Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
18 May 2010 to 9 June 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study compliant with GLP and test guidelines and fulfils requirements of Annexes VII - X of Regulation (EC) 1907/2006 of 18 December 2006 concerning REACH

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Qualifier:
according to guideline
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Principles of method if other than guideline:
The maximum practical dose concentration was used. The target dose of 5 mg/L was not possible to achieve from the atmosphere generating system and the mean achieved exposure was limited to 1.306 mg/L air (4 h)
GLP compliance:
yes
Test type:
standard acute method
Limit test:
no

Test material

Constituent 1
Reference substance name:
Fumaric acid
EC Number:
203-743-0
EC Name:
Fumaric acid
Cas Number:
110-17-8
IUPAC Name:
but-2-enedioic acid
Details on test material:
- Name of test material (as cited in study report): Fumaric acid (trans-butendiacid or Fumarsäure)
- Molecular formula (if other than submission substance): C3H4O4

- Physical state: white crystalline powder
- Analytical purity: 99.7%
- Impurities (identity and concentrations): not stated
- Composition of test material, percentage of components: 99.7% fumaricacid
- Purity test date: 22 October 2009
- Lot/batch No.: LPFS6A4011 for analysis, also used for inhalation test.
- Expiration date of the lot/batch: 30 June 2010
- Stability under test conditions: stable
- Storage condition of test material: room temperature
- Other: solubility in water = 5 g/L at20 °C

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Crl:CD(SD)IGS BR, SPF sprague-Dawley rats obtained from Charles RIver Deutschland GmbH, Sulzfeld, Germany
- Age at study initiation: 8-9 weeks
- Weight at study initiation: male mean = 399.4g (range = 385 - 412g); female mean = 231.8g (range = 213 - 254g).
- Fasting period before study: not applicable
- Housing: Type III Makrolon cages with wire mesh lids
- Diet (e.g. ad libitum): Ssniff R/M-H rat and mouse maintenance diet
- Water (e.g. ad libitum):acidified tap water (adjusted to >pH 3 with HCl) via automated system, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Meam 19.7°C range 19.2 to 20.2°C
- Humidity (%): Mean 64% range 52 to 79% RH
- Air changes (per hr): 12/h
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 18 May 2010 To: 9 June 2010

Administration / exposure

Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
clean air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Nose only apparatus, using a two chamber system with tota volume of 19L. There are 20 ports in the chamber for insertion of rat holding /exposure tubes. Food and water withdrawn from rats during exposure.
- Exposure chamber volume: 19L
- Method of holding animals in test chamber: inhalation restraining tubes attached to the chamber
- Source and rate of air: Obtained from central pressure pump and delivered at rate of 1040 L/h
- Method of conditioning air: humidity reduced to circa 10% and filtered oil-free
- System of generating particulates/aerosols: a dust generator from TSE Systems GmbH was used according to Budiman. A powder cake, (5g pressed into asteel cylinder at 5 tonnes force, was used to fill the dust generator from which a scraper removed particles to be picked up by a clean air stream.
- Method of particle size determination: The size of the dust particles was analysed with a cascade impactor (Berner-Impaktor Type LPI4/0,06/2 from Hauke KG, Gmunden, Austria). It contains nine steps with cut-off diameters from 0.06 µm to 16 µm. 5.65 litres/minute of the test substance/air mixture were passed through the impactor within 2 minutes and the amount which settled on the individual steps was determined gravimetrically. The site of collection was the same as for the analysis - breathing zone. Calculation of the mean particle size (mass median aerodynamic diameter, MMAD), used the probit of the fraction of masses smaller then the cut-off diameters plotted against the logarithm of the cut-off diameters and the linear regression of this graph was calculated, preferring the data around 50 %. The diameter, where the regression gives a probit of 5 (corresponding to a fraction of 50 %) is the mass median diameter. The quotient of the diameter at a probit of 6 (corresponding to a fraction of 84 %) to the MMAD is the geometric standard deviation (GSD). .
- Treatment of exhaust air: extracted via fume cupboard
- Temperature, humidity, pressure in air chamber:

TEST ATMOSPHERE
- Brief description of analytical method used: gravimetric analysis - dust collection 8 times during course of exposure
- Samples taken from breathing zone: yes. The dust was collected 8 times during the exposure in plastic pipette-tips filled with cottonwool which were inserted into the inhalation facility through a separate hole between two inhalation tubes. The site of collection was within the outer chamber. The inner diameter of the tips was 7 mm. Accurately measured volumes of air with the dust were collected at a rate of 1.4 litres per minute which means a velocity of 0.7 m/sec in the tips. The exact amount of collected air was measured by a gas meter (Labor- Experimentiergaszähler, size 00, 2 to 120 L/h, Ritter Apparatebau GmbH, D-44892 Bochum). Each filter-tip was dried and weighed before sampling. After sampling dry air (< 10 % humidity) was passed through them until the weight was constant. The difference in the weights before and after sampling divided by the volume of air sampled is the
concentration of the dust.

VEHICLE
- Composition of vehicle (if applicable): filtered air
- Concentration of test material in vehicle (if applicable): nominal concentration = 34.5 mg/L. Achieved concentration was 1.306 mg/L or 3.8% of output

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: Since a concentration of 5 mg/L could not be reached, no separator for large particles was used as this would reduce somewhat also the small particles. From preliminary experiments it was known that grinding of the test substance also does not reduce the MMAD.
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): The calculated mass median aerodynamic diameter (MMAD) of the test substance dust in the two determinations was 15.54 and 11.92 µm, with a geometric standard deviation (GSD) of 3.40 and 3.13.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
The actual concentration of the test substance aerosol was analysed 8 times during each 4-hour exposure period.
Duration of exposure:
4 h
Concentrations:
The dust was collected 8 times during the exposure in plastic pipette-tips filled with cottonwool which were inserted into the inhalation facility through a separate hole between two inhalation tubes. The site of collection was within the outer chamber. The inner diameter of the tips was 7 mm. Accurately measured volumes of air with the dust were collected at a rate of 1.4 litres per minute which means a velocity of 0.7 m/sec in the tips. The exact amount of collected air was measured by a gas meter (Labor- Experimentiergaszähler, size 00, 2 to 120 L/h, Ritter Apparatebau GmbH, D-44892 Bochum). Each filter-tip was dried and weighed before sampling. After sampling dry air (< 10 % humidity) was passed through them until the weight was constant. The difference in the weights before and after sampling divided by the volume of air sampled is the concentration of the dust.
No. of animals per sex per dose:
Five
Control animals:
no
Details on study design:
Preliminary investigation:
Prior to commencing the main study, atempts were made to produce a dust atmosphere containg up to 5 ng test substance/L air.
First powder cakes with a force of 10 tons were produced. They were too hard and the scraper could not remove the powder but blocked. Powder cakes pressed with 5 tons were found to be suitable. The highest technically feasible dust concentration was 0.9 to 1.0 mg/L. The mean aerodynamic diameter (MMAD) of the dust was 8 to 12 µm and therefore higher than 4 µm. It was tried to reduce the MMAD by grinding the test substance in a ball mill (Pulverisette, 2 balls, 180 rpm, 10 minutes) and sieving through a 0.15 mm sieve. But this pre treatment did not reduce the MMAD and was omitted for the main experiment. Obviously the hardness of the test substance is too low and the force during the grinding forms new agglomerates.
As the limit concentration of 5 mg/L was not reached, also no separator for large particles was used as this would decrease also the smaller particles.

Several rats were then exposed to varying concentrations to determine potentail toxicity and to provide dose-ranging information.

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Behaviour, reactions and physical signs of each of the animals were observed 1, 2, 3, 4, 5 and 6 hours after the start of exposure, twice a day on the first day after exposure, and then at least once a day for a total of 2 weeks.
The individual body weight of each animal was determined at receipt, immediately before exposure, and 1, 3, 7 and 14 days p.a. Body weight gain was calculated between the measurements and in relation to the weight before administration.
- Necropsy of survivors performed: yes
Statistics:
No data

Results and discussion

Effect levelsopen allclose all
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 1.306 other: mg fumaric acid/L air
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
other: limit dose of 5mg/L could not be achieved. 1.306 mg/L was maximum feasible atmosphere concentration.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 1.284 other: mg succinic acid/L air
Based on:
other: read-across from fumaric acid
Exp. duration:
4 h
Remarks on result:
other: A read-across of the result for fumaric acid to succinic acid was performed, considering the slightly different molecular masses.
Mortality:
There were no deaths from exposure to 1.306 mg fumaric acid/L air in a four hour nose-only exposure study.
Clinical signs:
other: No clinical signs of reaction to treatment observed for any rat throughout the study.
Body weight:
Animals lost weight on day after exposure, probably attributable to the period of time spent in the exposure holding tube rather than attributable to treatment-related effects. Subsequent bodyweight gains, recorded for all rats, were satisfactory.
Gross pathology:
No macroscopic abnormalities noted during necropsy.
Other findings:
No sex related differences observed in this study.

Applicant's summary and conclusion

Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Fumaric acid did not elicit any indications of systemic toxicity following exposure of rats to 1.306 mg/L for 4 hours.

A read-across of the result for fumaric acid to succinic acid was performed, considering the slightly different molecular masses. The calculated LC50 is >1.284 mg/L for succinic acid.
Executive summary:

Fumaric acid was administered as a dust by inhalation to Sprague Dawley rats for 4 hours in a nose-only inhalation device. The target concentration for a limit test, 5 mg/L, could not be achieved. The maximum technically feasible concentration was administered - the analysed concentration of the dust in the atmosphere was 1.306 mg/L air.

Five male and five female rats were used. The chamber was 19 L in size and was operated with 1040 L air/dust per hour. Chamber temperature ranged from 19.6 to 21.7 °C, the relative humidity from 7.0 to 22.0 %. Methods and investigations were performed in accordance with the OECD Guideline 403 and with the Council Regulation (EC) No 440/2008, Method B.2.

All animals survived till the end of the study and no clinical signs of reaction to treatment were obseved.

Body weights were recorded prior to and at intervals following the day of the exposure. The animals lost weight the first day after the exposure (possibly due to restraint during exposure). Subsequently all animals gained weight. All animals were normal at the scheduled necropsy.

The mean actual dust concentration was 1.306 mg/L. A higher output was not possible for a 4 hour operation period. The calculated mass median aerodynamic diameter (MMAD) of the test substance dust in the two determinations was 15.54 and 11.92 µm, with a geometric standard deviation (GSD) of 3.40 and 3.13. But as the limit concentration of 5 mg/L could not be reached, no separator for large particles was used as this would reduce somewhat also the small particles. Grinding of the test substance did also not reduce the MMAD.

The inhalation exposure of rats to fumaric acid at the maximal technically feasible concentration of 1.306 mg/L did not produce signs of toxicity. All animals survived and no adverse effects were observed during the 14-day observation period. The acute median lethal concentration for four hour inhalation exposure (LC50) of fumaric acid for male and female rats was greater than the maximum practically achievable atmosphere concentration of 1.306 mg/L.

A read-across of the result for fumaric acid to succinic acid was performed, considering the slightly different molecular masses. The calculated LC50 is >1.284 mg/L for succinic acid.