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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
disregarded due to major methodological deficiencies
Study period:
26.11.1999 - 14.03.2000
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: OECD guideline study, GLP. Irrgegularities in the analytical quanatification of the test substance (analysis of the biological control sample revealed the presence of test substance).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
adopted 1984
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
For the analysis of the actual test Item concentrations the following samples were taken:

Just before test start:
- duplicate samples from each test medium (without algae)
- duplicate samples from the control (without algae)

After 72 hours (stability samples):
- duplicate samples from each test medium (without algae)
- duplicate samples from the control (without algae)
For the 72-hour stability samples additional flasks with adequate volumes of the freshly prepared test media of all test concentrations and the control were Incubated under the same conditions as in the actual test but without algae.

The samples from the undiluted filtrate at test start were analyzed directly after sampling, while the dilutions and all stability samples after 72 hours were deep-frozen (at about -20 °C)
immediately after sampling.

The concentrations of the test substance were analyzed in all test medium samples from both sampling times (0 and 72 hours). From the control samples only one of the duplicate samples was analyzed from each of both sampling times.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

The undiluted filtrate of the supersaturated stock dispersion with the maximum concentration of dissolved test substance was used as the highest concentrated test medium. Additionally, adequate volumes of the filtrate were added to test water for the preparation of test media with lower test item concentrations. No additional dilution step was inserted. The following dilutions were tested: 1:16, 1:8, 1:4 and 1:2 (one part of undiluted filtrate added to 15, 7, 3 and one part of test water), resulting in loading rates of 6.25, 12.5, 25 and 50 mg/L. Additionally, a control was tested in parallel (test water without test item). The test media were prepared just before addition of algae (=start of the test).

- Controls: test water without test substance

Test organisms

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Scenedesmus subspicatus CHODAT
- Strain: No. 86.81 SAG
- Source (laboratory, culture collection): Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universltät Göttingen, D-37073 Göttingen, Germany
- Age of inoculum (at test initiation): The test was started (0 hours) by Inoculation of 10,000 algal cells per ml test medium. These cells were taken from an exponentially growing pre-culture, which was set up 4 days prior to the test at the same conditions as in the test. One day prior to the test the pre-culture was diluted fourfold to keep the algae in exponential growth.
- Method of cultivation: The algae were cultivated and tested In synthetic test water, prepared according to the mentioned test guidelines.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h

Test conditions

Hardness:
0.24 mmol/L (= 24 mg/L) as CaCO3
Test temperature:
22 °C
pH:
between 8.1 to 8.4
Dissolved oxygen:
No data.
Salinity:
No data.
Nominal and measured concentrations:
Nominal:
The undiluted filtrate of a supersaturated stock dispersion (loading rate of 100 mg/L) with the maximum concentration of dissolved test substance was used as the highest concentrated test medium. Additionally, the dilutions 1:16, 1:8, 1:4 and 1:2 of the filtrate with loading rates of 6.25, 12.5, 25 and 50 mg/l, and a control were tested.

Measured:
The analytically measured concentration of the test substance in the undiluted filtrate was 0.092 mg/L. This concentration was within the range of the water solubility of the test substance in the used test water. The measured concentrations in the lower concentrated test media with the loading rates of 6.25, 12.5, 25 and 50 mg/L were 0.006, 0.011, 0.021, and 0.043 mg/L at the start of the test. After the test period of 72 hours, the test substance concentrations had decreased, probably caused by adsorption of the test item to surfaces at the low concentrations. The biological results are related to the loading rates of 6.25 to 100 mg/L with total mean measured test substance concentrations of 0.004 to 0.070 mg/L (calculated as the average over all measurements in the respective filtrates during the test period).
Details on test conditions:
TEST SYSTEM
- Test vessel: 50 mL Erlenmeyer flasks.
- Material, size, headspace, fill volume: 15 mL
- Aeration: no data
- Initial cells density: 10,000 algal cells per mL test medium.
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The algae were cultivated and tested In synthetic test water.
- Culture medium different from test medium: no
- Intervals of water quality measurement: The pH was measured in each test concentration and the control at the start and end of the test. The water temperature was measured daily in an Erienmeyer flask filled with water and incubated under the same conditions as the test flasks. The appearance of the test media was recorded dally.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuously illuminated
- Light intensity and quality: about 8000 Lux


EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: electronic particle counter
- Chlorophyll measurement: no
- Other: In addition, after the test period of 72 hours, a sample was taken from the control and from the highest test concentration (undiluted filtrate). The shape of the algal cells was microscopically examined.

TEST CONCENTRATIONS
The test concentrations were based on the results of a range-finding test and the results of the first experiment.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 0.07
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
6.25
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.004
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape: not observed
- Colour differences: not observed
- Flocculation: not observed
- Adherence to test vessels: not observed
- Aggregation of algal cells: not observed

Results with reference substance (positive control):
Not applicable.
Reported statistics and error estimates:
NOEC determined by Dunnett´s test and EC50 with probit analysis.

Any other information on results incl. tables

Summary of analytical results:

Analysis of the aged biological control sample revealed a test item concentration of 0.0046 mg/L although, no test substance was present. There is no obvious explanation for this value.

 

Applicant's summary and conclusion