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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09.02.-23.02.2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
The samples of test solutions were prepared and delivered by Ecotoxicity group. The samples were analyzed on the day of delivery. The samples were analysed non-diluted.
Details on test solutions:
TThe stock solution of the test substance was prepared in the test medium. 100 mg of the test substance was weighed into 1000 mL of the test medium for the preliminary and limit test. The stock solutions for the preliminary and limit test were ultrasonicated for 15 minutes. The concentrations of test solutions used in the preliminary test were obtained by dilution of the stock solution with test medium.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM- Common name: Desmodesmus subspicatus- Source (laboratory, culture collection): Brinkmann 1953/SAG 86.81 obtained from the collection of autotrophic organisms of The Botanic Institute of the Czech Academy of Science, Třeboň on date 3.6.2014.- Age of inoculum (at test initiation): Inoculum culture was kept 3-4 days under conditions at which thetest was performed.- Method of cultivation: The strain culture was preinoculated from the stock solution and cultivated in flasks with the test medium on indirect daylight at laboratory temperature.ACCLIMATION- Acclimation period: 3-4 days- Culturing media and conditions: same as test
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Test temperature:
21-24°C±2°C
pH:
7.6-7.7
Conductivity:
5 µS.cm-1
Nominal and measured concentrations:
100, 50, 10, 5, 1 mg/l
Details on test conditions:
TEST SYSTEM- Test vessel: Erlenmayer flask with stoppers venting air- Material, size, headspace, fill volume: glass, 50 ml- Aeration: without- Initial cells density: 5000 cells in 1 ml- Control end cells density:-No. of vessels per concentration (replicates): 3OTHER TEST CONDITIONSTemperature: 21 - 24 °C controlled at ± 2 °CLighting: under continuous white light, 4 440 lux to 8 880 luxExposition time: 72 hoursVolume of tested mixture: 50 mLInitial concentration of algae culture: 5 000 cells in a 1 mLWithout aeration.Agitation of algae culture by shaking.The light intensity and temperature were measured every 24 hours. The density of algae culture wasevaluated microscopically at 24, 48 and 72 hour.The cell density was measured by direct counting ofliving cells in Burker´s counting chamber.
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 52.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
52.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 52.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
52.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Results with reference substance (positive control):
The sensitivity of the test species and correctness of test performance in our laboratory is periodically verified on a six-month period by testing the reference substance, potassium dichromate.The results of the verification test with K2Cr2O7, carried out in period from 10.10. to 13.10. 2016, are the following: 72 hour – ErC50 = 1.05 mg.L-1 (95% confidence limit: 0.54-1.26 mg.L-1)
Validity criteria fulfilled:
yes
Conclusions:
The inhibition of growth rate was 1.3 % and inhibition of yield was 4.5 % in the limit test.Therefore, exact values of ErC50 and EyC50 could not be calculated and the values of EC are given in the form of a range.The determination of NOEC value was done by ANOVA (Analysis of Variance) analysis. Used ANOVA method is the part of statistical software QC.Expert 2.5 © 1998-2000 (product of TriloByte Ltd., Czech Republic). Because concentrations of the test substance have not been satisfactorily maintained within ± 20 per cent of the nominal or measured initial concentration throughout the limit test, the geometric mean of measured concentrations was used for results presentation and evaluation. 72 hour – ErC50 > 52.8 mg·L-1 (geometric mean of measured concentrations)72 hour – NOECr = 52.8 mg·L-1 (geometric mean of measured concentrations)72 hour – EyC50 > 52.8 mg·L-1 (geometric mean of measured concentrations)72 hour – NOECy = 52.8 mg·L-1 (geometric mean of measured concentrations)
Executive summary:

The test substance, Acid Yellow 25, was tested for growth inhibition on algae Desmodesmus subspicatus.

The test was performed according tomethod C.3. -Freshwater Algae and Cyanobacteria, Growth Inhibition Test,Commission Regulation (EU) No. 2016/266.The preliminary test was performed in a range of the test substance nominal concentrations
1 – 100 mg·L-1.

The highest inhibition of growth rate was 1.4 % and the highest inhibition of yield was 5.0 % in the preliminary test. Based on no toxicity of the test substance found in the preliminary test, the limit test was performed subsequently.

The concentration of 100 mg·L-1was tested in the limit test. The inhibition of growth rate was 1.3 % and inhibition of yield was 4.5 %.

In the preliminary test, the precipitation of the test substance in the test medium was observed.The precipitation occurred during shaking the samples on a laboratory shaker.

The analytical results in the preliminary test were incorrect due to precipitation of the test substance solutions (in solution there were visible agglomerates of the test substance).
At the end of the test in the highest nominal concentration (100 mg·L-1) the concentration 170.6 mg·L-1was determined. This high value was probably due to the precipitation of the test substance (the portion of sample which contained precipitate was taken for analysis).

Therefore, in the limit test the samples for analysis were taken from the clear solutions.

The analytical results showed that the test substanceAcid Yellow 25was not stable in the test medium during preliminary and limit test.

The guideline specifies that if evidence is available to demonstrate that the concentration of the test substance in limit test has been satisfactorily maintained within ± 20 per cent of the nominal or measured initial concentration throughout the limit test, then the results can be based on nominal or measured initial values.

Because the concentrations of test substance have not been satisfactorily maintained within
± 20 per cent of the nominal or measured initial concentration throughout the limit test, the geometric mean of measured concentrations was used for results presentation and evaluation.

72 hour – ErC50 > 52.8 mg·L-1 (geometric mean of measured concentrations)

72 hour – NOECr = 52.8 mg·L-1  (geometric mean of measured concentrations)

72 hour – EyC50 > 52.8 mg·L-1  (geometric mean of measured concentrations)

72 hour – NOECy = 52.8 mg·L-1  (geometric mean of measured concentrations)

Description of key information

The test was performed according tomethod C.3. -Freshwater Algae and Cyanobacteria, Growth Inhibition Test,Commission Regulation (EU) No. 2016/266.

Only one study is available.

GLP study.

Klimish score 1.

Key value for chemical safety assessment

EC50 for freshwater algae:
52.8 mg/L

Additional information