) can be considered as a NOAEL for fertility and developmental

effects.

 

Based on the effects on body weight and food consumption in the 750 mg/kg group, 150 mg

Trichloromethylstannanel kg diet (equivalent to 6.2-11.7 mg/kg body weight/day) can be considered as a NOAEL for maternal toxicity.

 

Qverall conclusion:

The NOAEL for general sub-chronic toxicity and maternal toxicity, fertility and developmental effects was placed at 150 mg trichloromethylstannane per kg diet.