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Administrative data

Description of key information

Data obtained from the Combined Repeated-Dose and Reproduction / Developmental Screening Study in rats revealed an oral NOAEL of 50 mg/kg bw/day calculated as peroxide for parental effects. Therefore, the NOAEL is expected to be 100 mg/kg/ bwday for the test material (50% act. ingr. in Shellsol T). A sub-acute inhalation study in rats indicates a NOAEC value of 280 mg/cubic meter for the active ingredient. The NOAEC is calculated to be 560 mg/cubic meter for the test material (50% tert-butyl peroxyacetate in Shellsol T).

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2007-03-14 and 2008-05
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
1996
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OPPTS 870.3650 “Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test” (2000)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: RCC Ltd, Laboratory Animal Services, Wölferstrasse 4, 4414 Füllinsdorf / Switzerland
- Age at study initiation: males (8 weeks), females (10 weeks)
- Weight at study initiation: Males: 240 - 280 grams, Females: 175 - 215 grams
- Fasting period before study: none
- Housing: Animals were housed in Makrolon cages (type-3) with wire mesh tops and standard granulated softwood bedding (Lignocel, Schill AG, 4132 Muttenz/Switzerland).
- Diet: Pelleted standard Kliba 3433 rat/mouse maintenance diet (Provimi Kliba AG, 4303 Kaiseraugst/Switzerland) was available ad libitum (Batch No. 67/06).
- Water: Tap water from Füllinsdorf in bottles was available ad libitum.
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/ 3
- Humidity (%): 30 -70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
1%, medium viscosity
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The test item was weighed into a glass beaker on a tared precision balance and approximately 80% of the vehicle were added (w/v). Using an appropriate homogenizer a homogeneous mixture was prepared. Having obtained a homogeneous mixture, vehicle was added until the required final volume was achieved. Separate formulations were prepared for each concentration. During the daily administration period homogeneity of the test item in the vehicle was maintained using a magnetic stirrer.
Frequency of dose formulation: weekly, Storage of dose formulations: refrigerator (2 - 8 °C)

VEHICLE
- Justification for use and choice of vehicle: 1% CMC was used as the vehicle for the test item in the dose groups and was administered as the control item to the animals of the control group. The test item was miscible in saturated aliphatic and aromatic solvents, immiscible in water at 10°C.
- Lot/batch no.: 1119535 24604357
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples for determination of concentration, homogeneity and stability (4 hours and 7 days) of the dose formulations were taken during the first week of the administration period. Additionally, samples for determination of concentration and homogeneity were taken during the last week of the administration period. The test item concentrations were determined by HPLC coupled to an UV/VIS detector (VWR-Hitachi L-2400).
Duration of treatment / exposure:
Males: 42 days
Females: from 14 days prior to mating until day 4 post partum (54 days in total)
Frequency of treatment:
once daily
Remarks:
Doses / Concentrations:
0, 50, 150, 500 mg/kg bw/day
Basis:
other: as peroxide
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
The rat is a suitable rodent species for repeated dose and reproduction/developmental toxicity studies required by regulatory authorities. The oral route is one possible route for human exposure.
Dose levels were selected in agreement with the Sponsor, based on the results of a preliminary dose range finding study (RCC Study No. A42390), where 600 mg/kg/ day as peroxide was used as highest dose level.
Positive control:
no data
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- daily
- Cage side observations checked: unusual body movements, abnormal behavior and posture as well as resistance to removal

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once prior to the first test item administration and weekly thereafter.

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION:
- Mean daily food consumption were calculated by a computer program based on on-line record data.

COMPOUND INTAKE: not examined

FOOD EFFICIENCY: not examined

WATER CONSUMPTION AND COMPOUND INTAKE: not examined

OPHTHALMOSCOPIC EXAMINATION: not examined

HAEMATOLOGY: Yes
- Time schedule for collection of blood: males: on the day before or on the day of scheduled necropsy, females: on day 5 post partum
- Anaesthetic used for blood collection: Yes, light isoflurane anaesthesia
- Animals fasted: Yes (overnight)
- Parameters checked: Erythrocyte count, Haemoglobin, Haematocrit, Mean corpuscular volume, Red cell volume distribution width, Mean corpuscular haemoglobin, Mean corpuscular haemoglobin concentration, Haemoglobin concentration distribution width, Platelet count, Total leukocyte count, Differential leukocyte count, Coagulation: Thromboplastin time, Activated partial thromboplastin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: males: on the day before or on the day of scheduled necropsy, females: on day 5 post partum
- Anaesthetic used for blood collection: Yes, light isoflurane anaesthesia
- Animals fasted: Yes (overnight)
- Parameters checked: Glucose, Urea, Creatinine, Bilirubin (total), Cholesterol (total), Aspartate aminotransferase, Alanine aminotransferase, Bile acids, Alkaline phosphatase, Gamma-glutamyl-transferase, Sodium, Potassium, Chloride, Calcium, Phosphorus inorganic, Protein (total), Albumin, Globulin, Albumin/Globulin ratio

URINALYSIS: not examined

NEUROBEHAVIOURAL EXAMINATION: not examined

OTHER: organ weight, mortality rate, functional observational battery
Sacrifice and pathology:
Males were sacrificed after the first dams had reached day 4 post partum. Females were sacrificed on day 5 post partum after blood sampling.
GROSS PATHOLOGY: yes
HISTOPATHOLOGY: yes

Organ weights were determined from the following organs for five adult males and females (wet weight):
liver, spleen, adrenals*, brain, thymus, heart, kidneys* (*weight as pairs)
The testes* and epididymides* of all parental males were weighed.

Of all parental males the following tissues were preserved in neutral phosphate buffered 4% formaldehyde solution: prostate, testes (in Bouin's fixative), seminal, vesicles with coagulation gland and epididymides (in Bouin's fixative). Of all parental females ovaries were taken.

In addition, of the five males and females per group selected for organ weights, the following tissues were preserved in neutral phosphate buffered 4% formaldehyde solution: gross lesions, heart, brain, thymus, spinal cord, thyroid, small and large intestines (incl. Peyer's patches), trachea and lungs (preserved by inflation with fixative), stomach, uterus (with vagina), liver, urinary bladder, kidneys, lymph nodes (mandibular, mesenterial), adrenals, peripheral nerve, spleen and bone marrow.
Statistics:
- If the variables could be assumed to follow a normal distribution, the Dunnett t-test, based on a pooled variance estimate, was used for inter-group comparisons.
- The Steel test (rank test) was applied when the data could not be assumed to follow a normal distribution.
- Fisher's Exact test for 2x2 tables was applied if the variables could be dichotomized without loss of information.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Mortality:
mortality observed, treatment-related
Description (incidence):
see details on results
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
see details on results
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY: 500 mg/kg/day: 3 dead males, clinical signs in all animals (ruffled fur, irregular breathing, ventral recumbency, uncoordinated gait, cold to touch); 500 mg/kg/day and 150 mg/kg/day: signs of discomfort in all animals in the way that the animals moved their heads through the bedding material after the daily administration of test item

BODY WEIGHT AND WEIGHT GAIN: dose-dependently decreased in both genders

FOOD CONSUMPTION: Mean food consumption was decreased in female animals at 500 mg/kg during pre-pairing and gestation period. Compound intake was not examined.

HAEMATOLOGY: In males, a tendency of inflammatory response could be noted as increase of total leukocyte count, increase of absolute and relative neutrophils, and increase of platelets at 500 mg/kg.

CLINICAL CHEMISTRY: A few parameters were noted with statistically significantly values still within the range of the reference values. Some parameters were increased in all groups. Therefore, none of the parameters under investigation for clinical biochemistry was considered to be affected by exposure to the test item.

ORGAN WEIGHTS: In male animals of group 4 (500 mg/kg bw), the mean liver weight was statistically significantly increased. But for organ/body weight ratio, brain, heart, kidneys, adrenals, and spleen were statistically significantly increased in group 4 (500 mg/kg bw).The epididymides’ weight was statistically significantly decreased while the relative epididymides’ weight was statistically significantly increased. Mean testes weight ratio was statistically significantly increased in groups 3 (150 mg/kg bw) and 4 (500 mg/kg bw). In female animals of group 4 (500 mg/kg bw), the mean organ weights of liver, adrenals, and spleen were statistically significantly increased.

GROSS PATHOLOGY: at 500 mg/kg/day, the gastro-intestinal organs were noted with signs of adaptive defense (thickened mucosa, altered mesenteric lymph nodes), fibrin-like coatings or adherences

HISTOPATHOLOGY: adverse treatment-related changes noted in stomach and small intestine, liver, bone marrow, spleen, lymphatic organs, and in the males reproductive organs at 500 mg/kg/day; stomach was also affected in animals at 150 and 50 mg/kg/day, the duodenum and jejunum in animals at 150 mg/kg/day at 50 mg/kg/day

OTHER FINDINGS: Behavioral investigations (slight piloerection, reduced activity, reduced rectal body temperature, slightly reduced locomotor activity) were considered to be influenced by treatment at 500 mg/kg.
Key result
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: histological findings in the gastrotestinal tract
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: histological findings in the gastrointestinal tract
Critical effects observed:
not specified

Effects were noted on reproduction data at 500 mg/kg/day as reduction of implantations, small number of live pups/dam at birth and at day 4 of lactation, smaller litter weight and slightly smaller pup weight.

Conclusions:
Based on these data, it can be concluded that the No Observed Adverse Effect Level (NOAEL) for reproductive and developmental effects was 150 mg/kg body weight/day; and 50 mg/kg/day calculated as peroxide for parental effects based on histological findings in the gastro-intestinal tract. Therefore, the NOAEL of the test material (50% in Shellsol T) is expected to be 100 mg/kg/day for parental effects.
Executive summary:

TBPA was administered once daily orally (by gavage) at dosages of 0, 50, 150, and 500 mg/kg/day calculated as peroxide, to male rats for 42 days in total and to female rats throughout the pre-pairing, the pairing, the gestation and the lactation periods until day 4 post partum (last dosing). Treatment at 500 mg/kg/day was associated with death of three males, and severe clinical symptoms in all animals (ruffled fur, irregular breathing, ventral recumbency, uncoordinated gait, cold to touch). A sign of discomfort was noted in all animals at 500 mg/kg/day and 150 mg/kg/day in the way that the animals moved their heads through the bedding material after the daily administration of test item. Mean food consumption was decreased in female animals at 500 mg/kg during pre-pairing and gestation period. In males, mean food consumption was dose-dependently decreased at all test item treated groups. Mean body weights were dose-dependently decreased in both gender. Behavioral investigations (slight piloerection, reduced activity, reduced rectal body temperature, slightly reduced locomotor activity) were considered to be influenced by treatment at 500 mg/kg. In males, a tendency of inflammatory response could be noted as increase of total leukocyte count, increase of absolute and relative neutrophils, and increase of platelets at 500 mg/kg. After treatment at 500 mg/kg/day, the gastro-intestinal organs were noted with signs of adaptive defense (thickened mucosa, altered mesenteric lymph nodes), fibrin-like coatings or adherences. Treatment at 500 mg/kg/day was associated with increased liver and kidney weights. Histopathological effects were noted in stomach and small intestine, liver, bone marrow, spleen, lymphatic organs, and in the males reproductive organs at 500 mg/kg/day. The stomach was also affected in animals at 150 and 50 mg/kg/day, the duodenum and jejunum in animals at 150 mg/kg/day and the duodenum in females at 50 mg/kg/day. Based on these data, it can be concluded that the No Observed Adverse Effect Level (NOAEL) for parental effects was 50 mg/kg/day calculated as peroxide based on histological findings in the gastro-intestinal tract. Therefore, the NOAEL is expected to be 100 mg/kg/day for the test material (50% act. ingr. in Shellsol T).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
100 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
according to OECD guideline, GLP-conform

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04-1975 and 01-1976
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
non-GLP conform
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands
- Weight at study initiation: average bw 67 g
- Diet: ad libitum
- Water: ad libitum
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass exposure cylinder with stainless steel interior
- Method of holding animals in test chamber: each exposure cylinder accommodated five male and five female rats, seperated from one another
- Temperature: 23°C
- Air flow rate: 10 L/min

TEST ATMOSPHERE
- Brief description of analytical method used: TBPA was evaporated by passing a measured, filtered and dried air flow through glass evaporation columns, packed with chromosorb W (60 to 80 mesh) at 23°C.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of the air were taken at intervals and analysed by means of GLC, using a 0.35 m x 4 mm column of stainless steel tubing packed with QF-1 4.8% / diglycerol 0.13% on Chrom G-AW-DHCS and a flame ionization detector.
Duration of treatment / exposure:
28 days
Frequency of treatment:
7 hours/day, 5 days/week fo a four-week period
Remarks:
Doses / Concentrations:
0, 2, 11, 52 ppm
Basis:
other: active ingredient
Remarks:
Doses / Concentrations:
0, 10.9, 60.4, 285.6 mg/m3
Basis:
other: active ingredient
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Positive control:
no positive control
Observations and examinations performed and frequency:
BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded weekly.
- How many animals: 10 male and 10 female rats per doses

HAEMATOLOGY: Yes
Haematological data (haemoglobin content, haematocrit value, and erythrocyte and leucocyte counts) were collected in week 4.
- How many animals: 10 male and 10 female rats per doses

URINALYSIS: Yes
- Urine examinations were made, including appearance, pH, glucose, protein, occult blood, ketones, using test stripes, and microscopy of the sediment in pooled urine samples of each group.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
- The organ weight of the heart, kidneys, liver, spleen and lung was determined of groups of ten male and ten female rats after a test period of four weeks.
HISTOPATHOLOGY: Yes
- Tissue samples of the organs described before and of the head (after removal of the skin, brain and lower jaw) were taken.
Other examinations:
No other examinations
Statistics:
Student's t-test
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Growth of the males of the low and intermediate doses groups was slightly, but statistically significantly retarded during the first weeks (see table 1 other information on results)
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:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
slightly elevated lung weight in males and a slightly lowered spleen weight in females at the low exposure levele; see table 2 (other information on results)
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
GROSS PATHOLOGY: Infrequently occuring gross changes unrelated to the inhalation of the active ingredient included pale livers and kidneys, a striking lobular pattern of the liver surface and hydronephrosis.

HISTOPATHOLOGY: NON-NEOPLASTIC: Histopathological changes occurred to about the same degree and frequency in the various groups studied histologically and are, moreover, quite common findings in the strain of rats used.


Key result
Dose descriptor:
NOAEC
Effect level:
52 ppm
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed up to 52 ppm.
Dose descriptor:
NOAEC
Effect level:
0.28 mg/L air
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed
Dose descriptor:
NOAEC
Effect level:
104 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Dose descriptor:
NOAEC
Effect level:
0.56 mg/L air
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Critical effects observed:
not specified

Table 1: Mean body weights of groups of ten male and ten female rats

Animal

TBP [ppm]

Mean body weight at end of week

        0                      1                         2       3                    4

Males

1

0

67

124

147

188

221

2

2

66

111***

134***

173**

193*

3

11

67

111***

134***

176*

208

4

52

67

121

145

186

215

Females

1

0

66

105

117

136

149

2

2

67

103

117

138

152

3

11

67

106

118

142

158

4

52

67

106

119

136

149

*p < 0.05, **p < 0.01, ***p < 0.001

Table 2: Relative organ weights (in g/100 g body weight) and their standard deviations of groups of ten male and ten female rats after test period of four weeks

TBP [ppm]

heart

kidneys

liver

spleen

lung

Males

0

0.383 (0.015)

0.80 (0.02)

5.47 (0.20)

0.293 (0.009)

0.61 (0.02)

2

0.417 (0.029)

0.85 (0.06)

5.67 (0.45)

0.303 (0.022)

0.69* (0.04)

11

0.378 (0.011)

0.79 (0.01)

5.26 (0.12)

0.271 (0.012)

0.58 (0.01)

52

0.377 (0.013)

0.77 (0.01)

5.38 (0.11)

0.290 (0.007)

0.63 (0.03)

Females

0

0.397 (0.013)

0.80 (0.02)

4.62 (0.13)

0.300 (0.012)

0.66 (0.03)

2

0.393 (0.014)

0.79 (0.01)

4.76 (0.08)

0.269* (0.008)

0.65 (0.01)

11

0.405 (0.008)

0.78 (0.02)

4.71 (0.07)

0.297 (0.018)

0.64 (0.01)

52

0.386 (0.008)

0.79 (0.01)

4.48 (0.14)

0.288 (0.009)

0.65 (0.02)

*p < 0.05, according to Student's t-test

Conclusions:
The results of the present four-week inhalation study indicates that the NOAEC of the active ingredient is higher than 52 ppm (correspond to 0.28 mg/L). Hence, the NOAEC for the test material (50% act. ingr. in Shellsol T) is expected to be approx. 0.56 mg/L.
Executive summary:

The sub-acute inhalation toxicity of the test substance (50% active ingredient) was studied in rats by exposing them 7 hours/day, 5 days/week to atmosphere, containing the vapour of the active ingredient at the concentrations 0, 2, 11 and 52 ppm (~ 0.28 mg/L), for a four-week period. Observations were made of behaviour, general appearance, growth, haematology, urine composition, organ weights and gross as well as microscopic pathology. None of the criteria applied revealed treatment-related effects. From the results of the present four-week inhalation study with the test substance it appeared that no deleterious effects could be demonstrated in rats exposed repeatedly to concentrations of the active ingredient of up to 52 ppm (0.28 mg/L) during twenty exposure days. It is concluded, therefore, that the NOAEC is higher than 0.28 mg/L. Therefore, the NOAEC for the test material (50% act. ingr. in Shellsol T) is expected to be approx. 0.56 mg/L.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
560 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
equivalent to OECD guideline; acceptable experimental procedure for assessment

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04-1975 and 01-1976
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
non-GLP conform
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands
- Weight at study initiation: average bw 67 g
- Diet: ad libitum
- Water: ad libitum
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass exposure cylinder with stainless steel interior
- Method of holding animals in test chamber: each exposure cylinder accommodated five male and five female rats, seperated from one another
- Temperature: 23°C
- Air flow rate: 10 L/min

TEST ATMOSPHERE
- Brief description of analytical method used: TBPA was evaporated by passing a measured, filtered and dried air flow through glass evaporation columns, packed with chromosorb W (60 to 80 mesh) at 23°C.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of the air were taken at intervals and analysed by means of GLC, using a 0.35 m x 4 mm column of stainless steel tubing packed with QF-1 4.8% / diglycerol 0.13% on Chrom G-AW-DHCS and a flame ionization detector.
Duration of treatment / exposure:
28 days
Frequency of treatment:
7 hours/day, 5 days/week fo a four-week period
Remarks:
Doses / Concentrations:
0, 2, 11, 52 ppm
Basis:
other: active ingredient
Remarks:
Doses / Concentrations:
0, 10.9, 60.4, 285.6 mg/m3
Basis:
other: active ingredient
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Positive control:
no positive control
Observations and examinations performed and frequency:
BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded weekly.
- How many animals: 10 male and 10 female rats per doses

HAEMATOLOGY: Yes
Haematological data (haemoglobin content, haematocrit value, and erythrocyte and leucocyte counts) were collected in week 4.
- How many animals: 10 male and 10 female rats per doses

URINALYSIS: Yes
- Urine examinations were made, including appearance, pH, glucose, protein, occult blood, ketones, using test stripes, and microscopy of the sediment in pooled urine samples of each group.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
- The organ weight of the heart, kidneys, liver, spleen and lung was determined of groups of ten male and ten female rats after a test period of four weeks.
HISTOPATHOLOGY: Yes
- Tissue samples of the organs described before and of the head (after removal of the skin, brain and lower jaw) were taken.
Other examinations:
No other examinations
Statistics:
Student's t-test
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Growth of the males of the low and intermediate doses groups was slightly, but statistically significantly retarded during the first weeks (see table 1 other information on results)
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:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
slightly elevated lung weight in males and a slightly lowered spleen weight in females at the low exposure levele; see table 2 (other information on results)
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
GROSS PATHOLOGY: Infrequently occuring gross changes unrelated to the inhalation of the active ingredient included pale livers and kidneys, a striking lobular pattern of the liver surface and hydronephrosis.

HISTOPATHOLOGY: NON-NEOPLASTIC: Histopathological changes occurred to about the same degree and frequency in the various groups studied histologically and are, moreover, quite common findings in the strain of rats used.


Key result
Dose descriptor:
NOAEC
Effect level:
52 ppm
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed up to 52 ppm.
Dose descriptor:
NOAEC
Effect level:
0.28 mg/L air
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed
Dose descriptor:
NOAEC
Effect level:
104 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Dose descriptor:
NOAEC
Effect level:
0.56 mg/L air
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Critical effects observed:
not specified

Table 1: Mean body weights of groups of ten male and ten female rats

Animal

TBP [ppm]

Mean body weight at end of week

        0                      1                         2       3                    4

Males

1

0

67

124

147

188

221

2

2

66

111***

134***

173**

193*

3

11

67

111***

134***

176*

208

4

52

67

121

145

186

215

Females

1

0

66

105

117

136

149

2

2

67

103

117

138

152

3

11

67

106

118

142

158

4

52

67

106

119

136

149

*p < 0.05, **p < 0.01, ***p < 0.001

Table 2: Relative organ weights (in g/100 g body weight) and their standard deviations of groups of ten male and ten female rats after test period of four weeks

TBP [ppm]

heart

kidneys

liver

spleen

lung

Males

0

0.383 (0.015)

0.80 (0.02)

5.47 (0.20)

0.293 (0.009)

0.61 (0.02)

2

0.417 (0.029)

0.85 (0.06)

5.67 (0.45)

0.303 (0.022)

0.69* (0.04)

11

0.378 (0.011)

0.79 (0.01)

5.26 (0.12)

0.271 (0.012)

0.58 (0.01)

52

0.377 (0.013)

0.77 (0.01)

5.38 (0.11)

0.290 (0.007)

0.63 (0.03)

Females

0

0.397 (0.013)

0.80 (0.02)

4.62 (0.13)

0.300 (0.012)

0.66 (0.03)

2

0.393 (0.014)

0.79 (0.01)

4.76 (0.08)

0.269* (0.008)

0.65 (0.01)

11

0.405 (0.008)

0.78 (0.02)

4.71 (0.07)

0.297 (0.018)

0.64 (0.01)

52

0.386 (0.008)

0.79 (0.01)

4.48 (0.14)

0.288 (0.009)

0.65 (0.02)

*p < 0.05, according to Student's t-test

Conclusions:
The results of the present four-week inhalation study indicates that the NOAEC of the active ingredient is higher than 52 ppm (correspond to 0.28 mg/L). Hence, the NOAEC for the test material (50% act. ingr. in Shellsol T) is expected to be approx. 0.56 mg/L.
Executive summary:

The sub-acute inhalation toxicity of the test substance (50% active ingredient) was studied in rats by exposing them 7 hours/day, 5 days/week to atmosphere, containing the vapour of the active ingredient at the concentrations 0, 2, 11 and 52 ppm (~ 0.28 mg/L), for a four-week period. Observations were made of behaviour, general appearance, growth, haematology, urine composition, organ weights and gross as well as microscopic pathology. None of the criteria applied revealed treatment-related effects. From the results of the present four-week inhalation study with the test substance it appeared that no deleterious effects could be demonstrated in rats exposed repeatedly to concentrations of the active ingredient of up to 52 ppm (0.28 mg/L) during twenty exposure days. It is concluded, therefore, that the NOAEC is higher than 0.28 mg/L. Therefore, the NOAEC for the test material (50% act. ingr. in Shellsol T) is expected to be approx. 0.56 mg/L.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
560 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
equivalent to OECD guideline; acceptable experimental procedure for assessment

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available
Quality of whole database:
In accordance with column 2 of REACH Annex IX, the test repeated dose toxicity after dermal application (required in section 8.6) does not need to be conducted as repeated dose toxicity studies for oral as well as inhalation application are available. Thus, no test on repeated dose dermal toxicity has to be conducted and risk assessment is based on the oral and inhalation key studies.

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available
Quality of whole database:
In accordance with column 2 of REACH Annex IX, the test repeated dose toxicity after dermal application (required in section 8.6) does not need to be conducted as repeated dose toxicity studies for oral as well as inhalation application are available. Thus, no test on repeated dose dermal toxicity has to be conducted and risk assessment is based on the oral and inhalation key studies.

Additional information

Oral:

Key study

TBPA was administered once daily orally (by gavage) at dosages of 0, 50, 150, and 500 mg/kg bw/day calculated as peroxide, to male rats for 42 days in total and to female rats throughout the pre-pairing, the pairing, the gestation and the lactation periods until day 4 post partum (last dosing). Treatment at 500 mg/kg/day was associated with death of three males, and severe clinical symptoms in all animals (ruffled fur, irregular breathing, ventral recumbency, uncoordinated gait, cold to touch). A sign of discomfort was noted in all animals at 500 mg/kg bw/day and 150 mg/kg bw/day in the way that the animals moved their heads through the bedding material after the daily administration of test item. Mean food consumption was decreased in female animals at 500 mg/kg bw/day during pre-pairing and gestation period. In males, mean food consumption was dose-dependently decreased at all test item treated groups. Mean body weights were dose-dependently decreased in both genders. Behavioral investigations (slight piloerection, reduced activity, reduced rectal body temperature, slightly reduced locomotor activity) were considered to be influenced by treatment at 500 mg/kg bw/day. In males, a tendency of inflammatory response could be noted as increase of total leukocyte count, increase of absolute and relative neutrophils, and increase of platelets at 500 mg/kg bw/day. After treatment at 500 mg/kg bw/day, the gastro-intestinal organs were noted with signs of adaptive defense (thickened mucosa, altered mesenteric lymph nodes), fibrin-like coatings or adherences. Treatment at 500 mg/kg bw/day was associated with increased liver and kidney weights. Histopathological effects were noted in stomach and small intestine, liver, bone marrow, spleen, lymphatic organs, and in the males reproductive organs at 500 mg/kg bw/day. The stomach was also affected in animals at 150 and 50 mg/kg bw/day, the duodenum and jejunum in animals at 150 mg/kg/day and the duodenum in females at 50 mg/kg bw/day.

Based on these data, it can be concluded that the No Observed Adverse Effect Level (NOAEL) for parental effects was 50 mg/kg bw/day calculated as peroxide based on histological findings in the gastro-intestinal tract. Therefore, the NOAEL is expected to be 100 mg/kg bw/day for the test material (50% act. ingr. in Shellsol T).

Supporting study

In a non-GLP preliminary OECD 422 dose range finding study, the doses of 60, 180 and 600 mg/kg body weight (as peroxide) were administered to male wistar rats (for 28 days) and female wistar rats (until day 13 p.c.). For the main study, taking into consideration these results, especially the clinical symptoms observed following administration, the doses of 50, 150, and 500 mg/kg as peroxide are proposed for the combined repeated dose toxicity study with the reproduction / developmental toxicity screening test in the rat.

Inhalation:

Key study

The sub-acute inhalation toxicity of the test substance (50% active ingredient) was studied in rats by exposing them 7 hours/day, 5 days/week to atmosphere, containing the vapour of the active ingredient at the concentrations 0, 2, 11 and 52 ppm (~ 0, 0.011, 0.060 and 0.28 mg/L), for a four-week period. Observations were made of behaviour, general appearance, growth, haematology, urine composition, organ weights and gross as well as microscopic pathology. None of the criteria applied revealed treatment-related effects. From the results of the present four-week inhalation study with the test substance it appeared that no deleterious effects could be demonstrated in rats exposed repeatedly to concentrations of the active ingredient of up to 52 ppm (~ 0.28 mg/L) during twenty exposure days. It is concluded, therefore, that the NOAEC is higher than 0.28 mg/L. The NOAEC of the test material (50% act. ingr. in Shellsol T) is expected to be approx. 0.57 mg/L.

Disregarded Study

One literature study with limited documentation is available. Rats were exposed to the test material (50% tert-butyl peracetate in dimethyl phthalate) with concentrations of 0.15, 0.7 and 1.6 ppm 5 days a week for 28 days. Over the whole concentration range no mortality was observed. Clinical signs and body weight loss were indicated at 0.7 ppm (3.8 mg/cubic m) and 1.6 ppm (8.8 mg/cubic m).


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
only one reliable study

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
only one reliable study

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
only one reliable study

Justification for classification or non-classification

Based on the results of the oral and inhalation repeated dose toxicity studies, tert-butyl ethaneperoxoate was not classified and labelled according to Directive 67/548/EEC (DSD) and to Regulation (EC) No 1272/2008 (CLP).