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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:
8th December 2000 to 21 September 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted in a GLP compliant laboratory according to accepted regulatory guidelines
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
Method C3, 92/69/EEC.
Deviations:
no
GLP compliance:
yes
Details on sampling:
- Concentrations: 3.12, 6.25, 12.5, 25 and 50 mg/L
- Sampling method: Not specified. Duplicate water samples were taken from the control and each loading rate WAF test group (replicates Rl " R) pooled) at 0 and 72 hours for quantitative analysis.
- Sample storage conditions before analysis: Duplicate samples were taken at 0 hours and stored frozen (approximately -20"C) for further analysis if necessary, Sample volumes required for chemical analysis precluded the storage of duplicate samples at 72 hours.

Based on the variable results obtained from the pre-test recovery analyses, it was considered appropriate to analyse samples taken from the definitive study both untreated and after centrifugation (40000 G, 30 minutes), The results of the untreated analyses gave a measure of the total dissolved and dispersed test material in the WAF, whilst the centrifuged analyses gave a measure of the dissolved and hence bioavailable test material concentration.
Details on test solutions:
Identity and concentration of auxiliary solvent for dispersal: None
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Due to the low aqueous solubility and complex nature of the test material for the purposes of the definitive study the test material was prepared as a WAF.
Amounts of test material (15.6, 31.3, 31.3, 62.5, and 125 mg) were each separately smeared onto thc inside of a beaker prior to thc addition of culture medium (5, 5, 2.5, 2.5 and 2.5 litres respectively) to give the 3.12, 6.25, 12.5, 25 and 50 mg/L loading rates. These were then stirred by magnetic stirrer using a stirring rate such that a vortex depth of approximately 25% of the overall height of the water column was achieved. These were stirred for 23 hours. The stirring was
stopped after 23 hours and the mixtures allowed to stand for 1 hour prior to removing the aqueous phase or WAF by mid-depth siphoning to give the 3.12, 6.25,12.5,25 and 50mg/l loading rate WAFs.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Green algae
- Strain: Scenedesmus subspicatus CCAP 276/20
- Source (laboratory, culture collection): Reputable commercial supplier
- Age of inoculum (at test initiation): N./a
- Method of cultivation: Cultures were maintained in the laboratory by the periodic replenishment of culture medium

ACCLIMATION
- Acclimation period: N/a
- Culturing media and conditions (same as test or not): Yes
- Any deformed or abnormal cells observed: No
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Post exposure observation period:
Not applicable
Hardness:
100 mg CaCO3/L
Test temperature:
24 ± 1C
pH:
7.0 - 7.5
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL conical glass flasks
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 100 mL solution added
- Initial cells density:
- Control end cells density:
- No. of organisms per vessel:10000 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3
- No. of vessels per vehicle control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reverse osmosis purified deionised water
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine:
- Alkalinity:
- Ca/mg ratio:
- Conductivity:
- Culture medium different from test medium:
- Intervals of water quality measurement:

OTHER TEST CONDITIONS
- Sterile test conditions: yes/no
- Adjustment of pH:
- Photoperiod:
- Light intensity and quality:
- Salinity (for marine algae):

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]
- Chlorophyll measurement:
- Other:

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations:
- Results used to determine the conditions for the definitive study:
Key result
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
3.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Key result
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
3.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
3.12 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Validity criteria fulfilled:
yes
Conclusions:
The effect of the test material on lhe growth of Scenedesmus subspicatus has been investigated over a 72-Hour period and gave an EbL*50 (72 h) value of 3.4 mg/l loading rate WAF· and an ErL*50 (O-72h) value of 3.4mg/l loading rate ** . The No Observed Effect Loading rale at 72 hours was 3.12 mg/L loading rate WAF.
• EL = Effective Loading rate
•• It was not possible to calculate 95% confidence limits for the EL50 values as the data generated did not fit the models available for the calculation of confidence limits eg Litchfield and Wilcoxon, ProM, Logistic and Weilbulls models (Litchfield and Wilcoxon 1949).

Description of key information

Key value for chemical safety assessment

EC50 for freshwater algae:
3.4 mg/L
EC10 or NOEC for freshwater algae:
3.1 mg/L

Additional information

The EL50 and NOEL values refer to nominal loading levels resulting from water accomodated fractions (WAF). Water solubility of the substance is < 0.1 mg/l.