Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
a short-term toxicity study does not need to be conducted because exposure of humans via inhalation in production and/or use is not likely as based on the provided thorough and rigorous exposure assessment
other:
Justification for type of information:
Acute toxicity studies with rats are available for the oral, dermal and inhalation routes of exposure. The LD50 are 500 mg/kg bw and > 2,000 mg/kg bw for the oral and dermal route, respectively. The LC50 for the inhalation route is in the range between 1.04 - 5.12 mg/L.
Oral exposure appears to lead to more significant toxicity than exposure by dermal or inhalation routes.
Repeat toxicity testing is therefore considered to be more relevent if by the oral route
Cross-referenceopen allclose all
Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
June 11 - July 5, 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
GLP compliance:
yes
Test type:
standard acute method
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Route of administration:
inhalation
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
Air Supply: Filtered air was supplied by an air compressor (Powerex Model: SES05) to the spray atomization nozzle. Additional compressed mixing air supplied from the air compressor was introduced into the chamber to help uniformly distribute the test atmosphere by creating a vortex at the chamber inlet. Compressed airflow was measured with a Mass Flowmeter. Chamber airflow was monitored throughout the exposure period and recorded periodically
Duration of exposure:
< 4 h
Concentrations:
After establishing the desired generation procedures during pre-test trials, twenty healthy rats were selected for test and equally distributed (5 per sex/group) into two exposure levels. Exposure levels of 1.0 and 5.0 mg/L were selected for testing.
No. of animals per sex per dose:
10 Males and 10 Females. The females assigned to test were nulliparous and non-pregnant.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 1.04 - < 5.12 mg/L air
Based on:
test mat.
Exp. duration:
4 h

1.04 mg/L

The gravimetric and nominal chamber concentrations were 1.04 and 10.95 mg/L respectively. The mass median aerodynamic diameter was calculated to be 1.94 μm based on graphic analysis of the particle size distribution as measured with an ACFM Andersen Ambient Particle Sizing Sampler.

All animals survived exposure to the test atmosphere. Following exposure, all rats exhibited abnormal respiration. In addition, two males and one female showed signs of abdominal distention and/or ano-genital staining and reduced fecal volume. Ocular and nasal discharge and facial staining were also noted for this female. All five males and four female recovered from the above symptoms by Day 6. Although all rats lost body weight by Day 1 and/or through Day 3, all animals showed a weight gain over the course of the 14 day study. Apart from irregular respiration persisting through Day 14, the fifth female recovered from all other symptoms by Day 12 and showed improved health through weight gain by study termination (Day 14). No gross abnormalities were noted for any of the animals when necropsied at the conclusion of the 14-day observation period.

5.12 mg/L

The gravimetric and nominal chamber concentrations were 5.12 and 99.61 mg/L respectively. The mass median aerodynamic diameter was calculated to be 2.36 μm based on graphic analysis of the particle size distribution as measured with an ACFM Andersen Ambient Particle Sizing Sampler.

Following exposure to the test atmosphere, all rats exhibited clinical signs including abnormal respiration, hypoactivity, nasal and ocular discharge, ano-genital staining, tremors and/or an unthrifty appearance. One female was found dead on Day 1, three males and a second female died on Day 2 and a third female died by Day 3. The fifth male showed signs of opacity in one eye, a

Interpretation of results:
Category 4 based on GHS criteria
Remarks:
Migrated information
Conclusions:
Under the conditions of this study, the single exposure acute inhalation LC50 of the test substance is between 1.04 mg/L and 5.12 mg/L in male and female rats. Based on the results of this study, Formaldehyde, reaction product with ethylenediamine is assigned EC classification of risk phrase R-20 Harmful and meets the requirements of GHS Toxicity Category 4.
Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28 day oral - September 25 - October 30, 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Qualifier:
according to
Guideline:
EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
GLP compliance:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Route of administration:
oral: gavage
Vehicle:
water
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
28 days
Frequency of treatment:
Daily
Remarks:
Doses / Concentrations:
100 mg/kg bw/day
Basis:
nominal in water
Remarks:
Doses / Concentrations:
300 mg/kg bw/day
Basis:
nominal in water
Remarks:
Doses / Concentrations:
800 mg/kg bw/day
Basis:
nominal in water
No. of animals per sex per dose:
3 groups of test animals, one dose level per group, and a control group. The 4 groups comprised of 5 males and 5 females rats each one.
Control animals:
yes, concurrent vehicle
Observations and examinations performed and frequency:
- Body weight and food consumption: weekly.
- Clinical observations: at least once a day.
- Morbidity and mortality: twice daily, except on weekends and public holidays when observations were made once daily.
- Detailed cage side observations considering spontaneous activity, lethargy, recumbent position, convulsions, tremors, apnoea, asphyxia, vocalisation, diarrhoea, changes in skin and fur, eyes and mucous membranes (salivation, discharge), piloerection and pupil size were made outside the home cage in a standard arena once before the first administration and at least once a week thereafter.
- Ophthalmological examination, using an ophthalmoscope was made on all animals before the first administration and in the last week of the treatment period.
- Functional Observations: Once before the first exposure and once in the fourth week of exposure multiple detailed behavioural observations were made outside the home cage using a functional observational battery of tests.
- Haematology: Haematological parameters were examined at the end of the treatment prior to or as part of the sacrifice of the animals.
- Blood Coagulation: Coagulation parameters were examined at the end of the treatment prior to or as part of the sacrifice of the animals.
- Clinical Biochemistry: Parameters of clinical biochemistry were examined at the end of the treatment prior to or as part of the sacrifice of the animals.
- Urinalysis: A urinalysis was performed with samples collected from all animals prior to or as part of the sacrifice of the animals. Additionally, urine colour/ appearance were recorded.
Sacrifice and pathology:
- Pathology: One day after the last administration (study day 29) all surviving animals of the treatment period were sacrificed using anesthesia and subjected to a detailed gross necropsy which includes careful examination of the external surface of the body, all orifices and the cranial, thoracic and abdominal cavities and their contents.
- Organ Weight: The wet weight of the organs of all sacrificed animals was recorded as soon as possible. Paired organs were weighed separately. Organ weights of animals found dead or euthanised for animal welfare reasons were not recorded.
- Histopathology: The following organs were examined histopathologically after preparation of paraffin sections and haematoxylin-eosin staining: brain (cerebrum, cerebellum and pons), spinal cord, eye, liver, kidneys, adrenal glands, stomach, small and large intestines (including Peyer´s patches), thymus, thyroid glands, spleen, lung and trachea, mammary glands, skin.
Statistics:
A statistical assessment of the results of the body weight, food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test. These statistics were performed with GraphPad Prism V.5.01 (p<0.05 was considered as statistically significant).
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No mortality occurred in the control or any of the dose groups during the treatment period of this study. These clinical signs observed in MD and HD group were signs of discomfort which are – as individual findings – considered to be of toxicological relevance but not to be adverse.
Mortality:
mortality observed, treatment-related
Description (incidence):
No mortality occurred in the control or any of the dose groups during the treatment period of this study. These clinical signs observed in MD and HD group were signs of discomfort which are – as individual findings – considered to be of toxicological relevance but not to be adverse.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Slightly attenuated body weight gain. Not statistical significant.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Slight tendency towards attenuated food consumption. Not statistical significant.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Increase in: MCV, MCH, basophils and PLT.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Changes in ASAT and TBA.
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
High protein levels as well as a high specific gravity were found in one animal.
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Changes in brain, thymus, pituitary glands, adrenals, liver, spleen, ovaries and uteri.
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Kidney: mild or moderate tubular degeneration/regeneration at the inner cortex in all rats treated at 800 mg/kg/day. Stomach: esions indicative of a sustained local irritant effect of the test item formulation.
Histopathological findings: neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Stomach, as a consequence of local irritant effect of the test item formulation: erosions, ulcers, epithelial hyperplasia of the nonglandular mucosa, superficial hyperplasia/basophilia of the glandular mucosa.
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: The 300 mg/kg bw group showed signs of discomfort, but effects observed in the higher dose group were not confirmed at this dose.
Critical effects observed:
not specified
Conclusions:
NOAEL: 300 mg/kg bw/day.
Executive summary:

On the basis of this 28-Day Repeated Dose Oral Toxicity study with formaldehyde, reaction products with ethylenediamine in male and female Wistar rats with dose levels of 100, 300, and 800 mg/kg body weight day, the following conclusions can be made:

At 800 mg/kg BW increased clinical symptoms were found which indicate discomfort of the animals. Furthermore, decreased thymus weight as well as increased adrenal weight indicates stress of the animals, too. An additional tissue besides the stomach occurred probably due to the local irritant effect of the test item. Pathologically, test item-related lesions of toxicological significance were seen in the kidney and comprised mild or moderate tubular degeneration/regeneration at the inner cortex in all rats treated at 800 mg/kg/day. Hence, the dosage of 800 mg/kg BW is assumed to induce adverse effects within this study.

In MD group, clinical signs of discomfort were found. Furthermore, decreased thymus weights indicate stress, too. A non significant increase of TBA in serum is not confirmed by other liver markers or histopathology. Furthermore, histological analysis do not confirm the adverse kidney findings in this MD dose level. Hence, the MD dose is assumed to induce stress and discomfort of the animals. However, these findings were mostly related to the local irritant effects of the test item. Hence, an adverse effect, relevant in humans, cannot be stated. The No Observed Adverse Effect Level is assumed to be 300 mg/kg BW within this study (MD group).

Data source

Materials and methods

Results and discussion

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion