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Acute Toxicity: inhalation

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Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
Before April 1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
As most publications, this one does not present all the details that are normally presented in a standard study report. However, a thorough and extensive study is described in the publication that gives valuable information on the acute toxicity upon inhalation of the substance and can thus be used to address the requirement in question. Moreover, the publication is from a well-known peer-reviewed paper and the study was carried out by a laboratory with a good reputation.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1997

Materials and methods

Principles of method if other than guideline:
Rats were exposed nose-only to respirable iron particles (carbonyl iron) for 4 week, 5 days/week and 6 h/day. Endpoints studied were focused at inflammatory reactions in the respiratory tract and clearance.
GLP compliance:
not specified
Remarks:
It is not customary to provide information on GLP compliance in studies published in regulat scientific journals.
Test type:
other: Actually a repeated-dose study that, due to the high concentrations in the air of the test substance also provides meaningful information about the acute toxicity upon inhalation.
Limit test:
no

Test material

Constituent 1
Reference substance name:
carbonyl iron
IUPAC Name:
carbonyl iron
Details on test material:
- Name of test material (as cited in study report): carbonyl iron
- Substance type:
- Physical state: solid iron particles; particle-size range: 0.2-2.0 micrometer
- Analytical purity: no data
- Impurities (identity and concentrations): no data
- Composition of test material, percentage of components: metallic iron
- Isomers composition: not applicable
- Purity test date: no data
- Lot/batch No.: no data
- Expiration date of the lot/batch: no data
- Radiochemical purity (if radiolabelling): not applicable
- Specific activity (if radiolabelling): not applicable
- Locations of the label (if radiolabelling): not applicable
- Expiration date of radiochemical substance (if radiolabelling): not applicable
- Stability under test conditions: no data
- Storage condition of test material: no data
- Other:Commercial carbonyl iron was purchased from the GAF Corporation (New York), a well known source of this material. The production of carbonyl iron by reduction of iron pentocarbonyl results in a very high purity. A search on the internet made clear that carbonyl iron of GAF is used as a dietary iron supplement. We further refer two Gordeul et al., 1987, Am J Clin Nutr 46, 1029-1034. The following is a quote from this publication. "Carbonyl Fe, a pure form of elemental Fe used widely as a food additive (7, 8), has remarkably low toxicity and much larger doses are tolerated when compared with ionized forms of Fe such as FeSO4 (9, 10). Carbonyl does not refer to the composition ofthe Fe particles but rather to the manufacturing process in which the controlled heating of vaporized Fe pentacarbonyl leads to the deposition of uncharged elemental Fe as microscopic spheres of < 5 /L in diameter (11 ). Although not now approved by the Food and Drug Administration for pharmacologic use, recent studies have demonstrated that carbonyl Fe is effective for the treatment of Fe deficiency anemia (12) and for the replacement of Fe lost in blood donation (13). The low toxicity of carbonyl Fe has permitted a study to test if 10 times the usual daily amount of Fe would be therapeutically effective in a shorter time period." See original publication for references.

Test animals

Species:
rat
Strain:
other: Charles River Crl:CDBR
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, New York
- Age at study initiation: 7-8 weeks
- Weight at study initiation: not data
- Fasting period before study: not data
- Housing: no data
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum):no data
- Acclimation period:no data


ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data


IN-LIFE DATES: From: To: 180 days

Administration / exposure

Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
Animals were placed in cylindrical polycarbonate or stainless steel holders equipped with conical nose pieces. The restrainers were inserted into face plates on the exposure chambers such that the nose of each animal protruded into the chamber. Atmospheres of carbonyl iron were generated with a K-tron bin feeder equipped with twin feed screws. The dust was metered into a polycarbonate transfer tube where high pressures of air swept the test material into the exposure chamber. Chamber concentrations of silica or iron were maintained by controlling the dust-feed rate into the generation apparatus, or by varying the air-flow rate.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
For gravimetric analysis, samples of atmospheric carbonyl iron or silica were taken from the animal breathing zone at approximately 30-min intervals by drawing calibrated volumes of chamber atmosphere through preweighed glass-fiber filters. Filters were w
Duration of exposure:
28 d
Remarks on duration:
4 weeks, 5 days/week, 6 h/day
Concentrations:
Nominal: 0, 5, 50 and 250 mg/m^3; actual: 4.8, 51.8, 243.5 mg/m^3; MMAD (mass median aerodynamic diameter): 3.4, 3.2 and 2.9 micrometer, respectively.
No. of animals per sex per dose:
No data
Control animals:
yes
Details on study design:
The following endpoints were studied in the exposed rats.
- Presence of iron in the lungs;
- Particles in alveolar macrophages obtained by means of bronchoalveolar lavage (BAL);
- Measures for inflammation in BAL;
- Lactate dehydrogenase and protein in BAL;
- Cell proliferation in lungs by means of BrdU labelling;
- Lung clearance;
- Translocation of particles to tracheobronchial lymph nodes;
- Functional responses of alveolar marcophages;
- Lung histopathology.
Statistics:
Two-tailed Student t test

Results and discussion

Preliminary study:
No preliminary study performed
Effect levels
Sex:
male
Dose descriptor:
LC50
Effect level:
> 250 mg/m³ air
Exp. duration:
6 h
Remarks on result:
other: 6 h/day, 5 days/week, 4 weeks
Mortality:
Although the presentation of the results does not explicitly states anything about mortality, the exposure for 4 weeks, 5 days/week and 6 h/day to up to 250 mg/m^3 and the extensive description of sublethal effects clearly indicates that no treatment-related mortality occurred. From this study it can safely be concluded that 6 h exposure to 250 mg/m^3 respirable iron powder (carbonyl iron) does not result in mortality.
Clinical signs:
other: No data
Body weight:
No data
Gross pathology:
No data
Other findings:
- Histopathology: The publication presents an extensive and detailed descriptions of the histopathological observations made in the lungs, which was summarized in the abstract as follows (slightly modified). "Free granular pigment (CI = carbonyl iron) was present on the hypertrophic mucosal surfaces of bronchioles and bronchi, and particle-laden macrophages, found individually, were numerous throughout alveoli and within lymphoid tissues immediately after exposure. Aggregates of particle-laden macrophages were present within alveoli and alveolar ducts from 1 week postexposure through the entire 6-month recovery period. Macrophage accumulations increased in size and number from 1 week through 1 month postexposure and then appeared to remain constant through the remaining 5-month postexposure period. Minimal cellular hypertrophy and hyperplasia were evident at alveolar duct bifurcations adjacent to macrophage aggregates, and this effect was most prominent at 3 to 6 months postexposure." These observations were largely restricted to 50 mg/m^3 and 250 mg/m^3.
- Potential target organs: The study was focused entirely on the effects of the exposure on the lungs, in particular effects associated with clearance and inflammation.
- Other observations: What follows is a quote from the abstract of the publication (slightly modified). "Four-week exposures to Ti02 or CI particles at concentrations of 250 mg/m3 resulted in lung burdens of 12 mg titanium and 17 mg iron, respectively, with particle retention half-times ranging from 68 days for 5 mg/m3 Ti02 to approximately 330 days for 250 mg/m3. The impact of this Ti02 dust load and similar lung burdens of CI particles produced a sustained pulmonary inflammatory response measured through a period of 3-6 months postexposure concomitant with increases in BrdU cell labeling of terminal airway and pulmonary parenchymal cells. The impairment of particle clearance mechanisms was accounted for by deficits in in vitro phagocytic and chemotactic potential of alveolar macrophages recovered from the lungs of high-dose, TiO2 or CI-exposed rats." In another part of the publication the observations made are summarized as:
- Pulmonary inflammation;
- Enhanced proliferation of pulmonary cells;
- Impairment of particle clearance mechanisms;
- Deficits in in macrophage function;
- Macrophage aggregation.
No signs of inflammation or affected clearance were found at 5 mg/m^3, while these effects were found at 50 and 250 mg/m^3.

Applicant's summary and conclusion

Interpretation of results:
other: Highest concentration tested was 250 mg/m^3; so no final classification possible.
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
The study showed that exposure of rats to air concentrations of carbonyl iron up to 250 mg/m^ for 6 h/day, 5 days/week, 4 weeks, does not result in mortality. So an LC50 of > 250 mg/m^3 is established. Fifty and 250 mg/m^3 gave rise to clear effects on particle clearance and resulted in inflammation. Such effects were not found at 5 mg/m^3.
Executive summary:
Although not specifically aimed at establishing the acute inhalation toxicity of iron particles as is done in a standard guideline study, this study shows that up to 250 mg/m^3, respirable iron particles (carbonyl iron) do not cause mortality in males rats when the animals were exposed for 6 h/day, 5 days/week over a period of 4 weeks. So the LC50 lies clearly above the highest air concentration applied of 250 mg/m^3. The other aspects of the study are summarized adequately in the abstract of the original paper, which is, therefore, reproduced below.

This study was carried out to assess the time course of pulmonary clearance impairment and persistence of inflammation following high-dose inhalation exposures to titanium dioxide (Ti02) or carbonyl iron (CI) particles. Male rats were exposed to air, Ti02 or CI particles 6 hr/day, 5 days/week, for 4 weeks at concentrations of 5, 50, and 250 mg/m3and evaluated at selected intervals through 6 months postexposure. Indices of pulmonary inflammation as well as alveolar macrophage clearance functions (i.e., morphology, in vivo and in vitro phagocytosis, and chemotaxis), cell proliferation, and histopathology endpoints were measured at several post­exposure time periods through 6 months. In addition, amounts of TiO2 or CI in lungs and tracheobronchial lymph nodes were measured to allow an evaluation of particle clearance and translocation patterns. Four-week exposures to TiO2 or CI particles at concentrations of 250 mg/m3resulted in lung burdens of 12 mg titanium and 17 mg iron,respectively, with particle retention half-times ranging from 68 days for 5 mg/m3TiO2 to approximately 330 days for 250 mg/m3. The impact of this TiO2 dust load and similar lung burdens of CI particles produced a sustained pulmonary inflammatory response measured through a period of 3 -6 months postexposure concomitant with increases in BrdU cell labeling of terminal airway and pulmonary parenchymal cells. The impairment of particle clearance mechanisms was accounted for by deficits in in vitro phagocytic and chemotactic potential of alveolar macrophages recovered from the lungs of high-dose, TiO2or CI-exposed rats. Free granular pigment (TiO2or CI) was present on the hypertrophic mucosal surfaces of bronchioles and bronchi, and particle­laden macrophages, found individually,were numerous throughout alveoli and within lymphoid tissues immediately after exposure. Aggregates of particle-laden macrophages were present within alveoli and alveolar ducts from 1 week postexposure through the entire 6 -month recovery period. Macrophage accumulations increased in size and number from 1 week through 1 month postexposure and then appeared to remain constant through the remaining 5-month postexposure period. Minimal cellular hypertrophy and hyperplasia were evident at alveolar duct bifurcations adjacent to macrophage aggregates, and this effect was most prominent at 3 to 6 months postexposure. The results of this study clearly demonstrate that exposure to high dust concentrations of two different innocuous particle types produced sustained pulmonary inflammation, enhanced proliferation of pulmonary cells, impairment of particle clearance, deficits in macrophage function, and the appearance of macrophage aggregates at sites of particle deposition. In addition,the mass deposition rate determination appears to be a less sensitive indicator of"overload when compared to biomarkers of pulmonary toxicity, such as macrophage function and cellular inflammation and proliferation indices.

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