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Acute Toxicity: inhalation

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acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant, guideline study, unpublished report available, no restrictions, fully adequate for assessment

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guidelineopen allclose all
according to guideline
EPA OPP 81-3 (Acute inhalation toxicity)
equivalent or similar to guideline
OECD Guideline 403 (Acute Inhalation Toxicity)
equivalent or similar to guideline
EU Method B.2 (Acute Toxicity (Inhalation))
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:
solid: particulate/powder
migrated information: powder
Details on test material:
Sulfur Technical was supplied by Sandoz Agro Ltd. Three brown glass bottles with a total weight of 2.5 kg test material were received on November 25, 1993. Sulfur Technical is a yellow powder which has the following characteristics:
Batch no. : 1089 DLD
Purity : 100% (w/w)
Vapour pressure : 3.96 x 10-06 torr at 30.4°C
Density : 0.9 kg/l at 293 K
Storage conditions : dark at circa 20°C

Test animals

Details on test animals or test system and environmental conditions:
The animals used were male and female SPF-reared, Wistar derived (Crl:WI(WU)BR) rats delivered by Charles River, Wiga, Sulzfeld, FRG. Females were nulliparous and non-pregnant. The animals were 5-6 weeks old on arrival. On arrival they were taken in their unopened shipping containers to the quarantine room, and subsequently checked for overt signs of ill health and anomalies. Three days later the animals were transferred to the inhalation facilities and were housed in groups of five, separated by sex. The acclimatization period in the animal room was 31 days.
Just before the start of the exposures, the mean body weights of the male and female rats were respectively 295.8±2.0 g and 189.6±1.6 g. Before the start of the exposure the animals were coded with an earmark. From the arrival of the rats until the end of the study, the rats were fed the Institute's cereal-based, powdered stock diet for rats, mice and hamsters. Potable water for human consumption was supplied. Food and water were provided ad libitum. The nutrient composition of the diet and the levels of the various contaminants are regularly checked by the supplier in each batch of the stock diet and in samples of drinking water.

Before and after exposure the animals were housed under conventional conditions in suspended stainless steel cages fitted with wire-mesh floor and front. The number of air changes was about 10 per hour. The temperature was between 21 and 24°C, relative humidity was generally between 35 and 60%. A 12 hour light/dark cycle was maintained.

Administration / exposure

Route of administration:
Type of inhalation exposure:
nose only
other: unchanged (no vehicle)
Details on inhalation exposure:
Animals were exposed to the test atmosphere in a nose-only inhalation chamber. The chamber consisted of a cylindrical aluminium column, surrounded by a transparent PVC cylinder. The aluminium column had a volume of circa 50 liter, consisting of a top assembly with the entrance of the chamber, two mixing chambers, underneath the rodent tube section and at the bottom the base assembly with the exhaust port. The rodent tube section had 20 ports for animal exposure. Two or three ports were used for test atmosphere sampling, particle size analysis, and measurement of temperature and relative humidity. The animals were secured in plastic animal holders positioned radially through the outer cylinder around the central aluminium column. The rats were placed in an alternating order (male, female, male, etc). The remaining ports were closed. Only the nose of the rats protruded into the interior of the column.
The animal's body does not exactly fit in the animal holder which always results in some leak from high to low pressure side. By securing a positive pressure in the central colum and a slightly negative pressure in the outer cylinder, which enclosed the entire animal holder, dilution of test atmosphere by air leaking from the animal's thorax to the nose was prevented.
The air which passes through the nose-only chamber tank consisted of a mixture of air used in the generation device and air used for humidification. The pressure settings of the generation device (jetmill) were recorded 6 times, and they did not change during the exposure period (mill 4.5bar, jet 1.5 bar).
The mean total airflow was 80.7 l/min. Both temperature and relative humidity were monitored. They were recorded 5 times during exposure at regular intervals. Mean relative humidity and mean temperature were resp. 11.0±1.3% and 20.6±0.1°C.
The inhalation equipment was designed to expose rats to a continuous supply of fresh test atmosphere. A test atmosphere was generated by passing the test material using a dry material feeder tot a jet mill. The jet mill was operated with dry pressurized air. The test material was delivered using a slip stream of air conditioned room air. The slip stream accounted for about 36% of the total amount of air through the inhalation chamber.
The generated particles were passed to the exposure unit except for the fraction with larger particles which was captured using a settling chamber that was mounted to the mill. Before the inlet of the exposure unit the generated particles were mixed with humidified, pressurized air and the resulting test atmosphere was directed downward through the mixing chambers towards the animals. At the bottom of the unit the test atmosphere was exhausted.
Analytical verification of test atmosphere concentrations:
the actual concentration was determined 6 times during exposure at regular intervals by means of gravimetric analysis
Duration of exposure:
4 h
5.43 g/m3 (about 75% of the respirable particles had an aerodynamic diameter equal to or smaller than 4.2 µm (MMAD = 3.8 µm, GSD = 1.3))
No. of animals per sex per dose:
5 males and 5 females
Control animals:
Details on study design:
The study was started with one group of ten rats (5 males/5 females) that was exposed for 4 hours nose-only to a concentration of 5.43 g/m3. During exposure the animals were deprived of food and water and were housed individually in holders. Immediately after the exposure the fur around the head/neck area was cleaned gently using pressurized air. The animals were housed overnight in a hood seperately from the animal room. One day after exposure the animals were returned to their living cages, and were held there for the rest of the observation period (two weeks).

Observations and measurements:
Clinical observations: the rats were visually inspected just before exposure, for reactions to treatment during the exposure, shortly after exposure, and at least once daily during the observation period.
Body weights: recorded just prior to exposure (day 0), on days 2, 4, 7 and 14 and at death.
Complete gross necropsy (as no macroscopic changes were observed, the lungs were not processed for histopathological examination).
No statistical analysis was performed.

Results and discussion

Effect levels
Dose descriptor:
Effect level:
> 5.43 other: g/m3
Exp. duration:
4 h
Remarks on result:
other: 8 of 10 animals survived a 4 hour exposure of 5.43 g/m3
Two male animals died the first day of exposure (probably related to the very high, acute dust load).
Clinical signs:
other: A visually decrease in the breathing frequency was observed in all rats starting from the second hour of exposure. Irregular breathing was observed in all rats in the last two hours of exposure. In the first hour after exposure all rats showed choking beh
Body weight:
Male rats showed a decrease in body weight on day 2, 4 and 7, females on day 2 and 4. Animals recovered in the second week and on day 14 all surviving rats showed bodyweight gain.
Gross pathology:
Males that died within one day of exposure showed a hydrothorax and red discoloured patches on the lung. At autopsy on day 14 two female rats showed grey or pale discoloured lungs. No other macroscopic findings were observed.
Since no relevant macroscopic changes were observed at autopsy on day 14 no histopathological investigations were preformed.

Applicant's summary and conclusion

Interpretation of results:
not classified
Migrated information Criteria used for interpretation of results: EU
Exposure of rats to sulfur powdr for 4hrs at a concentration of 5.43 g/m3 did not result in any tretment related effects. The acute LC50 was > 5.43 g/m3.
Executive summary:

The 4 hour LC50 of sulfur was determined in a GLP study in accordance with EPA guideline OPP 81-3. One group of five male and five female rats was exposed to a concentration of 5.43 g/m3 during a single period of four hours. The animals were observed for 14 days after exposure, after which survivors were sacrificed and necropsied. Two animals died within the first day of exposure. The mortality was most probably due to the very high, acute, dust load and not the toxicity of the test material. The surviving animals recovered and within one week of exposure no clinical signs and changes in body weight gain were observed anymore. Since 8 of 10 treated animals survived, it was concluded that the 4 hour LC50 value was higher than 5.43 g/m3.