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Administrative data

short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2 (reliable with restrictions)

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 203 (Fish, Acute Toxicity Test)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Reference substance name:
2237 JG
2237 JG
Test material form:
other: white solid
Details on test material:
Batch 2237 JG; Expiry date: June 2012

Sampling and analysis

Analytical monitoring:
Details on sampling:
The stability of tested concentrations in WAF samples was checked in fresh and old media.

Test solutions

Details on test solutions:
A special solubilisation protocol was carried out before the experimentation according to guidance document OECD 23 on aquatic toxicity testing of difficult substances and mixtures. This method has consisted in stirring three test item nominal concentrations (1, 10 and 100 mg/L) in test dilution water during 24 hours, at obscurity and 20+/-2°C (each concentration was prepared by adding directly the test item in a flask containing test solution). Each test solution plus a dilution water control have been filtered through a membrane filter to remove suspended particles; solutions obtained were named WAF (Water Accommodated Fraction).

Test organisms

Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
Danio rerio (Hamilton Buchanan) commonly named Zebra fish, of a length of 25 to 30 mm, free from any apparent malformations and diseases.
Every batch of fish received in the laboratory is held in the dilution water for a minimum time period of 12 days before testing.

Study design

Test type:
Limit test:
Total exposure duration:
96 h
Post exposure observation period:

Test conditions

250 mg/L as CaCO3
Test temperature:
Dissolved oxygen:
79% to 95% of saturation during the test.
Nominal and measured concentrations:
Nominal concentration: 1 mg/L test item
Measured concentration at T0: < 0.075 mg/L APG
Measured concentration at T24h (before renewal) - biotic samples: < 0.075 mg/L APG (below LOQ)
Measured concentration at T24h (before renewal) - abiotic samples: < 0.075 mg/L APG (below LOQ)

Nominal concentration: 10 mg/L test item
Measured concentration at T0: 0.87 mg/L APG
Measured concentration at T24h (before renewal) - biotic samples: 0.71 mg/L APG (-18.4%)
Measured concentration at T24h (before renewal) - abiotic samples: 0.61 mg/L APG (-29.9%)

Nominal concentration: 100 mg/L test item
Measured concentration at T0: 5.88 mg/L APG
Measured concentration at T24h (before renewal) - biotic samples: 5.42 mg/L APG (-7.8%)
Measured concentration at T24h (before renewal) - abiotic samples: 4.88 mg/L APG (- 17.0%)
Details on test conditions:
Lethality was recorded every 24 hours and compared with control values.
The solutions were renewed at 24h, 48h and 72h.
The test contained a total of 5 aquariums, one for each test solutions, including both control.
For each aquarium containing 2.0 litres of test solution, 7 fish were used in order to respect a loading of 1 g/L (corresponding to the ratio of fish/volume of test solution recommended by the OECD guideline)
The dead fish were reported every 24 hours and removed as soon as possible. Fish is considered dead if there is no visible movement (e.g. gill movement) and if touching of the caudal peduncle produces no reaction. The visible abnormalities (loss of equilibrium, swimming behaviour, respiratory function, pigmentation, etc...) were reported if need be.
Temperature, pH and dissolved oxygen were measured every 24 hours in each aquarium during the test duration.
Reference substance (positive control):
potassium dichromate

Results and discussion

Effect concentrations
96 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
No dead fish was observed after 96h at the three nominal concentrations (1 mg/L, 10 mg/L and 100 mg/L).
Results with reference substance (positive control):
The toxic response obtained on the fish with the reference item (K2Cr2O7, batch 241011 ) gives a LC50-24h of 210 mg/L (173-254 mg/L) which was in good accordance with the historical data obtained by the laboratory.

Applicant's summary and conclusion

Validity criteria fulfilled:
In the conditions of the test (semi-static design with WAF approach), the LC50-96h on fish Danio Rerio was > 100 mg/L (nominal) for 2237 JG.
Executive summary:

In a semi-static acute toxicity test performed according to OECD guideline 203, zebra fish (Danio rerio) were exposed for 96 hours to 3 nominal concentrations ranging between 1 and 100 mg 2237 JG/L water.

Since the test item was insoluble in the tested medium, the use of Water Accommodated Fractions (WAF) for the preparation of the test media was necessary.

The test item stability was checked by the determination of Alkyl Poly Glucosid (APG) in fresh and old WAF (GC-FID; non GLP data). Actual concentrations of APG were in good agreement with the nominal concentrations.

After 96 hours of exposure, no mortality was observed at the three test concentrations.

The 96 hours LC50of 2237 JG to the zebra fish (Danio rerio) was > 100 mg/L (nominal).