Amides, adipic acid

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 26 February, 2020 to 12 March, 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 guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2001

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Directive 96/54/EEC

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

solid

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Sprague-Dawley derived, albino

Sex:

female

Details on test animals or test system and environmental conditions:

Animals
Number of Animals: 3
Sex: Female, nulliparous and non-pregnant.
Species/Strain: Rat/Sprague-Dawley derived, albino.
Age/Body Weight: Young adult (10 weeks)/213-221 grams at experimental start.
Source: Received from Charles River Laboratories on February 6, 2020.

Husbandry
Housing: The animals were housed in caging which conforms to the size recommendations in the most recent Guide for the Care and Use of Laboratory Animals (Natl. Res. Council, 2011). Animals were group housed, except on the day of administration, at which time they were single housed and until the animals were deemed acceptable, based on observations, to return to group housing. Enrichment (e.g., toy) was placed in each cage and litter was changed at least once per week.
Animal Room Temperature and Relative Humidity Ranges: 20-24ºC and 42-56%, respectively.
Animal Room Air Changes/Hour: 12. Airflow measurements are evaluated regularly and the records are kept on file at Product Safety Labs.
Photoperiod: 12-hour light/dark cycle
Acclimation Period: 20 or 21 days
Food: Envigo Teklad Global 16% Protein Rodent Diet® #2016. The diet was available ad libitum, except during fasting.
Water: Filtered tap water was supplied ad libitum.
Contaminants: There were no known contaminants reasonably expected to be found in the food or water at levels which would have interfered with the results of this study. Analyses of the food and water are conducted regularly and the records are kept on file at Product Safety Labs.

Identification
Cage: Each cage was identified with a cage card indicating at least the study number, dose level, identification, and sex of the animal.
Animal: A number was allocated to each rat on receipt and a stainless steel ear tag bearing this number was attached to the rat. This number, together with a sequential animal number assigned to study number 52369, constituted unique identification. Only the sequential animal number is presented in this report.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

The prepared test mixture was administered to the stomach using a stainless steel ball-tipped gavage needle attached to an appropriate syringe. Following administration, each animal was returned to its designated cage. Feed was replaced approximately 3-4 hours after dosing.

Doses:

5000 mg/kg bw limit dose

No. of animals per sex per dose:

3 female rats

Control animals:

no

Details on study design:

Procedure
Selection of Animals
Prior to each dosing, experimentally naive rats were fasted overnight by removing the feed from their cages. During the fasting period, the rats were examined for health and weighed (initial). Three healthy, naive female rats (not previously tested) were selected for test.

Preparation of Test Substance
The test substance, as received, was a solid. The test substance was administered as a 40% w/w mixture in distilled water. Preliminary sample preparation assessments conducted by PSL indicated that mixtures in excess of 40% (i.e., 50-80%) were too viscous to be administered properly. Each preparation was mixed well prior to use.

Dose Calculations
Individual doses were calculated based on the initial body weights, taking into account the density (determined by PSL) and concentration of the test mixture.

Dosing
The prepared test mixture was administered to the stomach using a stainless steel ball-tipped gavage needle attached to an appropriate syringe. Following administration, each animal was returned to its designated cage. Feed was replaced approximately 3-4 hours after dosing. An initial limit dose of 5000 mg/kg was administered to one healthy female rat by oral gavage. Due to the absence of mortality in this animal, two additional females received the same dose level, simultaneously.

In-life Observations
The animals were observed for mortality, signs of gross toxicity, and behavioral changes approximately 30 minutes post-dosing, during the first several hours post-dosing and at least once daily thereafter for 14 days after dosing. Observations included gross evaluation of skin and fur, eyes and mucous membranes, respiratory, circulatory, autonomic and central nervous systems, somatomotor activity and behavior pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhea, and coma.

Body Weights
Individual body weights of the animals were recorded prior to test substance administration (initial) and again on Days 7 and 14 (terminal) following dosing.

Necropsy
All rats were euthanized via CO2 inhalation at the end of the 14-day observation period. Gross necropsies were performed on all animals. Tissues and organs of the thoracic and abdominal cavities were examined.

Statistics:

Statistical analysis was limited to the calculation of the mean density value for dosing.

Results and discussion

Preliminary study:

All animals survived test substance administration, gained body weight, and appeared active and healthy during the study. There were no signs of gross toxicity, adverse clinical effects, or abnormal behavior. No gross abnormalities were noted for any of the animals when necropsied at the conclusion of the 14-day observation period.

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other:

Mortality:

No mortality observed

Clinical signs:

There were no signs of gross toxicity, adverse clinical effects, or abnormal behavior.

Body weight:

All animals survived test substance administration, gained body weight, and appeared active and healthy during the study.

Gross pathology:

No gross abnormalities were noted for any of the animals when necropsied at the conclusion of the 14-day observation period.

Any other information on results incl. tables

Table 1: Individual body weight

Animal No.	Sex	Dose Level (mg/kg)	Body Weight (g)			Dose*
			Initial	Day 7	Day 14	mL
3101	F	5000	214	236	265	2.5
3102	F		213	246	265	2.5
3103	F		221	250	280	2.6

*The test substance was administered as a 40% w/w mixture in distilled water. Density –1.06g/mL

Table 2: In-life observations

Animal Number

Animal Sex

Dose Level (mg/kg)

Observation

Day of Observation (x=observation is present)

0(0.5 hr)

0(3 hrs)

1

2

3

4

5

6

7

8

9

10

11

12

13

14

3101

F

5000

Active and healthy

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

3102

F

5000

Active and healthy

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

3103

F

5000

Active and healthy

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

Table 3: Necropsy observation

Animal Number	Animal Sex	Dose Level (mg/kg)	Organ / Tissue	Observation
3101	F	5000	All tissues and organs	No gross abnormalities
3102	F	5000	All tissues and organs	No gross abnormalities
3103	F	5000	All tissues and organs	No gross abnormalities

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the study conditions, the acute oral LD50 of the test substance was determined to be >5000 mg/kg bw in female rats.

Executive summary:

A study was conducted to determine the acute oral toxicity of the test substance in rats according to OECD Guideline 423 and EU Method B.1 tris, in compliance with GLP. Initially, a limit dose of 5000 mg/kg bw was administered to one healthy female rat by oral gavage. Due to the absence of mortality in this animal, two additional females received the same dose level, simultaneously. Since these animals survived, no additional animals were tested. All animals were observed for mortality, signs of gross toxicity and behavioural changes at least once daily for 14 days after dosing. Body weights were recorded prior to administration (initial) and again on Days 7 and 14. Necropsies were performed on all animals at terminal sacrifice. All animals survived test substance administration, gained body weight, and appeared active and healthy during the study. There were no signs of gross toxicity, adverse clinical effects or abnormal behaviour. No gross abnormalities were noted for any of the animals when necropsied. Under the study conditions, the acute oral LD50 of the test substance was determined to be >5000 mg/kg bw in female rats (Durando, 2020).