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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
one-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 26 - December 11, 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Qualifier:
according to
Guideline:
other: Health Effects guidelines, OPPTS 870.3550, Reproduction/ Developmental Toxicity Screening Test. EPA 712-C-00-367, July 2000
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: liquid
Details on test material:
Name: Formaldehyde, reaction product with ethylenediamine
CAS No.: 84066-92-2
Batch No.: PK280-61
Purity: 100%

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
The test item and vehicle were administered at a single dose to the animals by oral gavage. The application volume for all groups was 5 mL/kg body weight.
For each animal the individual dosing volume was calculated on the basis of the body weight most recently measured.
The following doses were evaluated:
Control: 0 mg/kg body weight
Low Dose: 100 mg/kg body weight
Medium Dose: 300 mg/kg body weight
High Dose: 700 mg/kg body weight
Details on mating procedure:
Mating was performed using a ratio of 1:1 (male to female). The vaginal smear of the females was checked every morning after the start of the mating period to confirm the pregnancy. The day of the vaginal plug and/or sperm was considered as day 0 of gestation. Females with unsuccessful mating will be allowed to mate with other male of the same group.
The cages were arranged in such a way that possible effects due to cage placement were minimised.
Duration of treatment / exposure:
The animals were treated with the test item formulation or vehicle on 7 days per week for a period of 54 days, i.e. during 14 days of pre-mating and 14 days of mating in both males and females, during the gestation period and up to post-natal day 3 in females. Males were dosed after the mating period until the minimum total dosing period of 28 days were completed.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 300, 700 mg/kg bw
Basis:
nominal conc.
No. of animals per sex per dose:
80 animals (40 males and 40 females) were included in the study (10 male and 10 female animals per group).
Control animals:
yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:
Clinical Observations
General clinical observations were made at least once a day, preferably at the same time each day and considering the peak period of anticipated effects after dosing. The health condition of the animals was recorded. Twice daily all animals were observed for morbidity and mortality except on weekends and public holidays when observations were made once daily.
Females showing signs of abortion or premature delivery prior to the scheduled scarification of the animals were sacrificed and subjected to a thorough macroscopic examination.
Once before the first exposure, and at least once a week thereafter, detailed clinical observations were made in all animals outside the home cage in a standard arena. Clinical observations included spontaneous activity, lethargy, recumbent position, convulsions, tremors, apnoea, asphyxia, vocalisation, diarrhoea, changes in the skin and fur, eyes and mucous membranes (salivation, discharge), piloerection and pupil size. Changes in gait, posture, response to handling as well as the presence of clonic or tonic movements, stereotypes, difficult or prolonged parturition or bizarre behaviour were recorded.
Litter observations:
Litter Observations
The duration of the gestation was recorded and was calculated from day 0 of the pregnancy. Each litter was examined as soon as possible after delivery of the dam to establish the number and sex of pups, stillbirths, live births, runts and the presence of gross abnormalities.
Live pups were counted and sexed and litters weighed within 24 hours of parturition (day 0 post-partum) and on day 4 post-partum. Live pups were identified by writing numbers on the back with the help of a permanent marker or by tattooing. In addition to the observations of parent animals, any abnormal behaviour of the offspring was recorded.

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Increased salivation post-dose, with greater incidence at higher dose levels.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Although females increased weight more than controls, th high dos group males showed reduced body wegith gain.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Reduction in food uptake with high dose group animals
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
not specified

Effect levels (P0)

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

Target system / organ toxicity (P0)

Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
> 700 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability

Target system / organ toxicity (F1)

Critical effects observed:
no

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

No mortality occurred in the control or any of the dose groups during the treatment period of this study.

Several clinical findings occurred in C and treatment groups and some of them were increased observed in MD and HD group. In male animals, common findings, were e.g. moving the bedding (10/10 HD; 10/10 MD), moderate salivation (10/10 HD; 2/10 MD), and severe salivation (6/10 HD; 1/10 MD).

In female animals, some of the most important findings were moving the bedding (10/10 HD; 10/10 MD), severe salivation (4/10 HD; 1/10 MD), and moderate salivation (10/10 HD; 4/10 MD).

In HD male animals a statistical significant decreased body weight could be found at pre-mating day 14, mating- and post-mating day 7, as well as at terminal sacrifice. During pre-mating days 1-7 as well as the overall periods pre-mating day 1- post-mating day 14 and pre-mating day 1 – terminal sacrifice, the body weight gain was significantly decreased for male HD animals.

Body weight gain of female animals was significantly increased during pre-mating days 7-14 as well as decreased during gestation days 0-7, and gestations days 0-20.

In correlation to the body weight and body weight change, the food consumption in male HD group was significantly decreased between pre-mating days 1-7 and pre-mating days 7-14 when compared to C group.

Significant decreased food consumption was observed for HD female animals, too. Between pre-mating days 1-7 and 7-14 as well as gestation days 7-14 a significant decrease in food consumption was observed when compared to C animals.

The total number of pubs born was reduced in tendency in the HD group. In particular 7.7 pubs (HD) and 9.6 pubs (C) were counted. In correlation to this, the number of female pubs was slightly reduced in HD group at PND 0, too. 3.7 (HD) and 5.3 (C) were counted. In HD group, 3 still births were observed at PND0 – two for female No. 71 and one for female No. 76. At PND 4, the number of live pubs was still reduced by tendency. 7.0 (HD) and 9.6 (C) were evaluated. Furthermore, number of female pubs at PND 4 was slightly reduced in HD group (3.5) when compared to C group (5.3).

Litter mean weight was slightly but not significantly reduced at PND 4 in HD group (10.24 (HD) as well as 11.34 (C) were counted). Due to the decreased number of pubs, the total litter mean weight was decreased in HD group, too. At PND 0, 45.6 g (HD) were measured in comparison to 61.52 g (C). The female litter weight was significantly decreased (p<0.05) at PND 0. In particular, 21.33 (HD) and 25.29 (C) were measured. At PND 4, the total lighter weight was reduced, too. HD animals exhibited 76.86 g and C animals exhibited 104.85 g. Again, a statistical significance was observed regarding the difference of female litter weight at PND 4. 37.24 g (HD) and 60.17 g (C) were evaluated.

No treatment-related effect was observed during the pre-coital interval or during the duration of gestation when compared with the control group. The values were comparable between the groups. All pregnancies resulted in normal births.

Percentage of pre-implantation loss was increased in HD group (26.44 %) when compared to C group (15.33 %). Furthermore, percentage of post-implantation loss was increased in HD group (16.53 %), too, when compared to C group (5.83 %). A tendency to an increased percentage of post-implantation loss was recognized for MD group (9.79%).

No test item related change was found for viability index, copulation index, delivery index, and fertility index.

No significant effect on survival of the pups from PND 0 to PND 4 was observed in any treatment group when compared with controls.

No treatment-related gross external findings were observed in any of the treated groups. Few incidences of external findings were observed in all groups which were considered to be spontaneous and not related to the test item.

Few specific gross pathological changes were recorded for the male and female animals in C, LD, and MD groups which were not considered to be treatment-related.

In male animals, absolute weights of prostates (including seminal vesicles and coagulating glands) were significantly decreased in LD group and HD group. MD animals exhibited a non significant decreased prostate weight. This could be confirmed for relative prostate weights (plus seminal vesicles and coagulating glands).

Relative weights of testes were significantly increased in male HD animals. This could be also seen by tendency for absolute testes weights.

No test item-related macroscopic or histopathological lesions were noted in male and female reproductive organs in this study.

One female treated at 100 mg/kg/day did not show any indication of recent pregnancy at terminal sacrifice. Histologically, physiological sexual cycling was observed in this animal and the unsuccessful mating of this isolated rat was therefore considered to be a chance event.

Applicant's summary and conclusion

Conclusions:
On the basis of this reproduction/ developmental toxicity screening test with Formaldehyde, reaction product with ethylenediamine in male and female Wistar rats with dose levels of 100, 300, and 700 mg/kg body weight day the following conclusions can be made:
In HD group, increased clinical symptoms were recognized for male and female animals which display discomfort of the animals. Furthermore, a decreased body weight development was recognized for male and female HD animals, which was confirmed by decreased food consumption. The total number of pubs born was reduced in HD group. In this regard, the total litter weight was decreased in HD group at PND0. An increased percentage of pre- and post-implantation loss was also recognized in HD animals. In MD animals, clinical symptoms were recognized which are attributed to animal´s discomfort. Since no other symptoms for female and male animals were recognized, the No Observed Adverse Effect Level (NOAEL) for parentals animals was considered to be 300 mg/kg BW (MD group).
A tendency to an increased percentage of post-implantation loss was recognized for MD group. A test item relation is possible and cannot be excluded. Hence, the NOAEL for reproductive/developmental toxicity is assumed to be 100 mg/kg (LD group).