Registration Dossier

Administrative data

Description of key information

Repeat dose exposure is discussed

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

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Endpoint:
chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1957
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study not conducted in compliance with GLP however, data is included for SIDS OECD works so is deemed reliable. Included as a supporting study, due to the age and route of administration (dietary, no analysis).
Qualifier:
according to
Guideline:
other: Not specified
Principles of method if other than guideline:
The test material (see concentrations above) was administered to rats (25/sex/group) for 64 weeks as a dietary study.
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
other: Carworth
Sex:
male/female
Details on test animals and environmental conditions:
Not specified
Route of administration:
oral: feed
Vehicle:
peanut oil
Details on oral exposure:
The test material was diluted with peanut oil and then added to the diets to product the desired concentrations.
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
Not specified
Duration of treatment / exposure:
Exposure period 64 weeks
Frequency of treatment:
Daily
Remarks:
Doses / Concentrations:
2,500, 5,000, and 10,000 ppm
Basis:
nominal in water
No. of animals per sex per dose:
25 per sex per group
Control animals:
yes, concurrent no treatment
Details on study design:
The test material (see concentrations above) was administered to rats (25/sex/group) for 64 weeks. The animals were individually housed and quarantined for 11 days prior to the start of the study. The control group was fed animal chow without the test material. The test material was diluted with peanut oil and then added to the diets to product the desired concentrations. Hematologic evaluations were conducted on 5 rats/sex/group at the initiation of the test and at interval of 3 months throughout the study. Animals were observed for growth. Histopathological exams were conducted on 88 animals which died during the study (14) or were sacrificed at the end of the weeks 51, 56, and 58. Those animals that were sacrificed were representative of the groups with respect to sex and dietary level.
Positive control:
Not used in the study
Observations and examinations performed and frequency:
Hematologic evaluations were conducted on 5 rats/sex/group at the initiation of the test and at interval of 3 months throughout the study. Animals were observed for growth.
Sacrifice and pathology:
Histopathological exams were conducted on 88 animals which died during the study (14) or were sacrificed at the end of the weeks 51, 56, and 58. Those animals that were sacrificed were representative of the groups with respect to sex and dietary level.
Other examinations:
None specified
Statistics:
Not specified
Clinical signs:
not specified
Mortality:
not specified
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Daily dietary administration significantly retarded growth in females at 2500 ppm and higher.
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:
not specified
Clinical biochemistry findings:
not specified
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Liver enlargement was noted at all concentrations in both sexes.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Degenerative changes in the liver were described as diffuse cloudy swellings and fatty metamorphosis of the cytoplasm of the hepatocyte.
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Details on results:
Daily dietary administration significantly retarded growth in females at 2500 ppm and higher. No effect on growth occurred in males at 2500 ppm.

Liver enlargement was noted at all concentrations in both sexes.

Diffuse hepatic degeneration was observed in all test animals. However, the severity of the liver changes were not treatment-related.

The degenerative changes in the liver were described as diffuse cloudy swellings and fatty metamorphosis of the cytoplasm of the hepatocyte. No compound-related hematopoietic changes were observed in any of the test groups.
Dose descriptor:
LOEL
Effect level:
2 500 ppm
Based on:
test mat.
Sex:
male/female
Critical effects observed:
not specified
Conclusions:
NOEL: Not Identified
LOEL: 2500 ppm
Executive summary:

Study not conducted in compliance with GLP and test guidelines are not referenced within the report, however, the data is included in SIDS OECD works. LOEL = 2500 ppm.

Daily dietary administration significantly retarded growth in females at 2500 ppm and higher. No effect on growth occurred in males at 2500 ppm. Liver enlargement was noted at all concentrations in both sexes. Diffuse hepatic degeneration was observed in all test animals. However, the severity of the liver changes were not treatment-related. The degenerative changes in the liver were described as diffuse cloudy swellings and fatty metamorphosis of the cytoplasm of the hepatocyte. No compound-related hematopoietic changes were observed in any of the test groups.

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
Not specified
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Guideline study, which is rated as reliability 2 because it is a read-across study.
Justification for type of information:
Please refer to IUCLID Section 13 for the read-across justification.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
not specified
GLP compliance:
yes
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Species: rat
Sex: male/female
Strain: Sprague-Dawley Crl:CD® (SD) IGS BR
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Not specified
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
Males: 43 days; Females: up to 54 days
Frequency of treatment:
Daily
Remarks:
Doses / Concentrations:
0, 50, 250 and 600 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
Ten males and ten females pr dosing group.
Control animals:
yes, concurrent vehicle
Details on study design:
Not specified
Positive control:
No data
Observations and examinations performed and frequency:
Clinical signs, behavioural assessments, bodyweight development, food and water consumption were monitored during the study. Haematology and blood chemistry were evaluated prior to mating on five selected males and females from each dose group.
Sacrifice and pathology:
Males were terminated on Day 43, followed by the termination of all surviving females and offspring on Day 5 post partum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed.
Other examinations:
Pairing of animals within each dose group was undertaken on a one male: one female basis on Day 15 of the study, to produce litters. Extensive functional observations were performed on five selected parental males from each dose group after the completion of the mating phase, and for five selected parental females from each dose group on Day 4 post partum.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Elevated alkaline phosphatase levels, reduced cholesterol levels.
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Elevated liver and adrenal weights,
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
liver and thyroid gland
Histopathological findings: neoplastic:
no effects observed
Details on results:
Mortality: No treatment-related deaths were detected.

Clinical Observations: No clinically observable signs of toxicity were detected.

Behavioural Assessments: No treatment-related effects were detected.

Functional Performance Tests: No treatment-related effects were detected.

Sensory reactivity Assessments: No treatment-related effects were detected.

Bodyweights: No adverse effect on bodyweight was observed for males throughout the treatment period, or for females during the maturation, gestation or lactation phases of the study.

Food Consumption: No adverse effect on dietary intake was detected for males throughout the treatment period, or for females during the maturation, gestation or lactation phases of the study.

Water Consumption: No overt intergroup differences were detected. Haematology: No treatment-related changes were detected prior to mating.

Blood Chemistry: Elevated alkaline phosphatase levels were detected for males treated with 600 mg/kg/day. Males treated with 600 and 250 mg/kg/day also showed reduced cholesterol levels. No such effects were detected for females treated with 600 or 250 mg/kg/day or for animals of either sex treated with 50 mg/kg/day.

Necropsy of Adults: No treatment-related macroscopic abnormalities were detected for the interim death female or for the remaining animals at terminal kill.

Organ Weights: Elevated liver and adrenal weights, both absolute and relative to terminal bodyweights, were detected for animals of either sex treated with 600 mg/kg/day.

Histopathology: Histopathological examination of adult tissue revealed the following treatment-related changes:
Liver: Centrilobular hepatocyte enlargement was observed for animals of either sex treated with 600 and 250 mg/g/day, with the effect extending into the female 50 mg/kg/day dose group.
Thyroid glands: Follicular cell hypertrophy was observed for males treated with 600 and probably also at 250 mg/kg/day. No such effects were detected for females at these dose levels, or for animals of either sex treated with 50 mg/kg/day.

These effects however, were considered entirely adaptive in nature, therefore the ‘No Observed Adverse Effect Level’ (NOAEL) was considered to be 600 mg/kg/day.
Dose descriptor:
NOAEL
Effect level:
600 other: mg/kg/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: overall effects
Critical effects observed:
not specified
Conclusions:
No Observed Adverse Effect Level (NOAEL) was considered to be 600 mg/kg/day.
Executive summary:

Study conducted to OECD test guidance in compliance with GLP and data included for OECD SIDS dossier. The oral administration of CAS No 68442-68-2 to rats by gavage, at dose levels of 600, 250 and 50 mg/kg/day, resulted in treatment-related effects at all dose levels. These effects however, were considered entirely adaptive in nature, therefore the ‘No Observed Adverse Effect Level’ (NOAEL) was considered to be 600 mg/kg/day. Read across to supporting substance, CAS No. 68442 -68 -2, by structural analogue.

This substance has been supported under Environmental Protection Agency’s (EPA’s) High Production Volume (HPV) Challenge Program. The American Chemical Councils RAPA Panel, has derived a “Substituted Diphenylamines” category of chemicals for this substance, please refer to EPA reference 201-14700A located at

 

http://www.epa.gov/hpv/pubs/summaries/subdipha/c13378rt.pdf

 

Relying on several factors specified in EPA’s guidance document on “Development of Chemical Categories in the HPV Challenge Program,” in which use of chemical categories is encouraged, the following closely related chemicals constitute a chemical category:

 

Structural Similarity. A key factor supporting the classification of these chemicals as a category is their structural similarity (see Figure 1). All share a common starting material; Diphenylamine (Benzenamine, N-phenyl-, CAS# 122-39-4), a common synthetic pathway, and all compounds in this category are diamines with various substitutions.

 

Similarity of Physicochemical Properties. The similarity of the physicochemical properties of these materials parallels their structural similarity. All are off-white to light brown solids or viscous liquids intended for use as antioxidants in finished rubber articles or as antidegradant additives that extend the useful life of heavy-duty industrial functional fluids used in high-speed, high-temperature and/or high-load applications. As a class, these amine-based antidegradant compounds are less migratory (more polymer-bound) and less staining than the Substituted p-Phenylenediamine antidegradants. The use of these materials requires that they be stable under high temperatures. Their low volatility is due to their low vapor pressure, highly viscous or solid form. The existing information for these materials indicates that they have low water solubility and high flash points.

 

Fate and Transport Characteristics. Members of this category have been shown to be not readily biodegradable, so additional testing is not needed. The lack of water solubility of the members of this category makes hydrolysis testing unnecessary. These materials have been shown not to partition to water or air if released into the environment due to their low water solubility and low vapor pressure.

 

Toxicological Similarity. Review of existing published and unpublished test data for Substituted Diphenylamines shows the aquatic and mammalian toxicity among the materials within this category are similar.

 

Mammalian Toxicology - Acute. Data on acute mammalian toxicity were reviewed, and the findings indicate a low concern for acute toxicity for all materials. Data are available for most members of the category indicating that the category has been well tested for acute mammalian effects. Therefore, for the purposes of the HPV Program, no

additional acute mammalian toxicity testing is proposed.

 

Mammalian Toxicology - Mutagenicity. Data from bacterial reverse mutation assays, in vitro and in vivo chromosome aberration studies, as well as additional supporting in vitro and in vivo genetic toxicity studies were reviewed, and the findings indicate a low concern for mutagenicity either for aryl or alkyl substituted materials. Similarly, the data for a mixed aryl/alkyl substituted molecule also indicates a lack of mutagenicity. Data are available for several members of the category or close structural analogs, and these data can be bridged to the other members of the category. Therefore, for the purposes of the HPV Program, the category has been adequately tested for mutagenicity, and no additional mutagenicity testing is proposed.

 

Mammalian Toxicology – Repeated Dose Toxicity. Data from repeated-dose toxicity studies were reviewed. Sufficient data are not available to adequately represent the Substituted Diphenylamines for the purposes of the HPV Program, and additional testing is proposed on the smallest aryl- and akyl-substituted materials.

 

Mammalian Toxicology - Reproductive and Developmental Toxicity. Data from reproductive and developmental toxicity studies were reviewed. Sufficient data are not available to adequately represent the Substituted Diphenylamines for the purposes of the HPV Program, and additional testing is proposed, and additional testing is proposed. It is proposed to test the smallest aryl- and akyl-substituted materials.

 

Conclusion. Based upon the data reviewed in “Substituted Diphenylamines” category of chemicals, the physicochemical and toxicological properties of the Substituted Diphenylamine category members are similar and follow a regular pattern as a result of that structural similarity. Therefore, the definition of a chemical category has been met, and read across is considered appropriate for the category of chemical.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
600 mg/kg bw/day
Study duration:
subacute
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: inhalation
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Two results are available; one for the substance itself (K2) study and one for read across purposes from equivalent substances. Both results indicate that this category of substances are not harmful by prolonged oral exposure. For the purposes of hazard assessment, the lower of the available NOAEL's is utilised as a worst case assessment. Information is as follows:

Feeding study on the substance itself:

Daily dietary administration for 64 weeks significantly retarded growth in females at 2500 ppm and higher. No effect on growth occurred in males at 2500 ppm was noted. Liver enlargement was noted at all concentrations in both sexes. Diffuse hepatic degeneration was observed in all test animals. However, the severity of the liver changes were not treatment-related. The degenerative changes in the liver were described as diffuse cloudy swellings and fatty metamorphosis of the cytoplasm of the hepatocyte. No compound-related hematopoietic changes were observed in any of the test groups.

Oral gavage study on read across substance:

The oral administration of CAS No 68442-68-2 to rats by gavage, at dose levels of 600, 250 and 50 mg/kg/day, resulted in treatment-related effects at all dose levels. These effects however, were considered entirely adaptive in nature, therefore the ‘No Observed Adverse Effect Level’ (NOAEL) was considered to be 600 mg/kg/day. Read across to supporting substance, CAS No. 68442 -68 -2, by structural analogue. A supporting paper to justify the read across is attached below.

Both substances have been supported under Environmental Protection Agency’s (EPA’s) High Production Volume (HPV) Challenge Program. The American Chemical Councils RAPA Panel, has derived a “Substituted Diphenylamines” category of chemicals for this substance, please refer to EPA reference 201-14700A located at

 

http://www.epa.gov/hpv/pubs/summaries/subdipha/c13378rt.pdf

 

Relying on several factors specified in EPA’s guidance document on “Development of Chemical Categories in the HPV Challenge Program,” in which use of chemical categories is encouraged, the chemicals constitute a chemical category on the following basis:

 

Structural Similarity. A key factor supporting the classification of these chemicals as a category is their structural similarity (see Figure 1). All share a common starting material; Diphenylamine (Benzenamine, N-phenyl-, CAS# 122-39-4), a common synthetic pathway, and all compounds in this category are diamines with various substitutions.

 

Similarity of Physicochemical Properties. The similarity of the physicochemical properties of these materials parallels their structural similarity. All are off-white to light brown solids or viscous liquids intended for use as antioxidants in finished rubber articles or as antidegradant additives that extend the useful life of heavy-duty industrial functional fluids used in high-speed, high-temperature and/or high-load applications. As a class, these amine-based antidegradant compounds are less migratory (more polymer-bound) and less staining than the Substituted p-Phenylenediamine antidegradants. The use of these materials requires that they be stable under high temperatures. Their low volatility is due to their low vapor pressure, highly viscous or solid form. The existing information for these materials indicates that they have low water solubility and high flash points.

 

Fate and Transport Characteristics. Members of this category have been shown to be not readily biodegradable, so additional testing is not needed. The lack of water solubility of the members of this category makes hydrolysis testing unnecessary. These materials have been shown not to partition to water or air if released into the environment due to their low water solubility and low vapor pressure.

 

Toxicological Similarity. Review of existing published and unpublished test data for Substituted Diphenylamines shows the aquatic and mammalian toxicity among the materials within this category are similar.

 

Mammalian Toxicology – Repeated Dose Toxicity. Data from repeated-dose toxicity studies were reviewed. Sufficient data are not available to adequately represent the Substituted Diphenylamines for the purposes of the HPV Program, and additional prolonged testing was proposed on the smallest aryl- and akyl-substituted materials. However the data available for the substance predicts that there are no effects associated with the substance for the purposes of REACH.

 

Conclusion. Based upon the data reviewed in “Substituted Diphenylamines” category of chemicals, the physicochemical and toxicological properties of the Substituted Diphenylamine category members are similar and follow a regular pattern as a result of that structural similarity. Therefore, the definition of a chemical category has been met, and read across is considered appropriate for the category of chemical.

 

The substance, CAS68442-68-2, is analogous to the substance to be registered, but does not contain the additional alkyl side chain groups noted in CAS 68921-45-9. Such alkyl groups are not known to effect repeated dose toxicity to any great degree; indeed their presence makes a substance less bioavailable on the basis of molecular size. 

 

CAS68442-68-2 is not classified for repeated dose or reproduction effects, and CAS 68921-45-9 is a larger molecule and is considered to be less bioavailable thanCAS68442-68-2 on the basis of the chemical structure. As a result of this, it is considered that CAS68442-68-2 would be more bioavailable, and as this substance is not classified for repeated dose effects, read across is considered appropriate on a “worst case” basis.

 

Such read across is also confirmed by the 64 week study on CAS68921-45-9 itself. Whilst this data is not of high quality, it does demonstrate the lack of effects in a chronic study for the repeated dose endpoint.

 


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Two results are available; one for the substance itself (K2) study and one for read across purposes from equivalent substances. Both results indicate that this category of substances are not harmful by prolonged oral exposure. For the purposes of hazard assessment, the lower of the available values is utilised as a worst case assessment.

Justification for classification or non-classification

The above studies have all been ranked reliability 1 or 2 according to the Klimisch et al system. This ranking was deemed appropriate because the feeding study was not conducted to GLP but sufficient dose ranges and numbers are detailed; hence it is appropriate for use based on reliability and animal welfare grounds. Read across to the Klimisch 1 OECD 422 study on the analogue is applicable, and this study was conducted recently to a recognised method and to GLP.

The above results triggered no classification under the Dangerous Substance Directive (67/548/EEC) and the CLP Regulation (EC No 1272/2008). No classification for repeated dose effects is therefore required.