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EC number: 284-902-1 | CAS number: 84989-13-9
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Toxicity to aquatic algae and cyanobacteria
Administrative data
Link to relevant study record(s)
- Endpoint:
- toxicity to aquatic algae and cyanobacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 13 July 2018 to 30 August 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.3 (Algal Inhibition test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Date of Inspection: 18 July 2017 - 20 July 2017 Date of Issue: 28 November 2017
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: Control (0), 1,0, 3.2, 10, 32 and 100 mg/L
- Sampling method:
Samples were taken from the control and each test group from the bulk test preparation at 0 hours and from the pooled replicates at 72 hours for quantitative analysis. A set of duplicate samples were taken at 0 and 72 hours and stored frozen for further analysis if necessary.
An additional sample of each test concentration containing no algal cells was incubated alongside the test to provide samples for uninoculated analysis at 72 hours.
- Sample storage conditions before analysis:
Samples were stored frozen prior to analysis. - Vehicle:
- yes
- Remarks:
- culture medium
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:
Nominal amounts of test item (194 and 62 mg) were each separately dissolved in culture medium with the aid of ultrasonication for approximately 5 minutes, and the volume adjusted to 1 liter to give 100 and 32 mgi/L stock solutions respectively. A series of dilutions was made from these stock solutions to give further stock solutions of 10, 3.2 and 1.0 mg/L. An aliquot (500 mL) of each of the stock solutions was separately inoculated with algal suspension (2.9 mL) to give the required test concentrations of 1.0, 3.2, 10, 32 and 100 mg/L.
All concentrations were corrected for a test item water content of 48.4% w/w.
- Controls:
- Chemical name of vehicle (organic solvent, emulsifier or dispersant):
Culture medium
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): None recorded - Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Scottish Marine Institute, Oban, Argyll, Scotland
- Age of inoculum (at test initiation): Not reported
- Method of cultivation: Master cultures were maintained in the laboratory by the periodic replenishment of culture medium. ). The master cultures were maintained under constant agitation by orbital shaker (approximately 150 rpm) and constant illumination at 24 ±1 °C.
- Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 72 h
- Test temperature:
- 24 ± 1 °C.
- pH:
- 7.5 - 8.8
- Nominal and measured concentrations:
- Nominal: 1.0, 3.2, 10, 32 and 100 mg/L.
Measured: 0.760/0.836*, 3.04, 7.26/8.65*, 37.0, and 121 mg/L
*duplicate samples stored frozen prior to analysis - Details on test conditions:
- TEST SYSTEM
- Test vessel:
- Type : closed
- Material, size, headspace, fill volume:
250 mL glass conical flasks, plugged with foam bungs
- Aeration:
N/A
- 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: 5.00E+03 cells per mL (nominal); 4.24E+04 cells per mL (measured)
- Control end cells density: 2.04E+06 cells per mL
- No. of vessels per concentration (replicates) 6
- No. of vessels per vehicle control (replicates): 3
GROWTH MEDIUM
- Standard medium used: yes (AAP-medium)
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reverse osmosis purified deionized water
OTHER TEST CONDITIONS
- Adjustment of pH: Adjusted to 7.5 with 0.1N NaOH or HCl
- Photoperiod: Continuous illumination
- Light intensity and quality: 7000 lux, provided by warm white lighting (380 to 730 nm)
- Other: constantly shaken at approximately 150 rpm for 72 hours
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic particle counter (measured at 24, 48, and 72 hours)
- Other: the shape and size of the algal cells was inspected microscopically and any abnormalities recorded
TEST CONCENTRATIONS
- Spacing factor for test concentrations: Approx 3.2
- Range finding study
- Test concentrations: of 0.10, 1.0, 10 and 100 mg/L. Due to a technical oversight, the concentrations employed were not corrected for the test item water content as was stated in the Study Plan. As the test item contains 48.4% w/w of water, the equivalent test item concentrations employed were 0.0516, 0.516, 5.16 and 51.6 mg/L. This oversight was considered to have had no adverse effect on the outcome of the range-finding test as it was still possible to assign dose levels for the definitive test from the data obtained.
- Results used to determine the conditions for the definitive study:
The results showed no effect on growth at the test concentrations of 0.0516, 0.516 and 5.16 mg/L. However, growth was observed to be reduced at 51.6 mg/L.
Based on this information test concentrations of 1.0, 3.2, 10, 32 and 100 mg/L were selected for the definitive test. - Reference substance (positive control):
- yes
- Remarks:
- potassium dichromate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 18 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.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 2.9 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- 19 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Details on results:
- - Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test):
- Unusual cell shape:
None observed
- Colour differences:
At the start of the test all control and test cultures were observed to be clear colourless solutions. After the 72-hour test period all control, 1.0, 3.2 and 10 mg/L test cultures were green dispersions, whilst the 32 and 100 mg/L test cultures were light green or pale green dispersions.
- Flocculation:
None observed
- Adherence to test vessels:
None observed
- Aggregation of algal cells:
None observed
- Other:
Some healthy cells and some cell debris were observed to be present in the test cultures at 100 mg/L
- Any stimulation of growth found in any treatment:
N/A
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values:
None observed
- Effect concentrations exceeding solubility of substance in test medium:
N/A - Results with reference substance (positive control):
- - Results with reference substance valid? Yes
- EC50:
ErC50 (72 h): 1.6 mg/L, 95% confidence limits 1.4 to 1.8 mg/L
EyC50 (72 h): 0.77 mg/L, 95% confidene limits 0.68 to 0.87 mg/L - Reported statistics and error estimates:
- Statistical analysis of the growth rate and yield data was carried out for the control and all test concentrations using one way analysis of variance incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf, 1981) and Dunnett's multiple comparison procedure for comparing several treatments with a control (Dunnett, 1955).
Growth Rate: There were no statistically significant differences (P≥0.05), between the control, 1.0 and 3.2 mg/L test concentrations, however all other test concentrations were significantly different (P<0.05) and, therefore the NOEC based on growth rate was 3.2 mg/L. Correspondingly the LOEC based on growth rate was 10 mg/L.
Yield: . There were no statistically significant differences (P≥0.05), between the control, 1.0 and 3.2 mg/L test concentration however all other test concentrations were significantly different (P<0.05) and, therefore the NOEC based on yield was 3.2 mg/L. Correspondingly the LOEC based on yield was 10 mg/L. - Validity criteria fulfilled:
- yes
- Conclusions:
- It was not possible to calculate an ErC50 value as no more than 39% inhibition of growth rate occurred at the maximum concentration employed of 100 mg/L. It was therefore also not possible to determine 95% confidence limits for growth rate. The ErC10 was estimated to be 18 mg/L.
The EyC50 value was determined to be 19 mg/L, 95% confidence limits of 14-25 mg/L. The EyC10 was estimate dto be 2.9 mg/L. - Executive summary:
A study was performed to assess the effect of the test item on the growth of the green algaPseudokirchneriellasubcapitata. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (2006) No 201, "Freshwater Alga and Cyanobacteria, Growth Inhibition Test" referenced as Method C.3 of Commission Regulation (EC) No 761/2009.
Following a preliminary range-finding test, Pseudokirchneriella subcapitata was exposed to an aqueous solution of the test item at concentrations of 1.0, 3.2, 10, 32 and 100 mg/L (three replicate flasks per concentration) for 72 hours, under constant illumination and shaking at a temperature of 24 ±1 °C. Samples of the algal populations were removed daily and cell concentrations determined for each control and treatment group, using a Coulter®Multisizer Particle Counter.
Analysis of the test preparations at 0 hours showed measured test concentrations to range from 84% to 121% of nominal. A concentration dependent decline in measured concentrations was observed at 72 hours in the range of 61% to 118% of nominal. Analysis of additional samples prepared at the start of the test with the omission of algal cells showed measured test concentrations at 72 hours to range from 88% to 137% of nominal suggesting that the losses observed in those samples where algae were present was due to adsorption of the test item to the algal cells present rather than instability under the conditions of the test.
Exposure of Pseudokirchneriella subcapitata to the test item gave the following results based on the nominal test concentrations:
Response Variable
EC50
(mg/L)EC10
(mg/L)95% Confidence Limits (mgL)
No Observed Effect Concentration (NOEC)
(mg/L)Lowest Observed Effect Concentration (LOEC)
(mg/L)Growth Rate
>100*
18
Not determined due to nature of data
3.2
10
Yield
19
2.9
14
-
25
3.2
10
*It was not possible to calculate an ErC50value as no more than 39% inhibition of growth rate occurred at the maximum concentration employed of 100 mg/L.
Reference
Range-finding Test
The cell densities and percentage inhibition of growth values from the exposure of Pseudokirchneriella subcapitata to the test item during the range-finding test are given in Table 1.
Table 1: Cell Densities and Percentage Inhibition of Growth from the Range‑finding Test
Nominal Concentration |
Cell Densities*(cells per mL) |
Inhibition Values (%) |
|||
0 Hours |
72 Hours |
Growth Rate |
Yield |
||
Control |
R1 |
6.37E+03 |
6.64E+05 |
- |
- |
R2 |
6.28E+03 |
1.24E+06 |
|||
Mean |
6.32E+03 |
9.50E+05 |
|||
0.0516 |
R1 |
6.28E+03 |
1.25E+06 |
[4] |
[28] |
R2 |
6.37E+03 |
1.19E+06 |
|||
Mean |
6.32E+03 |
1.22E+06 |
|||
0.516 |
R1 |
6.13E+03 |
1.08E+06 |
[1] |
[9] |
R2 |
6.48E+03 |
9.97E+05 |
|||
Mean |
6.31E+03 |
1.04E+06 |
|||
5.16 |
R1 |
6.57E+03 |
8.48E+05 |
4 |
7 |
R2 |
7.30E+03 |
9.22E+05 |
|||
Mean |
6.94E+03 |
8.85E+05 |
|||
51.6 |
R1 |
7.45E+03 |
3.78E+05 |
21 |
64 |
R2 |
5.81E+03 |
3.21E+05 |
|||
Mean |
6.63E+03 |
3.50E+05 |
* Cell densities represent the mean number of cells per mL calculated from the mean of the cell counts from 3 counts for each of the replicate flasks
R = Replicate
- = Not applicable
[ ]= Increase in growth compared to controls
The results showed no effect on growth at the test concentrations of 0.0516, 0.516 and 5.16 mg/L. However, growth was observed to be reduced at 51.6 mg/L.
Based on this information test concentrations of 1.0, 3.2, 10, 32 and 100 mg/L were selected for the definitive test.
Chemical analysis of the 0.516, 5.16 and 51.6 mg/L test preparations at 0 hours showed measured test concentrations to range from 87% to 99% of nominal. A concentration dependent decline in measured test concentrations was observed at 72 hours in the range of 66% to 99% of nominal. This was considered to be due to possible adsorption of the test item to the algal cells present.
Definitive Test
Verification of Test Concentrations
Analysis of the test preparations at 0 hours showed measured test concentrations to range from 84% to 121% of nominal. A concentration dependent decline in measured concentrations was observed at 72 hours in the range of 61% to 118% of nominal. Analysis of additional samples prepared at the start of the test with the omission of algal cells showed measured test concentrations at 72 hours to range from 88% to 137% of nominal suggesting that the losses observed in those samples where algae were present was due to adsorption of the test item to the algal cells present rather than instability under the conditions of the test.
Given this it was considered appropriate to calculate the results based on the nominal test concentrations only as this gives a more accurate measure of the levels of test item to which the algae were exposed.
Growth Data
Cell density values determined at each sampling time and pH values at 0 and 72 hours are given in Table 2. Daily specific growth rates for the control cultures are given in Table 3. Growth rate and yield values for the control and test cultures after 72 hours and percentage inhibition values are given in Table 4.
Table 2 Cell Densities and pH Values in the Definitive Test
Nominal Concentration |
pH |
Cell Densities* (cells per mL) |
pH |
|||
0 Hours |
24 Hours |
48 Hours |
72 Hours |
72 Hours |
||
Control |
R1 |
7.8 |
4.16E+04 |
2.66E+05 |
2.18E+06 |
8.6 |
R2 |
4.33E+04 |
2.54E+05 |
1.99E+06 |
|||
R3 |
4.43E+04 |
2.62E+05 |
2.04E+06 |
|||
R4 |
4.28E+04 |
2.58E+05 |
2.13E+06 |
|||
R5 |
4.05E+04 |
2.43E+05 |
1.96E+06 |
|||
R6 |
4.21E+04 |
2.40E+05 |
1.92E+06 |
|||
Mean |
4.24E+04 |
2.54E+05 |
2.04E+06 |
|||
1.0 |
R1 |
7.7 |
4.48E+04 |
2.14E+05 |
1.36E+06 |
8.6 |
R2 |
4.50E+04 |
2.77E+05 |
2.18E+06 |
|||
R3 |
4.22E+04 |
2.53E+05 |
1.82E+06 |
|||
Mean |
4.40E+04 |
2.48E+05 |
1.79E+06 |
|||
3.2 |
R1 |
7.6 |
4.07E+04 |
2.72E+05 |
1.80E+06 |
8.8 |
R2 |
4.34E+04 |
2.98E+05 |
1.84E+06 |
|||
R3 |
4.17E+04 |
2.72E+05 |
1.82E+06 |
|||
Mean |
4.19E+04 |
2.81E+05 |
1.82E+06 |
|||
10 |
R1 |
7.6 |
3.66E+04 |
2.43E+05 |
1.46E+06 |
8.6 |
R2 |
3.68E+04 |
2.54E+05 |
1.51E+06 |
|||
R3 |
3.68E+04 |
2.41E+05 |
1.34E+06 |
|||
Mean |
3.67E+04 |
2.46E+05 |
1.43E+06 |
|||
32 |
R1 |
7.5 |
2.39E+04 |
1.38E+05 |
7.40E+05 |
8.3 |
R2 |
2.15E+04 |
1.38E+05 |
7.93E+05 |
|||
R3 |
2.17E+04 |
1.13E+05 |
6.35E+05 |
|||
Mean |
2.24E+04 |
1.29E+05 |
7.22E+05 |
|||
100 |
R1 |
7.5 |
1.44E+04 |
5.29E+04 |
2.36E+05 |
7.8 |
R2 |
1.38E+04 |
4.76E+04 |
1.98E+05 |
|||
R3 |
1.42E+04 |
5.05E+04 |
2.17E+05 |
|||
Mean |
1.41E+04 |
5.03E+04 |
2.17E+05 |
* Cell densities represent the mean number of cells per mL calculated from the mean of the cell counts from 3 counts for each of the replicate flasks
R = Replicate
Table3 Daily Specific Growth Rates for the Control Cultures in the Definitive Test
|
Daily Specific Growth Rate (cells/mL/hour) |
|||
Day 0 to 1 |
Day 1 to 2 |
Day 2 to 3 |
||
Control |
R1 |
0.088 |
0.077 |
0.088 |
R2 |
0.090 |
0.074 |
0.086 |
|
R3 |
0.091 |
0.074 |
0.085 |
|
R4 |
0.089 |
0.075 |
0.088 |
|
R5 |
0.087 |
0.075 |
0.087 |
|
R6 |
0.089 |
0.073 |
0.087 |
|
Mean |
0.089 |
0.075 |
0.087 |
R = Replicate
Table 4 Inhibition of Growth Rate and Yield in the Definitive Test
Nominal Concentration |
Growth Rate (cells/mL/hour) |
Yield (cells/mL) |
|||
0 to 72 Hour |
% Inhibition |
0 to 72 Hour |
% Inhibition* |
||
Control |
R1 |
0.084 |
- |
2.18E+06 |
- |
R2 |
0.083 |
1.99E+06 |
|||
R3 |
0.083 |
2.03E+06 |
|||
R4 |
0.084 |
2.12E+06 |
|||
R5 |
0.083 |
1.96E+06 |
|||
R6 |
0.083 |
1.91E+06 |
|||
Mean |
0.083 |
2.03E+06 |
|||
SD |
0.001 |
1.01E+05 |
|||
1.0 |
R1 |
0.078 |
6 |
1.36E+06 |
|
R2 |
0.084 |
[1] |
2.17E+06 |
|
|
R3 |
0.082 |
1 |
1.82E+06 |
|
|
Mean |
0.081 |
2 |
1.78E+06 |
12 |
|
SD |
0.003 |
|
4.08E+05 |
|
|
3.2 |
R1 |
0.082 |
1 |
1.79E+06 |
|
R2 |
0.082 |
1 |
1.84E+06 |
|
|
R3 |
0.082 |
1 |
1.82E+06 |
|
|
Mean |
0.082 |
1 |
1.82E+06 |
11 |
|
SD |
0.000 |
|
2.36E+04 |
|
|
10 |
R1 |
0.079 |
5 |
1.45E+06 |
|
R2 |
0.079 |
5 |
1.50E+06 |
|
|
R3 |
0.078 |
6 |
1.33E+06 |
|
|
Mean |
0.079 |
5 |
1.43E+06 |
30 |
|
SD |
0.001 |
|
8.75E+04 |
|
|
32 |
R1 |
0.069 |
17 |
7.35E+05 |
|
R2 |
0.070 |
16 |
7.88E+05 |
|
|
R3 |
0.067 |
19 |
6.30E+05 |
|
|
Mean |
0.069 |
17 |
7.17E+05 |
65 |
|
SD |
0.002 |
|
8.06E+04 |
|
|
100 |
R1 |
0.054 |
35 |
2.31E+05 |
|
R2 |
0.051 |
39 |
1.93E+05 |
|
|
R3 |
0.052 |
37 |
2.12E+05 |
|
|
Mean |
0.052 |
37 |
2.12E+05 |
90 |
|
SD |
0.002 |
|
1.88E+04 |
|
* In accordance with the OECD test guideline only the mean value for yield for each test concentration is calculated
R = Replicate
SD =Standard Deviation
[ ] =Increase in growth compared to controls
- = Not applicable
Description of key information
72h ErC50 = > 100 mg/L, 72h ErC10 = 18 mg/L; OECD 201; Vryenhoef, 2018
Key value for chemical safety assessment
- EC10 or NOEC for freshwater algae:
- 18 mg/L
Additional information
A study was performed to assess the effect of the test item on the growth of the green alga Pseudokirchneriellasubcapitata. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (2006) No 201, "Freshwater Alga and Cyanobacteria, Growth Inhibition Test" referenced as Method C.3 of Commission Regulation (EC) No 761/2009.
It was not possible to calculate an EC50 based on inhibition of growth rate as no more than 39% inhibition of growth rate occurred at the maximum concentration employed. Therefore only the EC10 is reported.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
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