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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:
From 28 July 2016 to 5 August 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
The room temperature at 48 and 72 hours in the reference definitive test was above test conditions, by a maximum of 4.3°C. This deviation is not thought to have had a significant impact on the overall reference test result as it met the validity criteria.
Qualifier:
according to
Guideline:
ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)
Deviations:
no
GLP compliance:
yes
Test material information:
Composition 1
Analytical monitoring:
no
Details on sampling:
- Concentrations: 100, 180, 320, 560 and 1000 mg/L
- Sampling method: Test material stocks were prepared using dilution series - nominal concentrations were used.
- Sample storage conditions before analysis: Test material was stored at room temperature in the dark.
Vehicle:
yes
Remarks:
OECD culture medium
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Dilution series was used to prepare test substance.
- Eluate: N/A
- Differential loading: N/A
- Controls: A range-finding test was completed using this culture, this included controls
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N/A
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): N/A
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: 278/4
- Source (laboratory, culture collection): Stock laboratory cultures from SAMA Limited
- Age of inoculum (at test initiation): 2 - 4 days old
- Method of cultivation: Culture was prepared from stock laboratory cultures by inoculating autoclaved freshwater at 120 *C for 15 minutes with nutrient medium ISO 8692, 2 - 3 days before test commencement and incubated at approx. 21 +/- 2 *C under constant illumination. This was used as the inoculum source from subsequent toxicity tests.

ACCLIMATION
- Acclimation period: No acclimation period
- Culturing media and conditions (same as test or not): Yes, however the culture medium pH was adjusted if required
- Any deformed or abnormal cells observed: Not specified
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Remarks on exposure duration:
Algae growth observations were recorded at 48 hours and 72 hours
Post exposure observation period:
Not specified
Hardness:
Not specified
Test temperature:
21 - 24 *C
pH:
8.1 +/- 0.2control pH was increased during the test by no more than 1.5 relative to the pH of the growth medium
Dissolved oxygen:
Not specified
Salinity:
Not specified
Conductivity:
Not specified
Nominal and measured concentrations:
Nominal: 100, 180, 320, 560 and 1000 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 100ml borosilicate glass conical flasks
- Type (delete if not applicable): loosely covered with aluminium foil caps
- Material, size, headspace, fill volume: 50 ml
- Aeration: Some
- Type of flow-through (e.g. peristaltic or proportional diluter): N/A
- Renewal rate of test solution (frequency/flow rate): N/A
- Initial cells density: approximately 5 x 10(3) - 10(4) cells per millilitre
- Control end cells density: Not specified
- No. of organisms per vessel: approximately 5 x 10(3) - 10(4) cells per millilitre
- No. of vessels per concentration (replicates): 3 replicates per concentration
- No. of vessels per control (replicates): 6 controls

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: A 1000 mg/L stock was prepared in deionised water, and the resulting mixture was stirred for one hour and its behaviour assessed. As the material was observed to be soluble the material was prepared by dilution series. A main test material stock was made by weighing out an appropriate amount of test material, adding deionised water and stirring for approximately 1h or until completely dissolved. The test concentrations were then prepared by serial dilution of the main stock.
- Total organic carbon: Not specified
- Particulate matter: Not specified
- Metals: Not specified
- Pesticides: Not specified
- Chlorine: Not specified
- Alkalinity: Not specified
- Ca/mg ratio: Not specified
- Conductivity: Not specified
- Culture medium different from test medium: No
- Intervals of water quality measurement: Not specified

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: Yes
- If, at 0h the pH of the test material stock was out-with the pH range of 8.1±0.2, the pH was adjusted with either 1M HCl or NaOH as appropriate; following this, a stirring period was provided to ensure the pH remained constant.
- Photoperiod: 24 hour light
- Light intensity and quality: 4440-8880 lux

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: fluorimeter or haemocytometer or Sedgewick-Rafter chamber
- Chlorophyll measurement: Not measured

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.75
- Test concentrations: 1, 10, 100 and 1000 mg/L
- Results used to determine the conditions for the definitive study: Yes
- 72 hour response growth rate was used to determine conditions for definitive study
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
591 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
738 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
other: EC80
Effect conc.:
948 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
320 mg/L
Nominal / measured:
nominal
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): Not specified
- Unusual cell shape: Not specified
- Colour differences: Not specified
- Flocculation: Not specified
- Adherence to test vessels: Not specified
- Aggregation of algal cells: Not specified
- Any stimulation of growth found in any treatment: There was a slight increase of growth observed at 180 mg/L and 320 mg/L of glutaric acid
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not specified
- Effect concentrations exceeding solubility of substance in test medium: None
Results with reference substance (positive control):
- Results with reference substance valid? Yes - EC50: 1.35 mg/L
Reported statistics and error estimates:
48 hour and 72 hour EC50 values were calculated by non-linear regression within the CETIS suite of statistical analysis.

 Test concentration (mg/L)     48 h control growth rate  72 h control growth rate  48 hour test growth rate  72 h test growth rate
 100  A   1.79 1.71 1.75  1.76 
 B   1.79   1.71  1.73 1.72 
   C   1.79   1.71  1.66 1.74 
 180   1.79   1.71 1.72  1.80 
  B   1.79   1.71  1.73 1.80 
   C   1.79   1.71 1.75  1.73 
 320   1.79   1.71 1.75  1.81 
   B   1.79   1.71 1.78  1.81 
   C   1.79   1.71 1.77   1.75
 560  A   1.79   1.71 1.01  1.50 
   B   1.79   1.71 0.97   1.50
   C   1.79   1.71  0.90  1.41
 1000  A   1.79   1.71  0.00 0.00
    1.79   1.71  0.00  0.00 
   C   1.79   1.71  0.00  0.00
Validity criteria fulfilled:
yes
Conclusions:
The 72 hour EC50 was 738 mg/L. No growth of the algae was recorded at 1000 mg/L, when compared to the control groups. The test substance is not considered to be toxic to aquatic algae and cyanobacteria.
Executive summary:

The acute toxicity of the test material to aquatic algae was determined in accordance with the OECD Guideline 201 growth inhibition test. Nominal test concentrations of 100, 180, 320, 560 and 1000 mg/L of the test material were prepared based on the results of a dose-range finding study. The freshwater algae Pseudokirchneriella subcapitata was exposed to the test concentrations in order to determine the EC50 over 72 hours. The 72 hour EC50 of the test material was 738 mg/L. At 1000 mg/L, after 72 hours, the algae growth rate was recorded as 0, in comparison to the control groups. Glutaric acid is not considered to be toxic to aquatic algae and cyanobacteria.

Description of key information

The acute toxicity of the tested substance to aquatic algae was determined in accordance with the OECD Guidance 201 freshwater algae growth inhibition test. Nominal test concentrations of the test material were prepared over the range 100 to 1000 mg/l. The freshwater algae Pseudokirchneriella subcapitata was exposed to the test concentrations in order to determine the concentration which elicits a 50% response from the freshwater algae over 72 hours (EC50).

The lowest concentration of test material producing 0 growth rate was determine to be 1000 mg/L at 72 hours. All concentrations of test material produced some reduction in growth rate in this study. All parameters measured in this study lay within 95% confidence limits. The test material is not considered toxic to Pseudokirchneriella subcapitata within the conditions of this study. The registered substance is not considered to be toxic to aquatic algae based on the findings of this study.

Key value for chemical safety assessment

EC50/LC50 for freshwater algae:
738 mg/L
EC10, LC10 or NOEC for freshwater algae:
320 mg/L

Additional information