Registration Dossier

Administrative data

Description of key information

The median lethal dose of tert-Butyl peroxybenzoate (CAS# 641-45-9) after single oral administration to female rats, observed over a period of 14 days, is: LD50 (female rat): greater than 2000 mg/kg body weight.//Acute Oral Toxicity (LD50) Study in Rats Male Rats: 7084 (5289-9489) mg/kg Female Rats: 3382 (2697-4241) mg/kg Combined Male and Female Rats: 4838 (3662-6391) mg/kg

The median lethal dose of tert-butyl peroxybenzoate after single dermal administration to rats of both sexes, observed over a period of 14 days, is:

LD50 (rat): greater than 2000 mg/kg body weight

The LC50 of Tert-butyl Peroxybenzoate obtained in this study was estimated to be between 4.9 and 1.01 mg/L air (chemically determined mean aerosol concentration).

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD guideline 423 with full conformance with GLP.
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
other: RccHan:WIST(SPF)
Sex:
female
Details on test animals and environmental conditions:
Test System
Animals: Rat, RccHan: WIST(SPF)
Rationale: Recognized by international guidelines as a
recommended test system.
Breeder: Harlan Laboratories B.V.
Kreuzelweg 53
5961 NM Horst / The Netherlands
Number of Animals per Group: 3 females
Total Number of Animals: 6 females
Age (when treated): 11 weeks
Body Weight Range (when treated): 201.8 g - 217.2 g
Identification: Unique cage number and corresponding color-coded
spots on the tail. The animals were marked at
acclimatization start.
Randomization: Selected by hand at time of delivery.
Acclimatization: Under laboratory conditions, after health
examination. Only animals without any visible signs
of illness were used for the study.

Room Numbers: 136, Harlan Laboratories Ltd., Itingen
Conditions: Standard Laboratory Conditions. Air-conditioned
with 10 - 15 air changes per hour, and continuously
monitored environment with ranges for room
temperature 22 ± 3 °C and for relative humidity
between 30 - 70% (values above 70% during
cleaning process possible), automatically controlled
light cycle of 12 hours light and 12 hours dark,
music during the daytime light period.
Accommodation: In groups of three in Makrolon type-4 cages with
wire mesh tops and standard softwood bedding
(J. Rettenmaier & Söhne GmbH & Co. KG, 73494
Rosenberg / Germany, imported by Provimi Kliba
AG, 4303 Kaiseraugst / Switzerland) including paper
enrichment (Enviro-dri from Lillico, Biotechnology,
Surrey / UK).
Diet: Pelleted standard Harlan Teklad 2914C rodent
maintenance diet (Provimi Kliba AG,
4303 Kaiseraugst / Switzerland), batch no. 82/09,
was available ad libitum. Results of respective
analyses for contaminants are included.
Water: Community tap-water from Itingen was available ad
libitum in water bottles. Results of bacteriological
assay, chemical and contaminant analyses of
respective samples are included.
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Oral administration was considered to be an appropriate application method as it is a possible
route of human exposure during manufacture, handling and use of the test item.
Homogeneity of the test item in the vehicle was maintained during administration using a
magnetic stirrer.
The animals received a single dose of the test item by oral gavage administration at 2000 mg/kg
body weight after being fasted for approximately 17 hours (access to water was permitted). Food
was provided again approximately 3 hours after dosing.
The dosing volume was 10 mL/kg body weight.
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
6F
Control animals:
no
Details on study design:
Oral administration was considered to be an appropriate application method as it is a possible
route of human exposure during manufacture, handling and use of the test item.
Homogeneity of the test item in the vehicle was maintained during administration using a
magnetic stirrer.
The animals received a single dose of the test item by oral gavage administration at 2000 mg/kg
body weight after being fasted for approximately 17 hours (access to water was permitted). Food
was provided again approximately 3 hours after dosing.
The dosing volume was 10 mL/kg body weight.
Statistics:
none
Preliminary study:

5 CONCLUSION
Key result
Sex:
female
Dose descriptor:
LD0
Effect level:
2 000 mg/kg bw
Based on:
test mat.
Mortality:
none
Clinical signs:
One female (no. 2) showed slight somnolence and slightly ruffled fur 5 hours post treatment.
Otherwise, no clinical signs were observed during the course of the study.
Body weight:
The body weight of the animals was within the range commonly recorded for this strain and age.
Gross pathology:
No macroscopic findings were recorded at necropsy.
Other findings:
none
Interpretation of results:
not classified
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
The median lethal dose of tert-Butyl peroxybenzoate (CAS# 641-45-9) after single oral administration to female rats, observed over a period of 14 days, is: LD50 (female rat): greater than 2000 mg/kg body weight
Executive summary:

Two groups, each consisting of three female RccHan:WIST (SPF) rats, were treated with tert-Butyl peroxybenzoate (CAS# 641-45-9) by single oral gavage administration at a dosage of 2000 mg/kg body weight. The test item was formulated in corn oil at a concentration of 0.2 g/mL and administered at a dosing volume of 10 mL/kg. The animals were examined daily during the acclimatization period and mortality, viability and clinical signs were recorded. All animals were examined for clinical signs within the first 30 minutes and approximately 1, 2, 3 and 5 hours after treatment on day 1 and once daily during test days 2 - 15. Mortality/viability was recorded within the first 30 minutes and approximately 1, 2, 3 and 5 hours after administration on test day 1 (with the clinical signs) and twice daily during days 2 - 15. Body weights were recorded on day 1 (prior to administration) and on days 8 and 15. All animals were necropsied and examined macroscopically. All animals survived until the end of the study period. Slight clinical signs including somnolence and ruffled fur were noted in one female at the 5-hour observation. Otherwise, no clinical signs were observed during the course of the study. The body weight of the animals was within the range commonly recorded for this strain and age. No macroscopic findings were recorded at necropsy. The median lethal dose of tert-Butyl peroxybenzoate (CAS# 641-45-9) after single oral administration to female rats, observed over a period of 14 days, is: LD50 (female rat): greater than 2000 mg/kg body weight.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
discriminating dose
2 000 mg/kg bw
Quality of whole database:
A study, reliable without restrictions, is available.

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Conducted according to OECD 436, in full comformance with GLP. Gravimetric and analytical verification of test article concentrations in nose-only aerosol exposures for 4-hours. analytical verification of MMAD (2-3 u).
Qualifier:
according to
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
other: HanRcc:WIST(SPF)
Sex:
male/female
Details on test animals and environmental conditions:
Husbandry
Room Numbers: Animal room number 306 and 307 (Group 1) and
307 and 315 (Group 2), Harlan Laboratories Ltd,
Füllinsdorf.
Conditions: Optimal laboratory conditions behind a barrier
system. Air-conditioned with 10 - 15 air changes per
hour, continuously monitored environment with
temperature range of 22 ± 3 °C, a relative humidity
range of 30 - 70% and a 12 hour fluorescent light /
12 hour dark cycle. A radio program was played
during most of the light period.
Accommodation: Animals were housed in groups of 3 of the same sex
in Makrolon® type-IV cages with wire mesh tops
and standard softwood bedding ("Lignocel" J.
Rettenmaier & Söhne GmbH & Co KG, 73494
Rosenberg / Germany, imported by Provimi Kliba
AG, 4303 Kaiseraugst / Switzerland).
Diet: Animals had ad libitum access to a pelleted standard
Teklad rat maintenance diet (Provimi Kliba AG,
4303 Kaiseraugst, Switzerland) batch no. 82/09
except during the period when the animals were
restrained in exposure tubes. Results of the analyses
for contaminants and their limits of acceptability are
archived at Harlan Laboratories Ltd.
Water: Community tap water from Füllinsdorf ad libitum in
water bottles, except during the period when they
were restrained in exposure tubes. Results of
representative analyses for contaminants are
archived at Harlan Laboratories Ltd.
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
clean air
Details on inhalation exposure:
Treatment
Room: Inhalation laboratory no. 310, Harlan Laboratories
Ltd., Füllinsdorf
Method: Inhalation by nose-only, flow-past exposure.
Rationale for Method: Inhalation is a possible route of human exposure.
Frequency of Administration: Single, 4-hour exposure period.
Allocation and Target AerosolConcentration:
Males Females Target AerosolConcentration(mg/L air)
Group 1 1-3 4-6 slightly above 5
Group 2 7-9 10-12 slightly above 1

Rationale for Aerosol Concentration:
Group 1:
The target concentration of slightly above 5 mg/L air
for 4 hours is the recommended concentration for a
limit test (OECD 436, “Acute Inhalation Toxicity”).
Group 2:
The target concentration of 1 mg/L air for 4 hours is
the recommended concentration according to annex
3 d (OECD 436, “Acute Inhalation Toxicity”).
Duration of Observation Period: 14 days

Inhalation Exposure System
Inhalation exposure was performed using a system similar to that originally described by Sachsse
et al. (see References (1) and (2)). The animals were confined separately in restraint tubes which
were positioned radially around the flow-past, nose-only exposure chamber as described by
Cannon et al. (see References (3)). The design of this chamber is based upon the fluid dynamic
modeling of the test aerosol flow.
The exposure system ensured a uniform distribution and provided a constant flow of test material
to each exposure tube. The flow of air at each tube was 1.0 L/min, which is sufficient to
minimize re-breathing of the test atmosphere as it is more than twice the respiratory minute
volume of a rat.
Before commencement of the exposure of the groups, technical trials were conducted (without
animals) using the inhalation system foreseen for the study. The technical trials were conducted
using established procedures based on GLP, but were not inspected by the Harlan Laboratories
Ltd. Quality Assurance. Technical trial data are retained in the raw data.

Test Aerosol Generation
The test atmosphere was generated using a Hudson nebulizer connected to a syringe pump. The
polyethylene injector inside the nebulizer was replaced by a stainless steel injector.

Exposure System Monitoring
The aerosol concentration, the particle size distribution, relative humidity, temperature and
oxygen concentration were measured on test aerosol samples taken at a representative exposure
port.

Nominal Determination of Aerosol Concentration
The test item usage was measured by weighing the syringe and nebulizer reservoir containing the
test item before and after exposure to determine the quantity of test item used. The weight used
was then divided by the total air-flow volume to give the nominal concentration.

Gravimetric Determination of Aerosol Concentrations
Gravimetric determinations of aerosol concentration were performed four times during each
exposure. The samples were collected on a Millipore®durapore filter, Type HVLP loaded in a 47
mm in-line stainless steel filter sampling device. The filters were weighed before and
immediately after sampling using a calibrated balance. The test aerosol concentration was
calculated from the amount of test item present on the filter and the sample volume.

Chemical Determination of Aerosol Concentrations
Chemical determinations of aerosol concentration were performed four times during each
exposure.The samples were collected on a Millipore®durapore filter, Type HVLP loaded in a 47 mm inline
stainless steel filter sampling device. The filters were transferred into appropriate labeled
vials, forwarded in a cool box to the responsible scientist for analytical chemistry and stored at 2-
8 °C until analysis. The samples were analyzed using a HPLC method supplied by the Sponsor.

Particle Size Distribution and Mass Median Aerodynamic Diameter
The particle size distribution was measured gravimetrically three times during each exposure
using a 7 stage cascade Mercer Impactor (Model 02-130, In-Tox. Products Inc., Albuquerque,
New Mexico, U.S.A.). Mass Median Aerodynamic Diameters and Geometric Standard
Deviations were calculated on the basis of the results from the impactor, using Microsoft Excel
Software. The target range for the Mass Median Aerodynamic Diameter was 1 to 4 μm.

Oxygen Concentration
The oxygen concentration of the test atmosphere was measured continuously during each
exposure using a calibrated device. The results were recorded manually and are reported at 30
minute intervals from the start of exposure. The oxygen concentration was maintained above
19% during each exposure period.

Relative Humidity / Temperature
The temperature and relative humidity of the test atmosphere was measured continuously during
each exposure using a calibrated device. The results were recorded manually and are reported at
30 minute intervals from the start of exposure.

Airflow Rate
The actual airflow rate through the exposure chamber was recorded at approximately 30 minute
intervals from the start of the inhalation exposure.
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
1.01 mg/L and 4.9 mg/L
No. of animals per sex per dose:
3M & 3F
Control animals:
no
Details on study design:
Treatment
Room: Inhalation laboratory no. 310, Harlan Laboratories
Ltd., Füllinsdorf
Method: Inhalation by nose-only, flow-past exposure.
Rationale for Method: Inhalation is a possible route of human exposure.
Frequency of Administration: Single, 4-hour exposure period.
Allocation and Target AerosolConcentration:
Males Females Target AerosolConcentration(mg/L air)
Group 1 1-3 4-6 slightly above 5
Group 2 7-9 10-12 slightly above 1

Rationale for Aerosol Concentration:
Group 1:
The target concentration of slightly above 5 mg/L air
for 4 hours is the recommended concentration for a
limit test (OECD 436, “Acute Inhalation Toxicity”).
Group 2:
The target concentration of 1 mg/L air for 4 hours is
the recommended concentration according to annex
3 d (OECD 436, “Acute Inhalation Toxicity”).
Duration of Observation Period: 14 days

Inhalation Exposure System
Inhalation exposure was performed using a system similar to that originally described by Sachsse
et al. (see References (1) and (2)). The animals were confined separately in restraint tubes which
were positioned radially around the flow-past, nose-only exposure chamber as described by
Cannon et al. (see References (3)). The design of this chamber is based upon the fluid dynamic
modeling of the test aerosol flow.
The exposure system ensured a uniform distribution and provided a constant flow of test material
to each exposure tube. The flow of air at each tube was 1.0 L/min, which is sufficient to
minimize re-breathing of the test atmosphere as it is more than twice the respiratory minute
volume of a rat.
Before commencement of the exposure of the groups, technical trials were conducted (without
animals) using the inhalation system foreseen for the study. The technical trials were conducted
using established procedures based on GLP, but were not inspected by the Harlan Laboratories
Ltd. Quality Assurance. Technical trial data are retained in the raw data.

Test Aerosol Generation
The test atmosphere was generated using a Hudson nebulizer connected to a syringe pump. The
polyethylene injector inside the nebulizer was replaced by a stainless steel injector.

Exposure System Monitoring
The aerosol concentration, the particle size distribution, relative humidity, temperature and
oxygen concentration were measured on test aerosol samples taken at a representative exposure
port.

Nominal Determination of Aerosol Concentration
The test item usage was measured by weighing the syringe and nebulizer reservoir containing the
test item before and after exposure to determine the quantity of test item used. The weight used
was then divided by the total air-flow volume to give the nominal concentration.

Gravimetric Determination of Aerosol Concentrations
Gravimetric determinations of aerosol concentration were performed four times during each
exposure. The samples were collected on a Millipore®durapore filter, Type HVLP loaded in a 47
mm in-line stainless steel filter sampling device. The filters were weighed before and
immediately after sampling using a calibrated balance. The test aerosol concentration was
calculated from the amount of test item present on the filter and the sample volume.

Chemical Determination of Aerosol Concentrations
Chemical determinations of aerosol concentration were performed four times during each
exposure. The samples were collected on a Millipore®durapore filter, Type HVLP loaded in a 47 mm inline
stainless steel filter sampling device. The filters were transferred into appropriate labeled
vials, forwarded in a cool box to the responsible scientist for analytical chemistry and stored at 2-
8 °C until analysis. The samples were analyzed using a HPLC method supplied by the Sponsor.

Particle Size Distribution and Mass Median Aerodynamic Diameter
The particle size distribution was measured gravimetrically three times during each exposure
using a 7 stage cascade Mercer Impactor (Model 02-130, In-Tox. Products Inc., Albuquerque,
New Mexico, U.S.A.). Mass Median Aerodynamic Diameters and Geometric Standard
Deviations were calculated on the basis of the results from the impactor, using Microsoft Excel
Software. The target range for the Mass Median Aerodynamic Diameter was 1 to 4 μm.

Oxygen Concentration
The oxygen concentration of the test atmosphere was measured continuously during each
exposure using a calibrated device. The results were recorded manually and are reported at 30
minute intervals from the start of exposure. The oxygen concentration was maintained above
19% during each exposure period.

Relative Humidity / Temperature
The temperature and relative humidity of the test atmosphere was measured continuously during
each exposure using a calibrated device. The results were recorded manually and are reported at
30 minute intervals from the start of exposure.

Airflow Rate
The actual airflow rate through the exposure chamber was recorded at approximately 30 minute
intervals from the start of the inhalation exposure.
Statistics:
none
Key result
Sex:
male/female
Dose descriptor:
LC100
Effect level:
4.9 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Key result
Sex:
male/female
Dose descriptor:
LC0
Effect level:
1.01 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Mortality:
6/6 at 4.9 mg/L
Clinical signs:
Mortality and Clinical Signs

Group 1:
One male and one female were found dead on day 2 of the observation period. The remaining
two males were found dead on day 3. One female was found dead on day 11. The remaining
female was killed in extremis on day 15.
Salivation was recorded during and up to immediately after exposure in all animals. Decreased
activity, ruffled fur and labored breathing were observed in all animals after exposure on day 1
and day 2. Ruffled fur persisted up to day 5 in one female. Decreased activity and ruffled fur
were recorded up to day 6 in the other surviving female. Tachypnea was recorded on day 5 and 6
and decreased activity, hunched posture, ruffled fur and tachypnea were seen from day 13 until
necropsy in this female.

Group 2:
All animals survived the scheduled observation period.
Salivation was recorded during and/or immediately after exposure in all animals. Ruffled fur was
observed in all animals on day 1 and 2 of the observation period. Effects on breathing, like
labored breathing or tachypnea, were seen in all animals on the day of exposure up to day 2.
Decreased activity was recorded in all males 1 hour after exposure end. There were no clinical
signs from day 3 onwards.


Body weight:
Group 1:
From test day 1 to test day 2, marked body weight loss was noted in all surviving animals.
Further body weight loss was recorded in the surviving females until day 4. Thereafter body
weight gain was recorded in these animals although one of these females was found dead on day
9. The other female showed a further body weight loss from day 8 to day 15.
Group 2:
From test day 1 to test day 2, slight body weight loss was noted in all males. Stagnation of body
weight was recorded in two females. Thereafter normal body weight development was recorded
in all males and two females. Stagnation of body weight was observed in one female up to day 8.
Gross pathology:
Group 1:
Incompletely collapsed lung, dark red or reddish discoloration of the lung or the thymus, were
seen in several animals during necropsy.
Group 2:
There were no macroscopic findings.
Interpretation of results:
Toxicity Category IV
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
Treatment of HanRcc:WIST(SPF) rats with Tert-butyl Peroxybenzoate at a concentration of 4.9
and 1.01 mg/L for 4 hours resulted in effects on body weight and clinical signs such as ruffled
fur, decreased activity and breathing problems.
Incompletely collapsed lung and dark red discoloration of the lung, seen in the female that had to
be killed in extremis after exposure at 4.9 mg/L air, may be related to treatment with the test item.
All other macroscopic findings were considered to be due to the premature death of the animals.
Exposure at 4.9 mg/L air caused the premature death of 6 animals. The remaining female was
killed on day 15 due to marked body weight loss during the observation period.
In conclusion, the LC50 of Tert-butyl Peroxybenzoate obtained in this study was estimated to be
between 4.9 and 1.01 mg/L air (chemically determined mean aerosol concentration). Due to this
LC50 range the test item was classified in category 4 (GHS). There was no indication of relevant
sex-related differences in toxicity of the test item.
Executive summary:

Two groups of three male and three female albino rats [HanRcc:WIST(SPF)] were exposed by nose-only, flow-past inhalation for four hours to the test item at a chemically determined mean concentration of 4.9 and 1.01 mg/L air, respectively. All surviving animals were observed for clinical signs and mortality during the inhalation exposure and the observation period of 14 days. Body weights were recorded prior to exposure on test day 1, and during the observation period on test days 2, 4, 8 and 15 before necropsy. On day 15 all surviving animals were sacrificed and necropsied. The ranges of aerosol concentration, temperature, relative humidity, oxygen content and airflow rate measured during the exposure were considered to be satisfactory for a study of this type. In addition, the test item was considered to be respirable to rats. Group 1: One male and one female were found dead on day 2 of the observation period after exposure at 4.9 mg/L air. The remaining two males were found dead on day 3. One female was found dead on day 11. The remaining female was killed for humane reasons on day 15. At 4.9 mg/L air salivation was recorded during up to immediately after exposure in all animals. Decreased activity, ruffled fur and labored breathing were observed in all animals after exposure on day 1 and day 2. These signs persisted partly up to day 6. Decreased activity, hunched posture, ruffled fur and tachypnea were seen from day 13 until necropsy in this female. Marked effects on body weights were recorded in all animals. Group 2: All animals survived the scheduled observation period at 1.01 mg/L air. Salivation was recorded during and/or immediately after exposure in all animals. Decreased activity was recorded in all males 1 hour after exposure end. Ruffled fur and effects on breathing were recorded in most of the animals on days 1 and 2. There were no clinical signs from day 3 onwards. Effects on body weights were recorded in all animals. There were no macroscopic findings in groups 1 and 2 that were considered to be related to treatment with the test item. In conclusion, the LC50 of tert-butyl perbenzoate (CAS# 614-45-9) obtained in this study was estimated to be between 1.01 and 4.9 mg/L air (chemically determined mean aerosol concentration). Due to this LC50 range the test item was classified in category 4 (GHS). There was no indication of relevant sex-related differences in toxicity of the test item.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
1 010 mg/m³
Quality of whole database:
A study, reliable without restrictions, is available.

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Conducted according to OECD 402, in full comformance with GLP.
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
GLP compliance:
yes (incl. certificate)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
other: RccHan:WIST(SPF)
Sex:
male/female
Details on test animals and environmental conditions:
Test System
Animals: Rat, RccHanTM: WIST(SPF)
Rationale: Recognized by international guidelines as a
recommended test system.
Breeder: Harlan Laboratories B.V.
Kreuzelweg 53
5961 NM Horst / The Netherlands
Number of Animals per Group: 5 males and 5 females
Total Number of Animals: 5 males and 5 females
Age (when treated): Males: 9 weeks
Females: 12 weeks
Body Weight Range (when treated): 232.2 g - 259.0 g (males)
186.6 g - 196.6 g (females)
Identification: Unique cage number and corresponding color-coded
spots on the tail. The animals were marked at
acclimatization start.
Randomization: Selected by hand at time of delivery.
No computer generated randomization program.
Acclimatization: Under laboratory conditions, after health
examination. Only animals without any visible signs
of illness were used for the study.

Husbandry
Room Number: Harlan Laboratories Ltd. / Itingen, 1OG.133A
Conditions: Standard Laboratory Conditions. Air-conditioned
with 10 - 15 air changes per hour, and continuously
monitored environment with ranges for room
temperature 22 ± 3 °C and for relative humidity
between 30 - 70% (values above 70% during
cleaning process possible), automatically controlled
light cycle of 12 hours light and 12 hours dark,
music during the daytime light period.
Accommodation: During acclimatization in groups of five per sex in
Makrolon type-4 cages with standard softwood
bedding. Individually in Makrolon type-3 cages with
standard softwood bedding (‘Lignocel’ J. Rettenmaier
& Söhne GmbH & Co. KG, 73494 Rosenberg
/ Germany, imported by Provimi Kliba AG,
4303 Kaiseraugst / Switzerland) during treatment
and observation.
Diet: Pelleted standard Provimi Kliba 3433 rat/mouse
maintenance diet, batch no. 58/09 (Provimi Kliba
AG, 4303 Kaiseraugst / Switzerland) ad libitum.
Results of respective analyses for contaminants are
included.
Water: Community tap water from Itingen ad libitum.
Results of bacteriological assay, chemical and
contaminant analyses of respective samples are
included.
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
One day before treatment, the backs of the animals were clipped with an electric clipper,
exposing an area of approximately 10% of the total body surface.
Only those animals without injury or irritation on the skin were used in the test.
On test day 1, the test item was applied at a dose of 2000 mg/kg body weight evenly on the intact
skin with a syringe and covered with a semi-occlusive dressing. The dressing was wrapped
around the abdomen and fixed with an elastic adhesive bandage.
Application volume/kg body weight: 2 mL
Twenty-four hours after the application the dressing was removed and the skin was flushed with
lukewarm tap water and drapped off with disposable paper towels. Thereafter, the reaction sites
were assessed.
Duration of exposure:
24 h
Doses:
2000 mg/kg (2ml/kg)
No. of animals per sex per dose:
5M & 5F
Control animals:
no
Details on study design:
Preparation of Dose Formulations
Dose levels are in terms of the test item as supplied by the Sponsor.
The test item was applied undiluted as delivered from the Sponsor. The doses were calculated
based upon the rats’ body weight.

Test Item Administration
One day before treatment, the backs of the animals were clipped with an electric clipper,
exposing an area of approximately 10% of the total body surface.
Only those animals without injury or irritation on the skin were used in the test.
On test day 1, the test item was applied at a dose of 2000 mg/kg body weight evenly on the intact
skin with a syringe and covered with a semi-occlusive dressing. The dressing was wrapped
around the abdomen and fixed with an elastic adhesive bandage.

Application volume/kg body weight: 2 mL

Twenty-four hours after the application the dressing was removed and the skin was flushed with
lukewarm tap water and drapped off with disposable paper towels. Thereafter, the reaction sites
were assessed.

Rationale
Dermal administration was used as this is one possible route of human exposure during
manufacture, handling and use of the test item.

Observations
Viability / Mortality: Daily during the acclimatization period, within the
first 30 minutes and at approximately 1, 2, 3 and 5
hours after administration on test day 1 (with the
clinical signs) and twice daily during days 2 - 15.

Clinical Signs: Daily during the acclimatization period, within the
first 30 minutes and at approximately 1, 2, 3 and 5
hours after administration on test day 1. Once daily
during days 2 - 15.

Local Dermal Signs: Once daily during days 2 (following dressing
removal) through day 15 using the numerical scoring
system described in Appendix IV on p. .
Body Weights: On test days 1 (prior to administration), 8 and 15.
Pathology
Necropsy
All animals were killed at the end of the observation period by an intraperitoneal injection of
vetanarcol at a dose of at least 2.0 mL/kg body weight (equivalent to at least 324 mg sodium
pentobarbitone/kg body weight) and discarded after macroscopic examinations were performed.
An external examination and opening of the abdominal and thoracic cavities for examinations of
major organs were performed. The appearance of any macroscopic abnormalities was recorded.
No organs or tissues were retained.

Determination and Scoring of Clinical Signs and Local Dermal Signs
Observation
Data were summarized in tabular form, showing for each individual animal the clinical and local
signs at each measurement interval. All findings were described.
The skin reaction was assessed according to the numerical scoring system listed in the
Commission Regulation (EC) No 440/2008 B.4 (see Appendix IV on p. ).

Statistical Analysis
No statistical analysis was used.

Data Compilation
Body weights were recorded on-line.
Clinical and local dermal signs were recorded on data sheets.

Mortality/viability were compiled into the RCC Tox Computer System during recording and/or
recorded on data sheets.

Macroscopic findings were compiled into the RCC Tox Computer System during recording.
The RCC Tox Computer System (RCC-Tox-Lims) had been validated with respect to data
collection, storage and retrievability.
Statistics:
none
Key result
Sex:
male/female
Dose descriptor:
LD0
Effect level:
2 000 mg/kg bw
Based on:
test mat.
Mortality:
none
Clinical signs:
none
Body weight:
not affected
Gross pathology:
dermal irritation
Other findings:
Generalized erythema, slight in degree, was noted in all animals after removal of the semiocclusive
bandage 24 hours after administration. This finding was noted Initally in all males,
(persisting between two and five days) and in all females (persisting for one to five days). Slight
to moderate desquamation was noted in all five males and two of five females, with localized
necrotic areas in one male and one female which persisted until the last day of observation.
Interpretation of results:
not classified
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
The median lethal dose of tert-butyl peroxybenzoate after single dermal administration to rats of
both sexes, observed over a period of 14 days, is:
LD50 (rat): greater than 2000 mg/kg body weight
Executive summary:

Five male and five female RccHan:WIST (SPF) rats were treated with tert-butyl peroxybenzoate at 2000 mg/kg by dermal application. The test item was applied undiluted as delivered from the Sponsor at a volume dosage of 2 mL/kg. The application period was 24 hours. The animals were examined daily during the acclimatization period and mortality, viability and clinical signs were recorded. All animals were examined for clinical signs within the first 30 minutes and at approximately 1, 2, 3 and 5 hours after treatment on day 1 and once daily during test days 2 - 15. Local signs were noted once daily from test day 2 to 15. Mortality/viability was recorded within the first 30 minutes and at approximately 1, 2, 3 and 5 hours after administration on test day 1 (with the clinical signs) and twice daily during days 2 - 15. Body weights were recorded on day 1 (prior to administration) and on days 8 and 15. All animals were necropsied and examined macroscopically. No deaths occurred during the study. No clinical signs were evident during the course of the study. The body weight of the animals was within the range commonly recorded for this strain and age. Local effects included slight generalized erythema Initally in all males and females. This finding persisted up to five days in both sexes, although males seemed to be more susceptible. Slight to moderate desquamation, noted in all five males and two of five females, was accompanied by localized necrotic areas in one male and one female. This latter finding persisted until the last day of observation. At necropsy, dermal sores were noted on the treated skin of two males (nos. 1 and 4) and one female (no. 10) and were considered to be related to the treatment with the test item. The median lethal dose of tert-butyl peroxybenzoate after single dermal administration to rats of both sexes, observed over a period of 14 days, is: LD50 (rat): greater than 2000 mg/kg body weight

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
discriminating dose
2 000 mg/kg bw
Quality of whole database:
A study, reliable without restrictions, is available.

Additional information

The actual LD50s are GREATER THAN the referenced values.

The actual LC50 is between 1.01 and 4.9 mg/L

Justification for selection of acute toxicity – oral endpoint

Apparently well conducted GLP study.  Other studies support the conclusion.  

Justification for selection of acute toxicity – inhalation endpoint

Apparently well conducted GLP study.

Justification for selection of acute toxicity – dermal endpoint

Apparently well conducted GLP study.

Justification for classification or non-classification

Acute lethal values for oral and dermal exposure are above the minimum EU GHS classification levels therefore no classification is warranted.

The LC50 value is category 4 based on EU GHS classification.