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Diss Factsheets

Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
27 Jun - 01 Jul 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Version / remarks:
adopted Mar 2006, corrected 28 Jul 2011
Deviations:
no
Qualifier:
equivalent or similar to guideline
Guideline:
EU Method C.3 (Algal Inhibition test)
Version / remarks:
2009
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Series on Testing and Assessment, N° 23, "Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures"
Version / remarks:
15 Dec 2000
GLP compliance:
yes (incl. QA statement)
Remarks:
Hess. Ministerium für Umwelt, Energie, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany (24 Sep 2013)

Test material

Constituent 1
Chemical structure
Reference substance name:
4-(2-phenylpropan-2-yl)phenyl 4-oxo-3-(λ⁵-diazynylidene)-3,4-dihydronaphthalene-1-sulfonate
Cas Number:
23121-00-8
Molecular formula:
C25H20N2O4S
IUPAC Name:
4-(2-phenylpropan-2-yl)phenyl 4-oxo-3-(λ⁵-diazynylidene)-3,4-dihydronaphthalene-1-sulfonate
Test material form:
solid

Sampling and analysis

Analytical monitoring:
no

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A supersaturated stock solution of 100 mg test item/L was prepared by dissolving 49.9 mg test item in 499 mL test water. The stock solution was ultrasonicated for 15 min and shaken in an overhead shaker for 23.5 h. The non-dissolved fractions of the test item were separated from the test medium by membrane filtration (0.45 µm cellulose acetate filter). The test media were prepared just before introduction of the algae ( = start of the test).
- Controls: Yes, in the control test water (reconstituted water: OECD medium) was used without addition of the test item.
- Evidence of undissolved material: There were no remarkable observations after filtration.

Test organisms

Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: unicellular green microalga
- Strain: n° 61.81 SAG
- Source: Sammlung von Algenkulturen, Göttingen, Germany
- Age of inoculum (at test initiation): 4 d old, from an exponentially growing pre-culture grown under the same conditions as in the test.
- Method of cultivation: The algae were cultivated in the laboratories of Ibacon under standardised condition according to the test guidelines.

ACCLIMATION
- Culturing media and conditions (same as test or not): Culturing conditions same as test conditions.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h

Test conditions

Hardness:
0.24 mmol/L = 24 mg/L (in reconstituted water: OECD medium)
Test temperature:
21.4 - 22.5 °C
pH:
Control: 8.0 (0 h), 9.4 (72 h)
Treatment: 7.8 (0 h), 9.4 (72 h)
Nominal and measured concentrations:
Control and 100 mg/L (nominal)
Details on test conditions:
TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Type: Covered with glass dishes.
- Material, size, headspace, fill volume: 50 mL glass Erlenmeyer flasks filled with 50 mL.
- Initial cells density: 5000 cells/mL test medium
- Control end cells density: Mean (72 h) = 82.658E+04 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: Yes, reconstituted water (OECD medium).

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: No.
- Intervals of water quality measurement: The water temperature was measured daily in an Erlenmeyer flask filled with water and incubated under the same conditions as the test flasks. pH values were measured in the single test concentration and the control at the start and the end of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: The pH of the OECD medium was adjusted to pH 8.0 with 1 M HCl.
- Photoperiod: Continuous illumination.
- Light intensity and quality: Mean light intensity = 5815 lux (5550 - 6110 lux)
- Other: Test media in the Erlenmeyer flasks were continuously stirred with magnetic stirrers and incubated in a water bath. The flasks were placed in a random order and were repositioned each day to minimize differences in test conditions.

EFFECT PARAMETERS MEASURED :
- Determination of cell concentrations: spectrophotometric measurements after 24, 48 and 72 h

TEST CONCENTRATIONS
- Range finding study: Yes
- Results used to determine the conditions for the definitive study: Non-GLP pre-experiments were performed to determine a suitable concentration range and to establish suitable methods to prepare the test solutions.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 0.073 mg/L
Nominal / measured:
estimated
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other:
Remarks:
Value refers to the max. water solubility of the test item calculated with the program EPI-Suite.
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat. (dissolved fraction)
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
> 0.073 mg/L
Nominal / measured:
estimated
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other:
Remarks:
Value refers to the max. water solubility of the test item calculated with the program EPI-Suite.
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat. (dissolved fraction)
Basis for effect:
growth rate
Details on results:
- Exponential growth in the control (for algal test): Yes, a 165.3-fold increase was observed within 72 h.
- Observation of abnormalities: No
- Any observations that might cause a difference between measured and nominal values: There were no remarkable observations of the test medium after filtration.
Results with reference substance (positive control):
- Results with reference substance valid? Yes, dating Feb 2016.
- EC50 (72 h): 1.5 mg Potassium Dichromate/L, 95% confidence interval: 1.43 - 1.58 mg/L (nominal)
Reported statistics and error estimates:
Based on the calculated cell densiities, the 72 h ErC50 and the 72 h EyC50 could not be calculated by Probit analysis due to the lack of effects and were therefore determined directly from the raw data.
For the determination of the 72 h LOEC and NOEC, the calculated growth rates and yields at each test concentration were tested for significant differences compared to the control values with a two sample t-test.
The software used to perfrom the statistical analysis was ToxRat Professional, Version 3.2.1, ToxRat Solutions GmbH.

Applicant's summary and conclusion