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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:
1987
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study. All relevant validity criteria were met.
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
Equivalent OECD guideline as effective in 1988.
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
not specified
GLP compliance:
not specified
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0 (control), 10, 20, 35, 50 and 100 ppm. Samples from control replicates and test group replicates of equal nominal concentration were pooled for analysis.
- Sampling method: Samples were taken at 0 h, 24 h, 48 h, 72 h and 96 h for quantitative analysis. Samples were analysed using an electronic particle-counter after removal of a 2ml portion and dilution of the 2ml portion 10 times with Isoton II. At the sampling, flasks were shaken slowly to stir the contents and randomly repositioned each day to minimise the spatial differences on the incubation table.
- Sample storage conditions before analysis: Samples were analysed immediately.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Direct dissolution of test item in culture medium to prepare stock; serial dilution of stock to prepare test media.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Not applicable.
- Concentration of vehicle in test medium (stock solution and final test solution): Not applicable.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): No precipitate reported.
Test organisms (species):
other: Selenastrum Capricornutum and Chlorella Vulgaris - both recommended in the OECD test guideline.
Details on test organisms:
TEST ORGANISM
- Common name (first organism): Selenastrum Capricornutum
- Common name (second organism): Chlorella Vulgaris
- Source (laboratory, culture collection): Laboratory stock cultures: S. Capricornatum sourced from National Institute for Environmental Studies, Tsukuba, Japan; C. Vulgaris sourced from Applied Microbiology, University of Tokyo, Tokyo, Japan.
- Method of cultivation: Master cultures were maintained in the laboratory by incubation at 21 ± 1 ºC temperature and continuous illumination of 4000 lux, for 4 or 5 d in Gorham's medium (pH 7.5) with the addition of Gaffron trace element solution.

ACCLIMATION
- Acclimation period: No. However, prior to the start of the test exposure sufficient master culture was added to approximately 100 ml culture medium in flasks at 5x10^4 cells/mL.
- Culturing media and conditions (same as test or not): Yes.
Culture medium: (mg/L) - equivalent or similar to AAP medium, according to OECD 201, Annex 3. pH 7.5.
- Any deformed or abnormal cells observed: None reported.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Test temperature:
21 ± 1ºC
pH:
0-hours pH: 7.5
Nominal and measured concentrations:
Nominal Concentrations were: 0 (control), 10, 20, 35, 50 and 100 mg/L
Nominal concentrations were considered appropriate for effect level calculations.
Details on test conditions:
TEST SYSTEM
- Test vessel: 500 mL glass conical flasks
- Type (delete if not applicable): Closed – Static. vessels plugged with foam bungs.
- Headspace, fill volume: 100 mL fill volume, approximately 400 mL headspace
- Aeration: Vessel shaken continuously.
- Renewal rate of test solution (frequency/flow rate): Not applicable.
- Initial cells density: nominal: 5x10^4 cells/mL
- Control end cells density: Not specified.
- No. of vessels per concentration (replicates): 3 replicates of each test concentration
- No. of vessels per control (replicates): 3 replicates of the controls.
- No. of vessels per vehicle control (replicates): Not applicable.

GROWTH MEDIUM
- Standard medium used: Yes. Equivalent or similar to AAP medium, according to OECD 201, Annex 3. Gorham's medium with Gaffron trace element solution added, pH 7.5.

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: No.
- Intervals of water quality measurement: Not applicable.

OTHER TEST CONDITIONS
- Sterile test conditions: No; however culture flasks had been sterilised prior to use.
- Adjustment of pH: No.
- Photoperiod: 24 hours ; continuous
- Light intensity and quality: Intensity approximately 4000 lux; wavelength(s) not specified.
- Salinity (for marine algae): Not applicable.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Coulter(R) TA-II particle counter

TEST CONCENTRATIONS
- Spacing factor for test concentrations: definitive test concentrations justified from the results of the range finding study.
- Range finding study: Yes.
- Test concentrations: Three replicates per concentration were exposed to 10, 20, 35, 50 and 100 ppm and control based on nominal concentrations.
- Results used to determine the conditions for the definitive study: Yes. Growth observed to be reduced at 10 ppm level.
Reference substance (positive control):
not specified
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
38 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: S. Capricornutum
Remarks:
S. Capricornutum
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
29 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: C. vulgaris
Remarks:
C. vulgaris
Details on results:
- Exponential growth in the control (for algal test): Yes.
- Observation of abnormalities (for algal test): None reported.
- Unusual cell shape: Not reported.
- Colour differences: Not reported.
- Flocculation: Not reported.
- Adherence to test vessels: Not reported.
- Aggregation of algal cells: Not reported.
- Other:
- Any stimulation of growth found in any treatment: Not reported in the range finder or definitive test.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not reported.
- Effect concentrations exceeding solubility of substance in test medium: Not reported.
Results with reference substance (positive control):
- Results with reference substance valid?: Yes.
- EC50: The EC50 was determined to be 38mg/L for S. Capricornutum and 29mg/L for C. Vulgaris.
- Other: DMSO was used as a solvent vehicle to dissolve 3-chlorophenol and higher, at a maximum concentration of 0.3%. DMSO exhibited no observable effect on algal growth in confirmatory experiments.
Validity criteria fulfilled:
yes
Conclusions:
The test item 96h-EC50 was 38 mg/L (S. Capricornutum) and 29 mg/L (C. Vulgaris), based on analytically confirmed nominal concentrations.
Executive summary:

The algal growth inhibition to Selenastrum Capricornutum and Chlorella Vulgaris, was carried out according to OECD TG 201 Freshwater Alga and Cyanobacteria, Growth Inhibition Test, equivalent or similar to EU Method C.3 guidelines. The aim of the study was to assess the effects on growth rate and yield over a period of 96 hours. Following a range finding study, a definitive study was conducted at 0 (control), 10, 20, 35, 50 and 100 mg/L conditions with an initial cell density nominal of 5x10^4 cells/mL. Three replicates were tested for each test item concentration and three replicates for the control under constant illumination and shaking at a temperature of 21 ± 1 °C. Environmental conditions were within the acceptable limits. The validity criteria of the test guideline were fulfilled. After 96 hours with algae, the EC50 for growth rate reduction (96h-EC50) based on nominal concentrations was 38 mg/L for S. Capricornutum and 29 mg/L for C. Vulgaris.

Description of key information

EC50 (algae; growth rate) = 38 mg/L, 96hour-freshwater, OECD TG 201, 1988 (Selenastrum Capricornutum)

EC50 (algae; growth rate) = 29 mg/L, 96hour-freshwater, OECD TG 201, 1988 (Chlorella Vulgaris)

Key value for chemical safety assessment

EC50 for freshwater algae:
29 mg/L

Additional information

The algal growth inhibition to Selenastrum Capricornutum and Chlorella Vulgaris, was carried out according to OECD TG 201 Freshwater Alga and Cyanobacteria, Growth Inhibition Test, equivalent or similar to EU Method C.3 guidelines. The aim of the study was to assess the effects on growth rate and yield over a period of 96 hours. Following a range finding study, a definitive study was conducted at 0 (control), 10, 20, 35, 50 and 100 mg/L conditions with an initial cell density nominal of 5x10^4 cells/mL. Three replicates were tested for each test item concentration and three replicates for the control under constant illumination and shaking at a temperature of 21 ± 1 °C. Environmental conditions were within the acceptable limits. The validity criteria of the test guideline were fulfilled. After 96 hours with algae, the EC50 for growth rate reduction (96h-EC50) based on nominal concentrations was 38 mg/L for S. Capricornutum and 29 mg/L for C. Vulgaris.