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Description of key information

The LD 50 of tert-amyl hydroperoxide is 500 mg/kg in Sprague Dawley rats (Doubs, 1981) and is more generally between 450 mg/kg to 864 mg/kg in rodents. The dermal LD 50 is 446 mg/kg in rats (Doubs, 1995). The 4-hour LC 50 is 2425 mg/m3 [IC 95: 2100 -2750 mg/m3] (TNO, 1981).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1994-1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Young adults rats were received at SLS from Charles River Laboratories, lnc., Portage, Michigan. 200 to 300 g prior to the study
Method of identification: Upon receipt, metal ear tags displaying unique identification numbers were used to individually identify the animais.
Housing: The animals were housed individually in suspended stainless steel cages.
Environment: The animal room temperature and relative humidity ranges were 63-71° F and 35-55%, respectively. Environmental control equipment was monitored and adjusted as necessary to minimize fluctuations in the animal room environment. Light timers were set to maintain a 12-hour light/12-hour dark cycle. There were ten to twelve air changes in the animal room per hour. The animal room temperature and relative humidity were recorded a minimum of once daily.
Food and water ad libitum
Quarantine: 5 days at minimum
Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Doses:
0.27, 0.55 and 1.10 ml/kg (equivalent to 250, 500 and 1000 mg test mat./kg bw/day based on a density of 0.91)
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: two times on study day 0 (postdose) and daily thereafter (days 1- 14). A mortality check was performed twice daily, in the morning and afternoon.
- Weighing: prior to fasting (day -1), prior to dosing on day 0 and for all surviving animals on days 7 and 14 and at the time of death for any animal dying on study.
- All study animals which died spontaneously during the study or were euthanized (carbon dioxide inhalation) at study termination (day 14) were necropsied. Body cavities (cranial, thoracic, abdominal and pelvic) were opened and examined. No tissues were retained.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
500 mg/kg bw
Based on:
test mat.
95% CL:
418 - 598
Sex:
male
Dose descriptor:
LD50
Effect level:
518 mg/kg bw
Based on:
test mat.
95% CL:
383 - 700
Sex:
female
Dose descriptor:
LD50
Effect level:
483 mg/kg bw
Based on:
test mat.
95% CL:
357 - 652
Mortality:
Dose Males Females Males/females
250 0/5 0/5 0/10
500 2/5 3/5 5/10
1000 5/5 5/5 10/10

Rats died at day 1 or 2
Clinical signs:
Salivation was present in all groups on study day 0 only.
Abnormal breathing and urine stain were present in all groups on study days 0-2 (except for one animal in the 500 mg/kg dose group who had urine stain on study days.
Decreased activity, dehydration, reddish-colored urine, decreased/no defecation and dark material around the facial area were present in the 250 and 500 mg/kg dose groups on study days 0-3.
Rough hair coat was present in the 250 and 500 mg/kg dose groups on study days 0-6.
Hunched posture.
Piloerection and wobbly gait were present in the 500 mg/kg dose group on study days 0-2; partially closed eye lids was present in the 500 and
1000 mg/kg dose groups; tremors, lacrimation, apparent hypothermia, and prostration were only present in the animals that died during the study.
1-5)
Body weight:
No abnormal gain body weight.
Gross pathology:
The most notable gross internal findings were observed in the animals that died and included congested meningeal vessels in the brain, mottled livers, abnormal contents in the digestive tract, smooth mucosa, linear striations and dark red foci in the stomach, reddened mucosa in the small intestines, dark red and thickened serosa in the stomach, discolored thymus, abnormally colored fluid contents in the urinary bladder and thoracic cavity and musculature and viscera in the whole body stained orange.

                                                                                                                          5/5


 

 

 

 

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Under the conditions of this test, the acute oral LD50 of t-Amyl Hydroperoxide in the male rat was determined to be 518 mg/kg. ln the female rat, the oral LD50 was determined to be 483 mg/kg. ln the sexes combined, the oral LD50 was determined to be 500 mg/kg.
Executive summary:

The Acute oral toxicity of tert-amyl hydroperoxide was evaluated in rats according a method similar to OECD N°401 guideline (Acute Toxic Standard Method), GLP study compliant. Groups of 5 male and 5 female Sprague Dawley rats were given a single oral dose of tert-amyl hydroperoxide at doses of 0, 250, 500, 1000 mg/kg. Following treatment, rats were observed daily and weighted weekly. A gross necropsy examination was performed at the time of scheduled euthanasia (Day 14) or when animals were found dead.

No mortality occurred in the lowest dose group. 5 animals died before day 2 after administration of 500 mg/kg (2 males and 3 females). All animals of the highest dose group died one day after oral administration of the substance. Clinical signs observed were [hypoactivity, salivation, piloerection, abnormal breathing, urine stain, wobbly gate] for most of animals; and tremor, prostration, apparent hypothermia in animals who died.

The most notable gross internal findings were observed in the animals that died and included congested meningeal vessels in the brain, mottled livers, abnormal finding in the digestive tract, discolored thymus, abnormally colored fluid contents in the urinary bladder and thoracic cavity and musculature and viscera in the whole body stained orange.

Under these experimental conditions, the oral LD50 of tert-amyl hydroperoxide is 500 mg/kg in Sprague Dawley rats (517 mg/kg in males, 483 mg/kg in females).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
500 mg/kg bw
Quality of whole database:
Klimisch 1 study, GLP compliant.

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study acceptable for assessment. No data on analytical method was provided and there is no certificat of analysis.
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
GLP compliance:
not specified
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Rats were obtained from the Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands.
Just prior to exposure the weight variation of the animals was smaller than ± 20% of the mean body weight.
The animals were housed 5 to a cage. The cages were suspended in an open rack in an animal room. The animals received ad libitum the Institute's
stock diet for rats and bottled unfluoridated water. The temperature and the relative humidity in the animal room were set and controlled at 21 ± I°C
and 50 - 60% respectively.


Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Details on inhalation exposure:
Tert-Amylhydroperoxide aerosol was generated from a stainless (steel/glass air nebulizer, which was fitted with a baffle to remove the large droplets by impaction. The aerosol generated was passed into the inhalation chamber were it was mixed with an additional airflow.
The stainless steel exposure chamber, having a capacity of 1.5 m3, was used. The chamber was operated under dynamic airflow conditions. The
total airflow through the chamber ranged from 11 to 16 m3/hour. Through a glass door at the front of the exposure chamber the animals could be observed continuously. The front door is provided with several openings for taking samples of the test atmosphere at different locations.

The nominal concentration was measured by dividing the total quantity of the test material used by the total quantity of air which had been passed through the inhalation chamber during the exposure period.

The actual concentrations were determined: A measured quantity of test atmosphere was passed through an impinger filled with an absorbent (xylene). The quantity of tert-amylhydroperoxide in the absorbent was analysed by method no. Jo/73.6, d.d.10.07.1975, provided by the sponsor. The actual concentration in the test atmosphere was calculated from the quantity of Tert- Amylhydro­peroxide in the absorbent and the total volume of air sampled.

Particle size determinations and counts were carried out in samples taken from the test atmosphere with a cascade impactor.


Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
1580, 2100, 2230, 2375 and 2890 mg/m3
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
The animals were housed in wire mesh stainless steel cages which were located at one level in the exposure chamber. Individual housing was used to minimize filtration of inspired air by the animals fur as a result of crowding. During the exposure the animals were deprived of water and food.

After the exposure the survivals were returned to their living cages for an observation period of two weeks.

Observations:
During the exposure and the consecutive observation period all reactions to treatment or mortality were observed and recorded daily.
Body weights were recorded at days 1, 2, 4, 7 and 14. At the end of the
observation period the animals were killed by exsanquination of the abdominal aorta under ether anaesthesia, autopsied and examined for gross pathological changes.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
2 425 mg/m³ air (analytical)
Based on:
other: vapors of test mat.
95% CL:
2 100 - 2 750
Exp. duration:
4 h
Mortality:
All animals that died, died within a period of 2 days after starting the exposure, except for one male of the 2890 mg/m 3 air group, which died on day 3.
Clinical signs:
other: After about one hour of exposure the animals showed mouth breathing and after about 2 hours they showed also laboured respiration. Twelve hours after termination of the exposure all surviving animals showed normal respiration pattern.
Body weight:
Both males and females lost body weight during the first 24 hours after starting the exposure. Thereafter they gained weight in a normal way.
Gross pathology:
No abnormalities that were considered to be treatment-related, neither in died animal nor in no-died animals

Analytical data:

Exposure n° Nominal concentration (mg/m3) Actual concentration (mg/m3)* IC 95%
1 1620 1580 (1420 - 1740)
2 1990 2100 (1820 - 2260)
3 2240 2230 (1980 - 2360)
4 2320 2375 (2040 - 2660)
5 2520 2890 (2460 - 3120)

* All attempts to determine particle size distribution were unsuccesful most probably as a result of the relative high volatility of the test

material. Animals were most probably exposed to vapors.

Mortality:

Exposure n° Actual concentration (mg/m3) Mortality _
1 1580 0% _
2 2100 20% 10 % at the end of the exposure and 10% at the end of day 1
3 2230 60% 50 % at the end of day 1 and 10% at the end of day 2
4 2375 70% 10 % at the end of the exposure, 20% at the end of day 1, and 40% at the end of day 2
5 2890 50%  20% at the end of day 1, 20% at the end of day 2 and 10 % at the end of day 3
Interpretation of results:
Category 3 based on GHS criteria
Conclusions:
The 4-hour LC 50 of tert-amylhydroperoxide in rats was calculated to be 2425 mg/m3 air with 2100 and 2750 mg/m3 air as the 95 % confidence limits.
Executive summary:

Five groups of 5 male and 5 female rats were whole-body exposed for 4 hours to vapors of ter-amyl hydroperoxide. Dermal exposure was minimized. The exposure concentrations were 1580, 2100, 2230, 2375 and 2890 mg/m3. Mortality occured mainly after the end of the exposure. At necropsy, no gross abnormality was observerd in any rats. The 4-hour LC50 of tert-amylhydroperoxide in rats, whole-body exposed, was calculated to be 2425 mg/m3 air with 2100 and 2750 mg/m3 air as the 95% confidence limits.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
Value:
2 425 mg/m³
Quality of whole database:
Klimisch 2 study, no data on GLP compliance.

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:
1994-1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Young adults rats were received at SLS from Charles River Laboratories, lnc., Portage, Michigan. 200 to 300 g prior to the study
Method of identification: Upon receipt, metal ear tags displaying unique identification numbers were used to individually identify the animais.
Housing: The animais were housed individually in suspended stainless steel cages.
Environment: The animal room temperature and relative humidity ranges were 65-70° F and 18-65%, respectively. Environmental control equipment was monitored and adjusted as necessary to minimize fluctuations in the animal room environment. Light timers were set to maintain a 12-hour light/12-hour dark cycle. There were ten to twelve air changes in the animal room per hour. The animal room temperature and relative humidity were recorded a minimum of once daily.
Food and water ad libitum
Quarantine: 5 days at minimum
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: dorsal trunk after the fur was removed (on day 1)
- % coverage: 10% of the animal's body surface are
- Type of wrap if used :porous gauze dressing, which was backed by plastic wrap (occlusive binding)

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, after removal of the dressing

TEST MATERIAL
- Amount(s) applied (volume or weight with unit):
- Concentration (if solution):
- Constant volume or concentration used: yes/no
- For solids, paste formed: yes/no

VEHICLE
- Amount(s) applied (volume or weight with unit):
- Concentration (if solution):
- Lot/batch no. (if required):
- Purity:
Duration of exposure:
24 hours
Doses:
0.27; 0.55; 0.82 ml/kg bw (equivalent to 250, 500 and 750 mg/kg bw based on a density of 0.91)
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 : LD50 study animals were observed for clinical abnormalities a minimum of two times on study day 0 (postdose) and daily thereafter (days 1-14). A mortality check was performed twice daily, in the morning and afternoon.
-Frequency of dermal observation: LD50 study animals were examined for erythema and edema following patch removal on study day 1 and daily thereafter (days 2-14) according to the Dermal Grading System
- lndividual body weights were obtained for the LD50 study animals prior to dosing on day 0 and for all surviving animais on days 7 and 14 and at the time of death for any animal dying on study.
- Necropsy of survivors performed: yes. All animals which died spontaneously during the study or were euthanized (carbon dioxide inhalation) at study termination (day 14) were necropsied. Body cavities (cranial, thoracic, abdominal and pelvic) were opened and examined. No tissues were retained.
- Other examinations performed: clinical signs, body weight,organ weights, histopathology.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
446 mg/kg bw
Based on:
test mat.
95% CL:
378 - 526
Mortality:
All mortality occured day 4
Clinical signs:
at 250 mg/k: dark material around the mouth and urine stain and were noted on study days 0-1 and 0-2, respectively
For the surviving animals in the 500 and 750 mg/kg dose levels, clinical abnormalities included decreased activity, which was noted in two animals on study day 1 and in one animal on study days 2-6, and wobbly gait, which was noted in two animals.Additional clinical abnormalities noted for these animais included transient incidences of rough haircoat, dark material around the facial area, reddish colored urine, decreased defecation, urine stain, eye(s) appeared dark and decreased food consumption.

For the animals that died during the study, notable clinical abnormalities included reddish colored urine, urine stain, dark material around the facial area, eye(s) appear dark, lacrimation, eyelids partially closed, decreased activity, prostration, and fasciculation. Dermal irritation was also noted at the site of test article application.
Gross pathology:
For animals that died: discolored kidneys and/or spleen, reddened thymus, abnormal content in the small intestine, stomach, and urinary bladder, mottled lungs, congested meningeal vessels, reddened mucosa in the urinary bladder and reddened cervical lymph nades

Dose

Mortality in males

Mortality in females

Combined mortality

 

250

 

0/5

 

0/5

 

0/10

500

3/5

5/5

8/10

750

4/5

5/5

9/10

 

 

 

LD50

 

95%Confidence Interval

 

 

 

95% Confidence Interval

Sex

(mg/kg)

(mg/kg)

Slope

Slope

 

Male

 

491.8

 

361.1 - 669.8

 

1.5

 

1.3-1.8

Female

353.6

294.0 - 425.2

1.2

1.2-1.3

Combined

446.3

378.5 - 526.3

1.4

1.3-1.5

Interpretation of results:
Category 3 based on GHS criteria
Conclusions:
Under the conditions of this test, the acute dermal LD50 of t-Amyl Hydroperoxide in the male rat was determined to be 491.8 mg/kg. ln the female rat, the dermal LD50 was determined to be 353.6 mg/kg. ln the sexes combined, the acute dermal LD50 was determined to be 446.3 mg/kg
Executive summary:

An LD50 study was performed in which three groups of male and female rats received a single dermal administration of the test article at graded dosage levels. Following dosing, the LD50 study rats were observed daily and weighed weekly. A gross necropsy examination was performed on all LD50 study animals at the time of death or scheduled euthanasia (day 14).

All mortality occurred by study day 4.  The majority of clinical observations were observed in the 500 mg/kg and 750 mg/kg dose levels. In these group, for the surviving animals, clinical abnormalities included decreased activity, wobbly gait, transient incidences of rough haircoat, dark material around the facial area, reddish colored urine, decreased defecation, urine stain, eye(s) appeared dark and decreased food consumption.

For the animals that died during the study, notable clinical abnormalities included reddish colored urine, urine stain, dark material around the facial area, eye(s) appear dark, lacrimation, eyelids partially closed, decreased activity, prostration, and fasciculation. Dermal irritation was also noted at the site of test article application.

The most notable gross internal findings were observed in the animals that died and included discolored kidneys and/or spleen, reddened thymus, abnormal content in the small intestine, stomach, and urinary bladder, mottled lungs, congested meningeal vessels, reddened mucosa in the urinary bladder and reddened cervical lymph nodes.

Under the conditions of this test, the acute dermal LD50 of t-Amyl Hydroperoxide in the male rat was determined to be 491.8 mg/kg. ln the female rat, the dermal LD50 was determined to be 353.6 mg/kg. ln the sexes combined, the acute dermal LD50 was determined to be 446.3 mg/kg.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
446 mg/kg bw
Quality of whole database:
Klimisch 1 study, GLP compliant.

Additional information

Acute oral toxicity

In a key study (Doubs, 1981a) performed according to a method similar to OECD N°401 guideline and GLP compliant, groups of 5 male and 5 female Sprague Dawley rats were given a single oral dose of undiluted tert-amyl hydroperoxide at 0, 250, 500, 1000 mg/kg. The LD50 of tert-amyl hydroperoxide was 500 mg/kg in Sprague Dawley rats (517 mg/kg in males, 483 mg/kg in females). Animals died day one or two days after administration of the substance. Clinical signs observed were : hypoactivity, salivation, piloerection, abnormal breathing, urine stain, wobbly gate for most of animals; and tremor, prostration, apparent hypothermia in animals who died. At gross necropsy, in animals found dead, followings abnormalities were observed: congested meningeal vessels in the brain, mottled livers, abnormal findings in the digestive tract, discolored thymus, abnormally colored fluid contents in the urinary bladder and thoracic cavity and musculature and viscera in the whole body stained orange.

This value of oral LD50 is supported by another LD50 of 0.64 ml/kg bw in rats (Spanjers and Til, 1981).

Acute dermal toxicity

In a key study (OECD 402, GLP compliant), the acute dermal LD50 of tert-amyl hydroperoxide in rats was determined to be 446.3 mg/kg (Doubs, 1995b). Following a 24_hour exposure period, clinical observations included decreased activity, wobbly gait, transient incidences of rough haircoat, dark material around the facial area, reddish colored urine, decreased defecation, urine stain, eye(s) that appeared dark and decreased food consumption. For the animals that died during the study, notable clinical abnormalities included reddish colored urine, urine stain, dark material around the facial area, eye(s) appear dark, lacrimation, eyelids partially closed, decreased activity, prostration, and fasciculation. Dermal irritation was also noted at the site of test article application. At necropsy, following findings were reported: discolored kidneys and/or spleen, reddened thymus, abnormal content in the small intestine, stomach, and urinary bladder, mottled lungs, congested meningeal vessels, reddened mucosa in the urinary bladder and reddened cervical lymph nodes.

 

Acute inhalation toxicity

In a key acute inhalation test (TNO, 1981), 5 groups of 5 male and 5 female rats were whole-body exposed for 4 hours to vapors of tert-amyl hydroperoxide. Mortality occured mainly after the end of the exposure. At necropsy, no gross abnormality was observed in any rats. The 4-hour LC 50 was calculated to be 2425 mg/m3 [IC 95: 2100 -2750 mg/m3].

Justification for classification or non-classification

According to Regulation (EC) N0. 1272/2008 (CLP), the substance is classified category 4 for acute oral toxicity (H302), and in category 3 for acute inhalation toxicity (H331) and acute dermal toxicity (H311).