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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
Exposure: 20 Jul. 1984 - 19 Oct. 1984 / end observation: 15 Oct. 1985
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1987
Report Date:
1987
Reference Type:
publication
Title:
Unnamed
Year:
1991

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
Special modifications as compared with standard study:  Focus upon lung, respiratory  tract, and regional lymph nodes. Post-exposure recovery period up to one year.
Principles of method if other than guideline:
Comparative study including Aerosil 200 (pyrogenic, hydrophilic), Aerosil R 974 (pyrogenic, hydrophobic), Sipernat 22S (precipitated, hydrophilic) as well as quartz (crystalline).
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Aerosil 200:
CAS-Name: Silica, amorphous, fumed, crystalline-free,
CAS-No.: 112945-52-5
- Substance type: inorganic
- Physical state: solid
- Surface area (BET): 151 m2/g (Report p. 59 Specification Certificate)
- Analytical purity: >99.8 % (SiO2)
- Particle size: The range of the geometric agglomerate/aggregate size distribution was 1  to about 120 µm for the amorphous silicas 
with maxima at approx. 10 µm and 100 µm (Report 1987, p. 13)    
- Stability under test conditions: stable
- Storage condition of test material: room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Central Institute for Breeding of Laboratory Animals TNO, Zeist/NL
- Age at study initiation: 4 weeks
- Weight at study initiation: 50 - 70 g
- Fasting period before study: no
- Housing: single during exposure
- Diet: no access during exposure
- Water: no access during exposure
- Acclimation period: 10 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +-1
- Humidity (%): 50 - 70
- Air changes (per hr): 12x/h
- Photoperiod (hrs dark / hrs light): no data

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure:
whole body
Remarks on MMAD:
MMAD / GSD: no monitoring data due to technical difficulties (see above "Details on inhalation exposure")
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: stainless steel exposure chamber, multitiered (manufactered by Hazelton)
- Exposure chamber volume: 2.3 m3
- Method of holding animals in test chamber: single
- Source and rate of air: Aerosol entrance at top of the chamber
- Method of conditioning air: no data
- System of generating particulates/aerosols: Institute´s dust generator with compressed air operating atomizer
- Temperature, humidity, pressure in air chamber: av. 21 - 23 °C, minimum 19.1, max. 25.4 °C /
65 - 75 % rel. humidity, during extreme weather occasionally up to 95.5 % or down to 48 %.
- Air flow rate: approx. 40 m3/h
- Air change rate: 40 / 2.3 = ~17/h
- Method of particle size determination: due to electrostatic charge of the particles not measured:
technical failure of the 10-stage Mercer cascade impactor and the QCM cascade (Report p. 16)
- Treatment of exhaust air: filtered before release


TEST ATMOSPHERE
- Brief description of analytical method used: gravimetrically - Air samples are drawn through glass fiber filters (Sartorius)
and weighed (3 - 4 times per day)
- Samples taken from breathing zone: no data


Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
see Report Tables (Part 2), Table 1: Daily mean concentrations are documented:
based on 250 - 253 measurements: 1.26 (SEM 0.08) mg/m3; 5.88 (SEM 0.88) mg/m3; 31.04 (SEM 0.87) mg/m3
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours/day, 5 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
1.3, 5.9 or 31 mg/m3 (mean analytical values)
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
1, 6 and 30 mg/m3 (target concentrations)
Basis:
nominal conc.
No. of animals per sex per dose:
70
Aerosil 200: assigned dose groups B, C, and D, each sub-divided in 7 sub-groups a, b, c , d, e, f, and g
10 each (sacrificed after 13 wks),
50 each kept for a recovery period of at most 52 wks (13, 26, 39, and 52 wks). 
Control animals:
yes
Details on study design:
- Dose selection rationale: based on range findings (14 d)
- Rationale for selecting satellite groups: post-exposure recovery period for examination of reversibility of effects
- Post-exposure recovery period in satellite groups: 13, 26, 39, and 52 wks
Positive control:
Quartz (crystalline silica, 58 mg/m3) included (assigned Group G)

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: yes
- Time schedule: 2x/day, 1x/d (weekends)
- Cage side observations checked in table 3 and 4 (mortalities) were included.


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: see body weight


BODY WEIGHT: Yes
- Time schedule for examinations: start, weekly during exposure, 1x/wk during recovery
- Tables 5 and 6

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data


FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data


WATER CONSUMPTION: No data
- Time schedule for examinations:


OPHTHALMOSCOPIC EXAMINATION: No


HAEMATOLOGY: Yes
- Time schedule for collection of blood: week 13, 26, 39, 52, 65 (i.e. including recovery period)
- Anaesthetic used for blood collection: No (data)
- Animals fasted: No data
- How many animals: 10 males, 10 females
- Tables 7-16


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: week 14, 27, 40, 53, and 66
- Animals fasted: Yes overnight
- How many animals: 10 males, 10 females
- Parameters in tables 17 - 26


URINALYSIS: Yes
- Time schedule for collection of urine: week 13, 26, 40/41, 52, and 65
- Animals fasted: Yes
- Parameters in tables 27 - 36


NEUROBEHAVIOURAL EXAMINATION: No


OTHER: ---
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (Table 63 - 67)
Relative organ weights (Table 37 - 56)
HISTOPATHOLOGY: Yes (see table 68 - 73), in particular lung and lymph nodes
in addition:
Si contents of lung and lymph nodes (Tables 59 - 62)
Collagen content in lung (Tables 57/58)
Other examinations:
Relative organ weights (Table 37 - 56)
Statistics:
Body weights: analysis of co-variance followed by Dunnett´s test
Histopathological changes and mortality: Fisher´s exact probability test
Organ weights, blood parameter: analysis of variance and Dunnett´s test

Results and discussion

Results of examinations

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:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
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:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY
Respiration rate: concentration-related increase
No mortality

BODY WEIGHT AND WEIGHT GAIN
No effect in females at all dose levels (Tab. 6)
Depressive effect in males:
1 mg/m3: slightly at day 14 of exposure only (~ -5%)
6 mg/m3: slightly from day 49 to day 77 of exposure (~ - 6 to <5 %)
no more significant by end of exposure (day 91)
30 mg/m3: significantly throughout exposure: ~ -7 - -10 %, day 91: -7 %
Recovery: no difference from control at day 455 (52 weeks post-exposure)


HAEMATOLOGY
1 mg/m3: no effects
6 mg/m3: White blood cell count elevated in both males and females due to increases in the numbers of neutrophilic leukocytes,
but concentration-response relationship was poor.
After 3 months recovery, these blood parameters normalized in males and females.
30 mg/m3: Red blood cell count and hemoglobin were statistically higher in males, but not in females.
White blood cell count elevated in both males and females due to increases in the numbers of neutrophilic leukocytes,
at 3 months of recovery (days 176/177, Table 8/Table 13), but concentration-response relationship was poor.
In females, a slight increase above the control group apparently still existed after 6 months of recovery (day 275, Table 14).


CLINICAL CHEMISTRY
no significant effects

URINALYSIS
no significant effects

ORGAN WEIGHTS
No changes in heart, thyroid, thymus, adrenals, testes, brain, spleen, kidney
Treatment-related degrees of severity: swollen lungs and enlarged mediastinal lypmph nodes at the end of exposure
LUNG
1 mg/m3: no significant increase
6 mg/m3: mean increase in relative weight 1.7x (males), 1.4x (females)
30 mg/m3: mean increase in relative weight 2.3x (males), 2.0x (females)
LYMPH NODE: no weight data


PATHOLOGY
Swollen and spotted lungs and enlarged mediastinal lymph nodes, the degree of severity being treatment-related.
At 6 and 30 mg/m3, collagen content in the lungs was clearly increased, most pronounced in males.

The above-mentioned effects gradually subsided after the exposure period,
but in males exposed to 6 and 30 mg/m3 the collagen content was still above control values at the end of the study.


HISTOPATHOLOGY: NON-NEOPLASTIC
Accumulation of alveolar macrophages and granular material, cellular debris, polymorphonuclear leucocytes, increased septal cellularity.
Alveolar bronchialisation, focal interstitial fibrosis, cholesterol clefts and granuloma-like lesions in the lung.
The granuloma-like lesions were seen in a few animals at the end of exposure period and after 13 weeks of recovery. They did not show fibroblastic activity and hyalinization and regressed during recovery [not progressive, i.e. no silicogenic nodules formed (no silicosis)].

Accumulation of macrophages were seen in the mediastinal lymph nodes (disappeared after wk 39 post-exposure).
Treatment-related microscopic changes in the nasal region were occasionally found at the end of exposure period,
such as focal necrosis and slight atrophy of the olfactory epithelium.

Interstitial fibrosis was not noted directly after the exposure period, but appeared with a delay,
for the first time observed after 13 wks post-exposure: increasing incidence especially in 30-mg rats, and a few in the 6-mg group (Report p. 44),
but decreased in severity and frequency until the end of the study (Report p. 51).

All types of pulmonary lesions were more marked in males than in females.


The level of 1.3 mg/m3 induced only slight changes after 13-wk exposure (Table 68),
which generally recovered quickly (no differences from control after 13-wk post-exposure: Table 69).

Morphological changes after 13-wk exposure, that were considered statistically significant at 1.3 mg/m3 (Table 68):

males females ! males females
treated ! untreated controls
------------------------------------------------------------------------------------------------------------------------------
Accumulation of alveolar macrophages: slight in 10/10 (very) slight in 10/10 ! (very) slight 4/10 slight in 1/10
Intra-alveolar polymorphonuclear !
leukocytes: (very) slight in 6/10 (very) slight in 8/10 ! 0/10 0/10
Increased septal cellularity: ( very) slight in 10/10 (very) slight in 9/10 ! very slight 1/10 very slight 1/10
Olfactory epithelial atrophy: (very) slight in 5/10 (very) slight in 8/10 ! 0/10 0/10
Intracytoplasmic proteinaceous droplets !
-respiratory epithelium: in 8/10 in 9/10 ! 1/10 0/10
Mediastinal lymph node !
-macrophage accumulation: (very) slight in 8/10 (very) slight in 8/10 ! 0/10 0/10
------------------------------------------------------------------------------------------------------------------------------



HISTOPATHOLOGY: NEOPLASTIC
No particular findings


HISTORICAL CONTROL DATA (if applicable): no data


OTHER FINDINGS - SILICA DEPOSITION
Silica could be detected in lungs only in relatively small amounts at the end of the exposure period (Tables 59):
on the average 0.1 - 0.2 mg per lung of male animal groups (not dose-related), 0.05 - 0.21 mg per lung of female groups (dose-related).
Only one male exposed to 30 mg/m3 showed a small amount of silica in the regional lymph node.
90 days after termination of exposure (day 188), no silica could be recovered from any animal.
(see Chapter 7.1: Degussa 87-0004-DGT_ Aero200_inhal_Si-deposition, 13 wk, rat_key_RL2)

Effect levels

open allclose all
Dose descriptor:
NOAEC
Effect level:
1.3 mg/m³ air (analytical)
Sex:
male/female
Basis for effect level:
other: Histopathology: based on the slight and fully reversible pulmonary response noted at this exposure level (inflammation reaction) (see details below "Details on results")
Dose descriptor:
LOAEC
Effect level:
5.9 mg/m³ air (analytical)
Sex:
male/female
Basis for effect level:
other: (haematology); organ weights (hypertrophy lung); histopathology (collagen increase, sporadic focal fibrosis)
Dose descriptor:
NOEC
Effect level:
< 1.3 mg/m³ air (analytical)
Sex:
male/female
Basis for effect level:
other: Histopathology: based on the pulmonary response (inflammation reaction) (see details below "Details on results")

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion