Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
06 Jun 2017 - 04 Jul 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
December 2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
May 2008
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
December 2002
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF Guidelines
Version / remarks:
2000, including the most recent revisions
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Colourless solidified melt
Details on test material:
Expiry date: 25 October 2018

Test animals

Species:
rat
Strain:
other: Crl:WI (Han)
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS:
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: Young adult animals (approx. 8-9 weeks old).
- Weight at study initiation: Body weight variation was 152gr to 186gr.
- Fasting period before study: Animals were deprived of food overnight (for a maximum of 20 hours).
- Housing: Group housing of 3 animals per cage in labeled Macrolon cages.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 – 22
- Humidity (%): 45 - 68
- Air changes (per hr): ten or greater.
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 13 June 2017 to 04 July 2017

Administration / exposure

Route of administration:
oral: gavage
Details on oral exposure:
GAVAGE METHOD: plastic feeding tubes.

Frequency: single dosage, on Day 1.

VEHICLE:
Trial preparations were performed at the Test Facility to select the suitable vehicle and to establish a suitable formulation procedure.
Propylene glycol was the vehicle of choice.
Specific gravity: 1.036


MAXIMUM DOSE VOLUME APPLIED:
(10 mL/kg) body weight.


DOSAGE PREPARATION: Test item dosing formulations (w/w) were homogenized to visually acceptable levels at appropriate concentrations to meet dose level requirements.
The dosing formulations were kept at room temperature until dosing. The dosing formulations and vehicle were stirred until and during dosing.
Adjustment was made for specific gravity of the vehicle and test item. No correction was made for the purity/composition of the test item.
Any residual volumes were discarded.

Doses:
2000 mg/kg body weight
300 mg/kg body weight


No. of animals per sex per dose:
2000 mg/kg body weight: 3
300 mg/kg body weight: 6 (2 groups of three females in a stepwise manner)
Control animals:
no
Details on study design:
The toxicity of the test item was assessed by stepwise treatment of groups of 3 females. The absence or presence of mortality of animals dosed at one step determined the next step, based on the test procedure defined in the guidelines. The onset, duration and severity of the signs of toxicity were taken into account for determination of the time interval between the dose groups. The first group was treated at a dose level of 2000 mg/kg body weight. Based on the results, two additional groups were dosed at 300 mg/kg body weight.

Animals were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test substance. Water was available ad libitum.

- Duration of observation period following administration: 14 days.
- Frequency of observations: Postdose observations were performed at periodic intervals on the day of dosing (at least three times) and once daily thereafter.
Mortality/Viability: twice daily.
Body weights: Days 1 (pre-administration), 8 and 15.
Clinical signs: At periodic intervals on the day of dosing (Day 1) and once daily thereafter, until Day 15.
- Necropsy of survivors performed: All moribund animals and animals surviving to the end of the observation period were sacrificed by oxygen/carbon dioxide procedure. All animals assigned to the study were subjected to necropsy and descriptions of all internal macroscopic abnormalities were recorded.
- Other examinations performed: none.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Mortality:
At 2000 mg/kg body weight, two animals were sacrificed for humane reasons on Day 1.
At 300 mg/kg body weight, no mortality occurred.
Clinical signs:
At 2000 mg/kg body weight, comatose state, lethargy, flat posture, abnormal posture, lateral recumbency,
uncoordinated movements, hunched posture, rales, piloerection, watery discharge from the
eyes, ptosis, brown coloration of urine and/or hypothermia were noted for the animals on Day
1. Additionally, the surviving animal showed hunched posture and piloerection on Days 2
and/or 3.
At 300 mg/kg body weight, hunched posture, piloerection and/or ptosis were noted for all animals on Day 1 only.
Body weight:
The body weight gain shown by the surviving animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.
Gross pathology:
At 2000 mg/kg body weight, gelatinous contents of the gastrointestinal tract (stomach, duodenum and/or jejunum) and reddish contents of the urinary bladder were found in the animals sacrificed for humane reasons during the study, at macroscopic post mortem examination. No abnormalities were found in the remaining animal. At 300 mg/kg body weight, no abnormalities were found at macroscopic examination

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
The oral LD50 value of ethanol, 2,2’-[(3-methylphenyl)imino]bis- in Wistar rats was established to be within the range of 300-2000 mg/kg body weight.
According to CLP and GHS regulations the subtstance is classified as category 4, labelled with H302, (harmful if swallowed).
Executive summary:

An assessment of acute oral toxicity with ethanol, 2,2’-[(3-methylphenyl)imino]bis- in the rat (Acute Toxic Class Method) was performed according to OECD/EC test guidelines and GLP principles.The test item was administered by oral gavage to one groups three female Wistar rats at 2000 mg/kg body weight. Two animals were sacrificed for humane reasons on Day 1. At day 1 comatose state, lethargy, flat posture, abnormal posture, lateral recumbency, uncoordinated movements, hunched posture, rales, piloerection, watery discharge from the eyes, ptosis, brown coloration of urine and/or hypothermia were noted for the animals. Additionally, the surviving animal showed hunched posture and piloerection on Days 2 and/or 3. Based on the results, two additional groups were dosed at 300 mg/kg body weight. At 300 mg/kg body weight, no mortality occurred but hunched posture, piloerection and/or ptosis were noted for all animals on Day 1.

The body weight gain shown by the animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.

At 2000 mg/kg body weight, gelatinous contents of the gastrointestinal tract (stomach, duodenum and/or jejunum) and reddish contents of the urinary bladder were found in the animals sacrificed for humane reasons during the study, at macroscopic post mortem examination. No abnormalities were found in the remaining animal. At 300 mg/kg body weight, no abnormalities were found at macroscopic examination.

The oral LD50 value of Ethanol, 2,2’-[(3-methylphenyl)imino]bis- in Wistar rats was therefore established to be within the range of 300-2000 mg/kg body weight.

According to the OECD 423 test guideline, the LD50 cut-off value was considered to be 1000 mg/kg body weight.

Based on these results, Ethanol, 2,2’[(3-methylphenyl)imino]bis- is classified according to CLP and GHS regulations the subtstance is classified as category 4, labelled with H302 , (harmful if swallowed).