Amines, C12-14-alkyl, isooctyl phosphates

Endpoint summary

Description of key information

Acute Oral Toxicity (rat) = 200 - 2000 mg/kg bw, OECD 423 Report RCR/028/002075/AC (2000). According to the OECD 423 test guideline 1996, the LD50 cut-off value was considered to be 1000 mg/kg body weight

Acute Oral Toxicity (rat) = <5000 mg/kg bw, OECD 401 Report MB 84 -7101A (1984)

Acute Oral Toxicity (rat) = >5000 mg/kg bw, OECD 401 Report MB 84 -7206A (1984)

Acute Dermal toxiicty (rabbit) = 2000mg/kg bw OECD 402 Report 480 -2369 (1986)

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 November to 17 December 1999

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.

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Adopted 1996

Deviations:

yes

Remarks:

The low value recorded for humidity (26%) slightly below the range specified was not considered to have affected the integrity or validity of the study.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The low value recorded for humidity (26%) slightly below the range specified was not considered to have affected the integrity or validity of the study.

GLP compliance:

yes

Test type:

acute toxic class method

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Male and Female CD rats Sprague-Dawley origin (Hsd: Sprague-Dawley (CD) obtained from Harlan Laboratories UK Ltd., Bicester, Oxon, UK.
- Age at study initiation: five to seven weeks of age
- Weight at study initiation: 86 - 142 g .
- Fasting period before study: Overnight fast immediately before dosing and for approximately four hours after dosing
- Housing: The animals were housed in groups of three of the same sex in metal cages with wire mesh floors.
- Diet (e.g. ad libitum): Standard laboratory rodent diet (Special diet services RM1(E) SQC expanded pellet) was allowed throughout the study. Each batch of diet used for the study was analysed for certain nutrients, possible contaminants and micro-organisms
- Water (e.g. ad libitum): Free access to mains drinking water was allowed throughout the study. Results of routine physical and chemical examination of drinking water conducted by the supplier are made available to Huntington Life Sciences Ltd.
- Acclimation period: At least five days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Set to achieve limits of 20 to 23°C
- Humidity (%): Set to achieve limits of 26 - 50%
Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study.
- Air changes (per hr): At least fifteen changes per hour
- Photoperiod (hrs dark / hrs light): lighting was controlled by a time switch to give twelve hours continuous light (07:00 to 19:00) and twelve hours darkness.

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

VEHICLE
The test item was formulated at a concentration of either 2% (200mg/kg) or 20% (2000mg/kg) in corn oil.

MAXIMUM DOSE VOLUME APPLIED:
10 ml/kg

DOSAGE PREPARATION (if unusual):
The test item was prepared on the day of dosing.

PROCEDURE
The appropriate dose volume of the test substance was administerd to each rat by oral gavage using a plastic syringe and catheter.

Doses:

200 and 2000 mg/kg

No. of animals per sex per dose:

2000 mg/kg 3 female
200 mg/kg 3 female
200 mg/kg 3 male

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: The animals were observed for mortality twice daily. Animals were observed for clinical signs soon after dosing and thereafter at frequent intervals on the day of dosing. On subsequent days, surviving animals were observed at least once in the morning and again at the end of the experimental day.
- Individual bodyweights were recorded on Days 1 (prior to dosing) and eight and fifteen.
- Necropsy of survivors performed: yes; All surviving animals were killed on Day 15 by carbon dioxide asphyxiation. All animals were subjected to a macroscopic examination which consisted of opening the cranial, thoracic and abdominal cavities. The appearance of all examined organs was recorded.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 200 - <= 2 000 mg/kg bw

Based on:

test mat.

Key result

Sex:

male

Dose descriptor:

LD50

Effect level:

> 200 mg/kg bw

Based on:

test mat.

Mortality:

See attached background information
Two females dosed at 2000 mg/kg died within 3 days of dosing. There were no other deaths.

Clinical signs:

other: See attached background information Piloerection was observed soon after dosing in all animals treated with either 2000 or 200 mg/kg. All females treated with 2000 mg/kg also showed increased salivation, abnormal gait, ungroomed appearance, hunched postur

Gross pathology:

See attached background information
No abnormalities were observed in animals that survived treatment and were terminated on Day 15.
Macroscopic examination of the 2 females treated with 2000 mg/kg and which died within 3 days of dosing revealed congestion of the subcutaneous tissue and brain with atrophy of the spleen and pallor of the kidneys. Congestion, gaseous distension and fluid contents were also noted in the stomach and along the alimentary tract.

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

Under the conditions of the test the acute oral median lethal dose (LD50) of the test item in the Sprague-Dawley strain rat was within the range of 200-2000 mg/kg body weight. According to the OECD 423 test guideline 1996, the LD50 cut-off value was considered to be 1000 mg/kg body weight

Executive summary:

Introduction.

The study was performed to assess the acute oral toxicity of the test item following a single oral administration in the Sprague-Dawley strain rat. The method was designed to be compatible with the following:

- OECD Guidelines for the Testing of Chemicals No. 423 “Acute Oral Toxicity – Acute Toxic Class Method” (adopted 22 March 1996)

- Method B1trisAcute Toxicity (Oral) of EEC Methods for the determination of toxicity, Annex to Directive 96/54/EEC

Method. 

A group of three fasted females was treated with a single oral gavage dose of the test item formulated in corn oil, at a dose level of 2000 mg/kg bodyweight. As a result of mortalities at this dose level indicating the acute lethal oral dose of the test item to be less than 2000 mg/kg bodyweight, a further group of three fasted females was dosed at 200 mg/kg. A group of three fasted males was then dosed at 200 mg/kg to confirm the results at 200mg/kg.

Mortality, clinical signs and bodyweight development were monitored during the study. All animals were subjected to gross necropsy.

Mortality. 

Two females dosed at 2000 mg/kg died within 3 days of treatment. Slight bodweight loss was observed in both animals. Macroscopic examination of both animals revealed congestion of the subcutaneous tissue and brain with atrophy of the spleen and pallor of the kidneys. Congestion, gaseous distension and fluid contents were also noted in the stomach and along the alimentary tract.

Clinical Observations. 

Clinical signs of reaction to treatment comprised piloerection, seen in all rats treated with test item. This sign was accompanied in all rats at 2000 mg/kg by increased salivation, abnormal gait, ungroomed appearance, hunched posture and pink staining on muzzle and uro/genital area with partially closed eyelids in two females at 2000 mg/kg.

Bodyweight. 

The surviving animals showed satisfactory gains in bodyweight over the study period.

Necropsy. 

No abnormalities were observed for animals that survived treatment to study termination on Day 15.

Conclusion. 

The acute oral median lethal dose (LD₅₀) of the test item in the Sprague-Dawley strain rat was estimated to be greater than 200 mg/kg bodyweight but less than 2000 mg/kg bodyweight, (Globally Harmonised Classification System – Acute Oral Toxicity Category 4). According to the OECD 423 test guideline 1996, the LD50 cut-off value was considered to be 1000 mg/kg body weight

The test item was classified as Acute Oral Toxicity Category 4 according to the Regulation (EC) No. 1272/2008. The Signal Word ‘Warning’ and the Hazard Statement ‘H302: Harmful if swallowed’ are therefore required.

Reference 2

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31.05.1984 to 14.06.1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

yes

Remarks:

A multi dose LD50 was not performed

GLP compliance:

yes

Test type:

standard acute method

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Male and Female Wistar rats obtained from Ace Animals.
- Weight at study initiation: 221 - 286 g males. 223 - 259g females.
- Fasting period before study: Overnight fast immediately before dosing and for approximately four hours after dosing
- Housing: The animals were housed 5 /sex/ cage. Cages with wire mesh floors.
- Diet : Fresh Purina Rat Chow (Diet 5012) was freely available except for 16 - 20 hours prior to dosing.
- Water was freely available at all times.
- Acclimation period: At least five days.

ENVIRONMENTAL CONDITIONS
The animal room, reserved exclusivley for rats on acute tests was temperature and humidty controlled, had a 12 hour light/dark cycle and was kept clean and vermin free.

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

PROCEDURE
The appropriate dose volume of the test substance was administerd to each rat by syringe and dosing needle.

Doses:

A multi dose LD50 was not performed. A single dose level of 5g/kg was used.

No. of animals per sex per dose:

5 Males and 5 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations : The animals were observed 1, 2 and 4 hours post dose and once each morning and afternoon for mortality and toxicity.
- Individual bodyweights were recorded on the day of dosing, weekly thereafter, at death and at termination
- Necropsy : All animals were examined for gross pathology. Abnormal tissues were preserved in 10% buffered formalin for possible future microscopic examination.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Mortality:

4 of the 5 females died by day 3. There were no deaths in the males.

Clinical signs:

other: The four females that died exhibited chromorhinorrhea, lethargy, ataxia, tremors, diarrhea, ptsosis, dyspnea, brown staining of body areas and red staining of nose / mouth. Clinical signs of animals that survived included diarrhea, piloerection, ataxia,

Gross pathology:

Necropsy of survivors revealed abnormalities of the stomach in two males and alopecia of the anogenital area in two other males. One male and the one surviving female appeared normal. Necropsy of females that died revealed abnormalities of the lungs, spleen, gastrointestinal tract and red / brown staining of the nose / mouth and brown staining of the anogenital area.

Interpretation of results:

GHS criteria not met

Conclusions:

The study indicated the acute oral LD50 to be greater than 5000mg/kg

Executive summary:

Introduction

The study was performed to determine the acute oral toxicity of the test item following a single oral administration in male and female Wistar strain rat. The study was designed to comply with OECD 401 (adopted 5/12/81)

Method

A group of five fasted males and five fasted females were treated with a single oral gavage dose of the test item.

Mortality, clinical signs and bodyweight development were monitored during the study. All animals were subjected to gross necropsy.

Mortality

Four females died by day 3 of the study.

Clinical observations

The four females that died exhibited chromorhinorrhea, lethargy, ataxia, tremors, diarrhea, ptsosis, dyspnea, brown staining of body areas and red staining of nose / mouth.

Clinical signs of animals that survived included diarrhea, piloerection, ataxia, ptsosis, dyspnea, chromorhinorrhea, lethargy, brown staining of body areas, red staining of nose / mouth area and alopecia surrounding the anogenital area.

Bodyweight

Bodyweight increase in survivors were normal.

Necropsy

Necropsy of survivors revealed abnormalities of the stomach in two males and alopecia of the anogenital area in two other males. One male and the one surviving female appeared normal. Necropsy of females that died revealed abnormalities of the lungs, spleen, gastrointestinal tract and red / brown staining of the nose / mouth and brown staining of the anogenital area.

Conclusion

The LD50 was greater than 5000mg/kg.

Reference 3

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13.03.1984 to 27.03.1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

yes

Remarks:

A multi dose LD50 was not performed

GLP compliance:

yes

Test type:

standard acute method

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Male and Female Wistar rats obtained from Ace Animals.
- Weight at study initiation: 243 - 259 g males. 218 - 240g females.
- Fasting period before study: Overnight fast immediately before dosing and for approximately four hours after dosing
- Housing: The animals were housed 5 /sex/ cage. Cages with wire mesh floors.
- Diet : Fresh Purina Rat Chow (Diet 5012) was freely available except for 16 - 20 hours prior to dosing.
- Water was freely available at all times.
- Acclimation period: At least five days.

ENVIRONMENTAL CONDITIONS
The animal room, reserved exclusivley for rats on acute tests was temperature and humidty controlled, had a 12 hour light/dark cycle and was kept clean and vermin free.

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Procedure
The appropriate dose volume of the test substance was administerd to each rat by syringe and dosing needle.

Doses:

A multi dose LD50 was not performed. A single dose level of 5g/kg was used.

No. of animals per sex per dose:

5 Males and 5 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations : The animals were observed 1, 2 and 4 hours post dose and once each morning and afternoon for mortality and toxicity.
- Individual bodyweights were recorded on the day of dosing, weekly thereafter, at death and at termination
- Necropsy : All animals were examined for gross pathology. Abnormal tissues were preserved in 10% buffered formalin for possible future microscopic examination.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

< 5 000 mg/kg bw

Based on:

test mat.

Mortality:

3 of the 5 males died and all females died

Clinical signs:

other: Two males and five females that died by Day 3 had signs of dyspnea, tremors, ptosis, chromodacryorrhea, ataxia, lethargy, chromorhinorrhea, piloerection, convulsions diarrhea, brown staining of body areas, wetness of nose / mouth area and hyperactivity.

Gross pathology:

Necropsy of animals that died revealed abnormalities of the lungs, liver, kidneys, and gastrointestinal tract as well as red or brown staining of body areas and yellow staining of body fat.
Necropsy of survivors were normal in one animal. Adhesions in the peritoneal cavity were noted in two survivors.

Interpretation of results:

study cannot be used for classification

Conclusions:

The study indicated the acute oral LD50 to be less than 5000mg/kg

Executive summary:

Introduction

The study was performed to determine the acute oral toxicity of the test item following a single oral administration in male and female Wistar strain rat. The study was designed to comply with OECD 401 (adopted 5/12/81)

Method

A group of five fasted males and five fasted females were treated with a single oral gavage dose of the test item.

Mortality, clinical signs and bodyweight development were monitored during the study. All animals were subjected to gross necropsy.

Mortality

Two male and five females died by day 3 of the study.

Clinical observations

Animals that died had signs of dyspnea, tremors, ptosis, chromodacryorrhea, ataxia, lethargy, chromorhinorrhea, piloerection, convulsions, diarrhea, brown staining of body areas, wetness of nose / mouth area and hyperactivity. Clinical signs noted in survivors were similiar to those noted in animals which died. Two survivors appeared normal by Day 14. Brown staining of the anogenital area and alopecia of the anogenital area were noted in one surviving male on Day 14.

Bodyweight

Bodyweight increase in survivors were normal.

Necropsy

Necropsy of animals that died revealed abnormalities of the lungs, liver, kidneys, and gastrointestinal tract as well as red or brown staining of body areas and yellow staining of body fat. Necropsy of survivors were normal in one animal. Adhesions in the peritoneal cavity were noted in two survivors.

Conclusion

The LD50 was less than 5000mg/kg.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

1 000 mg/kg bw

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08/04/1986 to 22/04/1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

yes

Remarks:

A multi dose LD50 was not performed

GLP compliance:

yes

Test type:

standard acute method

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Young adult male and female albino New Zealand rabbits obtained from Clerco Research Farm, CincinNati, OH, USA
- Housing: The animals were housed individually in stainless steel cages with wire mesh floors.
- Diet (e.g. ad libitum): Purina Certified Rabbit Chow 5322 was fed throughout the study.
- Water (e.g. ad libitum): Free access to filtered tap water was provided through an automatic watering system.
- Acclimation period: 7 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 67 ± 5 F
- Humidity (%): 30 - 70%
- Ventilation: The animal room was well ventilated and air conditioned
- Photoperiod (hrs dark / hrs light): lighting was controlled by automatic timers to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.

Type of coverage:

occlusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
Approximately 24 hours before treatment the dorsal and lateral trunk (approximately 10% of the body surface area) of each animal were clipped free of hair.

On the day of treatment bodyweights were recorded and doses calculated. An impervious binder consistng of plastic wrap and adhesive tape was applied around each animal's trunk and a measured volume of the test article was introduced under the binder and spread over the application site. The entire trunk of each animal was then wrapped with additional adhesive tape and masking tape.

After a 24 Hour exposure period each binder was removed and the test site of each animal was wiped with gauze sponges moistened with propylene glycol to remove remaining test item.

Duration of exposure:

24 hours

Doses:

2000mg/kg. A multi dose LD50 was not performed

No. of animals per sex per dose:

5 males and 5 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days

- Frequency of observations and weighing:
The animals were observed frequently on the day of treatment for deaths or overt signs of toxicity and subsequently once daily for fourteen days.

Individual bodyweights were recorded prior to application of the test item on Day 0 and on Days 7 and 14.

- Necropsy of survivors performed: yes
At the end of the study the animals were killed by intravenous injection of barbiturate and exanguinated prior to necropsy. Animals found dead during the study were necropsied as soon as possible after death. 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.

Key result

Sex:

male/female

Dose descriptor:

approximate LD50

Effect level:

ca. 2 000 mg/kg bw

Based on:

test mat.

Mortality:

One female was found dead on Days 2 and 6. Two males and one females were found dead on Day 7.

Clinical signs:

other: Observations noted during the study were erythema, edema, black / brown disclouration, atonia, eschar formation, fissures and or disquamation of the test sites, muscle tremors, few / no stools, loose stools, no urine, food appeared undisturbed, emaciation

Gross pathology:

Necropsy of animals found dead revealed treated skins that were discolored red, black, or red black, thickened, crusted and / or stiff; black disclourations on the mucosa of stomachs; dark contents of cecums; tan disclourations on the surface of the left lateral lobe of one liver; fluid content of small intestine; and / or urinary bladder distended with gelatinous amber fluid.

Necropsy of surviving animals revealed linear abrasions, crusting sloughing epidermis, ulcerative dermatitis, thickened areas with hyperemic borders, multiple scabs, and / or underlying granulation tissue on the treated area of the back

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

Under the conditions of this study, the acute dermal LD50 of the test material was considered to be approximately 2000 mg/kg body weight.

Executive summary:

Method

The test material was applied topically to five male and five female New Zealand White Rabbits at a dose level of 2000 mg/kg body weight for a period of 24 hours. The animals were observed for pharmacotoxic signs and mortality for 14 days following the day of treatment.

Mortality

Two males and three females died following treatment. Deaths ocurred on Days 2, 6, and 7 after test material application.

Clinical signs and Bodweight

Observation noted during the study were erythema, edema, black / brown disclouration, atonia, eschar formation, fissures and or disquamation of the test sites, muscle tremors, few / no stools, loose stools, no urine, food appeared undisturbed, emaciation, lethargy, salivation, ataxia, red stained fur around test sites poor coat quality.

All surviving animals except one gained weight and one female had no overall weight gain

Necropsy

Necropsy of animals found dead revealed treated skins that were discolored red, black, or red black, thickened, crusted and / or stiff; black disclourations on the mucosa of stomachs; dark contents of cecums; tan disclourations on the surface of the left lateral lobe of one liver; fluid content of small intestine; and / or urinary bladder distended with gelatinous amber fluid. Necropsy of surviving animals revealed linear abrasions, crusting sloughing epidermis, ulcerative dermatitis, thickened areas with hyperemic borders, multiple scabs, and / or underlying granulation tissue on the treated area of the back.

Conclusion

Under the conditions of this study, the acute dermal LD50 of the test material was considered to be approximately 2000 mg/kg body weight.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Additional information

The substance was tested for acute oral toxicity in three different studies. One study according to acute toxic class method (OECD 423 1996) indicated an LD50 of >200 <2000 mg/kg bw. According to the OECD 423 test guideline 1996, the LD50 cut-off value was considered to be 1000 mg/kg body weight. Two of studies used the OECD 401 method at one fixed dose of 5000mg/kg bw and reported LD50's of <5000mg/kg bw and >5000mg/kg bw. The OECD 423 study was the most recent study and was considered the most reliable and was used for classification.

An acute dermal toxicity study according to OECD 402 reported an LD50 of 2000mg/kg.

Justification for classification or non-classification

 According to the criteria for classification and labelling of substances and mixtures (Regulation (EC) No. 1272/2008), the substance qualifies for classification as “Acutely Toxic Category 4” (H302) and (H312).