Registration Dossier

Administrative data

Description of key information

No experimental data on repeated dose toxicity is available for AOH itself. Results from a related total petroleum hydrocarbon aromatic mixture (>EC9 to EC16 TPH fraction) and from a structure-related tar oil are used to characterise the repeated dose toxicity of AOH. Oral repeated dose toxicity is based on a TDI value derived for the aromatic TPH fraction while for inhalation repeated dose toxicity, a NOAEC of the structure related tar oil creosote is adopted.
Dominant human health effect of AOH is not repeated dose toxicity but carcinogenicity due to its content of benzo[a]pyrene. Therefore, risk assessment will not be based on repeated dose toxicity but on the carcinogenic effect of benzo[a]pyrene.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: oral
Remarks:
other: subchronic and chronic
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Peer-reviewed data, evaluation of current database for deriving permissible exposure levels
Qualifier:
no guideline required
Principles of method if other than guideline:
RIVM project 711701 "Risk in relation to Soil Quality", RIVM Bilthoven/NL to derive Maximum Permissible Risk levels (MPR) for the oral route and in isolated cases for inhalation. The MPR(human) is defined as the amount of a substance (usually a chemical substance) that any human individual can be exposed to daily during lifetime without significant health risk..... For genotoxic carcinogens the MPR has been defined as the excess lifetime cancer risk of 1 in 10000 (1:10^4) (RIVM 2001, Chap. 2, p. 9). MPRs are expressed as Tolerable Daily Intake (TDI) or Tolerable Concentration in Air (TCA) [see Results]. For background information on MPR see Janssen and Speijers 1997.
GLP compliance:
not specified
Species:
other: rat and mouse
Sex:
male/female
Details on test animals and environmental conditions:
no data
Route of administration:
other: oral and inhalation
Details on oral exposure:
not applicable
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
not applicable
Frequency of treatment:
not applicable
No. of animals per sex per dose:
not applicable
Details on study design:
not applicable
Positive control:
not applicable
Observations and examinations performed and frequency:
not applicable
Sacrifice and pathology:
not applicable
Other examinations:
Evaluation of databases (see "Any other information..")
Statistics:
--
Details on results:
not applicable
Dose descriptor:
NOAEL
Remarks:
for aromatic >EC9 to EC16 fraction from TPH
Effect level:
120 other: mg/kg bw/day, mean for the TPH fraction
Based on:
other: evaluation of toxicity database of petroleum products, in this case aromatic >EC9 to EC16 fraction
Basis for effect level:
other: TDI is derived to be 0.04 mg/kg bw/day (see below). Factor used to calculate individual TDI values from NOAEL values is 3000 (Baars et al. 2001 (RIVM), page 292). Using this factor. a mean NOAEL for the aromatic >EC9 to EC16 fraction can be estimated
Dose descriptor:
other: TDI (Tolerable Daily Intake)
Effect level:
0.04 other: mg/kg bw/d (lifelong for any human)
Based on:
other: evaluation of toxicity database of petroleum products
Sex:
male/female
Basis for effect level:
other: non-neoplastic effects, derived as mean/representative for the aromatic >EC9 to EC 16 fraction of TPH (total petroleum hydrocarbons)
Dose descriptor:
other: TCA (Tolerable Concentration in Air)
Effect level:
0.2 other: mg/m3 (lifelong for any human)
Based on:
other: evaluation of toxicity database of petroleum products
Sex:
male/female
Basis for effect level:
other: non-neoplastic effects; value is derived from the oral TDI above, assuming an absorption rate of 100% in either case: TCA = (TDI x 70 kg/bw) / (20 m³/d)
Critical effects observed:
not specified

--

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
120 mg/kg bw/day
Species:
mouse
Quality of whole database:
NOAEL is calculated from a peer-reviewed TDI derived from an comprehensive evaluation of available experimental data (dataset for TPH - total petroleum hydrocarbons) using the uncertanty factor of 3000 applied in establaishing RfD values from experimental NOAEL (US EPA IRIS) that are used in establishing the group TDI.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-guideline study
Qualifier:
according to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
- Detailed examination for clinical signs of toxicity was performed weekly instead of daily.
Qualifier:
according to
Guideline:
EPA OPP 82-4 (90-Day Inhalation Toxicity)
Qualifier:
according to
Guideline:
EU Method B.29 (Sub-Chronic Inhalation Toxicity:90-Day Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:CD® BR VAF/PLUS®
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, Michigan, USA
- Age at study initiation: Approx. 6 weeks
- Weight at study initiation: Males: 181-211 g; females: 130-149 g
Route of administration:
inhalation
Type of inhalation exposure:
whole body
Vehicle:
clean air
Remarks on MMAD:
MMAD / GSD: MMAD [µm] (± GSD [µm])
Low dose: 3.0 (1.92)
Mid dose: 2.2 (1.99)
High dose: 2.4 (1.91)
Details on inhalation exposure:
--
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Nominal concentration: Calculated from the amount of test compound used during the exposure period (by weighing the reservoir before and after the exposure period) and dividing the total creosote consumed by the total air volume passed through the chamber.
Analytical aerosol concentration: Determined by gravimetric determination of the oil amount adsorbed onto a 25-mm glass-fibre filter pad, divided by the sample volume.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 h/d, 5 days per week
Remarks:
Doses / Concentrations:
Mean nominal concentration [mg/m³]: 22, 128, and 221
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
Mean aerosol concentration [mg/m³]: 5.4, 49, 106
Basis:
analytical conc.
No. of animals per sex per dose:
20/sex/group
An additional group (5/sex) was sacrificed pre-test to define a baseline for clinical chemistry and haematological values.
Control animals:
yes, sham-exposed
Details on study design:
--
Positive control:
none
Observations and examinations performed and frequency:
Mortality: Yes, twice weekly

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: pre-test and weekly

FOOD CONSUMPTION: Yes - weekly

FOOD EFFICIENCY: No data

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes, prior to terminal necropsy and prior to recovery necropsy.

HAEMATOLOGY: Yes
Number of animals: 5/sex at pre-test, 10/sex/group at termination of study and on surviving recovery animals at the end of the recovery period.
Parameters: haematocrit, haemoglobin, erythrocyte count, total and differential leukocyte count, platelet count, reticulocytes, MCV, MCH, MCHC

CLINICAL CHEMISTRY: Yes
Number of animals: 5/sex at pre-test, 10/sex/group at termination of study and on surviving recovery animals at the end of the recovery period.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Organ weights:
Organs: adrenal, brain, ovary, testis with epididymis, heart, kidney, liver, lung, mammary gland, thymus, thyroid/parathyroid, trachea

Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals: external examination; contents of abdominal, thoracic and cranial cavities were examined both in situ and after removal and dissection.

HISTOPATHOLOGY: Yes
All animals: heart, thyroid, nasal tissues, trachea, and lungControl and high dose group, animals that died on study: adrenal, aorta, auditory sebaceous gland, bone with bone marrow (femur), bone marrow smear, brain (fore, mid and hind), eye including optic nerve and contiguous Harderian gland, gastrointestinal tract, gonads, heart, kidney, larynx, lachrymal gland, liver, lung, lymph nodes, mammary gland (females only), nasal tissues, pancreas, pituitary, prostate and seminal vesicle, salivary gland, sciatic nerve, skeletal muscle (thigh), skin, spinal cord, spleen, thymus, thyroid/parathyroid, trachea, tracheal bifurcation, urinary bladder, vagina, uterus and cervix
Other examinations:
Ten animals per dose group and sex were subject to a six-week post-exposure period after which they were sacrificed and examined macroscopically for reversibility of eventual effects.
Statistics:
Analysis of body weights, food consumption, clinical pathology laboratory tests and organ weights were performed as follows:Generally, when the number of animals in any one group was ≤10, non-parametric analysis was conducted using the KRUSKAL-WALLIS one-way analysis of variance, followed by the MANN-WHITNEY U test, where appropriate. In those cases where the number of animals in all groups was greater than ten an the measurements were on at least an interval scale (continuous data), parametric analysis was conducted utilizing BARTLETT’s chi-square test for homogeneity of variance, followed by an analysis of variance and then, where appropriate, by DUNNETT’s t-test.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
CLINICAL SIGNS AND MORTALITY:
One male in the mid-dose group died during the exposure phase (week 4) of this study.

BODY WEIGHT AND WEIGHT GAIN
Body weight decreases were observed in the two highest exposure groups. They were significant in both sexes of the high-dose group and in mid-dose females during exposure week 6. Mean weights for the low-level group were similar to the control group.

FOOD CONSUMPTION:
With the exception of week 1 (all groups significantly less than controls) and week 3 (males in the high-dose group significantly less than controls), food consumption during the 13-week exposures and the six-week post-exposure recovery period was comparable to that of controls.

HAEMATOLOGY:
Several haematological parameters showed significant changes in the two highest exposure levels at the terminal sacrifice including: decreased erythrocytes, haemoglobin, and haematocrit; increased numbers of reticulocytes. These changes were not detectable at the end of the recovery period.

CLINICAL CHEMISTRY:
Serum cholesterol was significantly increased in males of the mid-dose group and in females of the two highest exposure groups. This change was not evident at the end of the recovery period (see table below).

ORGAN WEIGHTS:
Terminal sacrifice: There was a test-article-related and statistically significant increase in the lung/trachea/body weight ratio in males and females of the high-dose group when compared to the respective control values. These increases correlated with the macroscopic observation of grey discolouration of the lungs, and the microscopic observation of pigmented macrophages within the lungs of the animals in the affected groups. In mid- and high-dose males, the adrenal/body weight ratio was increased. No macroscopic or microscopic changes were associated with these changes.
In males, increases in liver weight (mid-dose group) and liver/body weight ratio (mid- and high-dose group) were noted. Relative liver weights were significantly increased at the mid and high dose (about +20 and +25 %, respectively).
In females, there were increases in liver weight (high-dose group), liver/body weight ratio (mid- and high-dose group) and liver/brain weight ratio (mid- and high-dose group) when compared to controls. No macroscopic or microscopic changes were associated with these changes, thus, their toxicbiological significance is uncertain.

Recovery sacrifice: There were no test-article-related changes in organ weight or weight ratios for males in any dose group. In mid-dose females, the mean adrenal weight was significantly decreased compared to controls. No macroscopic or microscopic observations were associated with that finding. Thus, it was deemed not test article related.

GROSS PATHOLOGY:
At the terminal sacrifice, a deposition of the test article consisting of a grey discolouration of the lung was seen at the two highest exposure levels. The discolouration persisted through the recovery period. The control and low-dose group

HISTOPATHOLOGY: NON-NEOPLASTIC:
Microscopic changes were observed in the hearts, lungs, nasal tissues, and thyroid glands of male and female rats at the time of terminal sacrifice. Heart lesions were found in one male of the mid-dose group that died on the study, and in one male and one female of the high-dose group (diffuse myocardial degeneration affecting mainly the right side of the heart). Associated to this change was diffuse arterial medial hypertrophy of small arterioles in the lung, brown pigment within the epithelial cells of the convoluted tubules of the kidney, and in the animal that died on study, alveolar macrophages containing brown pigment consistent with haemosiderin (“heart failure cells”) within the lung and diffuse centrilobular fibrosis within the liver. No heart lesions were found in any dose group at the end of the recovery period.
Note: Cardiac pathology (ie: hemorrhage, lymphocytic infiltration and cardiomyopathy) was noted in all animals of all groups (including controls).

Test-article-related changes in the lung were seen in all animals of the exposed groups (small black pigment granules within alveolar macrophages). Alveolar macrophages containing pigment granules could be detected in all lobes of the lungs indicating a uniform dispersion of the test article throughout the alveolar spaces of the lungs. There were no other changes within the lungs which could be associated with the presence of the granules, such as inflammation, increased number of pulmonary macrophages and/or Type-II pneumocyte hyperplasia. These findings were also seen at recovery sacrifice.

Nasal cavity: Small cystic spaces, containing basophilic mucoid material, within the olfactory epithelium at all levels of the nasal tissues examined and in both sexes, and were considered test article related. Mucoid cysts were seen in mid- and high-dose males, and in low-, mid-, and high-dose females. Similar findings were made at recovery sacrifice.
Other histological changes within the nasal tissues: Squamous metaplasia of respiratory and/or olfactory epithelium and naso­lachrymal duct epithelium, inflammatory cell infiltrates, and glandular dilation within the lamina propria/submucosa of the nasal cavity. These additional changes showed no definitive test article relationship.

Hypertrophy of thyroid follicular cells which resulted in a reduction in the amount of colloid present within the thyroid follicles was seen in both male and female rats of all exposure groups. This anomaly was considered test-article related. No test-article-related effects on the thyroid glands were detected at recovery sacrifice. There was no measurable effect on the mass of the thyroid gland (no changes in absolute and relative weights).

Dose descriptor:
NOAEC
Remarks:
systemic
Effect level:
5.4 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on the death of one male, decreases in body-weight gain (>10 %) as well as on the increase in liver weight (>=20 %), and on the increase in hypertrophy of follicular cells of the thyroid gland
Dose descriptor:
NOAEC
Remarks:
local, nasal
Effect level:
5.4 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
NOAEC
Remarks:
systemic
Effect level:
22 mg/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
NOAEC
Remarks:
local, nasal
Effect level:
22 mg/L air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
LOAEC
Remarks:
systemic
Effect level:
49 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
LOAEC
Remarks:
local, nasal
Effect level:
49 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
LOAEC
Remarks:
systemic
Effect level:
128 mg/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
LOAEC
Remarks:
local, nasal
Effect level:
128 mg/L air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Critical effects observed:
not specified

Results of clinical chemistry and haematology

Parameter changed

Unit

Controls
0 mg/m³

Low dose
5.4 mg/m³

Medium dose
49 mg/m³

High dose
106 mg/m³

Time point

Terminal

Recovery

Terminal

Recovery

Terminal

Recovery

Terminal

Recovery

Males

Haemoglobin

g/dl

15.5

15.2

15.4

15.3

14.3**

15.0

14.6

15.1

Haematocrit

%

44.7

40.0

43.5

40.1

39.7*

39.7

40.3

39.8

Reticulocytes

/ 100 RBC

2.8

2.1

2.8

1.8

4.0

1.6

5.2

1.6

Phosphorus

mg/dl

7.6

7.0

8.9

6.9

8.3

6.7

8.5**

6.8

ALT

U/L

34

33

28

35

28

37

25*

31

Cholesterol

mg/dl

52

70

52

72

74*

69

70

78

Females

Haemoglobin

g/dl

15.4

15.0

15.3

15.5

14.6

15.5

13.5**

15.5

Haematocrit

%

41.4

38.8

40.9

39.6

37.8

39.6

35.0**

39.7

Reticulocytes

/ 100 RBC

2.9

2.3

3.0

2.2

3.4

2.0

7.8*

1.5

g-GT

U/L

1

3

1

1

3*

1

4*

2

Cholesterol

mg/dl

77

94

76

103

109**

99

116**

77

*  p≤ 0.05;              **p≤ 0.01

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
22 mg/m³
Study duration:
subchronic
Species:
rat

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-guideline study
Qualifier:
according to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
- Detailed examination for clinical signs of toxicity was performed weekly instead of daily.
Qualifier:
according to
Guideline:
EPA OPP 82-4 (90-Day Inhalation Toxicity)
Qualifier:
according to
Guideline:
EU Method B.29 (Sub-Chronic Inhalation Toxicity:90-Day Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:CD® BR VAF/PLUS®
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, Michigan, USA
- Age at study initiation: Approx. 6 weeks
- Weight at study initiation: Males: 181-211 g; females: 130-149 g
Route of administration:
inhalation
Type of inhalation exposure:
whole body
Vehicle:
clean air
Remarks on MMAD:
MMAD / GSD: MMAD [µm] (± GSD [µm])
Low dose: 3.0 (1.92)
Mid dose: 2.2 (1.99)
High dose: 2.4 (1.91)
Details on inhalation exposure:
--
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Nominal concentration: Calculated from the amount of test compound used during the exposure period (by weighing the reservoir before and after the exposure period) and dividing the total creosote consumed by the total air volume passed through the chamber.
Analytical aerosol concentration: Determined by gravimetric determination of the oil amount adsorbed onto a 25-mm glass-fibre filter pad, divided by the sample volume.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 h/d, 5 days per week
Remarks:
Doses / Concentrations:
Mean nominal concentration [mg/m³]: 22, 128, and 221
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
Mean aerosol concentration [mg/m³]: 5.4, 49, 106
Basis:
analytical conc.
No. of animals per sex per dose:
20/sex/group
An additional group (5/sex) was sacrificed pre-test to define a baseline for clinical chemistry and haematological values.
Control animals:
yes, sham-exposed
Details on study design:
--
Positive control:
none
Observations and examinations performed and frequency:
Mortality: Yes, twice weekly

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: pre-test and weekly

FOOD CONSUMPTION: Yes - weekly

FOOD EFFICIENCY: No data

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes, prior to terminal necropsy and prior to recovery necropsy.

HAEMATOLOGY: Yes
Number of animals: 5/sex at pre-test, 10/sex/group at termination of study and on surviving recovery animals at the end of the recovery period.
Parameters: haematocrit, haemoglobin, erythrocyte count, total and differential leukocyte count, platelet count, reticulocytes, MCV, MCH, MCHC

CLINICAL CHEMISTRY: Yes
Number of animals: 5/sex at pre-test, 10/sex/group at termination of study and on surviving recovery animals at the end of the recovery period.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Organ weights:
Organs: adrenal, brain, ovary, testis with epididymis, heart, kidney, liver, lung, mammary gland, thymus, thyroid/parathyroid, trachea

Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals: external examination; contents of abdominal, thoracic and cranial cavities were examined both in situ and after removal and dissection.

HISTOPATHOLOGY: Yes
All animals: heart, thyroid, nasal tissues, trachea, and lungControl and high dose group, animals that died on study: adrenal, aorta, auditory sebaceous gland, bone with bone marrow (femur), bone marrow smear, brain (fore, mid and hind), eye including optic nerve and contiguous Harderian gland, gastrointestinal tract, gonads, heart, kidney, larynx, lachrymal gland, liver, lung, lymph nodes, mammary gland (females only), nasal tissues, pancreas, pituitary, prostate and seminal vesicle, salivary gland, sciatic nerve, skeletal muscle (thigh), skin, spinal cord, spleen, thymus, thyroid/parathyroid, trachea, tracheal bifurcation, urinary bladder, vagina, uterus and cervix
Other examinations:
Ten animals per dose group and sex were subject to a six-week post-exposure period after which they were sacrificed and examined macroscopically for reversibility of eventual effects.
Statistics:
Analysis of body weights, food consumption, clinical pathology laboratory tests and organ weights were performed as follows:Generally, when the number of animals in any one group was ≤10, non-parametric analysis was conducted using the KRUSKAL-WALLIS one-way analysis of variance, followed by the MANN-WHITNEY U test, where appropriate. In those cases where the number of animals in all groups was greater than ten an the measurements were on at least an interval scale (continuous data), parametric analysis was conducted utilizing BARTLETT’s chi-square test for homogeneity of variance, followed by an analysis of variance and then, where appropriate, by DUNNETT’s t-test.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
CLINICAL SIGNS AND MORTALITY:
One male in the mid-dose group died during the exposure phase (week 4) of this study.

BODY WEIGHT AND WEIGHT GAIN
Body weight decreases were observed in the two highest exposure groups. They were significant in both sexes of the high-dose group and in mid-dose females during exposure week 6. Mean weights for the low-level group were similar to the control group.

FOOD CONSUMPTION:
With the exception of week 1 (all groups significantly less than controls) and week 3 (males in the high-dose group significantly less than controls), food consumption during the 13-week exposures and the six-week post-exposure recovery period was comparable to that of controls.

HAEMATOLOGY:
Several haematological parameters showed significant changes in the two highest exposure levels at the terminal sacrifice including: decreased erythrocytes, haemoglobin, and haematocrit; increased numbers of reticulocytes. These changes were not detectable at the end of the recovery period.

CLINICAL CHEMISTRY:
Serum cholesterol was significantly increased in males of the mid-dose group and in females of the two highest exposure groups. This change was not evident at the end of the recovery period (see table below).

ORGAN WEIGHTS:
Terminal sacrifice: There was a test-article-related and statistically significant increase in the lung/trachea/body weight ratio in males and females of the high-dose group when compared to the respective control values. These increases correlated with the macroscopic observation of grey discolouration of the lungs, and the microscopic observation of pigmented macrophages within the lungs of the animals in the affected groups. In mid- and high-dose males, the adrenal/body weight ratio was increased. No macroscopic or microscopic changes were associated with these changes.
In males, increases in liver weight (mid-dose group) and liver/body weight ratio (mid- and high-dose group) were noted. Relative liver weights were significantly increased at the mid and high dose (about +20 and +25 %, respectively).
In females, there were increases in liver weight (high-dose group), liver/body weight ratio (mid- and high-dose group) and liver/brain weight ratio (mid- and high-dose group) when compared to controls. No macroscopic or microscopic changes were associated with these changes, thus, their toxicbiological significance is uncertain.

Recovery sacrifice: There were no test-article-related changes in organ weight or weight ratios for males in any dose group. In mid-dose females, the mean adrenal weight was significantly decreased compared to controls. No macroscopic or microscopic observations were associated with that finding. Thus, it was deemed not test article related.

GROSS PATHOLOGY:
At the terminal sacrifice, a deposition of the test article consisting of a grey discolouration of the lung was seen at the two highest exposure levels. The discolouration persisted through the recovery period. The control and low-dose group

HISTOPATHOLOGY: NON-NEOPLASTIC:
Microscopic changes were observed in the hearts, lungs, nasal tissues, and thyroid glands of male and female rats at the time of terminal sacrifice. Heart lesions were found in one male of the mid-dose group that died on the study, and in one male and one female of the high-dose group (diffuse myocardial degeneration affecting mainly the right side of the heart). Associated to this change was diffuse arterial medial hypertrophy of small arterioles in the lung, brown pigment within the epithelial cells of the convoluted tubules of the kidney, and in the animal that died on study, alveolar macrophages containing brown pigment consistent with haemosiderin (“heart failure cells”) within the lung and diffuse centrilobular fibrosis within the liver. No heart lesions were found in any dose group at the end of the recovery period.
Note: Cardiac pathology (ie: hemorrhage, lymphocytic infiltration and cardiomyopathy) was noted in all animals of all groups (including controls).

Test-article-related changes in the lung were seen in all animals of the exposed groups (small black pigment granules within alveolar macrophages). Alveolar macrophages containing pigment granules could be detected in all lobes of the lungs indicating a uniform dispersion of the test article throughout the alveolar spaces of the lungs. There were no other changes within the lungs which could be associated with the presence of the granules, such as inflammation, increased number of pulmonary macrophages and/or Type-II pneumocyte hyperplasia. These findings were also seen at recovery sacrifice.

Nasal cavity: Small cystic spaces, containing basophilic mucoid material, within the olfactory epithelium at all levels of the nasal tissues examined and in both sexes, and were considered test article related. Mucoid cysts were seen in mid- and high-dose males, and in low-, mid-, and high-dose females. Similar findings were made at recovery sacrifice.
Other histological changes within the nasal tissues: Squamous metaplasia of respiratory and/or olfactory epithelium and naso­lachrymal duct epithelium, inflammatory cell infiltrates, and glandular dilation within the lamina propria/submucosa of the nasal cavity. These additional changes showed no definitive test article relationship.

Hypertrophy of thyroid follicular cells which resulted in a reduction in the amount of colloid present within the thyroid follicles was seen in both male and female rats of all exposure groups. This anomaly was considered test-article related. No test-article-related effects on the thyroid glands were detected at recovery sacrifice. There was no measurable effect on the mass of the thyroid gland (no changes in absolute and relative weights).

Dose descriptor:
NOAEC
Remarks:
systemic
Effect level:
5.4 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on the death of one male, decreases in body-weight gain (>10 %) as well as on the increase in liver weight (>=20 %), and on the increase in hypertrophy of follicular cells of the thyroid gland
Dose descriptor:
NOAEC
Remarks:
local, nasal
Effect level:
5.4 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
NOAEC
Remarks:
systemic
Effect level:
22 mg/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
NOAEC
Remarks:
local, nasal
Effect level:
22 mg/L air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
LOAEC
Remarks:
systemic
Effect level:
49 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
LOAEC
Remarks:
local, nasal
Effect level:
49 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
aerosolic fraction
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Dose descriptor:
LOAEC
Remarks:
systemic
Effect level:
128 mg/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see effects above under NOAEC systemic, analytical effect levels
Dose descriptor:
LOAEC
Remarks:
local, nasal
Effect level:
128 mg/L air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on local chronic inflammation reactions in the nasal cavity in both sexes
Critical effects observed:
not specified

Results of clinical chemistry and haematology

Parameter changed

Unit

Controls
0 mg/m³

Low dose
5.4 mg/m³

Medium dose
49 mg/m³

High dose
106 mg/m³

Time point

Terminal

Recovery

Terminal

Recovery

Terminal

Recovery

Terminal

Recovery

Males

Haemoglobin

g/dl

15.5

15.2

15.4

15.3

14.3**

15.0

14.6

15.1

Haematocrit

%

44.7

40.0

43.5

40.1

39.7*

39.7

40.3

39.8

Reticulocytes

/ 100 RBC

2.8

2.1

2.8

1.8

4.0

1.6

5.2

1.6

Phosphorus

mg/dl

7.6

7.0

8.9

6.9

8.3

6.7

8.5**

6.8

ALT

U/L

34

33

28

35

28

37

25*

31

Cholesterol

mg/dl

52

70

52

72

74*

69

70

78

Females

Haemoglobin

g/dl

15.4

15.0

15.3

15.5

14.6

15.5

13.5**

15.5

Haematocrit

%

41.4

38.8

40.9

39.6

37.8

39.6

35.0**

39.7

Reticulocytes

/ 100 RBC

2.9

2.3

3.0

2.2

3.4

2.0

7.8*

1.5

g-GT

U/L

1

3

1

1

3*

1

4*

2

Cholesterol

mg/dl

77

94

76

103

109**

99

116**

77

*  p≤ 0.05;              **p≤ 0.01

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
22 mg/m³
Study duration:
subchronic
Species:
rat

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

No data on repeated dose toxicity have been identified for the substance distillates (coal tar), heavy oils (anthracene oil high (> 50 ppm) BaP, AOH) itself. Data located for oral exposure relate to an aromatic mixture of the >EC9 to EC16 fraction (EC = equivalent carbon number index, see above Baars et al. (RIVM) 2001, page 281) from total petroleum hydrocarbons (TPH). Data on inhalation exposure originate from the closely related tar oil creosote.

Oral exposure

AOH is a UVCB and consists of a complex combination of polycyclic aromatic hydrocarbons (PAH). It comprises mainly 3- and 4-ring aromatic compounds and to a much lesser extent PAHs with 5 rings (see Chapter 1.). Main components are phenanthrene, anthracene (3-ring PAH), fluoranthene, pyrene, benz[a]anthracene, and chrysene (4-ring PAH) each accounting for ca. 3 to 10 %.

The >EC9 to EC16 TPH fraction also consists mostly of aromatic substances with carbon numbers higher than 9. This includes 3- and 4-ring PAH. As repeated dose toxicity is the relevant endpoint, PAH identified as carcinogenic are excluded from the toxicological evaluation. Representative substances used for characterising the repeated dose toxicity of the >EC9 to EC16 fraction are (amongst others) acenaphthene, fluorene, anthracene, fluoranthene and pyrene (3- and 4-ring PAH). These substances are also present in AOH and can be used to describe the repeated dose toxicity of AOH.

In evaluating the toxicity of the most relevant components in the aromatic >EC9 to EC16 TPH fraction based on expert judgement, Baars et al. derived an oral TDI value (tolerable daily intake) of 0.04 mg/kg bw/day. This value characterises the repeated dose toxicity of the total fraction including all the constituents in the mixture. This value is adopted to represent the repeated dose toxicity of AOH as a whole since relevant components in both mixtures are the same.

Basis for deriving the TDI value for the mixture as total were RfD values (oral reference doses, equivalent to TDI values) of individual substances obtained from animal (mouse) experimental data. Evaluation of the toxicity of these substances resulted in the average TDI for the total mixture. The uncertainty factor used for deriving individual RfD values was in any case 3000. Thus, it is possible to derive an average NOAEL for AOH by recalculating the NOAEL from the TDI value of 0.04 mg/kg bw/day. This approach results in an average NOAEL for AOH of 120 mg/kg bw/day.

Inhalation exposure

AOH and creosote are produced in a similar production process (fractionated distillation of coal tar using overlapping conditions). Composition of both tar substances correspond to each other. Major components are mid-range PAH for both substances (3- and 4-ring PAH) with some additional 2-ring PAH in creosote but not in AOH.

As 2-ring PAH are more volatile than 3- and 4-ring PAH and naphthalene is known to induce hyperplasia and metaplasia in nasal tissues, inhalation repeated dose toxicity of AOH may be overestimated when effects are based on creosote as supporting substance. Therefore, use of creosote in identifying the inhalation repeated dose toxicity of AOH is considered worst case allowing the use of creosote as supporting substance.

Low-grade toxicity was noted in rats exposed to an aerosol of creosote for 90 days. The aerosol MMAD was between 2.2 and 3.0 µm, potentially completely respirable. Inhalation of creosote aerosol caused nasal irritation and chronic inflammation in the nasal cavity of exposed rats. No particular systemic effects were observed but treatment-related hypertrophic changes of the thyroidal gland, the liver and lung as well as lung depositions of test material (Hilaski 1995). The NOAEC is reported as 22 mg/m³ (nominal) and includes both the particle and gaseous fraction of creosote. Analytically measured NOEC was determined to be 5.4 mg/m³. The nominal concentration is assessed to be more relevant and representative of the actual exposure as the analytical method only registered material deposited on filter pads, thus very likely underrating exposure because gas phase concentrations were not collected and analysed in the measurements.

Dermal exposure

No toxicological data are available from a dermal repeated dose toxicity study.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Value represents the characteristic repeated dose toxicity of an aromatic mixture/fraction composed of >EC9 to EC 16 aromatic components.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
No data for AOH itself. Results relate to the structure-related tar oil creosote providing evidence of adaptive response in liver and thyroid

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
No data for AOH itself. Results relate to the structure-related tar oil creosote providing evidence of chronic local adverse effects (inflammatory response in the nasal cavity)

Repeated dose toxicity: inhalation - systemic effects (target organ) digestive: liver; glandular: thyroids; respiratory: lung; respiratory: nose

Justification for classification or non-classification

There is no need for classification for non-neoplastic organ-specific toxicity.

The effects noted are of low severity and unspecific character (compare also the structure-related anthracene oil, B(a)P <50 ppm (AOL, CAS no. 90640 -80 -5). Moreover, the conservative estimates for a TDI (Tolerable Daily Intake) and a TCA (Tolerable Concentration in Air) of phenanthrene and related PAH, which are key components in AOH, are distinctly higher than the DMEL values (oral, inhalation, dermal) of benzo[a]pyrene for workers and general population. The need for classification for carcinogenicity based on the BaP content will overrule general and potential target-organ toxicity.