2-ethyl-N-(3-methoxyphenyl)-N-methylbutanamide

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was conducted between 05 June 2018 and 03 July 2018.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Reliability 1 is assigned because the study conducted according to OECD TG 420 in compliance with GLP, without deviations that influence the quality of the results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Test material

Test material information

Constituent 1

Reference substance name:

2-ethyl-N-(3-methoxyphenyl)-N-methylbutanamide

Cas Number:

2101947-22-0

Molecular formula:

C14H21NO2

IUPAC Name:

2-ethyl-N-(3-methoxyphenyl)-N-methylbutanamide

Test material form:

liquid

Specific details on test material used for the study:

Identification: FRET 15-0735
Physical state / Appearance: Clear colorless liquid
Storage Conditions: Room temperature in the dark

Test animals

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

Animal Information
Female Wistar (RccHan™:WIST) strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non pregnant. After an acclimatization period of at least 5 days the animals were selected at random and given a number unique within the study by indelible ink marking on the tail and a number written on a cage card. At the start of the study the animals were 8 to 12 weeks of age. The body weight variation did not exceed ±20% of the mean body weight of any previously treated animals.

Animal Care and Husbandry
The animals were housed in groups of up to four in suspended solid floor polypropylene cages furnished with wood flakes. With the exception of an overnight fast immediately before dosing and for approximately 3 to 4 hours after dosing, free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

arachis oil

Details on oral exposure:

Test Item Preparation and Analysis
For the purpose of the study the test item was freshly prepared, as required, as a solution in arachis oil BP. Arachis oil BP was used because the test item did not dissolve/suspend in distilled water.
The test item was formulated within 2 hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation. This is an exception with regard to GLP and has been reflected in the GLP compliance statement.

Doses:

300 mg/kg and 50 mg/kg

No. of animals per sex per dose:

300 mg/kg: 5 females
50 mg/kg: 5 females

Control animals:

no

Details on study design:

In the absence of data regarding the toxicity of the test item, 300 mg/kg was chosen as the starting dose.
A single animal was treated at 300 mg/kg.
In the absence of mortality at a dose level of 300 mg/kg, an additional group of 4 animals were treated.
A total of five animals were therefore treated at a dose level of 300 mg/kg in the study.
Due to mortality and signs of systemic toxicity at a dose level of 300 mg/kg, an additional animal was treated at 50 mg/kg.
In the absence of mortality at a dose level of 50 mg/kg, an additional group of 4 animals were treated.
A total of five animals were therefore treated at a dose level of 50 mg/kg in the study.
All animals were dosed once only by gavage, using a metal cannula attached to a graduated syringe. The volume administered to each animal was calculated according to the fasted body weight at the time of dosing. Treatment of animals was sequential. Sufficient time was allowed between each dose group to confirm the survival of the previously dosed animals.
Clinical observations were made 30 minutes, 1, 2, and 4 hours after dosing and then daily for up to 14 days. Morbidity and mortality checks were made twice daily, early and late during normal working days, and once daily at weekends and public holidays.
Individual body weights were recorded on Day 0 (the day of dosing) and on Days 7 and 14 or at death.
At the end of the observation period the surviving animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Results and discussion

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

>= 50 - <= 300 mg/kg bw

Based on:

test mat.

Mortality:

300 mg/kg:
Four animals were killed for humane reasons during the day of dosing, due to the occurrence of clinical signs of toxicity that approached the severity limit set forth in the UK Home Office Project License.
50 mg/kg:
There were no deaths.

Clinical signs:

other: 300 mg/kg: Signs of systemic toxicity commonly noted were hunched posture, ataxia and body tremors or occasional body tremors. Additional signs of systemic toxicity noted in three animals were vocalization, increased respiratory rate and increased activi

Gross pathology:

300 mg/kg:
Patchy pallor of the liver was noted at necropsy of one animal. No abnormalities were noted at necropsy of the remaining three animals that were humanely killed during the study or the animal that was killed at the end of the study.
50 mg/kg:
No abnormalities were noted at necropsy.

Any other information on results incl. tables

Individual Clinical Observations and Mortality Data - 300 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Effects Noted After Dosing (Hours)				Effects Noted During Period After Dosing (Days)
		½	1	2	4	1	2	3	4	5	6	7	8	9	10	11	12	13	14
300	1-0 Female	0	HA	HA	HATo	H	H	0	0	0	0	0	0	0	0	0	0	0	0
	2-0 Female	VTRiIX*	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-
	2-1 Female	HToI	HATo	VTRiIX*	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-
	2-2 Female	HToA	HATo (VCt)X*	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-
	2-3 Female	HToA	HATo	VTRiIX*	-	-	-	-	-	-	-	-	-	-	-	-	-	-	-

0       =  No signs of systemic toxicity                           H =       Hunched posture                                A =        Ataxia                         

T     =   Body tremors                                                    To =       Occasional body tremors                  Ri =       Increased respiratory rate                               

I       =  Increased activity                                              V =        Vocalization                                      Ct =       Tonic convulsions

X*   =   Animal killed for humane reasons due to the occurrence of clinical signs of toxicity that approached the severity limit set forth in the UK Home Office Project

             Licence

-       =   No data, animal dead

()    =  Additional observations noted prior to being humanely killed

Individual Body Weights and Body Weight Changes - 300 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Body Weight (g) at Day			Body Weight (g) at Death	Body Weight Gain (g) During Week
		0	7	14		1	2
300	1-0 Female	156	184	197		28	13
	2-0 Female	186	-	-	184	-	-
	2-1 Female	187	-	-	180	-	-
	2-2 Female	163	-	-	160	-	-
	2-3 Female	183	-	-	179	-	-

- =  Animal dead

Individual Necropsy Findings - 300 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Time of Death	Macroscopic Observations
300	1-0 Female	Killed Day 14	No abnormalities detected
	2-0 Female	Humanely killed Day 1	No abnormalities detected
	2-1 Female	Humanely killed Day 1	No abnormalities detected
	2-2 Female	Humanely killed Day 1	No abnormalities detected
	2-3 Female	Humanely killed Day 1	Liver: Patchy pallor

Individual Clinical Observations and Mortality Data - 50 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Effects Noted After Dosing (Hours)				Effects Noted During Period After Dosing (Days)
		½	1	2	4	1	2	3	4	5	6	7	8	9	10	11	12	13	14
50	3-0 Female	0	H	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4-0 Female	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4-1 Female	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4-2 Female	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	4-3 Female	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0

0 =  No signs of systemic toxicity

H = Hunched posture

Individual Body Weights and Body Weight Changes -50 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Body Weight (g) at Day			Body Weight Gain (g) During Week
		0	7	14	1	2
50	3-0 Female	161	182	193	21	11
	4-0 Female	190	201	217	11	16
	4-1 Female	168	175	185	7	10
	4-2 Female	190	197	212	7	15
	4-3 Female	171	173	187	2	14

Individual Necropsy Findings -50 mg/kg

Dose Level (mg/kg)	Animal Number and Sex	Time of Death	Macroscopic Observations
50	3-0 Female	Killed Day 14	No abnormalities detected
	4-0 Female	Killed Day 14	No abnormalities detected
	4-1 Female	Killed Day 14	No abnormalities detected
	4-2 Female	Killed Day 14	No abnormalities detected
	4-3 Female	Killed Day 14	No abnormalities detected

Applicant's summary and conclusion

Interpretation of results:

Category 3 based on GHS criteria

Conclusions:

The acute oral toxicity test showed an LD50 of between 50-300 mg/kg bw.

Executive summary:

The study was performed to assess the acute oral toxicity of FRET 15 -0735 in the Wistar strain rat. Following a sighting test at a dose level of 300 mg/kg, a further group of four fasted females were given a single oral dose of test item, as a solution in arachis oil BP, at a dose level of 300 mg/kg body weight. Due to mortality and signs of systemic toxicity at a dose level of 300 mg/kg, an additional sighting test was performed at a dose level of 50 mg/kg. This was followed by a further group of four fasted females at a dose level of 50 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.The acute oral LD50 for the substance in female rats was determined to be between 50 -300 mg/kg bw.