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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
A teratological assessment of four trihalomethanes in the rat
Author:
Ruddick JA, Villeneuve DC, Chu I, Valli VE.
Year:
1983
Bibliographic source:
Journal of Environmental Science and Health, Part B 1983; 18(3):333-349

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
No guideline was provided in the study and the age of the study (1983) predates the OECD 414 guideline. The study does; however, cover many of the endpoints included in the OECD 414 guideline, although exposure covered days 6 to 15 of gestation rather than 5 to 15 specified in the OECD 414 guideline.
GLP compliance:
no
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Bromoform
EC Number:
200-854-6
EC Name:
Bromoform
Cas Number:
75-25-2
Molecular formula:
CHBr3
IUPAC Name:
bromoform
Test material form:
liquid
Details on test material:
- Name: Bromoform
- Source Aldrich Chemical Company, Milwaukee, Wisconsin.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Biobreeding Ltd., Montreal, Quebec.
- Age at study initiation: Not stated.
- Weight at study initiation: 150-175 g
- Fasting period before study: Not stated.
- Housing: Each female rat was caged individually following pairing with a sire.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 40-60%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 hours light /12 hours dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
Pregnant rats were administered the test substance daily by oral intubation beginning on the 6th day of gestation and continuing until the 15th day of gestation.
Analytical verification of doses or concentrations:
no
Details on mating procedure:
Female rats were then paired overnight with a proven sire and the day on which sperm were seen in a vaginal smear was considered day 1 of gestation. The pregnant rats were then randomly divided into 4 groups (including the control) with each group having 15 rats.
Duration of treatment / exposure:
Rats were adminstered test substance from day 6 to day 15 day of gestation.
Frequency of treatment:
Daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Vehicle Control
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
200 mg/kg bw/day (nominal)
No. of animals per sex per dose:
15 rats per dose group.
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:

BODY WEIGHT: Yes
- Time schedule for examinations: The females were weighed on day 1 and days 6-15 of gestation as well as before and after caesarean section on day 22.


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 22
- Organs examined: Maternal rats were subject to gross pathological assessment and the liver, hear, brain spleen and one kidney were removed and weighed. The following tissue samples were taken from each animal and fixed: brain, heart, pituitary, thyroid, parathyroid, thymus, lungs, trachea and bronchi, bronchial node, liver, kidney, adrenal gland, spleen, skeleton muscle, peripheral nerve, salivary gland, skin, bone marrow, ovaries, uterus and bladder.

- OTHER EXAMINATIONS: Materal blood samples were taken fro haematology, blood chemistry. A liver sampels was taken following gross pathology for analysis of liver enzyme activiity.


Fetal examinations:
At sacrifce on gestation day 22 foetuses were removed, weighed individually and examined for viability and external malformations.

Two pups from each dam were fixed in formalin for histological evaluation. Approximately two-thirds of the remaining live foetuses from each litter were placed in absolute ethanol for future staining of the skeleton with Alizarin red and subsequent examination for skeleton abnormalities. The rest of the foetuses were fixed in Bouin's fluid and studied for visceral changes.

Statistics:
An analysis of variance was carried out on all measurements where applicable, and when a significant difference occurred (P<0.05), Duncan's Multiple Range Test (SPSS version 8.1) was applied to determine the group or groups that were different from the control.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
not specified
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
No efffects on measured liver enzyme activity was reported (i.e. it was not positively stated that there had been no changes and it is assumed that no effects occurred).

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
effects observed, treatment-related
Description (incidence and severity):
There was some of evidence of dose related increase in sternebra aberrations and observations of interparietal anomalies in foetuses. These were considered to be caused by a foetal toxicity effect.
Changes in pregnancy duration:
not specified
Changes in number of pregnant:
no effects observed

Effect levels (maternal animals)

Key result
Dose descriptor:
NOEL
Effect level:
200 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: No effects observed in maternal animals.

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
effects observed, treatment-related
Changes in sex ratio:
not specified
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
effects observed, treatment-related
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
There was some of evidence of dose related increase in sternebra aberrations and observations of interparietal anomalies in foetuses. These were considered to be caused by foetal toxic and not indicative of teratogenicity.
Visceral malformations:
no effects observed
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Mild hilar dilation was observed in the kidneys of some treated foetuses but this was considered to spontaneous in nature.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: Evidence of foetal toxicity.

Fetal abnormalities

Key result
Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
The test substance was not considered teratogenic to Sprague-Dawley rats. The study No Observed Effect Level (NOEL) for teratogenicity was 200 mg/kg bw/day (highest dose administered). There was some evidence of a foetal toxicity caused by the test substance. The study No Observed Adverse Effect Level (NOAEL) for foetal toxicity was considered to be 100 mg/kg bw/day. 
Executive summary:

Introduction

The teratogenic potential of the test substance toSprague-Dawley ratswas assessed using a study design that pre-dated the OCED 414 guideline, but which nevertheless covered many of the endpoints included in the guideline. 

 

Method

 Test substance was administered in corn oil by gavage to pregnant Sprague-Dawley rats from day 6 to day 15 of gestation at doses of 50, 100 or 200 mg/kg/day. A control group was administered corn oil only. Each group consisted of 15 rats and each rat was housed individually with free access to food and water.

On day 22 of gestation rates sacrificed and their viscera including the uteri were examined for pathological changes. The foetuses were removed, weighed individually and examined for viability and external malformations. Two pups from each dam were fixed in formalin for histological evaluation. Approximately two-thirds of the remaining live foetuses from each litter were placed in absolute ethanol for future staining of the skeleton with Alizarin red and subsequent examination for skeleton abnormalities. The rest of the foetuses were fixed in Bouin's fluid and studied for visceral changes.

Maternal rats were subject to gross pathological assessment and the liver, hear, brain spleen and one kidney were removed and weighed. The following tissue samples were taken from each animal and fixed: brain, heart, pituitary, thyroid, parathyroid, thymus, lungs, trachea and bronchi, bronchial node, liver, kidney, adrenal gland, spleen, skeleton muscle, peripheral nerve, salivary gland, skin, bone marrow, ovaries, uterus and bladder.

 

Maternal blood samples were taken for haematological and clinical chemistry assessments.

Following gross pathological examination a liver sample was taken from maternal animals for liver protein aniline hydroxylase (AH) and aminopyrine demethylase (APDM) activity.

Results

Body and Organ Weights: There were no effects on maternal weight, liver, kidney or spleen weights resulting from treatment with test substance.

Foetal effects: There were no effects or litter size, incidence of resorptions, foetal weight or incidence of visceral anomalies resulting from treatment with test substance.

Histopathology: There were no dose related histological, changes in mother or foetuses resulting from treatment with test substance. 

Haematology: Maternal haematological parameters assessed showed no affects from treatment with test substance.

Clinical Chemistry: Maternal clinical chemistry parameters assessed showed no affects from treatment with test substance.

Liver Enzymes: Maternal liver enzyme parameters assessed showed no affects from treatment with test substance.

Foetal toxicity: There was some of evidence of dose related increase in sternebra aberrations and observations of interparietal anomalies in foetuses at 100 and 200 mg/kg bw/day. These were considered to be the result of a foetal toxicity effect and not teratogenicity related.

 

Conclusion

The test substance was not considered teratogenic to Sprague-Dawley rats. The study No Observed Effect Level (NOEL) for teratogenicity was 200 mg/kg bw/day (highest dose administered).  

There was some evidence of a foetal toxicity caused by the test substance. The study No Observed Adverse Effect Level (NOAEL) for foetal toxicity was considered to be 100 mg/kg bw/day.