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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:
19 September 2009 to 5 January 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study according to internationally recognised guideline
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder

Test animals

Species:
rat
Strain:
other: Crl:CD(SD
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: a well established supplier
- Age at study initiation: approximately 65 days
- Weight at study initiation: 226 to 250 g
- Housing: P2000 or 2154 cages with solid flooring. Wood shavings (Lignocel type 3-4 wood flakes) were supplied as bedding material. In the mating period, the animals were maintained in RB3 modified cages (gridded flooring), suspended over trays covered with absorbent paper which was changed at appropriate intervals
- Diet: free access to standard rodent diet
- Water: free access to potable water taken from the public supply
- Acclimation period: : five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 23°C
- Humidity (%): 40 to 70%
- Air changes (per hr): not applicable (each animal room was kept at positive pressure with respect to the outside by its own supply of filtered fresh air, which was passed to atmosphere and not re-circulated).
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 28 October 2009 (animal arrival) To: 26 November 2009 (necroscopy)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 0.5% w/v carboxymethylcellulose sodium salt
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Justification for use and choice of vehicle (if other than water): not reported
- Concentration in vehicle: 0,10,50 &100 mg/ml
- Amount of vehicle (if gavage): 10ml
- Purity: 0.5% w/v carboxymethylcellulose sodium salt.

All formulations were prepared freshly each week and stored refrigerated (approximately 4°C) and allowed to reach room temperature before dosing.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Before treatment commenced, the suitability of the proposed mixing procedure was determined and specimen formulations were analysed to assess the homogeneity and stability of the test material in the liquid matrix. Specimen formulations were prepared at concentrations of 1.0 and 100 mg/mL.

Samples of each formulation (4 x 1 mL) prepared for administration in the first and last weeks of the dosing period were taken and two assays from each group were analysed for achieved concentration of the test substance. A representative sample of test formulation was accurately weighed and dissolved in a suitable volume of diluent. The extract was diluted using diluent to provide a solution containing BPS-MAE at an expected concentration within the range 2 μg/mL to 4 μg/mL. The concentration of BPS-MAE in the final solution was quantified by high performance liquid chromatography.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1:1
- Length of cohabitation: 2 days
- Proof of pregnancy: vaginal plug and sperm in vaginal smear
Duration of treatment / exposure:
Day 1 to Day 19 (after mating)
Frequency of treatment:
Once each day at approximately the same time each day
Duration of test:
19 days
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 10,50, 100 mg/mL
Basis:
nominal conc.
dose volume 10mL/kg
No. of animals per sex per dose:
22 females at each dose level
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: results from a preliminary study, in which animals receiving the highest dose only, showed slightly low bodyweight gain following daily oral gavage administration from Day 1 to 19 of gestation. Embryo-fetal survival, growth and development were unaffected by treatment. It was considered that the same doses, 0, 100, 500 and 1000 mg/kg/day and the same treatment regimen was suitable for main study.

- Rationale for animal assignment (if not random): Females were allocated to group and cage position in sequence of mating, thus ensuring that animals mated on any one day were evenly distributed amongst the groups.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean diet consumption calculated as g food/animal/day for the periods 0-2; 3-5; 6-9; 10-13; 14-17; 18-19

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: Any abnormality in the appearance or size of any organ and tissue was recorded. The gravid uterus, including ovaries, was weighed prior to dissection.

OTHER: All external features and orifices were examined visually + full macroscopic examination of the tissues.
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 (external examination)
Statistics:
Statistical analysis were applied where there was indication of possible meaningful intergroup differences. All statistical analyses were carried out using the individual animal (or litter) as the basic experimental unit.

Statistical tests used for bodyweight, food consumption, gravid uterus weight, corpora lutea, implantations, live young, placental, litter and fetal
weights data:

1) A parametric analysis was performed if Bartlett's test for variance homogeneity (Bartlett 1937) was not significant at the 1% level.
2) A non-parametric analysis was performed if Bartlett's test was still significant at the 1% level following both logarithmic and square-root transformations.

For gravid uterus weight, corpora lutea, implantations, live young, placental, litter and fetal weights data, if 75% of the data (across all groups) were the same value, Fisher’s Exact tests (Fisher 1973) were performed.

Pre/post implantation loss and sex ratio were analysed by generalised mixed linear model with binomial errors, a logit link function and litter as a random effect (Lipsitz et al 1991). Each treated group was compared to control using a Wald chi-square test.

For resorptions, each treated group was compared to control by exact Wilcoxon rank sum test (Wilcoxon 1945).
Indices:
Mean numbers of corpora lutea, implantations, male, female and total live young, early, late and total resorptions, mean sex ratio (percentage male), or pre and post implantation loss.
Historical control data:
None reported

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No significant effects

Effect levels (maternal animals)

Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No significant effects

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: fetotoxicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
1000 mg/kg/day test substance is the maternal no-observed effect-level (NOEL) and the fetal no-observed-adverse-effect-level (NOAEL).