Salts of 2-ethylhexyl phosphate esters with (Z)- octadec-9-enylamine

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 October - 08 November 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted in accordance with international guidelines and in accordance with GLP. All guideline validity criteria were met.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2001

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)

Version / remarks:

Commission Regulation (EC) No. 440/2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Salts of 2-ethylhexyl phosphate esters with (Z)- octadec-9-enylamine

EC Number:

949-054-5

Molecular formula:

not applicable, UVCB substance

IUPAC Name:

Salts of 2-ethylhexyl phosphate esters with (Z)- octadec-9-enylamine

Test material form:

liquid: viscous

Test animals

Species:

rat

Strain:

Wistar

Remarks:

RccHan:WIST

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Envigo RMS (UK) Limited, Oxon, UK.
- Females (if applicable) nulliparous and non-pregnant: Yes
- Age at study initiation:
8-12 weeks
- Weight at study initiation:
Body weight variation did not exceed ±20% of the mean body weight.
- Fasting period before study:
Yes, overnight fast prior to dosing.
- Housing: The animals were housed in groups of up to four in suspended solid-floor polypropylene cages furnished with softwood flake bedding.
- Diet (e.g. ad libitum):
Free access to feed (apart from during fasting period); 2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK.
- Water (e.g. ad libitum):
Free access to mains drinking water throughout the study.
- Acclimation period:
At least 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C):
19-25 ºC
- Humidity (%):
30-70 % RH
- Air changes (per hr):
≥15 air changes per hour.
- Photoperiod (hrs dark / hrs light):
12:12 h

IN-LIFE DATES: Not reported.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

VEHICLE
- Concentration in vehicle: N/A - test item applied undiluted and as supplied.
- Amount of vehicle (if gavage): N/A
- Justification for choice of vehicle: N/A
- Lot/batch no. (if required): N/A
- Purity: N/A

MAXIMUM DOSE VOLUME APPLIED: 2.16 mL/kg.

DOSAGE PREPARATION (if unusual): N/A - test item applied undiluted and as supplied.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: N/A - fixed dose limit test.

Doses:

2000 mg/kg

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: 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.
- Necropsy of survivors performed: Yes
- Other examinations performed: clinical signs, body weight, gross necropsy (consisting of an external examination and opening of the abdominal and thoracic cavities. Macroscopic abnormalities were recorded)

Statistics:

N/A

Results and discussion

Preliminary study:

A single animal was treated at 2000 mg/kg as the starting dose. In the absence of mortality at a dose level of 2000 mg/kg, an additional group of 4 animals was treaed at 2000 mg/kg.

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

One animal was killed for humane reasons, 8 days after dosing, due to the occurrence of clinical signs of toxicity that approached the severity limit set forth in the UK Home Office Project Licence.

Clinical signs:

other: Hunched posture was noted in all animals. Staining around ano-genital region was also noted in three animals. Ataxia, lethargy and pilo-erection were confined to one animal.

Gross pathology:

Abnormalities noted at necropsy of the animal that was humanely killed during the study were small left kidney (in comparison to the right), epithelial sloughing and thickened non-glandular epithelium of the stomach, gaseous distension of the small and large intestines and light brown fluid filled abdominal cavity. No abnormalities were noted at necropsy of animals that were killed at the end of the study.

Any other information on results incl. tables

Table 2       Individual clinical observations and mortality data

 

Dose level (mg/kg)	Animal number and sex	Effects noted after dosing (hours)					Effects noted during period after dosing (days)
		0.5	1	2	4	1		2	3	4	5	6	7	8	9	10	11	12	13	14
2000	1-0 F	0	0	0	0	0		0	0	0	0	0	HSa	HSa	H	H	H	H	0	0
	2-0 F	0	H	H	0	0		0	0	0	0	0	0	0	0	0	0	0	0	0
	2-1 F	H	H	H	0	0		0	0	0	HSa	HSa	H	H	H	H	0	0	0	0
	2-2 F	H	H	H	0	0		0	0	0	0	0	0	0	0	0	0	0	0	0
	2-3 F	0	H	H	0	0		0	0	0	HSa	HSa	HPA	HLAX

0: no sign of systemic toxicity

A: ataxia

H: hunched posture

L: lethargy

P: pilo-erection

Sa: staining around ano-genital region

X: animal humanely killed due to occurrence of clinical signs of toxicity that approached the severity limit set forth in the UK Home Office Project Licence

 

Table 3       Individual body weight and body weight changes

 

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
2000	1-0 F	152	149	167	-	-3	18
	2-0 F	158	169	180	-	11	11
	2-1 F	158	149	170	-	-9	21
	2-2 F	178	186	205	-	8	19
	2-3 F	165	136	-	129	-29	-

 

Table 4       Individual necropsy findings

 

Dose level (mg/kg)	Animal number and sex	Time of death	Macroscopic observations
2000	1-0 F	Killed Day 14	No abnormalities detected
	2-0 F	Killed Day 14	No abnormalities detected
	2-1 F	Killed Day 14	No abnormalities detected
	2-2 F	Killed Day 14	No abnormalities detected
	2-3 F	Humanely killed on Day 8	Kidneys: small left kidney, compared to right. Non-glandular epithelium of the stomach: epithelial sloughing; thickened. Abdominal cavity: fluid filled (light brown colour). Small intestine: gaseous distention. Large intestine: gaseous distention.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2000 mg/kg body weight. Test item does not meet the GHS criteria for classification in accordance with CLP Regulation (EC) No. 1272/2008.

Executive summary:

OECD 420 (2018) - In an acute oral toxicity study (OECD 420), five fasted, 8-12 week old, female Wistar strain rats were given a single oral dose of test item at dose level of 2000 mg/kg bw (limit test). 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.  After the 14 days observation, all surviving animals underwent necropsy.

Mortality

 

One animal was humaely killed for humane reasons on Day 8 due to occurrence of toxicity that approached the severity limit set forth in the UK Home Office Project Licence. No mortality was observed in the other 4 animals.

 

Clinical observations

 

Hunched posture was noted in all animals. Staining around the ano-genital region was noted in 3/5 animals. Ataxia, lethargy and pilo-erection were confined to the one animal that was humanely killed on Day 8.

 

Body weight

 

Two animals showed body weight loss during the first week with expected body weight gain in the second. Two animals showed expected body weight gain.

 

Necropsy

 

Abnormalities were confined to the animal that was humanly killed on Day 8. Abnormalities included; small left kidney (compared to the right); epithelial sloughing and thickened non-glandular epithelium of the stomach; gaseous distention of the large and small intestines an light-brown filled abdominal cavity. No abnormalities were noted in the other 4 animals.

 

The acute oral median dose level (LD50) of the test item was estimate to be greater than 2000 mg/kg bw. The test item does not meet the GHS criteria for classification in accordance with CLP Regulation (EC) No. 1272/2008.