Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2020
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2020
Report date:
2020

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
(ATC method)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
acute toxic class method
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
N1,N2-dimethyl-N1-{2-[methyl(propan-2-yl)amino]ethyl}-N2-(propan-2-yl)ethane-1,2-diamine
EC Number:
950-627-7
Cas Number:
1042950-30-0
Molecular formula:
C13H31N3
IUPAC Name:
N1,N2-dimethyl-N1-{2-[methyl(propan-2-yl)amino]ethyl}-N2-(propan-2-yl)ethane-1,2-diamine
Test material form:
liquid

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Sex:
female
Details on test animals or test system and environmental conditions:
Body weight
(at start of 1st administration) 213.4 – 307.7 g
Age (at start of administration) Approx. 9 – 10 weeks
Identification of animals By coloured marks and cage label

Duration of experiment
At least 5 adaptation days
1 test day
2 recovery weeks

Diet
Commercial diet, ssniff® R/M-H V1534 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany; see Appendix 3: Composition of the diet) served as food. Tap water was available ad libitum.
Periodic analysis of the food for contaminants based on EPA/USA is conducted by LUFA-ITL (see Appendix 3: Limitation for contaminants in the diet). Certificates of analysis of the composition and for contaminants were provided by the manufacturer and are QAU archived.
Housing
Granulated textured wood (Granulat A2, J. Brandenburg, 49424 Goldenstedt, Germany) was used as bedding material for the cages. The cages were changed and cleaned twice a week.
Periodic analysis of the bedding material for contaminants based on EPA/USA is conducted by LUFA-ITL (see Appendix 3: Limitation for contaminants in the bedding material).
During the 14-day observation period the animals were singly in MAKROLON cages (type III plus) at a room temperature of 22 °C ± 3 °C (maximum range) and a relative humidity of 55 % ± 10 % (maximum range). Deviations from the maximum range caused for example during cleaning procedures are dealt with in SOPs.
The rooms were lit (about 150 lux at approx. 1.50 m room height) and darkened for periods of 12 hours each.
Drinking water
Drinking water in bottles was offered ad libitum.
Drinking water is examined according to the 'Deutsche Trinkwasserverordnung 2001' [German Regulations on drinking water 2001] by the Hamburger Wasserwerke, 20539 Hamburg, Germany, at least four times a year.
In addition, drinking water samples taken at LPT are analysed by LUFA-ITL once a year for means of bacteriological investigations according to the 'Deutsche Trinkwasserverordnung 2001.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
Selection of route of administration

According to the OECD/EC guidelines

Vehicle
The test item was diluted to concentrations of 2000 and 300 mg/kg b.w. using tap water

Administration volume
10 mL/kg b.w. per application

Dose levels
2000 and 300 mg/kg b.w.
Doses:
Dose levels
2000 and 300 mg/kg b.w.
No. of animals per sex per dose:
Number of animals

Limit Test
3 animal at the dose level of 2000 mg/kg b.w.

First and second step
6 animals at the dose level of 300 mg/kg b.w.
Control animals:
no
Details on study design:
Principle of the ATC-test method

This procedure permits the identification of the 'acute-toxic-class' (ATC), a measurement of the acute toxicity by the oral route.
The test item is administered orally by gavage at a single dose level to a group of experimental animals. The dose used is selected from a series of defined dose levels. Due to the small number of animals used with this method, there is no need to perform a range finding test.
The test item is tested using a stepwise procedure, each step or dose level uses three female animals. The results of each step determine if:
o no further testing is needed,
o the next step will be performed with the same dose,
o the next step will be performed at the next higher or next lower dose level.

Starting at 2000 mg/kg b.w.
o Testing at 2000 mg/kg b.w.:
Three animals of one sex (preferably females) are treated at 2000 mg/kg b.w. (first step).
If two to three animals die, testing at 300 mg/kg b.w. should be performed.
If no to one animal dies, the test item should be retested (second step) with 2000 mg/kg b.w., using three animals of the same sex.
If, in this second step, two to three animals die, testing at 300 mg/kg b.w. should be performed. If, in this second step, no to one animal dies, no further testing is necessary.

o Testing at 300 mg/kg b.w.:
If the results of the test at 2000 mg/kg b.w. indicate the need for further testing at a lower dose level.
Three female animals are treated at 300 mg/kg b.w. (first step).
If two or three animals die, testing at 50 mg/kg b.w. should be performed.
If fewer than two animals die, the test item should be retested (second step) with 300 mg/kg b.w., using three animals of the same sex.
If, in this second step, two or three animals die, testing at 50 mg/kg b.w. should be performed. If, in this second step, no to one animal dies, no further testing is necessary.


o Testing at 50 mg/kg b.w.:
If the results of the test at 300 mg/kg b.w. indicate the need for further testing at a lower dose level.
Three female animals treated at 50 mg/kg b.w. (first step).
If two or three animals die, testing at 5 mg/kg b.w. should be performed.
If fewer than two animals die, the test item should be retested (second step) with 50 mg/kg b.w., using three animals of the same sex.
If, in this second step, two or three animals die, testing at 5 mg/kg b.w. should be performed. If, in this second step, no to one animal dies, no further testing is necessary.

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:
Under the present test conditions, two administrations at an interval of 1 hour of 2000 mg of the test item/kg b.w. revealed slightly to moderately reduced motility, slight to moderate ataxia, slightly reduced muscle tone, slight dyspnoea and slight ptosis in three animals employed. All three animals died prematurely (2/3 animals) or were humanely killed (1/3 animals) after gavage of the second dose (within 3 hours after start of the experiment).
Two administrations at an interval of 1 hour of 300 mg the test item b.w. revealed slightly reduced motility, slight ataxia and slight dyspnoea in six animals employed (first and second step). One animal died prematurely after gavage of the second dose (within 3 hours after starting the experiment).

Clinical signs:
Under the present test conditions, two administrations at an interval of 1 hour of 2000 mg of the test item/kg b.w. revealed slightly to moderately reduced motility, slight to moderate ataxia, slightly reduced muscle tone, slight dyspnoea and slight ptosis in three animals employed. Two administrations at an interval of 1 hour of 300 mg the test item b.w. revealed slightly reduced motility, slight ataxia and slight dyspnoea in six animals employed (first and second step). One animal died prematurely after gavage of the second dose (within 3 hours after starting the experiment).
Body weight:
All surviving animals gained the expected body weight.
Gross pathology:
No pathological changes were observed at necropsy.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Under the present test conditions, two administrations at an interval of 1 hour of 2000 mg PU-2019-872/kg b.w. revealed slightly to moderately reduced motility, slight to moderate ataxia, slightly reduced muscle tone, slight dyspnoea and slight ptosis in three animals employed. All three animals died prematurely or were humanely killed after gavage of the second dose (within 3 hours after starting the experiment).
Two administrations at an interval of 1 hour of 300 mg PU-2019-872/kg b.w. revealed slightly reduced motility, slight ataxia and slight dyspnoea in six animals employed (first and second step). One animal died prematurely after gavage of the second dose (within 3 hours after starting the experiment).
All surviving animals gained the expected body weight.
According to the Globally Harmonized Classification System (GHS) the test item requires labelling of category 4 for acute oral toxicity (as LD50 is between 300 - 2000 mg/kg b.w.).
Executive summary:

Under the present test conditions, two administrations at an interval of 1 hour of 2000 mg PU-2019-872/kg b.w. revealed slightly to moderately reduced motility, slight to moderate ataxia, slightly reduced muscle tone, slight dyspnoea and slight ptosis in three animals employed. All three animals died prematurely or were humanely killed after gavage of the second dose (within 3 hours after starting the experiment).

Two administrations at an interval of 1 hour of 300 mg PU-2019-872/kg b.w. revealed slightly reduced motility, slight ataxia and slight dyspnoea in six animals employed (first and second step). One animal died prematurely after gavage of the second dose (within 3 hours after starting the experiment).

All surviving animals gained the expected body weight.

According to the Globally Harmonized Classification System (GHS) the test item requires labelling of category 4 for acute oral toxicity (as LD50 is between 300 - 2000 mg/kg b.w.).