Menthane, monohydroperoxy derivative

Endpoint summary

Description of key information

No studies are available on the test substance.
On the structural analogue cumenehydroperoxide a 90-day inhalation study in rats was performed with test concentrations of 1, 6, 31 and 124 mg/m3. All rats died or were killed in extremis at 124 mg/m3 before day 12 of the study showing tissue irritation at the site of contact and included ulceration and inflammation of the cornea and eyes, nasal turbinates and lining of the stomach. Secondary effects included thymic atrophy, depletion of lymphoid tissue in the germinal centers of some lymph nodes and the spleen, decreased lipid content of the liver and decreased number of circulating white blood cells.  The exposure of 1, 6 or 31 mg/m3 cumene hydroperoxide induced no substance related statistically significant effects on haematology, urinalysis, clinical chemistry, body weights, organ weights and pathology.
NTP has performed repeated dose studies with cumene hydroperoxide in mice and rats with topical application of the test substance. In the 13-week studies at 6 mg/kg bw and above treatment related effects were found at the application site which included, hyperplasia of the sebaceous glands inflammation of the dermis and degeneration, exudate, ulcer, necrosis, hyperkeratosis and squamous hyperplasia of the epidermis. The effects increased in severity with increasing dose. No treatment related systemic effects were found.  The NOAEL is 3 mg/kg bw  (equivalent to 0.6%/ 0.018mg/cm2 in rats and 0.15%/ 0.009 mg/cm2 in mice).
In addition data on dermal irritation at different dose levels are available from 2 sensitization studies with the test substance. The NOAEL for dermal irritation of 1% is based on the lowest non irritant concentration as assessed in the CIT maximisation study (1994)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study

Qualifier:

no guideline followed

Principles of method if other than guideline:

10 rats were exposed to 4 concentrations of the test substance 5d/wk, over a period of 90 d.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: CDF (Fischer 344 derived)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA
- Weight at study initiation: males:245-308 g; females: 140-160 g
- Diet (e.g. ad libitum): Purina rat chow ad libitum
- Water (e.g. ad libitum): ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C):
- Humidity (%):
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light):

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass and stainless steel in construction, rectangular in cross section, 160 l in volume with regular pyramid top and bottom
- System of generating particulates/aerosols: the cumene hydroperoxide atmosphere was generated by aerosoling the test material with a modified DeVelbies nebulizer; the concentrated aerosol was deluted to the desired concentration with the chamber air at the inlet to the exposure chamber
- Temperature, humidity, pressure in air chamber: the chambers were operated under dynamic air flow conditions (40-50 l/min) with temperature and humidity control
- Method of particle size determination: the median particle diameter and geometric standard deviation were calculated using computer program RCUM. CLIST (Computations Laboratory, Dow Chemical Company)

TEST ATMOSPHERE
- Brief description of analytical method used: the chamber atmosphere was sampled by drawing known volumes of air (10 or 20l) through glass inpingers containing 50 ml n-octanol; cumene hydroperoxide in the n-octanol was assayed colorimetrically by the ferrous thiocynate method for organic peroxide

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Cumene hydroperoxide was assayed colorimetrically in n-propanol containing air samples

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

5 days/week, for approximately 3 months (males: 60 exposures in 88 days, females: 61 exposures in 89 days); 1 mg/ cubic m group was started with a delay of 15 days resulting in 50 exposure days

Remarks:

Doses / Concentrations:
1, 6, 31, 124 mg/cubic m aerosol
Basis:

No. of animals per sex per dose:

10

Control animals:

yes, concurrent no treatment

Details on study design:

Exposure of 124 mg/cubic m was terminated after 5 days due to toxicity and all surviving rats of this group were killed and examined on the 12th day of the study.

Positive control:

no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes, daily

DETAILED CLINICAL OBSERVATIONS: Yes, daily

BODY WEIGHT: Yes
- Time schedule for examinations: twice per week for the first 2 weeks and once per week thereafter

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at or near termination of the study
- How many animals: all animals exposed to 1, 6, 31 mg/square m and controls
- Parameters: red, white and differential blood cell counts, hemoglobin, concentration and packed cell volume

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at or near termination of study
- How many animals: all animals exposed to 1, 6, 31 mg/square m and controls
- Parameters: blood urea nitrogen, serum glutamic pyruvic transaminase, serum alkaline phosphatase

URINALYSIS: Yes
- Time schedule for collection of urine: at or near termination of study
- Metabolism cages used for collection of urine: No data
- Parameters: pH, specific gravity, sugar, protein, ketones, bilirubin, occult blood, urobilinogen

NEUROBEHAVIOURAL EXAMINATION: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table 1)
HISTOPATHOLOGY: Yes (see table 2)

Other examinations:

Frozen sections of adrenal glands and liver were stained with Oil Red G (from 3 males and 3 females from control group and from groups 31 and 124 mg/cubic m) in order to determine lipid content

Statistics:

Hematology, clinical chemistry, urine specific gravity, organ weight and body weight data were evaluated using an analysis of variance and Dunnett's test; the level of significance was for all cases p<0.05

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
124 mg/cubic m: males and females had difficulty breathing (gasping for air) after the first two days of exposure; subsequent daily exposure through 5 days resulted in progressive deterioration of health indicated by irritation to skin and mucous membranes (gangrene of extremities, exudate around eyes and nose) and breathing difficulties; 6/10 males and 3/10 females died within the first 12 days of the study
1, 6, 31 mg/cubic m: no clinical signs of irritation or toxicity
Mortality see Table 3

BODY WEIGHT AND WEIGHT GAIN
124 mg/square m: significant decrease in body weight
1, 6, 31 mg/square m: no statistically significant effects


HAEMATOLOGY
124 mg/square m: generalized decrease in PCV, RBC count, hemoglobin (it is difficult to assess whether this was a significant treatment related effect, because no control rats were killed simultaneously on day 12th of the study); the WBC count was also decreased (considered highly significant in comparison with historical controls) and coincided with the severe debilated state of surviving rats on day 12 of the study
1, 6, 31 mg/square m: no statistically significant effects


CLINICAL CHEMISTRY
1, 6, 31 mg/square m: no toxicologically significant effects



URINALYSIS
1, 6, 31 mg/square m: no treatment related effects


ORGAN WEIGHTS
1, 6, 31 mg/square m: no treatment related effects

GROSS PATHOLOGY
124 mg/square m: urine and fecal staining in the perineal region; debris around the eyes, external nares or extremities; porphyrin pigment around the nose, eyes, or both; cyanotic or hyperemic appearance of the extremities; dry gangrene of the feet, limbs, or tail; depletion of adipose tissue; variations in color (usually darker, hyperemic or congested) of several internal organs such as liver, kidney, lungs and nasal mucosa; decreased contents in the gastrointestinal tract; edema, ulcerations or erosions of the stomach; thymic athrophy; corneal cloudiness of the eyes
1, 6, 31 mg/cubic m: no treatment related effects


HISTOPATHOLOGY: NON-NEOPLASTIC
124 mg/cubic m: upper respiratory tract changes, including inflammation, erosions or ulcerations of the trachea and nasal turbinates, focal epithelial hyperplasia of tracheal mucosa and squamous metaplasia of the mucosa of the nasal turbinates (males); an apparent decrease in the amount ot secretory material in the pancreatic acini (more prominent in males than in females); changes in nonglandular stomach, including submucosal edema, hyperkeratosis, acanthosis, inflammation, erosions and ulcerations (prominent in males, but not in females); hyperkeratosis of the esophageal mucosa in 2/10 males; depletion of cortical and/or paracortical lymphocytes in the mesenteric lymph nodes; depletion of the germinal centers in the spleen; thymic athrophy with or without necrotic debris and hemorrhage; extremities with necrosis, inflammation, ulcerations, fibrin thrombi in blood vessels, and edema (more common in females than in males); eyes with increased corneal vascularization, corneal ulceration and erosions, and corneal inflammation (more common in males than in females)
1, 6, 31 mg/cubic m: no treatment related effects


OTHER FINDINGS
124 mg/cubic m: decreased lipid content in livers
31 mg/cubic m: no effect on lipid content in livers

Dose descriptor:

NOAEC

Effect level:

31 mg/m³ air

Sex:

male/female

Basis for effect level:

other: absence of recognized treatment related effects on hematology, urinalysis, clinical chemistry, body weights, organ weights and pathology in rats exposed to 1, 6, or 31 mg/cubic m cumene hydroperoxide

Critical effects observed:

not specified

Table 3: Mortality

Exposure group (mg/cubic m)	No. alive at start of the study                     m f	No. dying or killed moribund                    m f	No. terminated after 12 days m                 f	No. presented alive or killed at the end of the study m           f
0	10           10	0              0	0                   0	10              10
1^*	10            10	0               0	0                  0	10               10
6	10             10	0                0	0                  0	10               10
31	10             10	0                0	0                  0	10               10
124**	10             10	6                3	4                  7	0                  0

*: The 1 mg/cubic m exposure group was started 15 days later than the other groups and, therefore, received 10 fewer exposures than the 6, 31 mg/cubic m and control rats

**: The 124 mg/cubic m exposure group received 5 consecutive daily exposures and because of the high toxicity it was terminated on day 12^th of the study

Conclusions:

The highest no adverse effect concentration in this study was judged to be 31 mg/cubic m which is equivalent to 5 ppm vapor exposure.

Executive summary:

Male and female rats were exposed daily (6 h/day, 5 days/week) to 1, 6, 31 and 124 mg/cubic m cumene hydroperoxide delivered as aerosol over a period of 3 months. Inhalation of 124 mg/cubic m cumene hydroperoxide for 5 consecutive days resulted in decresed body weight and death of 6/10 males and 3/10 females, therefore exposure of this group was terminated after 5 day and all surviving rats were killed on day 12 of the study. Pathologic alterations in rats that died as well as those that survived at 124 mg/cubic m exposure level appeared to be the result of tissue irritation at the site of contact and included ulceration and inflammation of the cornea and eyes, nasal turbinates and lining of the stomach. Primary toxicologic effects following inhalation of124 mg/cubic m cumene hydroperoxide were consistent with those caused by a primary tissue irritant, other changes were judged by the authors as secondary effects and as caused by stress. These included thymic atrophy, depletion of lymphoid tissue in the germinal centers of some lymph nodes and the spleen, decreased lipid content of the liver and decreased circulating white blood cells. The exposure of 1, 6 or 31 mg/cubic m cumene hydroperoxide induced no substance related statistically significant effects on hematology, urinalysis, clinical chemistry, body weights, organ weights and pathology.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

31 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

The derived NOAEC is for local effects, but is considered sufficient to protect against systemic effects. The NOAEC is based on a study with a structural analogue substance (see attachment chapter 13)

Repeated dose toxicity: inhalation - local effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study

Qualifier:

no guideline followed

Principles of method if other than guideline:

10 rats were exposed to 4 concentrations of the test substance 5d/wk, over a period of 90 d.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: CDF (Fischer 344 derived)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA
- Weight at study initiation: males:245-308 g; females: 140-160 g
- Diet (e.g. ad libitum): Purina rat chow ad libitum
- Water (e.g. ad libitum): ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C):
- Humidity (%):
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light):

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

whole body

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass and stainless steel in construction, rectangular in cross section, 160 l in volume with regular pyramid top and bottom
- System of generating particulates/aerosols: the cumene hydroperoxide atmosphere was generated by aerosoling the test material with a modified DeVelbies nebulizer; the concentrated aerosol was deluted to the desired concentration with the chamber air at the inlet to the exposure chamber
- Temperature, humidity, pressure in air chamber: the chambers were operated under dynamic air flow conditions (40-50 l/min) with temperature and humidity control
- Method of particle size determination: the median particle diameter and geometric standard deviation were calculated using computer program RCUM. CLIST (Computations Laboratory, Dow Chemical Company)

TEST ATMOSPHERE
- Brief description of analytical method used: the chamber atmosphere was sampled by drawing known volumes of air (10 or 20l) through glass inpingers containing 50 ml n-octanol; cumene hydroperoxide in the n-octanol was assayed colorimetrically by the ferrous thiocynate method for organic peroxide

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Cumene hydroperoxide was assayed colorimetrically in n-propanol containing air samples

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

5 days/week, for approximately 3 months (males: 60 exposures in 88 days, females: 61 exposures in 89 days); 1 mg/ cubic m group was started with a delay of 15 days resulting in 50 exposure days

Remarks:

Doses / Concentrations:
1, 6, 31, 124 mg/cubic m aerosol
Basis:

No. of animals per sex per dose:

10

Control animals:

yes, concurrent no treatment

Details on study design:

Exposure of 124 mg/cubic m was terminated after 5 days due to toxicity and all surviving rats of this group were killed and examined on the 12th day of the study.

Positive control:

no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes, daily

DETAILED CLINICAL OBSERVATIONS: Yes, daily

BODY WEIGHT: Yes
- Time schedule for examinations: twice per week for the first 2 weeks and once per week thereafter

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at or near termination of the study
- How many animals: all animals exposed to 1, 6, 31 mg/square m and controls
- Parameters: red, white and differential blood cell counts, hemoglobin, concentration and packed cell volume

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at or near termination of study
- How many animals: all animals exposed to 1, 6, 31 mg/square m and controls
- Parameters: blood urea nitrogen, serum glutamic pyruvic transaminase, serum alkaline phosphatase

URINALYSIS: Yes
- Time schedule for collection of urine: at or near termination of study
- Metabolism cages used for collection of urine: No data
- Parameters: pH, specific gravity, sugar, protein, ketones, bilirubin, occult blood, urobilinogen

NEUROBEHAVIOURAL EXAMINATION: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table 1)
HISTOPATHOLOGY: Yes (see table 2)

Other examinations:

Frozen sections of adrenal glands and liver were stained with Oil Red G (from 3 males and 3 females from control group and from groups 31 and 124 mg/cubic m) in order to determine lipid content

Statistics:

Hematology, clinical chemistry, urine specific gravity, organ weight and body weight data were evaluated using an analysis of variance and Dunnett's test; the level of significance was for all cases p<0.05

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
124 mg/cubic m: males and females had difficulty breathing (gasping for air) after the first two days of exposure; subsequent daily exposure through 5 days resulted in progressive deterioration of health indicated by irritation to skin and mucous membranes (gangrene of extremities, exudate around eyes and nose) and breathing difficulties; 6/10 males and 3/10 females died within the first 12 days of the study
1, 6, 31 mg/cubic m: no clinical signs of irritation or toxicity
Mortality see Table 3

BODY WEIGHT AND WEIGHT GAIN
124 mg/square m: significant decrease in body weight
1, 6, 31 mg/square m: no statistically significant effects


HAEMATOLOGY
124 mg/square m: generalized decrease in PCV, RBC count, hemoglobin (it is difficult to assess whether this was a significant treatment related effect, because no control rats were killed simultaneously on day 12th of the study); the WBC count was also decreased (considered highly significant in comparison with historical controls) and coincided with the severe debilated state of surviving rats on day 12 of the study
1, 6, 31 mg/square m: no statistically significant effects


CLINICAL CHEMISTRY
1, 6, 31 mg/square m: no toxicologically significant effects



URINALYSIS
1, 6, 31 mg/square m: no treatment related effects


ORGAN WEIGHTS
1, 6, 31 mg/square m: no treatment related effects

GROSS PATHOLOGY
124 mg/square m: urine and fecal staining in the perineal region; debris around the eyes, external nares or extremities; porphyrin pigment around the nose, eyes, or both; cyanotic or hyperemic appearance of the extremities; dry gangrene of the feet, limbs, or tail; depletion of adipose tissue; variations in color (usually darker, hyperemic or congested) of several internal organs such as liver, kidney, lungs and nasal mucosa; decreased contents in the gastrointestinal tract; edema, ulcerations or erosions of the stomach; thymic athrophy; corneal cloudiness of the eyes
1, 6, 31 mg/cubic m: no treatment related effects


HISTOPATHOLOGY: NON-NEOPLASTIC
124 mg/cubic m: upper respiratory tract changes, including inflammation, erosions or ulcerations of the trachea and nasal turbinates, focal epithelial hyperplasia of tracheal mucosa and squamous metaplasia of the mucosa of the nasal turbinates (males); an apparent decrease in the amount ot secretory material in the pancreatic acini (more prominent in males than in females); changes in nonglandular stomach, including submucosal edema, hyperkeratosis, acanthosis, inflammation, erosions and ulcerations (prominent in males, but not in females); hyperkeratosis of the esophageal mucosa in 2/10 males; depletion of cortical and/or paracortical lymphocytes in the mesenteric lymph nodes; depletion of the germinal centers in the spleen; thymic athrophy with or without necrotic debris and hemorrhage; extremities with necrosis, inflammation, ulcerations, fibrin thrombi in blood vessels, and edema (more common in females than in males); eyes with increased corneal vascularization, corneal ulceration and erosions, and corneal inflammation (more common in males than in females)
1, 6, 31 mg/cubic m: no treatment related effects


OTHER FINDINGS
124 mg/cubic m: decreased lipid content in livers
31 mg/cubic m: no effect on lipid content in livers

Dose descriptor:

NOAEC

Effect level:

31 mg/m³ air

Sex:

male/female

Basis for effect level:

other: absence of recognized treatment related effects on hematology, urinalysis, clinical chemistry, body weights, organ weights and pathology in rats exposed to 1, 6, or 31 mg/cubic m cumene hydroperoxide

Critical effects observed:

not specified

Table 3: Mortality

Exposure group (mg/cubic m)	No. alive at start of the study                     m f	No. dying or killed moribund                    m f	No. terminated after 12 days m                 f	No. presented alive or killed at the end of the study m           f
0	10           10	0              0	0                   0	10              10
1^*	10            10	0               0	0                  0	10               10
6	10             10	0                0	0                  0	10               10
31	10             10	0                0	0                  0	10               10
124**	10             10	6                3	4                  7	0                  0

*: The 1 mg/cubic m exposure group was started 15 days later than the other groups and, therefore, received 10 fewer exposures than the 6, 31 mg/cubic m and control rats

**: The 124 mg/cubic m exposure group received 5 consecutive daily exposures and because of the high toxicity it was terminated on day 12^th of the study

Conclusions:

The highest no adverse effect concentration in this study was judged to be 31 mg/cubic m which is equivalent to 5 ppm vapor exposure.

Executive summary:

Male and female rats were exposed daily (6 h/day, 5 days/week) to 1, 6, 31 and 124 mg/cubic m cumene hydroperoxide delivered as aerosol over a period of 3 months. Inhalation of 124 mg/cubic m cumene hydroperoxide for 5 consecutive days resulted in decresed body weight and death of 6/10 males and 3/10 females, therefore exposure of this group was terminated after 5 day and all surviving rats were killed on day 12 of the study. Pathologic alterations in rats that died as well as those that survived at 124 mg/cubic m exposure level appeared to be the result of tissue irritation at the site of contact and included ulceration and inflammation of the cornea and eyes, nasal turbinates and lining of the stomach. Primary toxicologic effects following inhalation of124 mg/cubic m cumene hydroperoxide were consistent with those caused by a primary tissue irritant, other changes were judged by the authors as secondary effects and as caused by stress. These included thymic atrophy, depletion of lymphoid tissue in the germinal centers of some lymph nodes and the spleen, decreased lipid content of the liver and decreased circulating white blood cells. The exposure of 1, 6 or 31 mg/cubic m cumene hydroperoxide induced no substance related statistically significant effects on hematology, urinalysis, clinical chemistry, body weights, organ weights and pathology.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

31 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

The test is performed with a structural analogue. A rationale for the read-across is provided in a separate document attached to chapter 13.

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: standard NTP test protocol, not all standard parameters assessed The nature of the effects (local at the application site) allows conclusions on the NOAEL

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)

Deviations:

yes

Remarks:

no biochemistry, urinalysis, behavioural effects

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Type of coverage:

not specified

Vehicle:

ethanol

Details on exposure:

- Area of exposure: 10% of total surface (based on an average BW of 200 g and a conversion factor of 5.9 (Kg to m2)*: 0.1 times 0.034 m2 = 34 cm2)

TEST MATERIAL: no detailed data

VEHICLE: ethanol (indicated in the result section of the micronucleus test) 0.5 mL/ kg


* Freireich, EJ, et al. Quantitative comparison of toxicity of anticancer agents in mouse, rat, dog, monkey and man. Cancer Chemother Rep.1966;50(4):219-244

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

93 days

Frequency of treatment:

daily on week days

Remarks:

Doses / Concentrations:
0, 0.75, 1.5, 3.0, 6.0 and 12 mg/kg bw
Basis:
nominal per unit body weight

No. of animals per sex per dose:

10/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

After a 10- to 14-day quarantine period, animals are assigned at random to treatment groups. The study includes five treatment groups each administered a different concentration of the test article plus a control group. Each group contains 10 animals per sex per species. The animals receive the subject chemical by a designated route of exposure. Controls receive vehicle alone.Animals are exposed five times per week, weekdays only until the day prior to necropsy. All rats are housed individually.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once weekly

BODY WEIGHT: Yes
- Time schedule for examinations: on day 1 and weekly thereafter

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
on day 93: Erythrocyte count, Mean corpuscular volume, Hemoglobin, Packed cell volume, Mean corpuscular haemoglobin, Mean corpuscular hemoglobin concentration, Erythrocyte morphologic assessment, Leukocyte count, Leukocyte differential, Reticulocyte count, Platelet count and morphologic assessment

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

ORGAN WEIGHTS:
Liver, thymus, right kidney, right testis, heart, and lung

GROSS PATHOLOGY: Yes
Adrenal glands, Oral cavity, larynx and pharynx, Brain Ovaries, Clitoral glands, Pancreas, Esophagus, Parathyroid glands, Eyes, Pituitary gland, Femur, Preputial glands, Prostate, Gross lesions, Salivary glands, Harderian glands, Seminal vesicles, Heart and aorta, Skin, site of application (dermal studies), Large intestine (cecum, colon, rectum), Spinal cord, Small Intestine (duodenum, jejunum, ileum), Spleen, Kidneys, Stomach (forestomach and glandular), Liver, Testes, epididymides and vaginal tunics of testes, Lungs and mainstem bronchi, Thymus, Lymph nodes - mandibular and mesenteric, Thyroid gland , Tissue masses, Tongue, Mammary gland with adjacent skin, Trachea, Muscle (thigh), Urinary bladder, sciatic Nerve, Uterus, Nasal cavity and nasal turbinates, Vagina, Zymbal gland

HISTOPATHOLOGY: Yes
A complete histopathologic evaluation inclusive of treatment-related gross lesions on all early death animals regardless of dose group, all control animals and all animals in the highest treatment group. In addition all gross lesions irrespective of the dose group.

Statistics:

not yet performed

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

1 moribund

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

at 6 mg/kg bw and above

Mortality:

mortality observed, treatment-related

Description (incidence):

1 moribund

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

slightly decreased in high dose males

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

primary effect on skin at application site

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

skin lesions

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS AND MORTALITY
1 male died at 6 mg/kg bw on day 33 (reduced bodyweight(gain) and hyperplasia at application site), no mortality in females

BODY WEIGHT AND WEIGHT GAIN
males: slightly decreased at 12 mg/kg bw
females: no treatment related effects

HAEMATOLOGY
males: no treatment related effects
females: increased platelets numbers at 6 and 12 mg/kg bw

ORGAN WEIGHTS
males: slightly decreased absolute liver, lung, testis and thymus weights at 12 mg/kg bw (not relative to bodyweight)
females: no treatment related effects

HISTOPATHOLOGY: NON-NEOPLASTIC
epididymus:
inflammation and/or cellular infiltration of lymphocytes in 3/10 males at 12 mg/kg bw

at the application site
sebaceous glands:
hyperplasia in all males and females at 12 mg/kg bw and in 1 male at 6 mg/kg bw

dermis:
inflammation in 10 males and 9 females at 12 mg/kg bw; in 2/10 males at 6 mg/kg bw; in 1/10 males at 1.5 mg/kg bw

epidermis:
degradation, exudate, ulcer, necrosis, hyperkeratosis and and squamous hyperplasia in males and females at 12 mg/kg bw
minimal hyperkeratosis and squamous hyperplasia in males at 6 mg/kg bw

Effects increased in incidence and severity with increasing dose levels

Dose descriptor:

NOAEL

Effect level:

3 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: local effects on the application site due to the corrosive nature of the test substance

Critical effects observed:

not specified

No treatment related systemic effects were identified.

Conclusions:

The NOAEL under the conditions of the study is 3 mg/kg bw

Executive summary:

Ten rats per sex/dose and 10/sex for controls were treated dermally with the test substance for 13 weeks

at 0, 0.75, 1.5, 3.0, 6.0 and 12 mg/kg bw/day. One male died in the 6 mg/kg bw group. Body weights were slightly decreased in males at 12 mg/kg bw. Treatment related effects were found at the application site which included, hyperplasia of the sebaceous glands inflammation of the dermis and degeneration, exudate, ulcer, necrosis, hyperkeratosis and squamous hyperplasia of the epidermis. The effects increased in severity with increasing dose. No treatment related systemic effects were found. The NOAEL is 3.0 mg/kg bw/day (0.6% or 0.018 mg/cm2 per day).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

3 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

The test is performed with a structural analogue. A rationale for the read-across is provided in a separate document attached to chapter 13.

Repeated dose toxicity: dermal - local effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: standard NTP test protocol, not all standard parameters assessed The nature of the effects (local at the application site) allows conclusions on the NOAEL

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)

Deviations:

yes

Remarks:

no biochemistry, urinalysis, behavioural effects

GLP compliance:

not specified

Limit test:

no

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Type of coverage:

not specified

Vehicle:

ethanol

Details on exposure:

TEST SITE
- Area of exposure: 10% of total surface (based on an average BW of 25 g and a conversion factor of 3 (Kg to m2)*: 0.1 times 0.0083 m2 = 8.3 cm2)

TEST MATERIAL: no detailed data

VEHICLE: ethanol (indicated in the result section of the micronucleus test) 2.0 mL/ kg


* Freireich, EJ, et al. Quantitative comparison of toxicity of anticancer agents in mouse, rat, dog, monkey and man. Cancer Chemother Rep.1966;50(4):219-244

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

93 days

Frequency of treatment:

daily on week days

Remarks:

Doses / Concentrations:
0, 0.75, 1.5, 3.0, 6.0 and 12 mg/kg bw
Basis:
nominal per unit body weight

No. of animals per sex per dose:

10/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

After a 10- to 14-day quarantine period, animals are assigned at random to treatment groups. The study includes five treatment groups each administered a different concentration of the test article plus a control group. Each group contains 10 animals per sex per species. The animals receive the subject chemical by a designated route of exposure. Controls receive vehicle alone.Animals are exposed five times per week, weekdays only until the day prior to necropsy. All mice are housed individually.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once weekly

BODY WEIGHT: Yes
- Time schedule for examinations: on day 1 and weekly thereafter

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
on day 93: Erythrocyte count, Mean corpuscular volume, Hemoglobin, Packed cell volume, Mean corpuscular haemoglobin, Mean corpuscular hemoglobin concentration, Erythrocyte morphologic assessment, Leukocyte count, Leukocyte differential, Reticulocyte count, Platelet count and morphologic assessment

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

ORGAN WEIGHTS:
Liver, thymus, right kidney, right testis, heart, and lung

GROSS PATHOLOGY: Yes
Adrenal glands, Oral cavity, larynx and pharynx, Brain Ovaries, Clitoral glands, Pancreas, Esophagus, Parathyroid glands, Eyes, Pituitary gland, Femur, Preputial glands, Prostate, Gross lesions, Salivary glands, Harderian glands, Seminal vesicles, Heart and aorta, Skin, site of application (dermal studies), Large intestine (cecum, colon, rectum), Spinal cord, Small Intestine (duodenum, jejunum, ileum), Spleen, Kidneys, Stomach (forestomach and glandular), Liver, Testes, epididymides and vaginal tunics of testes, Lungs and mainstem bronchi, Thymus, Lymph nodes - mandibular and mesenteric, Thyroid gland , Tissue masses, Tongue, Mammary gland with adjacent skin, Trachea, Muscle (thigh), Urinary bladder, sciatic Nerve, Uterus, Nasal cavity and nasal turbinates, Vagina, Zymbal gland

HISTOPATHOLOGY: Yes
A complete histopathologic evaluation inclusive of treatment-related gross lesions on all early death animals regardless of dose group, all control animals and all animals in the highest treatment group. In addition all gross lesions irrespective of the dose group.

Statistics:

not yet performed

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

1 moribund at 1.5 mg/kg bw

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

at 6 mg/kg bw and above

Mortality:

mortality observed, treatment-related

Description (incidence):

1 moribund at 1.5 mg/kg bw

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

slightly decreased at 6 and 12mg/kg bwin females

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

primary effect on skin at application site

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

slight skin lesions

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS AND MORTALITY
1 female died at 1.5 mg/kg bw on day 8 (accidental kill), no mortality in males

BODY WEIGHT AND WEIGHT GAIN
males: no treatment related effects
females: slightly decreased at 6 and 12 mg/kg bw

HAEMATOLOGY
males: no treatment related effects
females: no treatment related effects

ORGAN WEIGHTS
males: no treatment related effects
females: no treatment related effects

HISTOPATHOLOGY: NON-NEOPLASTIC
at 12 mg/kg bw in 3 females cysts in the thymus

at the application site
dermis:
inflammation in 5/10 males and 10/10 females at 12 mg/kg bw; in 3/10 males and 7/10 females at 6 mg/kg bw
epidermis:
hyperplasia in 9/10 males and 10/10 females at 12 mg/kg bw; in 5/10 males and 9/10 females at 6 mg/kg bw

Effects increased in incidence and severity with increasing dose levels

Dose descriptor:

NOAEL

Effect level:

3 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: local effects on the application site due to the corrosive nature of the test substance

Critical effects observed:

not specified

No treatment related systemic effects were identified.

Conclusions:

The NOAEL under the conditions of the study is 3 mg/kg bw

Executive summary:

Ten mice per sex/dose and 10/sex for controls were treated dermally with the test substance for 13 weeks

at 0, 0.75, 1.5, 3.0, 6.0 and 12 mg/kg bw/day. One female accidently died in the 1.5 mg/kg bw group. Body weights were slightly decreased in females at 6 and 12 mg/kg bw. Treatment related effects were found at the application site which included inflammation of the dermis and squamous hyperplasia of the epidermis. The effects increased in severity with increasing dose. No treatment related systemic effects were found. The NOAEL is 3.0 mg/kg bw/day (0.15% or 0.009 mg/cm2 per day, see conversion table).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

0.009 mg/cm²

Study duration:

subchronic

Species:

rat

Quality of whole database:

The NOAEL is expressed in mg/kg bw (application volume not known). The test is performed with a structural analogue. A rationale for the read-across is provided in a separate dicument attached to chapter 13.

Additional information

The structural analogue substance cumene hydroperoxide is considered sufficiently similar to the test substance based on reactivity, toxicokinetic and toxicodynamic properties (see document on read-across attached to chapter 13)

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
no exposure of the general public/consumers foreseen

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
Both the NTP studies in rats and mice are suitable to be used as starting point. Both lead to the same NOAEL.

Justification for selection of repeated dose toxicity dermal - local effects endpoint:
Both the NTP studies in rats and mice are suitable to be used as starting point. Both lead to the same NOAEL

Repeated dose toxicity: inhalation - systemic effects (target organ) respiratory: other

Justification for classification or non-classification

Based on the outcome of the repeated dose inhalation study, the test substance has to be classified with STOT RE2 (H373) based on an effect concentration between 0.031 and 0.124 mg/L with the respiratory tract as target organ.