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Description of key information

Available studies result to GHS classification Category 3 for acute oral toxicity (LD50 between 50 and 300 mg/kg bw) with a LD50 cut-off of 200 mg/kg bw. No data available on acute toxicity via inhalation or dermal route.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
March 2016 - April 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
2002
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nousan, Notification No 8147, November 2000, including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Date certificate 3 November 2015
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
other: Crl:WI (Han)
Sex:
female
Details on test animals and environmental conditions:
- 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 within +/- 20% of the sex mean (141 - 190 grams).
- Fasting period before study: Animals were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test item.
- Housing: Group housing of 3 animals per cage in labeled Macrolon cages containing sterilized sawdust as bedding material and paper as cage-enrichment.
- 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
set to maintain:
- Temperature (°C): 18 – 24
- Humidity (%): 40 - 70
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 22 March 2016 to 21 April 2016
Route of administration:
oral: gavage
Vehicle:
other: no vehicle (undiluted) at 300 mg/kg bw, corn oil (50 mg/kg bw)
Details on oral exposure:
GAVAGE METHOD: plastic feeding tubes.

Frequency: single dosage, on Day 1.

VEHICLE
The vehicle was selected based on trial preparations performed at WIL Research Europe and on test item data supplied by the Sponsor. There was no information available regarding the solubility or stability in vehicle.

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION:
The test item (preparations) (w/w) were kept at room temperature and were dosed within 4 hours after adding the vehicle to the test item. Homogeneity was assessed by visual inspection of the solutions and the formulations were stirred during dosing, which ensures homogeneity sufficient for these kinds of studies. Adjustment was made for specific gravity of the vehicle and test item. No correction was made for purity of the test item.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Based on anticipated mortality at the next higher dose 300 mg/kg bw was selected as the starting dose.
Doses:
300 mg/kg bw
50 mg/kg bw
No. of animals per sex per dose:
300 mg/kg bw: 3
50 mg/kg bw: 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 first group was treated at a dose level of 300 mg/kg bw. 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.

Duration of observation period following administration: 14 days
Frequency of observations and weighing:
- Mortality/Viability: Twice daily. Animals showing pain, distress or discomfort, which was considered not transient in nature or was likely to become
more severe, were sacrificed for humane reasons based on OECD guidance document on humane endpoints (ENV/JM/MONO/2000/7). The time of death was recorded as precisely as possible.
- Body weights: Days 1 (pre-administration), 8 and 15 and at death.
- Clinical signs: At periodic intervals on the day of dosing (Day 1) and once daily thereafter, until Day 15.
- Necropsy performed: yes, all animals
- Other examinations performed: none.
Statistics:
No statistical analysis was performed (The method used is not intended to allow the calculation of a precise LD50 value).
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 50 - < 300 mg/kg bw
Based on:
test mat.
Mortality:
At 300 mg/kg bw, on Day 3, one animal was found dead and two animals were sacrificed for humane reasons.
At 50 mg/kg bw, no mortality occurred.
Clinical signs:
At 300 mg/kg bw lethargy, hunched posture, slow and shallow breathing, piloerection, diarrhoea,
chromodacryorrhoea in both eyes, lean appearance and/or ptosis were noted for the animals
between Days 1 and 3.
At 50 mg/kg bw, hunched posture was noted for all animals on Day 1.
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 300 mg/kg bw, abnormalities of the spleen (reduced in size), thymus (reduced in size) and/or GI-tractus (contents: gelatinous) were noted for the animals, at macroscopic post mortem examination.
At 50 mg/kg bw, macroscopic examination did not reveal any abnormalities.
Other findings:
Autolysis was noted for the animal found dead. This was considered not toxicologically relevant.
Interpretation of results:
Category 3 based on GHS criteria
Conclusions:
In an acute oral toxicity study with Diamine methylated in rats, performed according to OECD/EC test guidelines, an LD50 within the range of 50-300 mg/kg bw was determined, with a LD50 cut-off value of 200 mg/kg bw.
Executive summary:

Assessment of acute oral toxicity with Diamine methylated in the rat (Acute Toxic Class Method) was performed according to OECD/EC guidelines and in accordance with GLP principles. Initially, Diamine methylated was administered by oral gavage to three female Wistar rats at 300 mg/kg bw. In a stepwise procedure two additional groups of three females were dosed at 50 mg/kg bw.

At 300 mg/kg bw, on Day 3, one animal was found dead and two animals were sacrificed for humane reasons. At 50 mg/kg bw, no mortality occurred. At 300 mg/kg lethargy, hunched posture, slow and shallow breathing, piloerection, diarrhoea, chromodacryorrhoea in both eyes, lean appearance and/or ptosis were noted for the animals between Days 1 and 3. At 50 mg/kg bw, hunched posture was noted for all animals on Day 1. 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. At 300 mg/kg bw, abnormalities of the spleen (reduced in size), thymus (reduced in size) and/or GI-tractus (contents: gelatinous) were noted for the animals, at macroscopic post mortem examination. At 50 mg/kg bw, macroscopic examination did not reveal any abnormalities.

The oral LD50 value of Diamine methylated in Wistar rats was established to be within the range of 50-300 mg/kg bw. According to the OECD 423 test guideline, the LD50 cut-off value was considered to be 200 mg/kg bw.

Based on these results:

- according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2015) (including all amendments), Diamine methylated should be classified as: Toxic if swallowed (Category 3) for acute toxicity by the oral route;

- according to the Regulation (EC) No 1272/2008 on classification, labelling and packaging of items and mixtures (including all amendments), Diamine methylated should be classified as Category 3 and should be labeled as H301: Toxic if swallowed.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
200 mg/kg bw
Quality of whole database:
Guideline (OECD 423, ACT) study performed under GLP.

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Acute toxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Oral:

Assessment of acute oral toxicity with Diamine methylated in the rat (Acute Toxic Class Method) was performed according to OECD/EC guidelines and in accordance with GLP principles. Initially, Diamine methylated was administered by oral gavage to three female Wistar rats at 300 mg/kg bw. In a stepwise procedure two additional groups of three females were dosed at 50 mg/kg bw.

At 300 mg/kg bw, on Day 3, one animal was found dead and two animals were sacrificed for humane reasons. At 50 mg/kg bw, no mortality occurred. At 300 mg/kg lethargy, hunched posture, slow and shallow breathing, piloerection, diarrhoea, chromodacryorrhoea in both eyes, lean appearance and/or ptosis were noted for the animals between Days 1 and 3. At 50 mg/kg bw, hunched posture was noted for all animals on Day 1. 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. At 300 mg/kg bw, abnormalities of the spleen (reduced in size), thymus (reduced in size) and/or GI-tractus (contents: gelatinous) were noted for the animals, at macroscopic post mortem examination. At 50 mg/kg bw, macroscopic examination did not reveal any abnormalities.

The oral LD50 value of Diamine methylated in Wistar rats was established to be within the range of 50-300 mg/kg bw. According to the OECD 423 test guideline, the LD50 cut-off value was considered to be 200 mg/kg bw.

 

Inhalation:

There is no study on inhalation toxicity available forDiamine methylated.

REACH stipulates that testing by the inhalation route is appropriate if exposure of humans via inhalation is likely taking into account the vapour pressure of the substance and/or the possibility of exposure to aerosols, particles or droplets of an inhalable size. REACH guidance R.7.a, chapter. 7.4 Acute toxicity indicates that in principle no inhalation studies are needed when vp < 0.1 Pa at 20°C or particle size > 100 µm.Diamine methylatedis a liquid with no inhalable particles and a vapour pressure less than 4.5E-04 Pa at 25 °C. Also the use of this substance will not result in aerosols, particles or droplets of an inhalable size, so exposure to humans via the inhalation route will be unlikely to occur, and no acute inhalation test was performed.

 

Diamine methylatedis a liquid at ambient temperatures around 20°C and lower. No information on viscosity is available at 40°C, and hence the hazard for aspiration cannot be adequately assessed.

 

Dermal:

There is no dermal LD 50 value for acute skin toxicity ofDiamine methylated, and due to the corrosive nature of the substance it is not ethical to carry out this animal study.

Justification for classification or non-classification

The acute oral LD50 of Diamine methylated is within the range of 50-300 mg/kg bw. Hence the product should be classified according to GHS for acute toxicity as Cat.3, with hazard statement H301: Toxic if swallowed.

 

Acute dermal testing with corrosive materials is not justified. As a consequence no classification can be made for acute dermal toxicity. Effects will be characterised by local tissue damage. Systemic uptake via skin is likely to be very limited.

No classification for acute dermal toxicity is therefore indicated.

 

Also for acute inhalation toxicity information for classification is lacking, and is testing not justified.

 

Aspiration: Diamine methylatedis a liquid at ambient temperatures around 20°C and lower. No information on viscosity is available at 40°C, and hence the hazard for aspiration cannot be adequately assessed.

 

No classification STOT-SE Cat.3 needed:

The active substance is not structurally related to any known class of neurotoxic chemicals. In addition, repeated dose studies did not show indications of specific neurotoxicity, in specific neurotoxicity measures as sensory activity, grip strength, and motor activity assessment.