Phenol, heptyl derivs.

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Between 05 January 2012 and 07 February 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP and in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not effect the quality of the relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The bodyweight at death of one animal treated at a dose level of 2000 mg/kg was not recorded.

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The bodyweight at death of one animal treated at a dose level of 2000 mg/kg was not recorded.

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Phenol, heptyl derivs.

EC Number:

276-743-1

EC Name:

Phenol, heptyl derivs.

Cas Number:

72624-02-3

Molecular formula:

C13H20O

IUPAC Name:

4-(2-methylhexyl)phenol; 4-heptylphenol

Details on test material:

Description : amber coloured viscous liquid
Purity : not supplied
Date received : 01 December 2011
Expiry date : 01 December 2012
Storage conditions: room temperature in the dark

The integrity of supplied data relating to the identity, purity and stability of the test item is the responsibility of the Sponsor.

A Certificate of Analysis supplied by the Sponsor is given in Appendix 3 (attachment 1)

Test animals

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

Female Wistar (RccHan™:WIST) strain rats were supplied by Harlan Laboratories UK Ltd., Oxon, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non-pregnant. After an acclimatisation period of at least five 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 eight to twelve weeks of age. The bodyweight variation did not exceed ±20% of the bodyweight of the initially dosed animal.
The animals were housed in groups of up to four in suspended solid floor polypropylene cages furnished with woodflakes. With the exception of an overnight fast immediately before dosing and for approximately three to four hours after dosing, free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Harlan Laboratories UK Ltd., Oxon, UK) was allowed throughout the study. The diet, drinking water and bedding were routinely analysed 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. Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve 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:

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 bodyweight at the time of dosing. Treatment of animals was sequential. Sufficient time was allowed between each dose level to confirm the survival of the previously dosed animals.

Doses:

Using available information on the toxicity of the test item, 2000 mg/kg was chosen as the starting dose.

A single animal was treated as follows:
Dose Level (mg/kg) Specific Gravity Dose Volume (ml/kg) Number of Rats (female)
2000 0.948 2.11 1

In the absence of mortality in the initial animal at a dose level of 2000 mg/kg, an additional group of animals was treated as follows:
Dose Level (mg/kg) Specific Gravity Dose Volume (ml/kg) Number of Rats (female)
2000 0.948 2.11 4

Due to mortality and signs of systemic toxicity at a dose level of 2000 mg/kg, an additional animal was treated as follows:
Dose Level (mg/kg) Concentration (mg/ml) Dose Volume (ml/kg) Number of Rats (female)
300 30 10 1

In the absence of mortality at a dose level of 300 mg/kg, an additional group of animals was treated as follows:
Dose Level (mg/kg) Concentration (mg/ml) Dose Volume (ml/kg) Number of Rats (female)
300 30 10 4

No. of animals per sex per dose:

5 female at 2000 mg/kg
5 females at 300 mg/kg

Control animals:

no

Details on study design:

Experimental Preparation
For the purpose of the 2000 mg/kg dose level, the test item was used as supplied. The specific gravity was determined and used to calculate the appropriate dose volume for the required dose level. For the purpose of the 300 mg/kg dose level, 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 two 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.

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 bodyweight at the time of dosing. Treatment of animals was sequential. Sufficient time was allowed between each dose level to confirm the survival of the previously dosed animals.
Clinical observations were made ½, 1, 2, and 4 hours after dosing and then daily for up to fourteen days. Morbidity and mortality checks were made twice daily.
Individual bodyweights were recorded on Day 0 (the day of dosing) and on Days 7 and 14 or at death. Due to a technician error, the bodyweight at death of one animal treated at a dose level of 2000 mg/kg was not recorded. This deviation from the Study Plan was considered not to affect the purpose or integrity of the study.
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

Preliminary study:

None

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: 95% confidence limits not given in study report.

Mortality:

Dose Level - 2000 mg/kg
Four animals were found dead one or two days after dosing.

Dose Level - 300 mg/kg
There were no deaths.

Clinical signs:

other: Dose Level - 2000 mg/kg Signs of systemic toxicity noted were increased salivation, hunched posture, pilo erection, ataxia, gasping or noisy respiration, lethargy, hypothermia, ptosis, diarrhoea and dehydration. Thickening of the skin on the top of the h

Gross pathology:

Dose Level - 2000 mg/kg
Individual necropsy findings are given in Table 3.
Abnormalities noted at necropsy of animals that died during the study were abnormally red lungs, dark liver, dark kidneys, gaseous stomach, haemorrhage and epithelial sloughing of the gastric mucosa, haemorrhagic and reddened non glandular epithelium of the stomach, gaseous and fluid filled small intestine and off white substance adhered to the lining of the gastric mucosa and non glandular epithelium of the stomach. Ulcerated non glandular epithelium of the stomach was noted at necropsy of the animal that was killed at the end of the study.

Dose Level - 300 mg/kg
Individual necropsy findings are given in Table 6.
No abnormalities were noted at necropsy.

Other findings:

None

Any other information on results incl. tables

Data evaluations included the relationship, if any, between the animals' exposure to the test item and the incidence and severity of all abnormalities including behavioural and clinical observations, gross lesions, bodyweight changes, mortality and any other toxicological effects. If possible the signs of evident toxicity were also identified. Evident toxicity is defined as the toxic effects which are of a severity such that administration at the next highest level could result in mortality.

Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD₅₀) of the test item was made.

Table 1              Individual Clinical Observations and Mortality Data - 2000mg/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
2000	1-0 Female	0	0	0	0	0	0	0	0	0	0	0*	0*	0*	0*	Fl*	Fl^·	Fl^·	Fl^·
	2-0 Female	S	H	H	H	HPA	X
	2-1 Female	0	0	0	0	X
	2-2 Female	RnS	RnH	RnRg SH	HAPL	X
	2-3 Female	RnS	RnH	H	H	HoPPtDL AHDhX

0=     No signs of systemic toxicity

S =      Increased salivation

H =      Hunched posture

P =      Pilo-erection

A =      Ataxia

Rg =    Gasping respiration

Rn =    Noisy respiration

L =      Lethargy

Ho =    Hypothermia

Pt =     Ptosis

D =      Diarrhoea

Dh =    Dehydration

* =       Thickening of the skin on the top of the head and left ear

·=       Redness of the area where the thickening of the skin was observed

Fl =     Fur loss

X =      Animal dead

Table 2              Individual Bodyweights and Bodyweight Changes - 2000mg/kg

Dose Level mg/kg	Animal Number and Sex	Bodyweight (g) at Day			Bodyweight (g) at Death	Bodyweight Gain (g) During Week
		0	7	14		1	2
2000	1-0 Female	159	177	199		18	22
	2-0 Female	165	-	-	139	-	-
	2-1 Female	178	-	-	163	-	-
	2-2 Female	189	-	-	178	-	-
	2-3 Female	185	-	-	Ä	-	-

Table 3              Individual Necropsy Findings - 2000mg/kg

Dose Level mg/kg	Animal Number and Sex	Time of Death	Macroscopic Observations
2000	1-0 Female	Killed Day 14	Non-glandular epithelium of the stomach: ulcerated
	2-0 Female	Found dead Day 2	Liver: dark Kidneys: dark Stomach: gaseous Gastric mucosa: haemorrhagic                          epithelial sloughing                          off white substance adhered to stomach lining Non-glandular epithelium of the stomach:   haemorrhagic                                                               reddened
	2-1 Female	Found dead Day 1	Lungs: abnormally red Liver: dark Kidneys: dark Stomach: gaseous Gastric mucosa: epithelial sloughing                          off white substance adhered to stomach lining Non-glandular epithelium of the stomach:   reddened Small intestine: gaseous                        fluid filled (clear yellow)

2000	2-2 Female	Found dead Day 1	Lungs: abnormally red Liver: dark Kidneys: dark Stomach: gaseous Gastric mucosa: epithelial sloughing Non-glandular epithelium of the stomach: reddened                                                             off white substance adhered to stomach lining Small intestine: gaseous                        fluid filled (opaque, off white)
	2-3 Female	Found dead Day 1	Liver: dark Kidneys: dark Stomach: gaseous Gastric mucosa: haemorrhagic                          epithelial sloughing Non-glandular epithelium of the stomach: reddened

Table 4              Individual Clinical Observations and Mortality Data - 300mg/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	3-0 Female	0	0	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

Table 5              Individual Bodyweights and Bodyweight Changes - 300mg/kg

Dose Level mg/kg	Animal Number and Sex	Bodyweight (g) at Day			Bodyweight Gain (g) During Week
		0	7	14	1	2
300	3-0 Female	186	208	221	22	13
	4-0 Female	151	180	196	29	16
	4-1 Female	159	183	198	24	15
	4-2 Female	184	210	221	26	11
	4-3 Female	170	189	203	19	14

Table 6              Individual Necropsy Findings - 300mg/kg

Dose Level mg/kg	Animal Number and Sex	Time of Death	Macroscopic Observations
300	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 4 based on GHS criteria

Remarks:

Migrated information

Conclusions:

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be in the range of 300 - 2000 mg/kg bodyweight (Globally Harmonised Classification System - Category 4).

Executive summary:

Introduction.

The study was performed to assess the acute oral toxicity of the test item in the Wistar strain rat. The method was designed to be compatible with the following:

 OECD Guidelines for Testing of Chemicals No 420 “Acute Oral Toxicity - Fixed Dose Method” (2001)

 Method B1bisAcute Toxicity (Oral) of CommissionRegulation (EC) No. 440/2008

Method.

Following a sighting test at dose levels of2000 mg/kg and300 mg/kg, a further group of four fasted females was given a single oral dose of test item, as asolutioninarachis oil BP, at a dose level of300 mg/kg bodyweight. Clinical signs and bodyweight development were monitored during the study. All animals were subjected to gross necropsy.

Mortality.

Four animals treated at a dose level of 2000 mg/kg were found dead one or two days after dosing. There were no deaths noted at a dose level of 300 mg/kg.

Clinical Observations.

Signs of systemic toxicity noted in animals treated at a dose level of 2000 mg/kg were increased salivation, hunched posture, pilo-erection, ataxia, gasping or noisy respiration, lethargy, hypothermia, ptosis, diarrhoea and dehydration. Thickening of the skin on the top of the head and left ear, fur loss and redness were noted in one animal treated at a dose level of 2000 mg/kg. There were no signs of systemic toxicity noted at a dose level of 300 mg/kg.

Bodyweight.

Surviving animals showed expected gains in bodyweight.

Necropsy.

Abnormalities noted at necropsy of animals that died during the study were abnormally red lungs, dark liver, dark kidneys, gaseous stomach, haemorrhage and epithelial sloughing of the gastric mucosa, haemorrhagic and reddened non-glandular epithelium of the stomach, gaseous and fluid filled small intestine and off white substance adhered to the lining of the gastric mucosa and non‑glandular epithelium of the stomach. Ulcerated non-glandular epithelium of the stomach was noted at necropsy of one animal treated at a dose level of 2000 mg/kg that was killed at the end of the study. No abnormalities were noted at necropsy of animals treated at a dose level of 300 mg/kg.

Conclusion. 

The acute oral median lethal dose (LD₅₀) of the test item in the female Wistar strain rat was estimated to be in the range of 300 ‑ 2000 mg/kg bodyweight (Globally Harmonised Classification System-Category 4).