Registration Dossier

Toxicological information

Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 413 (90-Day (Subchronic) Inhalation Toxicity Study
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
Starting material: Aluminium chloride anhydrous ground (PRD 30041207); solid / yellowish white
Batch No. 000STD77L0
purity: > 99 %
Expiry date: 16 Oct 2019
The stability of the test substance under storage conditions over the test period was guaranteed by the sponsor, and the sponsor holds this responsibility.
As the test substance is highly hygroscopic, the test substance has been dissolved in ultrapure water, where it reacts violently with water and forms a stable AlCl3 hexahydrate and after appropriate dilution, sprayed as aqueous solution. A detailed explanation can be found attached in sectin "overall remarks, attachments" as well as in the corresponding CSR.

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Crl:WI(Han)
Details on species / strain selection:
Rats were selected since this rodent species is recommended in the respective test guidelines. Wistar rats were selected since there is extensive experience available in the laboratory with this strain of rats.
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH; Sandhofer Weg 7, 97633 Sulzfeld
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation:
- Weight at study initiation:
- Fasting period before study:
- Housing: The rats were housed together (up to 5 animals per cage) in ca. 2065 cm2 polysulfone cages. Bedding in the Polycarbonate cages were dust-free bedding. Motor activity measurements were conducted in Polycarbonate cages with wire covers floor area about 800 cm2 and small amounts of absorbent material. For enrichment wooden gnawing blocks were added.
- Diet (e.g. ad libitum): mouse and rat maintenance diet, GLP, 12 mm pellets
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period:

DETAILS OF FOOD AND WATER QUALITY:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24°C
- Humidity (%): 45-65%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
clean air
Mass median aerodynamic diameter (MMAD):
< 1.39 µm
Remarks on MMAD:
All cascade impactor measurements resulted in mass median aerodynamic diameters (MMADs) between 0.90 and 1.39 µm with geometric standard deviation (GSDs) <3.
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The inhalation atmosphere was maintained inside aerodynamic exposure systems consisting of a cylindrical inhalation chamber made of stainless steel sheeting and cone shaped outlets and inlets. The exposure systems were located in exhaust hoods in an air conditioned room.
- Method of holding animals in test chamber: The rats were restrained in glass exposure tubes. Their snouts projected into the inhalation chamber and thus they inhaled the aerosol.
- Source and rate of air:
- Method of conditioning air: activated charcoal filtered air conditioned to about 30% to 70% relative humidity and 20°C to 24°C
- System of generating particulates/aerosols: For each concentration, a respective constant amount of the test substance solution was supplied to a two-component atomizer by means of a metering pump and sprayed with compressed air into a glass mixing stage. The so generated aerosol was mixed with conditioned air and passed after appropriate dilution, via a cyclone separator into the inhalation system. The desired concentrations in test groups 1 and 2 were achieved by substituting appropriate amounts of aerosol by conditioned supply air.
- Temperature, humidity, pressure in air chamber: 20-24 °C; 30-70% relative humidity. A positive pressure was maintained inside the exposure systems by adjusting the air flow of the exhaust air system. This ensured that the aerosol in the breathing zones of the animals was not diluted by laboratory air.
- Method of particle size determination: The particle size analysis was carried out with a cascade impactor.
- Treatment of exhaust air: The exhaust air was filtered and conducted into the exhaust air of the building.

TEST ATMOSPHERE
- Brief description of analytical method used: The concentrations of the inhalation atmospheres were analyzed by gravimetric determination of filter samples. This analytical method is judged to be valid because the test substance itself does not possess an appreciable vapor pressure.
- Samples taken from breathing zone: yes

VEHICLE (if applicable)
- Justification for use and choice of vehicle: To adjust the exposure condition in the control group, the air was humidified with ultrapure
water. To humidify the atmosphere of the control group, ultrapure water was sprayed in the same way like the test substance into the inhalation system. The amount of dosed water was comparable with those in highest test substance concentration.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The concentrations of the inhalation atmospheres were analyzed by gravimetric determination of filter samples. This analytical method is judged to be valid because the test substance itself does not possess an appreciable vapor pressure.

For test groups 1 and 2, one sample per week or one daily sample was taken. Thus, no daily means were calculated for these two test groups. Daily means were calculated based on 3 measured samples per concentration and exposure in test group 3. From the daily mean values of each concentration, mean concentrations and standard deviations for the entire study were derived.

In these groups, the constancy of concentrations in each inhalation system was continuously monitored using scattered light photometers.

In the control group (test group 0) no sample was analyzed.

The gravimetric analyses were carried out at the Laboratory for Inhalation Toxicology of the Experimental Toxicology and Ecology of BASF SE.

Additional analyses (Al NMR) to assess the identity of the dissolved test substance were carried out as a separate study at the test facility Competence Centre Analytics, BASF SE, under the responsibility of the study director of this test facility. The study was carried out in compliance with the principles of Good Laboratory Practice.
Duration of treatment / exposure:
6h/d
Frequency of treatment:
5d/week
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/m³ air
Remarks:
humidified air
Dose / conc.:
0.1 mg/m³ air
Dose / conc.:
0.5 mg/m³ air
Dose / conc.:
2 mg/m³ air
No. of animals per sex per dose:
10
Control animals:
yes
Details on study design:
- Dose selection rationale: Based on available data (pre-study)
Positive control:
No

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: The animals were examined for evident signs of toxicity or mortality twice a day (in the morning and in the late afternoon) on working days and once a day (in the morning) on Saturdays, Sundays and public holidays.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: All animals were subjected to detailed clinical observations outside their cages once before the beginning, once at mid-term and once at the end of the exposure period. The examinations were generally performed in the morning.
The scope of examinations and the scoring of the findings that are observed includes but was not limited to the following parameters listed:

1. abnormal behavior during handling
2. fur
3. skin
4. posture
5. salivation
6. respiration
7. activity/arousal level
8. tremors
9. convulsions
10. abnormal movements
11. impairment of gait
12. lacrimation
13. palpebral closure
14. exophthalmus
15. feces (appearance/consistency)
16. urine
17. pupil size


BODY WEIGHT: Yes
- Time schedule for examinations: The body weight of the animals was determined at the start of the pre-exposure, at the start of the exposure period and then, as a rule, on Mondays and Fridays as well as prior to gross necropsy. As a rule, the animals were weighed at the same time of the day.

Body weight change of the first week was calculated as the difference between body weight on Friday and that on day 0. For the following weeks, the body weight change of the respective week was calculated as the difference of Friday to the previous Monday.

In general, the last weighing was on the day before final sacrifice. In examination groups 2, 3 and 4, body weight change was calculated as difference of the last weighing and the previous Monday. For examination group 1, the last weighing was on a Monday. Thus, no body weight change was calculated. Group means were derived from the individual differences.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
Food consumption was determined weekly and calculated as mean food consumption in grams per animal and day.
The animals were maintained in social-housing cages, with 5 animals per cage, during the whole study period. Therefore, the food consumption was determined cage-wise. The food consumption per animal and day was calculated by dividing food consumption of the day of a respective cage by the 5 animals per cage. As the animals of each test group were housed in only two cages per sex, no statistical evaluation of food consumption is possible.

OPHTHALMOSCOPIC EXAMINATION: Yes
Before the start of the exposure period, the eyes of male and female animals were examined for any changes in the refracting media with an ophthalmoscope after administration of a mydriatic. This was study day -5 for examination group 1, study day -6 for examination group 2, study day -7 for examination group 3 and study day -8 for examination group 4.

To detect any treatment-related effects in eyes, ophthalmological examination was performed again at the end of the study, in control and high concentration group animals. This was study day 87 for examination group 1, study day 86 for examination group 2, study day 85 for examination group 3 and study day 84 for examination group 4. the eyes of the animals of test group 0 (control group), test group 2 and test group 4 (high concentration) were examined.


HAEMATOLOGY: Yes
In the morning blood was taken from the retro-bulbar venous plexus from fasted animals.

The following parameters were determined in blood :
Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration,(MCHC), Platelet count (PLT), Differential blood count, Reticulocytes (RETA)
Clotting tests were carried out using a ball coagulometer.

CLINICAL CHEMISTRY: Yes
Parameters examined: Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (ALP), gamma-Glutamyltransferase (GGT), Sodium (NA), Potassium (K), Chloride (CL), Inorganic phosphate (INP), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Albumin (ALB), Globulins (GLOB), Triglycerides (TRIG), Cholesterol (CHOL)

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
A functional observation battery (FOB) was carried out on the assigned animals
Home cage observations:

The animals were observed in their closed home cages; any disturbing activities (touch¬ing the cage or rack, noise) were avoided during these examinations in order not to in¬fluence the behavior of the animals. Attention was paid to:

1. posture
2. tremor
3. convulsions
4. abnormal movements
5. impairment of gait

Open field observations:

The animals were transferred to a standard arena (50 x 50 cm with sides of 25 cm high) and observed for at least 2 minutes. Following parameters were examined:

1. behavior when removed from cage
2. fur
3. skin
4. salivation
5. nasal discharge
6. lacrimation
7. eyes/pupil size
8. posture
9. palpebral closure
10. respiration
11. tremors
12. convulsions
13. abnormal movements/ stereotypies
14. impairment of gait
15. activity/arousal level
16. feces (appearance/consistency) within two minutes
17. urine (amount/color) within two minutes
18. number of rearings within two minutes

Sensorimotor Tests/Reflexes:

The animals were removed from the open field and subjected to following sensorimotor or reflex tests:

19. approach response
20. touch response
21. vision ("visual placing response")
22. pupillary reflex
23. pinna reflex
24. audition ("startle response")
25. coordination of movements ("righting response")
26. behavior during "handling"
27. vocalization
28. pain perception ("tail pinch")
29. grip strength of forelimbs
30. grip strength of hindlimbs
31. landing foot-splay test

Motor activity was measured on the same day and with the same animals as FOB was performed.

BRONCHOALVEOLAR LAVAGE FLUID (BALF): Yes
Parameters examined: CYTOLOGY: Total cell count, Macrophages, Polymorphonuclear neutrophils, Lymphocytes, Eosinophils, Monocytes,
Epithelial
PROTEIN AND ENZYMES: gamma-Glutamyltransferase, Protein, Lactate dehydrogenase, Alkaline phosphatase, N-acetyl-b-Glucosaminidase
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
The following weights were determined in all animals sacrificed on schedule:

1. Anesthetized animals (terminal body weight)
2. Adrenal glands (fixed)
3. Brain
4. Epididymides
5. Heart
6. Kidneys
7. Liver
8. Lungs
9. Ovaries (fixed)
10. Spleen
11. Testes
12. Thymus (fixed)
13. Thyroid glands (with parathyroid glands) (fixed)
14. Uterus with cervix

All paired organs were weighed together (left and right).

HISTOPATHOLOGY: Yes
1. All gross lesions
2. Adrenal glands
3. Aorta
4. Bone marrow
5. Brain
6. Cecum
7. Cervix
8. Coagulating glands
9. Colon
10. Duodenum
11. Epididymides
12. Esophagus
13. Eyes with optic nerve
14. Extraorbital lacrimal gland
15. Femur with knee joint
16. Harderian glands
17. Heart
18. Ileum
19. Jejunum
20. Kidneys
21. Larynx (3 levels)
22. Liver
23. Lungs
24. Lymph nodes (tracheobronchial and mediastinal)
25. Lymph nodes (mesenteric)
26. Mammary gland (female)
27. Nasal cavity (4 levels)
28. Olfactory bulb
29. Ovaries
30. Pancreas
31. Pharynx
32. Parathyroid glands
33. Pituitary gland
34. Prostate
35. Rectum
36. Salivary glands (mandibular and sublingual glands)
37. Sciatic nerve
38. Seminal vesicles
39. Skeletal muscle
40. Skin
41. Spinal cord (cervical, thoracic and lumbar cord)
42. Spleen
43. Sternum with marrow
44. Stomach (forestomach and glandular stomach)
45. Teeth
46. Testes
47. Thymus
48. Thyroid glands
49. Trachea
50. Urinary bladder
51. Uterus
52. Vagina
Statistics:
Means, medians and standard deviations of each test group were calculated for several parameters:
Body weight, body weight change: Comparison of each group with the control group was performed using DUNNETT test (two-sided) for the hypothesis of equal means.
Feces, rearing, grip strength length forelimbs, grip strength length hindlimbs, foot-splay test, motor activity: Non-parametric one-way analysis using the KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal to or less than 0.05, a pair-wise com¬parison of each dose group with the control group was performed using WILCOXON test (two-sided) for the hypothesis of equal medians.
Blood parameters: For parameters with bidirectional changes: Non-parametric one-way analysis using KRUSKAL-WALLIS test. If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (two-sided) for the hypothesis of equal medians. For parameters with unidirectional changes: Pairwise comparison of each dose group with the control group using the WILCOXON-test (one-sided) for the hypothesis of equal medians.
Broncho-alveolar lavage fluid (BALF): Pairwise comparison of each dose group with the control group using the WILCOXON-test (one-sided) for the hypothesis of equal medians.
Weight parameters in pathology: Non-parametric one-way analysis using KRUSKAL-WALLIS H test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each test group with the control group was performed using WILCOXON-test (two-sided) for the equal medians.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
During the pre-exposure period and the exposure period the animals showed no clinical signs and findings different from normal. The detailed clinical observations did not reveal any abnormalities in any of the exposed animals.
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):
not examined
Ophthalmological findings:
effects observed, non-treatment-related
Description (incidence and severity):
The ophthalmologic examinations did not show any impairment of the refracting media.
Spontaneous findings such as remainders of the pupillary membrane or corneal stippling were observed in several animals of all test groups and the control group without any concentration-response relationship.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
At the end of the administration period, in males and females of test groups 2 and 3 (0.5 and 2.0 mg/m3) absolute and relative neutrophil counts were significantly increased and relative neutrophil counts were significantly decreased. These alterations were regarded as treatment-related and adverse.

Additionally, in males of test group 1 (0.1 mg/m3) absolute and relative neutrophil counts were significantly increased and relative lymphocyte counts were significantly decreased. However, absolute neutrophil counts were within the historical control range (males, absolute neutrophils 0.91-1.32 Giga/L). Relative neutrophil and lymphocyte counts were marginally beyond the historical control ranges (males, relative neutrophils 17.4-24.9 %, relative lymphocytes 70.3-78.4 %). Therefore, the mentioned changes in males of test group 1 were regarded as incidental and not treatment-related.

In males of test groups 1 and 2 (0.1 and 0.5 mg/m3) relative monocyte counts were significantly increased, and in females of test groups 1, 2 and 3 (0.1, 0.5 and 2.0 mg/m3) absolute monocyte counts were significantly higher compared to controls. However, all changed values were within historical control ranges (males, relative monocytes 1.6-2.3 %; females, absolute monocytes 0.04-0.09 Giga/L). Therefore, these changes were regarded as incidental and not treatment-related.

In females of test groups 1, 2 and 3 (0.1, 0.5 and 2.0 mg/m3) mean corpuscular volume (MCV) and mean corpuscular hemoglobin content (MCH) were significantly decreased. However, the measured red blood cell parameters (i.e., red blood cell (RBC) counts, hemoglobin and hematocrit values) were not changed. Therefore, the changes of only the calculated MCH and MCV indices were regarded as incidental and not treatment-related.
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
When compared to the control group 0 (=100%), the mean absolute lung weights were significantly increased in test group 3 (males 114%, females 113%).
When compared to the control group 0 (=100%), the following mean relative organ weights were significantly increased or decreased in one or more test groups: male animals: increased relative lung weight in group 3 animals (113%), decreased relative spleen weight in all dose groups (group 1: 86%, group 2: 89%, group 3: 89%)
female animals: increased relative kidney weight in group 2 and 3 animals (109% and 107%, respectively), increased relative liver weight in group 3 animals (106%), increased relative lung weight in group 3 animals (114%).
All other mean relative weight parameters did not show significant differences when compared to the control group 0.
The increased mean absolute and relative lung weights in males and females of test group 3 are regarded to be treatment-related.
The increased mean relative weights of the kidneys in female test group 2 and 3 animals was regarded to be equivocally treatment-related: both the mean relative weights of test group 2. (0.687%) and of test group 3 (0.673%) were above the historical control range (0.59% - 0.657%). There was, however, no clear dose response and no histopathological correlate in test group 3.
The decreased mean relative spleen weights in males of all treatment groups were regarded to be questionably treatment-related, the mean relative weight of the control group (0.193%) was slightly above historical controls (0.180% - 0.190%) while the treated test groups were below the historical control range (test group 1: 0.167%, test group 2: 0.172%, test group 3: 0.173%). There was, however, no dose response and no histopathological correlate in test group 3.
The statistically significantly increased mean relative liver weight in test group 3 females (2.575%) was assumed to be incidental as it fell within the historical control range (2.397% - 2.59%) and there was no histopathological correlate.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
The mediastinal and tracheobronchial lymph nodes were macroscopically enlarged in group 3 animals as well as in all dose groups in female animals.
All other findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.
Neuropathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Treatment-related findings were observed in the larynx (level I), lungs, nasal cavity (level IV), trachea, and the mediastinal and tracheobronchial lymph nodes.

Larynx, level I:
Epithelial alteration was seen with increased incidence and severity in treated animals of both sexes. This finding was characterized by a focal loss of cilia and flattening of the surface cells, primarily at the base of the epiglottis (incidences see table attached).

Lungs:
Several treatment-related findings were noted in the lungs with dose-related increase in incidence and/or severity in both male and female animals:
In the bronchus associated lymphoid tissue (BALT), there were aggregates of macrophages which contained yellow-brown 1-3 µm diameter particles in their cytoplasm which is assumed to be the test substance (described as solid yellowish white in the study plan).
In some alveoli, cell fragments and free 1-3 µm diameter particles (diagnosed as debris) were detected, this was interpreted as most-likely ruptured macrophages.
Histiocytosis, alveolar with particles was characterized by an increased number of histiocytes (macrophages) compared to controls. These macrophages mostly contained the same yellow-brown particles already described in macrophages having migrated to the BALT. This finding constitutes a continuum to “particles within histiocytes” which was diagnosed when the amount of histiocytes (macrophages) in alveoli did not exceed control levels, but some of these cells contained yellow-brown particles.
Infiltration, lymphoid, (multi)focal was characterized by nodules of mainly lymphocytes randomly distributed in the lung tissue. They were sometimes admixed with very few macrophages, some of them with ingested particles.
Inflammation, mixed cell was centered in the alveoli around bronchioles and terminal bronchioles and was composed mainly of neutrophils and macrophages. It was assumed to be a reaction to the inhaled test substance.

Lymph nodes (mediastinal and tracheobronchial):
The mediastinal and tracheobronchial lymph nodes showed macrophage aggregates as already described in the BALT. This correlated with the macroscopic finding “enlarged” in many of the affected lymph nodes. No further histopathological findings were seen in the lymph nodes.

Nasal cavity, level IV:
In the nasal cavity, 4 out of 10 male animals of test group 3 showed a focal loss of the respiratory epithelium in the ventral region of the nasal cavity which was diagnosed as erosion/ulcer. There was no or only minimal inflammatory reaction and no signs of regeneration could be detected.

Trachea:
The tip of the carina showed a minimal flattening of the epithelium with loss of cilia with increased incidence in treated animals of both sexes.

All other findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.



Histopathological findings: neoplastic:
no effects observed
Other effects:
effects observed, treatment-related
Description (incidence and severity):
BALF: At the end of the administration period, in the BAL of males and females of test groups 2 and 3 (0.5 and 2.0 mg/m3) total cell counts (apart from test group 2 females) as well as absolute and relative lymphocyte, neutrophils and monocytes were significantly increased. Relative macrophage counts were significantly decreased. Additionally, in females of test group 1 (0.1 mg/m3) absolute and relative neutrophil counts were significantly higher compared to controls. These alterations were regarded as treatment-related and adverse.
In males of test group 1 (0.1 mg/m3) relative neutrophil counts were significantly higher compared to controls. In females of this test group relative macrophage counts were significantly decreased whereas in males of test group 1 relative macrophage counts were significantly increased. These changes were only marginal and concerning the macrophages contrarily different between the sexes. Therefore, these changes were regarded as incidental and not treatment-related.

At the end of the administration period, in the BAL of males and females of test groups 2 and 3 (0.5 and 2.0 mg/m3) total protein values as well as lactate dehydrogenase (LDH; apart from females of test group 2), alkaline phosphatase (ALP) and γ-glutamyl transferase (GGT) activities were significantly increased. Additionally, in males of test group 3 -N-acetyl glucosaminidase (NAG) activities were significantly higher compared to controls. These alterations were regarded as treatment-related and adverse.

In males and females of test group1 (0.1 mg/m3) total protein levels and ALP activities were significantly higher compared to controls. However, the increases were only marginal (≤ 1.5-fold). Therefore, the alterations were regarded as maybe treatment-related, but non-adverse, apart from the ALP increase in females of test group 1, which is regarded in combination with the neutrophil count increases in BAL as treatment-related and adverse (See "any other information about results incl. tables" for more information).

Effect levels

open allclose all
Dose descriptor:
LOAEC
Effect level:
0.1 mg/m³ air
Based on:
test mat.
Sex:
female
Basis for effect level:
other: BALF findings
Remarks on result:
other: local effects
Dose descriptor:
NOAEC
Effect level:
0.1 mg/m³ air
Based on:
test mat.
Sex:
male
Basis for effect level:
other: BALF findings
Remarks on result:
other: local effects
Dose descriptor:
NOAEC
Effect level:
0.1 mg/m³ air
Based on:
test mat.
Sex:
male/female
Basis for effect level:
haematology
Remarks on result:
other: systemic toxicity

Target system / organ toxicity

Critical effects observed:
yes
Lowest effective dose / conc.:
0.1 mg/m³ air
System:
other: respiratory system
Organ:
lungs
Treatment related:
yes
Dose response relationship:
yes

Any other information on results incl. tables

Changes in mean absolute cell counts in BAL (x-fold of concurrent control) in male and female rats on study day 92:

Analyte

males

females

 

Gr. 1

0.1 mg/m3

Gr. 2

0.5 mg/m3

Gr. 3

2.0 mg/m3

Gr. 1

0.1 mg/m3

Gr. 2

0.5 mg/m3

Gr.3

2.0 mg/m3

Total Cells

0.7

1.5*

3.6**

0.9

0.9

2.9*

Macrophages

0.7

0.8

0.5

0.9

0.6

0.7

Lymphocytes

1.2

6.7**

24.9**

1.6

6.5**

51.5**

Neutrophils

0.8

13.8**

57.1**

30.3**

393.0**

2632.2**

Monocytes

+

+*

+**

+

+*

+*

Eosinophils

1.0

2.1

0.8

1.4

4.0

4.0

Epithelial cells

0.1

0.5

0.4

0.5

1.8

1.1

Changes in mean total protein and enzyme levels in BAL (x-fold of concurrent control) in male and female rats on study day 92:

Analyte

males

females

 

Gr. 1

0.1 mg/m3

Gr.2

0.5 mg/m3

Gr.3

2.0 mg/m3

Gr.1

0.1 mg/m3

Gr. 2

0.5 mg/m3

Gr. 3

2.0 mg/m3

Total Protein

1.3**

1.8**

3.1**

1.2*

1.4**

2.8**

LDH

1.0

2.0**

4.7**

1.1

1.3

3.3**

ALP

1.4**

1.7**

2.2**

1.5**

1.9**

2.1**

NAG

0.8

1.2

1.6**

0.8

0.8

1.3

GGT

1.1

2.1**

3.1**

1.2

1.7**

3.3**

Incidence of gross lesions:

 

Male animals

Female animals

Test group

(mg/m³)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

No. of animals

10

10

10

10

10

10

10

10

Mediastinal lymph node, enlarged

0

0

0

6

0

1

3

10

Tracheobronchial lymph node, enlarged

0

0

0

8

0

0

0

7

Incidences and gradings of histological changes in the larynx:

Larynx, level I

Male animals

Female animals

Test group

(mg/m³)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

No. of animals

10

10

10

10

10

10

10

10

Epithelial alteration

1

10

9

10

1

6

9

10

  • Grade 1

1

10

9

5

1

6

8

5

  • Grade 2

 

 

 

5

 

 

1

5

Incidences and gradings of histological changes in lung:

Lungs

Male animals

Female animals

Test group

(mg/m³)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

0

(0)

1

(0.1)

2

(0.5)

3

(2)

No. of animals

10

10

10

10

10

10

10

10

BALT: macrophage aggregates with particles

0

0

0

10

0

0

0

7

  • Grade 1

 

 

 

4

 

 

 

1

  • Grade 2

 

 

 

6

 

 

 

6

Debris, alveolar

0

0

7

10

0

0

0

8

  • Grade 1

 

 

7

3

 

 

 

5

  • Grade 2

 

 

 

6

 

 

 

3

  • Grade 3

 

 

 

1

 

 

 

 

Histiocytosis, alveolar with particles

0

1

10

10

0

0

10

10

  • Grade 1

 

1

10

7

 

 

10

 

  • Grade 2

 

 

 

3

 

 

 

5

  • Grade 3

 

 

 

 

 

 

 

5

Infiltration, lymphoid, (multi)focal

0

0

6

9

0

0

0

7

  • Grade 1

 

 

6

7

 

 

 

4

  • Grade 2

 

 

 

1

 

 

 

2

  • Grade 3

 

 

 

1

 

 

 

1

Inflammation, mixed cell

0

0

10

10

0

0

7

10

  • Grade 1

 

 

10

 

 

 

7

1

  • Grade 2

 

 

 

5

 

 

 

6

  • Grade 3

 

 

 

5

 

 

 

3

Particles within histiocytes

0

10

0

0

0

9

0

0

  • Present

 

10

 

 

 

9

 

 

Applicant's summary and conclusion