Cyclohexanamine, 4,4’-methylenebis[N-(2,2-dimethyl-3-(4-morpholinyl)propylidene)]-

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2021-04-13 to 2021-04-29

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Report date:

2021

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Version / remarks:

30 May 2008

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

adopted 17th december 2001

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Version / remarks:

December 2002

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(1E)-N-[4-({4-[(E)-[2,2-dimethyl-3-(morpholin-4-yl)propylidene]amino]cyclohexyl}methyl)cyclohexyl]-2,2-dimethyl-3-(morpholin-4-yl)propan-1-imine

EC Number:

954-590-8

Cas Number:

2522560-40-1

Molecular formula:

C31H56N4O2

IUPAC Name:

(1E)-N-[4-({4-[(E)-[2,2-dimethyl-3-(morpholin-4-yl)propylidene]amino]cyclohexyl}methyl)cyclohexyl]-2,2-dimethyl-3-(morpholin-4-yl)propan-1-imine

Test animals

Species:

rat

Strain:

Wistar

Remarks:

Han:Wist

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Toxi-Coop Zrt. 1103 Budapest, Cserkesz u. 90.
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: Young adult rats, 8 weeks old in group 1 and group 2
- Weight at study initiation: 176-185 g in group 1 and 179-182 g in group 2
- Fasting period before study: One day before the treatment, only food was withheld
- Housing: 3 animals/sex/cage, Type III polypropylene/polycarbonate; rat type cages with a solid floor, stainless steel wire covers and self-feeding baskets, laboratory bedding
- Diet: ssniff® SM R/M-Z+H complete diet produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany, ad libitum
- Water: Tap water from watering bottles, ad libitum
- Acclimation period: 5 days in first group and 6 days in second group
- Microbiological status when known: SPF at arrival and kept in a good conventional environment during the study

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): > 10 by central aircondition system
- Photoperiod (hrs dark / hrs light): Artificial light, from 6 am. to 6 pm

IN-LIFE DATES: From: 2021-06-08 To: 2021-04-15

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Helianthi annui oleum raffinatum (sunflower oil)

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 200 mg/mL
- Lot/batch no.: 8006332001
- Purity: not specified

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

CLASS METHOD
- Rationale for the selection of the starting dose: The test item was not expected to be lethal at 2000 mg/kg bw. A limit test was performed.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

3 females per group

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Inspection for signs of morbidity and mortality were made twice daily at the beginning and end of the working day; animals were observed individually after dosing once during the first 30 minutes, then 1 h, 2 h, 3 h, 4 h, after the treatment and once per day for 14 days thereafter
- Necropsy of survivors performed: Yes
- Clinical signs including body weight: Individual observations were performed on the skin and fur, eyes and mucous membranes and also respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma
- Other examinations performed: pathology (surviving animals were exsanguinated under isoflurane anaesthesia. After examination of the external appearance the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs were observed. All gross pathological changes were recorded for each animal on the post mortem record sheets)

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

All female rats in step 1 (2000 mg/kg bw dose) survived until the end of the 14-day observation period.

In step 2, one rat dosed at 2000 mg/kg bw died on Day 1. The death might be a consequence of the systemic toxicity of the test item.

Clinical signs:

other: In group 1 treated with 2000 mg/kg bw of the test item clinical signs of reaction comprised of decreased activity (16 cases of 57 observations), bedding chewing (4/57), closed eyes (14/57), piloerection (5/57), diarrhoea (1/57) and bloody diarrhoea (2/5

Gross pathology:

One rat treated with 2000 mg/kg bw dose of the test item spontaneously died during the study. Five animals treated with 2000 mg/kg bw dose of test item survived until the scheduled necropsy on Day 15. The stomach wall was attenuated and the intestines were full of mucous and bloody content in deceased animal. No pathological changes were found related to the test item during the macroscopic examination of surviving animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In an acute oral toxicity study according to OECD guideline 423, the LD50 was greater than 2000 mg/kg bw (LD50 ≥ 2000 mg/kg bw).

Executive summary:

In this acute oral toxicity study, two groups of female rats (Han:WIST) were given a single oral dose of the test item at a concentration of 2000 mg/kg bw. The starting dose was selected on the basis of the available information about the test item. The acute toxic class method according to OECD Guideline No. 423 was carried out involving a stepwise procedure with the use of 2000 mg/kg bw of the test item as the starting dose in three female rats. No animal died in the first step at 2000 mg/kg bw dose level. Therefore, three additional female rats were treated with 2000 mg/kg bw in a second step. Since only one animal died in the second step, no further testing was required. The stopping criteria of Annex 2d of OECD Guideline No. 423 were met. Animals were weighed, observed for lethality and toxic symptoms for 14 days after the treatment. Gross pathological examination was carried out on the Day 1 in deceased animal and on the 15th day after the treatment in surviving animals.

 

Lethality, Clinical symptoms and Body weight:
One of six animals treated with 2000 mg/kg bw of the test item died on Day 1. In the first step, CNS - and emotional symptoms (decreased activity, bedding chewing, closed eyes) and disturbances of the autonomic functions (diarrhoea/bloody diarrhoea, piloerection) were observed in animals between 30 minutes and 4 hours after the treatment and between Day 1 and Day 3, as well. In the second step, CNS - and emotional symptoms (decreased activity, bedding chewing, closed eyes) and disturbances of the autonomic functions (diarrhoea, piloerection) were observed in animals on the treatment day between 2 and 4 hours after the treatment and on Day 1, as well. The mean body weight of both groups corresponded to their species and age throughout the study.

Gross pathology:
Altogether 1 animal died, and 5 animals were sacrificed as scheduled during the
study. Treatment related gross pathological changes as attenuated stomach wall and mucous and bloody content in the intestines were found in deceased animal. All organs of the surviving animals treated with 2000 mg/kg bw dose proved to be free of treatment related gross pathological changes.