Registration Dossier

Administrative data

Description of key information

The most relevant exposure route is the inhalation route. No adverse effects have been reported in literature for both the oral and dermal routes.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not reported
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Qualifier:
no guideline followed
Principles of method if other than guideline:
The study was designed to test species differences and the influence of surface area. Particle retention kinetics, inflammation, and histopathology were examined in female rats, mice, and hamsters exposed for 13 weeks to high surface area Cb (HSCb) at doses of 0, 1, 7, and 50 mg/m3. Rats were also exposed to 50 mg/m3 low surface area Cb (LSCb). Groups of animals were sacrificed immediately after 13 weeks of exposure, and after 3 and 11 months of recovery for bronchoalveolar lavage analysis, as well as for measurements of lung burdens and lung histopathology
GLP compliance:
not specified
Limit test:
no
Species:
other: rat, mouse, hamster
Strain:
other: F344, B6C3F1, F1B Syrian golden hamster
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: rats from Harlan (Indianapolis, IN), B6C3F1 mice from Charles River (Wilmington, MA), F1B Syrian golden hamsters from BioBreeders (Watertown, MA)
- Age at study initiation: 5 weeks
- Weight at study initiation: not reported
- Housing: AAALAC-accredited barrier facility with 12-h light/dark cycle
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least two weeks
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
whole body
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: The HSCb (Printex 90, surface area 300 m2/g) aggregate aerosols had aerodynamic diameters ranging from 1.2 - 2.4 µm (geometric standard deviations [GSD]: 2.0 - 3.1); the LSCb (Sterling V, surface area 37 m2/g) aggregate aerosols had aerodynamic diameters of 0.6 - 0.9 µm (GSD: 3.0 - 3.7)
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: All exposures took place in compartmentalized, horizontal flow whole-body inhalation chambers (ca.. 300 L).
- Method of holding animals in test chamber: not reported. Each chamber can hold up to 32 rats or hamsters or up to 64 mice.
- Source and rate of air: not reported
- Method of conditioning air: not reported
- System of generating particulates/aerosols: the particle-containing exposure atmospheres were generated using a Wright dust feeder (low and mid dose carbon black) or a Venturi jet generator (high dose cabon black). Particles were deionized by passing the aerosols through an 83Kr source.
- Temperature, humidity, pressure in air chamber: not reported.
- Air flow rate: total flow through the chambers was ca 100 L/min.
- Air change rate: not reported.
- Method of particle size determination: mass concentration and particle size were periodically measured using gravimetric and impactor sampling, respectively.
- Treatment of exhaust air: not reported.

TEST ATMOSPHERE
- Brief description of analytical method used: Aerosol concentration was continuously monitored by a RAS-2 for the mid and high-dose or by a RAM-1 for the low dose (Monitoring Instruments for the Environment [MIE], Inc., Bedford, MA).

TEST CONCENTRATION ADAPTATION
5 weeks into the exposures of mice and hamsters, it was determined that the retained burden of HSCb was lower than was found for rats, which were exposed first. The concentrations for the mouse and hamster exposures were accordingly increased from 7 to 15 mg/m3 for the mid-dose and from 50 to 75 mg/m3 for the high-dose to produce equivalent predicted normalized lung burdens. For hamsters, the low dose also had to be increased from 1 to 1.1 mg/m3.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Aerosol concentrations were continuously monitored by a RAS-2 for the mid and high-dose or by a RAM-1 for the low dose (Monitoring Instruments for the Environment [MIE], Inc., Bedford, MA).
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours per day; 5 days per week
Remarks:
Doses / Concentrations:
0, 1, 7 and 50 mg/m3 (HSCb), 50 mg/m3 (LSCb)
Basis:
nominal conc.
No. of animals per sex per dose:
Groups of five to six females were exposed to filtered air, or the three dose levels of HSCb or 50 mg/m3 LSCb. For the histopathological and particle dosimetry analyses, groups of six animals were used; for all other endpoints, groups of five animals were used.
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: the doses were chosen to span a no observed adverse effects level (NOAEL) to particle overload
- Rationale for animal assignment (if not random): females were chosen because they were previously shown to be more sensitive to the induction of lung tumors by poorly soluble, low toxicity particles than males
- Section schedule rationale (if not random): additional sentinels were euthanized throughout the study to monitor pathogen status
Positive control:
no
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
DETAILED CLINICAL OBSERVATIONS: Yes
BODY WEIGHT: Yes
- Time schedule for examinations: body weights were obtained every 2 weeks.
HAEMATOLOGY: No
CLINICAL CHEMISTRY: Yes, in lavage fluid
Measurements were performed immediately after exposure and 3 and 11 months post-exposure; carbon black retention was also evaluated after 5 weeks of exposure.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: lungs
Other examinations:
- Determination of carbon black lung burden
- Measurement of cellular and biochemical parameters in lavage fluid
- Pulmonary morphometry
- Cell proliferation assessment (BrdU method)
Statistics:
Tukey t-tests uing SigmaStat (SPSS Science, Chicago, IL). The two factors for the ANOVAs were exposure dose and time. Data were appropriately transformed (e.g., base 10 logarithm, natural logarithm) if an analysis of residuals suggested deviations from the asumptions of normality and equal variance. Comparisons were considered statistically significant when p<= 0.05
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
not specified
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY
No rats died during exposure to carbon black. Nine control rats died in the post-exposure phase due to a blocked water line. One each in the 1 and 50 mg/m3 HSCb groups also died in the post-exposure phase. Thirteen mice died during exposure, seven of which were in the control group; two were in the low-dose gorup, three in the mid-dose group, and one in the high-dose group. In the post-exposure phase, six controls, one low-dose, two mid-dose, and six high-dose mice died. One hamster (high-dose group) died during exposuer. Two hamsters each from the control, low-dose, and high-dose groups and four from the mid-dose group died in the post-exposure phase of the study. These deaths were most likely due to a change in housing conditions from wire-bottom cages (during exposure) to plastic cages (post-exposure) and reflects the heightened sensitivity of hamsters to environmental changes. Screening of sentinels (all three species) did not reveal parasitic, bacterial, or viral pathogens that would explain the unscheduled deaths. The authors conclude that the unscheduled deaths were not related to exposure, i.e., not associated with carbon black dose or time after exposure. Out of the total number of animals exposed, ca. 4% of the animals per species died prematurely.

BODY WEIGHT AND WEIGHT GAIN
Significant decreases in body weight during exposure occurred only in hamsters exposed to 50 mg/m3 HSCb. Rats of the high-dose groups lost weight during the exposure (not statistically significant). Body weights returned to control values during the recovery period for both species.

ORGAN WEIGHTS
Lung weights were increased in high-dose exposed animals, but this persisted only in rats and mice up to the end of he study period.
For rats, significant elevations in lung weight (up to more than twice the control lung weight) were found for the high-dose HSCb at the end of exposure and at all post-exposure time points. The lung weights of rats exposed to LSCb were between those for the high- and mid-dose HSCb in magnitude and were significantl y elevated 1 day and 3 months after expsoure.
In mice, lung weights were found to be elevated 1 day and 3 months post-exposure in the high- and mid-dose groups; at the end of the recovery period, i.e., 11 onths post-exposure, significant elevations were found only for the high-dose group, and these changes persisted up to 3 months post-exposure; after 11 months of recovery, no significant difference were found.
Thus, the changes in lung weight were resolved most rapidly in hamsters.

HISTOPATHOLOGY: NON-NEOPLASTIC
Lung inflammation and histopathology were more severe and prolonged in rats than in mice and hamsters; both were similar in rats exposed to mid-dose HSCb and LSCb.
No adverse histological effects of exposure were found at any post-exposure time point in the lungs of rats, mice or hamsters exposed to low-dose HSCb. The lungs of these rats, mice or hamsters were similar to those of the control rats, with the exception that some alveolar macrophages containing small amounts of carbon black were widely scattered throughout the alveolar air spaces of the pulmonary parenchyma. However, the number and morphological character of these particle-containing macrophages were not different from those in the controls.

CELLULAR AND BIOCHEMICAL PARAMETERS IN LAVAGE FLUID
Total bronchoalveolar lavage (BAL) cell numbers were relevated at 1 day post-exposure and all through the recovery period in the high-dose groups for all three species in comparison to all other exposure groups. Whereas total cell number gradually decreased over time in mice andhamsters, it stayed elevated in the rats exposued to high-dose HSCb and LSCb. The presence of polymorphonuclear leukocytes (PMN) in lavage fluid was used as a sensitive indicator of lung inflammation. PMN were elevated in the mid- and high-dose HSCb and LSCb groups of rats at the end of exposure and throughout the recovery period. In rats, the PMN responses to high-dose HSCb and LSCb were similar. The PMN remained elevated in rats from the high-dose HSCb and LSCb groups through the end of the study. At the end of exposure in mice and hamsters, the mid- and high-dose groups were also elevated compared to the ocntrols and low-dose groups. The magnitudes of response were similar for the two species. During the recovery period, the percentage of PMN from the mid- and high-dose mice remained different from each other and the rest of the exposure groups. In hamsters, the PMN response decreased between 3 and 11 months post-exposure such that only the high-dose group retained a signifcant response at the termination of the study. The magnitude and duration of the PMN response was the highest in rats.. There were no significant elevations in cellular or biochemical parameters for any of the animals exposed to low-dose HSCb.
OTHER FINDINGS
Equivalent or similar mass burdens were achieved in rats exposed to high-dose HSCb and LSCb. Surface area burdens were equivalent for 7 mg/m3 HSCBc and 50 mg/m3 LSCb.
Prolonged retention was found in rats exposed to mid- and high-dose HSCb and to LSCb, but LSCb was cleared faster than HSCb. Retention was also prolonged in mice exposed to mid- and high-dose HSCb, and in hamsters exposed to high-dose HSCb.










Key result
Dose descriptor:
NOEC
Remarks:
(rat, mouse)
Effect level:
1 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: pulmonary inflammation
Key result
Dose descriptor:
LOEC
Remarks:
(rat, mouse)
Effect level:
7 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: histopathology; lung inflammation; prolonged carbon black retention in lungs
Key result
Dose descriptor:
NOEC
Remarks:
(hamster)
Effect level:
7 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: pulmonary inflammation
Key result
Dose descriptor:
LOAEC
Remarks:
(hamster)
Effect level:
50 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: histopathology; prolonged carbon black retention in lung; lung inflammation
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
7 mg/m³ air
System:
respiratory system: lower respiratory tract
Organ:
lungs
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Conclusions:
The results show that hamsters have the most efficient clearance mechanisms and least severe responses of the threee species tested. The results from rats also showe that particle surfae area is an important determinant of target tissue dose and, therefore, effects. From these results, a subchronic NOAEL of 1 mg/m3 respirable HSCb (Printex 90) can be assigned to female rats, mice, and hamsters.
Executive summary:

Particle retention kinetics, inflammation, and histopathology were examined in female rats, mice, and hamsters exposed for 13 weeks to high surface area Cb (HSCb) at doses chosen to span a no observable adverse effects level (NOAEL) to particle overload (0, 1, 7, 50 mg/m3, nominal concentrations). Rats were also exposed to low surface area Cb (50 mg/m3, nominal; LSCb). Retention and effects measurements were performed immediately after exposure and 3 and 11 months post-exposure; retention was also evaluated after 5 weeks of exposure. Significant decreases in body weight during exposure occurred only in hamsters exposed to high-dose HSCb. Lung weights were increased in high-dose Cb-exposed animals, but this persisted only in rats and mice up to the end of the study period. Equivalent or similar mass burdens were achieved in rats exposed to high-dose HSCb and LSCb, whereas surface area burdens were equivalent for mid-dose HSCb and LSCb. Prolonged retention was found in rats exposed to mid- and high-dose HSCb and to LSCb, but LSCb was cleared faster than HSCb. Retention was also prolonged in mice exposed to mid- and high-dose HSCb, and in hamsters exposed to high-dose HSCb. Lung inflammation and histopathology were more severe and prolonged in rats than in mice and hamsters, and both were similar in rats exposed to mid-dose HSCb and LSCb. The results show that hamsters have the most efficient clearance mechanisms and least severe responses of the three species. The results from rats also show that particle surface area is an important determinant of target tissue dose and, therefore, effects. From these results, a subchronic NOAEL of 1 mg/m3 respirable HSCb (Printex 90) can be assigned to female rats, mice, and hamsters.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not reported
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Qualifier:
no guideline followed
Principles of method if other than guideline:
The study was designed to test species differences and the influence of surface area. Particle retention kinetics, inflammation, and histopathology were examined in female rats, mice, and hamsters exposed for 13 weeks to high surface area Cb (HSCb) at doses of 0, 1, 7, and 50 mg/m3. Rats were also exposed to 50 mg/m3 low surface area Cb (LSCb). Groups of animals were sacrificed immediately after 13 weeks of exposure, and after 3 and 11 months of recovery for bronchoalveolar lavage analysis, as well as for measurements of lung burdens and lung histopathology
GLP compliance:
not specified
Limit test:
no
Species:
other: rat, mouse, hamster
Strain:
other: F344, B6C3F1, F1B Syrian golden hamster
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: rats from Harlan (Indianapolis, IN), B6C3F1 mice from Charles River (Wilmington, MA), F1B Syrian golden hamsters from BioBreeders (Watertown, MA)
- Age at study initiation: 5 weeks
- Weight at study initiation: not reported
- Housing: AAALAC-accredited barrier facility with 12-h light/dark cycle
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least two weeks
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
whole body
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: The HSCb (Printex 90, surface area 300 m2/g) aggregate aerosols had aerodynamic diameters ranging from 1.2 - 2.4 µm (geometric standard deviations [GSD]: 2.0 - 3.1); the LSCb (Sterling V, surface area 37 m2/g) aggregate aerosols had aerodynamic diameters of 0.6 - 0.9 µm (GSD: 3.0 - 3.7)
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: All exposures took place in compartmentalized, horizontal flow whole-body inhalation chambers (ca.. 300 L).
- Method of holding animals in test chamber: not reported. Each chamber can hold up to 32 rats or hamsters or up to 64 mice.
- Source and rate of air: not reported
- Method of conditioning air: not reported
- System of generating particulates/aerosols: the particle-containing exposure atmospheres were generated using a Wright dust feeder (low and mid dose carbon black) or a Venturi jet generator (high dose cabon black). Particles were deionized by passing the aerosols through an 83Kr source.
- Temperature, humidity, pressure in air chamber: not reported.
- Air flow rate: total flow through the chambers was ca 100 L/min.
- Air change rate: not reported.
- Method of particle size determination: mass concentration and particle size were periodically measured using gravimetric and impactor sampling, respectively.
- Treatment of exhaust air: not reported.

TEST ATMOSPHERE
- Brief description of analytical method used: Aerosol concentration was continuously monitored by a RAS-2 for the mid and high-dose or by a RAM-1 for the low dose (Monitoring Instruments for the Environment [MIE], Inc., Bedford, MA).

TEST CONCENTRATION ADAPTATION
5 weeks into the exposures of mice and hamsters, it was determined that the retained burden of HSCb was lower than was found for rats, which were exposed first. The concentrations for the mouse and hamster exposures were accordingly increased from 7 to 15 mg/m3 for the mid-dose and from 50 to 75 mg/m3 for the high-dose to produce equivalent predicted normalized lung burdens. For hamsters, the low dose also had to be increased from 1 to 1.1 mg/m3.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Aerosol concentrations were continuously monitored by a RAS-2 for the mid and high-dose or by a RAM-1 for the low dose (Monitoring Instruments for the Environment [MIE], Inc., Bedford, MA).
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours per day; 5 days per week
Remarks:
Doses / Concentrations:
0, 1, 7 and 50 mg/m3 (HSCb), 50 mg/m3 (LSCb)
Basis:
nominal conc.
No. of animals per sex per dose:
Groups of five to six females were exposed to filtered air, or the three dose levels of HSCb or 50 mg/m3 LSCb. For the histopathological and particle dosimetry analyses, groups of six animals were used; for all other endpoints, groups of five animals were used.
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: the doses were chosen to span a no observed adverse effects level (NOAEL) to particle overload
- Rationale for animal assignment (if not random): females were chosen because they were previously shown to be more sensitive to the induction of lung tumors by poorly soluble, low toxicity particles than males
- Section schedule rationale (if not random): additional sentinels were euthanized throughout the study to monitor pathogen status
Positive control:
no
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
DETAILED CLINICAL OBSERVATIONS: Yes
BODY WEIGHT: Yes
- Time schedule for examinations: body weights were obtained every 2 weeks.
HAEMATOLOGY: No
CLINICAL CHEMISTRY: Yes, in lavage fluid
Measurements were performed immediately after exposure and 3 and 11 months post-exposure; carbon black retention was also evaluated after 5 weeks of exposure.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: lungs
Other examinations:
- Determination of carbon black lung burden
- Measurement of cellular and biochemical parameters in lavage fluid
- Pulmonary morphometry
- Cell proliferation assessment (BrdU method)
Statistics:
Tukey t-tests uing SigmaStat (SPSS Science, Chicago, IL). The two factors for the ANOVAs were exposure dose and time. Data were appropriately transformed (e.g., base 10 logarithm, natural logarithm) if an analysis of residuals suggested deviations from the asumptions of normality and equal variance. Comparisons were considered statistically significant when p<= 0.05
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
not specified
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY
No rats died during exposure to carbon black. Nine control rats died in the post-exposure phase due to a blocked water line. One each in the 1 and 50 mg/m3 HSCb groups also died in the post-exposure phase. Thirteen mice died during exposure, seven of which were in the control group; two were in the low-dose gorup, three in the mid-dose group, and one in the high-dose group. In the post-exposure phase, six controls, one low-dose, two mid-dose, and six high-dose mice died. One hamster (high-dose group) died during exposuer. Two hamsters each from the control, low-dose, and high-dose groups and four from the mid-dose group died in the post-exposure phase of the study. These deaths were most likely due to a change in housing conditions from wire-bottom cages (during exposure) to plastic cages (post-exposure) and reflects the heightened sensitivity of hamsters to environmental changes. Screening of sentinels (all three species) did not reveal parasitic, bacterial, or viral pathogens that would explain the unscheduled deaths. The authors conclude that the unscheduled deaths were not related to exposure, i.e., not associated with carbon black dose or time after exposure. Out of the total number of animals exposed, ca. 4% of the animals per species died prematurely.

BODY WEIGHT AND WEIGHT GAIN
Significant decreases in body weight during exposure occurred only in hamsters exposed to 50 mg/m3 HSCb. Rats of the high-dose groups lost weight during the exposure (not statistically significant). Body weights returned to control values during the recovery period for both species.

ORGAN WEIGHTS
Lung weights were increased in high-dose exposed animals, but this persisted only in rats and mice up to the end of he study period.
For rats, significant elevations in lung weight (up to more than twice the control lung weight) were found for the high-dose HSCb at the end of exposure and at all post-exposure time points. The lung weights of rats exposed to LSCb were between those for the high- and mid-dose HSCb in magnitude and were significantl y elevated 1 day and 3 months after expsoure.
In mice, lung weights were found to be elevated 1 day and 3 months post-exposure in the high- and mid-dose groups; at the end of the recovery period, i.e., 11 onths post-exposure, significant elevations were found only for the high-dose group, and these changes persisted up to 3 months post-exposure; after 11 months of recovery, no significant difference were found.
Thus, the changes in lung weight were resolved most rapidly in hamsters.

HISTOPATHOLOGY: NON-NEOPLASTIC
Lung inflammation and histopathology were more severe and prolonged in rats than in mice and hamsters; both were similar in rats exposed to mid-dose HSCb and LSCb.
No adverse histological effects of exposure were found at any post-exposure time point in the lungs of rats, mice or hamsters exposed to low-dose HSCb. The lungs of these rats, mice or hamsters were similar to those of the control rats, with the exception that some alveolar macrophages containing small amounts of carbon black were widely scattered throughout the alveolar air spaces of the pulmonary parenchyma. However, the number and morphological character of these particle-containing macrophages were not different from those in the controls.

CELLULAR AND BIOCHEMICAL PARAMETERS IN LAVAGE FLUID
Total bronchoalveolar lavage (BAL) cell numbers were relevated at 1 day post-exposure and all through the recovery period in the high-dose groups for all three species in comparison to all other exposure groups. Whereas total cell number gradually decreased over time in mice andhamsters, it stayed elevated in the rats exposued to high-dose HSCb and LSCb. The presence of polymorphonuclear leukocytes (PMN) in lavage fluid was used as a sensitive indicator of lung inflammation. PMN were elevated in the mid- and high-dose HSCb and LSCb groups of rats at the end of exposure and throughout the recovery period. In rats, the PMN responses to high-dose HSCb and LSCb were similar. The PMN remained elevated in rats from the high-dose HSCb and LSCb groups through the end of the study. At the end of exposure in mice and hamsters, the mid- and high-dose groups were also elevated compared to the ocntrols and low-dose groups. The magnitudes of response were similar for the two species. During the recovery period, the percentage of PMN from the mid- and high-dose mice remained different from each other and the rest of the exposure groups. In hamsters, the PMN response decreased between 3 and 11 months post-exposure such that only the high-dose group retained a signifcant response at the termination of the study. The magnitude and duration of the PMN response was the highest in rats.. There were no significant elevations in cellular or biochemical parameters for any of the animals exposed to low-dose HSCb.
OTHER FINDINGS
Equivalent or similar mass burdens were achieved in rats exposed to high-dose HSCb and LSCb. Surface area burdens were equivalent for 7 mg/m3 HSCBc and 50 mg/m3 LSCb.
Prolonged retention was found in rats exposed to mid- and high-dose HSCb and to LSCb, but LSCb was cleared faster than HSCb. Retention was also prolonged in mice exposed to mid- and high-dose HSCb, and in hamsters exposed to high-dose HSCb.










Key result
Dose descriptor:
NOEC
Remarks:
(rat, mouse)
Effect level:
1 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: pulmonary inflammation
Key result
Dose descriptor:
LOEC
Remarks:
(rat, mouse)
Effect level:
7 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: histopathology; lung inflammation; prolonged carbon black retention in lungs
Key result
Dose descriptor:
NOEC
Remarks:
(hamster)
Effect level:
7 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: pulmonary inflammation
Key result
Dose descriptor:
LOAEC
Remarks:
(hamster)
Effect level:
50 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: histopathology; prolonged carbon black retention in lung; lung inflammation
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
7 mg/m³ air
System:
respiratory system: lower respiratory tract
Organ:
lungs
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Conclusions:
The results show that hamsters have the most efficient clearance mechanisms and least severe responses of the threee species tested. The results from rats also showe that particle surfae area is an important determinant of target tissue dose and, therefore, effects. From these results, a subchronic NOAEL of 1 mg/m3 respirable HSCb (Printex 90) can be assigned to female rats, mice, and hamsters.
Executive summary:

Particle retention kinetics, inflammation, and histopathology were examined in female rats, mice, and hamsters exposed for 13 weeks to high surface area Cb (HSCb) at doses chosen to span a no observable adverse effects level (NOAEL) to particle overload (0, 1, 7, 50 mg/m3, nominal concentrations). Rats were also exposed to low surface area Cb (50 mg/m3, nominal; LSCb). Retention and effects measurements were performed immediately after exposure and 3 and 11 months post-exposure; retention was also evaluated after 5 weeks of exposure. Significant decreases in body weight during exposure occurred only in hamsters exposed to high-dose HSCb. Lung weights were increased in high-dose Cb-exposed animals, but this persisted only in rats and mice up to the end of the study period. Equivalent or similar mass burdens were achieved in rats exposed to high-dose HSCb and LSCb, whereas surface area burdens were equivalent for mid-dose HSCb and LSCb. Prolonged retention was found in rats exposed to mid- and high-dose HSCb and to LSCb, but LSCb was cleared faster than HSCb. Retention was also prolonged in mice exposed to mid- and high-dose HSCb, and in hamsters exposed to high-dose HSCb. Lung inflammation and histopathology were more severe and prolonged in rats than in mice and hamsters, and both were similar in rats exposed to mid-dose HSCb and LSCb. The results show that hamsters have the most efficient clearance mechanisms and least severe responses of the three species. The results from rats also show that particle surface area is an important determinant of target tissue dose and, therefore, effects. From these results, a subchronic NOAEL of 1 mg/m3 respirable HSCb (Printex 90) can be assigned to female rats, mice, and hamsters.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
1 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
1

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Additional information

Extensive, but old studies in mice with carbon black application by the oral and dermal routes have not indicated any adverse effects, even at excessive dose levels (10% in feed, 20% suspensions for dermal application). A study investigating effects of ingested carbon black in rats and mice has also not found any effect on tissues or organs after 2 years of exposure to a dose level equivalent to 52 or 137 mg/kg bw/day in rats and mice, respectively and no adverse effects were reported at 1000 mg/kg bw/day from a 13-week oral toxicity study in rats conducted on carbon black following OECD guideline 408. A relevant systemic exposure after oral and dermal administration is not expected. In rats, the most sensitive species, no pathological or biochemical changes were found in the lungs at 1.0 mg/m3 after repeated inhalation of a carbon black for 13 weeks.

Justification for classification or non-classification

Based on available toxicological data and chemical-physical properties (insolubility, low absorption potential), specific target organ toxicity is not expected after repeated dermal or oral exposure. Several repeated dose inhalation toxicity studies (90-day studies) conducted in rats indicate a No Observed Adverse Effect Concentration (NOAEC) of 1.0 mg/m3(respirable). Target organ effects at higher doses are lung inflammation, hyperplasia, and fibrosis (Carter et al., 2006; Elder et al., 2005; Driscoll et al., 1996). However, this response in rats under conditions of lung overload is principally a species-specific response that is not relevant to humans. As stated in ECETOC (2013)“the rat represents a particularly sensitive model concerning the development of pulmonary non-neoplastic lesions (inflammation and fibro-proliferation) and, moreover, a unique model with regard to lung neoplastic responses under conditions of lung overload.”Results of epidemiological studies of carbon black production workers suggest that cumulative exposure to carbon black may result in small, non-clinical decrements in lung function. A U.S. respiratory morbidity study suggested a 27 mL decline in FEV1 from a 1 mg/m3 8-hour TWA daily (inhalable fraction) exposure over a 40-year period {Harber et al., 2003). An earlier European investigation suggested that exposure to 1 mg/m3 (inhalable fraction) of carbon black over a 40-year working lifetime would result in a 48 mL decline in FEV1 (Gardiner et al., 2001). However, the estimates from both studies were only of borderline statistical significance. Normal age-related decline among healthy male non-smokers is approximately 30 mL/year, resulting in approximately 1200 mLdecline over a 40-year working lifetime (Burrows et al., 1986;Sherrillet al., 1992). In the U.S. study, 9% of the highest non-smokers exposure group (in contrast to 5% of the unexposed group) reported symptoms consistent with chronic bronchitis. In the European study, methodological limitations in the administration of the questionnaire limit the conclusions that can be drawn about reported symptoms. This study, however, indicated a link between carbon black and small opacities on chest films, with negligible effects on lung function.The paucityof high profusion radiographic findings,the absence of progression to extensive radiographic abnormality,and the absence of restrictive physiologic abnormalities show that carbon black does not lead to a fibrotic pneumoconiosis.

Applying the guidelines of self-classification under GHS, carbon black is therefore not classifiable under STOT-RE for effects on the lung. Classification is not warranted on the basis of the unique response of rats resulting from the “lung overload” following exposure to poorly soluble particles such as carbon black. The pattern of pulmonary effects in the rat, such as inflammation and fibrotic responses, are not observed in other rodent species, non-human primates, or humans under similar exposure conditions. Lung overload does not appear to be relevant for human health. Overall, the epidemiological evidence from well-conducted investigations has shown no causative link between carbon black exposure and the risk of non-malignant respiratory disease in humans.