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EC number: 613-782-9 | CAS number: 65355-32-0
- 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:
- 2018-04-17 to 2018-06-27
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.3 (Algal Inhibition test)
- Version / remarks:
- 2016
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Alga, Growth Inhibition Test)
- Version / remarks:
- 2011
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Analytical monitoring:
- yes
- Details on sampling:
- All concentration levels and the control were analytically verified via LC-MS/MS after 0 (start of exposure) and 72 hours (end of the exposure) with algae. The samples and the control were diluted (factor 2) with acetonitrile directly after sampling. Further dilutions were made prior to analysis with dilution medium. All original samples were stored at room temperature before preparation. Prepared samples were stored in an auto-sampler at room temperature until analysis.
- Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION
A saturated solution (100 mg test item/L were weighed out) was freshly prepared with dilution water one day prior to the start of the exposure (day -1). The stock solution was stirred for 24 hours at approx. 1100 rpm at room temperature. After a separation phase of 1 hour at room temperature the test solution was removed from the bottom of the flask. The test item solution was checked via laser beam (Tyndall effect) for undissolved test item. A filtration step was added, because undissolved test item was confirmed by Tyndall effect. Undissolved particles were removed by membrane filtration (membrane filter 0.45 µm). The filter was saturated in order to avoid adsorption during the filtration. The first 25 mL of the filtrate were discarded. The filtration was interrupted for 10 minutes to allow adsorption and saturation of the filter material with dissolved test item. Thereafter, the filtration was continued. The next 25 mL were discarded. During filtration, the filter will always be kept covered. This saturated solution was used for the test. The saturated solution and four concentration levels were tested in a geometrical series with a dilution factor of 2: 6.25, 12.5, 25.0, 50.0 and 100 %. The concentration levels are based on the results of a preliminary range finding test (non-GLP). - Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: green alga
- Source: Culture Collection of Algae and Protozoa (CCAP), SAMS Research Services Ltd, Dunstaffnage Marine Laboratory, Dunbeg, OBAN; Argyll PA37 1QA; Scotland, UK
- Cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounted to 2590 - 5180 lux corresponding to 35 - 70 µE • m-2 • s-1 for 24 hours per day.
- Culture medium: Nutrient medium Z according to LÜTTGE et al. (1994) - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 72 h
- Test temperature:
- 21 - 24 °C controlled at ± 2°C
- pH:
- 7.64 - 9.65
- Nominal and measured concentrations:
- Geometric mean measured: 1.41, 0.808, 0.385, 0.204 and 0.117 mg/L; Nominal conc.: 100, 50, 25, 12.5 and 6.25 % of the saturated solution
- Details on test conditions:
- TEST SYSTEM
- Test vessel: Sterile glass Erlenmeyer flasks, volume: 250 mL, sealed with cotton wool plugs
- Type: closed
- Initial cells density: Nominal: approximately 5 x 103 - 104 cells/mL Actual: 6442 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: medium according to OECD TG 201
- Intervals of water quality measurement: The pH-values at the start of exposure were measured in one additional replicate of each concentration level and the control. At the end of exposure, the pH-values were measured from pooled samples of the test item concentrations and the control. The room temperature was measured continuously. Light intensity was measured prior to the start of exposure.
- Photoperiod: 24 hours/day light
- Light intensity and quality: Approximately 4440 to 8880 lux, corresponding to 60 to 120 µE*m-2*s-1
EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: fluorometer
- Chlorophyll measurement: The cell density was measured daily via Chlorophyll a-fluorescence, excitation at 436 nm, emission at 685 nm. Dilution water was used as a background signal. No self-fluorescence was observed in the preliminary range finding test at the saturated solution.
- Microscopic evaluation: Microscopic evaluation of the cells at the start and the end of the incubation period was carried out. The cells were checked for unusual cell shapes, colour differences, differences in chloroplast morphology, flocculation, adherence of algae to test containers and agglutination of algae cells.
TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 1.0 – 100 % of saturated solution
- Results used to determine the conditions for the definitive study: yes - Reference substance (positive control):
- yes
- Remarks:
- potassium dichromate
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 1.41 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EC20
- Effect conc.:
- 1.29 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: 95 % confidence intervals (1.15 - > 1.41) mg/L
- Key result
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 0.785 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: 95 % confidence intervals (0.697 - 0.898) mg/L
- Duration:
- 72 h
- Dose descriptor:
- LOEC
- Effect conc.:
- 0.385 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- 0.204 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EC50
- Effect conc.:
- 0.87 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: 95 % confidence intervals (0.764 - 0.999) mg/L
- Duration:
- 72 h
- Dose descriptor:
- EC20
- Effect conc.:
- 0.462 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: 95 % confidence intervals (0.366 - 0.555) mg/L
- Duration:
- 72 h
- Dose descriptor:
- EC10
- Effect conc.:
- 0.308 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Remarks on result:
- other: 95 % confidence intervals (0.200 - 0.424) mg/L
- Duration:
- 72 h
- Dose descriptor:
- NOEC
- Effect conc.:
- 0.204 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Duration:
- 72 h
- Dose descriptor:
- LOEC
- Effect conc.:
- 0.385 mg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- other: Yield
- Results with reference substance (positive control):
- Regularly, a positive control test to check the test system is carried out with potassium dichromate.
Growth rate: ErC50 = 1.09 mg/L (95% confidence interval: 0.842 - 1.31 mg/L)
Yield inhibition: EyC50 = 0.548 mg/L (95% confidence interval: 0.439 - 0.660 mg/L) - Validity criteria fulfilled:
- yes
- Conclusions:
- Under the test conditions, the EC50-vaiues for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were > 1.41 mg/L and 0.870 (C.I: 0.764 - 0.999) mg/L, respectively. The EC10 for growth rate was 0.785 mg/L.
- Executive summary:
The toxicity of the test item to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined according to the principles of OECD 201 and Council Regulation (EC) No. 266/2016/Method C.3. The aim of the study was the determination of the effects on the growth rate and yield over a period of 72 hours. The study was conducted under static conditions with an initial cell density of 6442 cells/mL. A saturated solution with a nominal concentration of 100 mg/L was prepared with dilution water. Out of the saturated solution five concentration levels were tested in a geometrical series with a dilution factor of 2: 6.25 - 12-5 - 25.0 - 50.0 - 100 % (1.41, 0.808, 0.385, 0.204 and 0.117 mg/L). Three replicates were tested for each test item concentration level and six replicates for the control. The environmental conditions were within the acceptable limits. The measured concentrations in the test solutions at the start of the exposure were 2 % of the nominal concentrations. At the end of exposure after 72 hours, the measured concentrations were in the range of 1 to 2 % of the nominal concentrations. All validity criteria were met. Under the test conditions, the EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were > 1.41 mg/L and 0.870 (C.I: 0.764 - 0.999) mg/L, respectively. The EC10 for growth rate was 0.785 mg/L.
Reference
Biological Data
The growth inhibition effects of the test item are summarized in Table 1. Cell densities are listed in Table 2. Evaluations of growth rate inhibition and yield inhibition are given in Table 3 (calculated based on cell density values). Section-by-section and average specific growth rate are given in Table 4.
Microscopic evaluation of the cells at the start and end of exposure revealed no morphological abnormalities. All effect values are given based on the nominal test item concentrations.
Table 1: NOEC, LOEC, ECx-values of the test item (0 - 72 hours): (based on the geometric mean measured test item concentrations, with 95% confidence intervals)
|
Growth Rate Inhibition [mg/L] |
NOEC |
0.204 |
LOEC |
0.385 |
ErC10 |
0.785 (0.697 - 0.898) |
ErC20 |
1.29 (1.15 - > 1.41) |
ErC50 |
> 1.41 |
|
Yield Inhibition [mg/L] |
NOEC |
0.204 |
LOEC |
0.385 |
ErC10 |
0.308 (0.200 - 0.424) |
ErC20 |
0.462 (0.366 - 0.555) |
ErC50 |
0.870 (0.764 - 0.999) |
Table 2: Cell Densities
Geometric mean measured test item concentration [mg/L] |
Replicate |
Cell density [cells/mL] |
|||
1.41 |
No. |
0 hours |
24 hours |
48 hours |
72 hours |
1 |
6442 |
20543 |
136719 |
638015 |
|
2 |
6442 |
19214 |
145556 |
692531 |
|
3 |
6442 |
12787 |
110076 |
521710 |
|
Mean |
6442 |
17515 |
130784 |
617419 |
|
0.808 |
1 |
6442 |
21483 |
174165 |
1249355 |
2 |
6442 |
26084 |
189397 |
1067487 |
|
3 |
6442 |
29370 |
207450 |
1277110 |
|
Mean |
6442 |
25646 |
190337 |
1197984 |
|
0.385 |
1 |
6442 |
34030 |
238461 |
1790590 |
2 |
6442 |
31979 |
281011 |
1716165 |
|
3 |
6442 |
29849 |
239852 |
2020916 |
|
Mean |
6442 |
31953 |
253108 |
1842557 |
|
0.204 |
1 |
6442 |
38342 |
325955 |
2055175 |
2 |
6442 |
38521 |
317117 |
2163362 |
|
3 |
6442 |
33543 |
279730 |
2267726 |
|
Mean |
6442 |
36802 |
307601 |
2162088 |
|
0.117 |
1 |
6442 |
37369 |
253972 |
2141615 |
2 |
6442 |
38283 |
293057 |
1954038 |
|
3 |
6442 |
38134 |
320731 |
2072552 |
|
Mean |
6442 |
37929 |
289253 |
2056068 |
|
Control |
1 |
6442 |
40611 |
297317 |
2133324 |
2 |
6442 |
39263 |
312152 |
2222495 |
|
3 |
6442 |
36289 |
296314 |
2335350 |
|
4 |
6442 |
38039 |
284437 |
2161525 |
|
5 |
6442 |
41204 |
314853 |
2127416 |
|
6 |
644 |
33841 |
286255 |
2196975 |
|
Mean |
6442 |
38208 |
29855 |
2196181 |
Table 3: Evaluation after 72 hours: Statistically significant differences of growth rates and yield compared to control values are marked (+), not significant differences are marked (-).
Geometric mean measured test item concentration [mg/L] |
Replicate No. |
Growth Rate [d-1] |
Inhibition of Growth Rate [%] |
Yield [cells/mL] |
Inhibition of yield [%] |
1.41 |
1 |
1.53 |
21 |
631573 |
71 |
2 |
1.56 |
20 |
686089 |
69 |
|
3 |
1.47 |
25 |
515268 |
76 |
|
Mean |
(+) 1.52 |
22 |
(+) 610977 |
72 |
|
0.808 |
1 |
1.76 |
10 |
1242913 |
43 |
2 |
1.70 |
12 |
1061045 |
52 |
|
3 |
1.76 |
9 |
1270668 |
42 |
|
Mean |
(+) 1.74 |
10 |
(+) 1191542 |
46 |
|
0.385 |
1 |
1.88 |
3 |
1784148 |
19 |
2 |
1.86 |
4 |
1709723 |
22 |
|
3 |
1.92 |
1 |
2014474 |
8 |
|
Mean |
(+) 1.89 |
3 |
(+) 1836115 |
16 |
|
0.204 |
1 |
1.92 |
1 |
2048733 |
6 |
2 |
1.94 |
0 |
2156920 |
2 |
|
3 |
1.96 |
-1 |
2261284 |
-3 |
|
Mean |
(-) 1.94 |
0 |
(-) 2155646 |
2 |
|
0.117 |
1 |
1.94 |
0 |
2135173 |
2 |
2 |
1.91 |
2 |
1947596 |
11 |
|
3 |
1.93 |
1 |
2066110 |
6 |
|
Mean |
(-) 1.92 |
1 |
(-) 2049626 |
6 |
|
Control |
1 |
1.93 |
|
2126882 |
|
2 |
1.95 |
2216053 |
|||
3 |
1.96 |
2328908 |
|||
4 |
1.94 |
2155083 |
|||
5 |
1.93 |
2120974 |
|||
6 |
1.94 |
2190533 |
|||
Mean |
1.94 |
2189739 |
Table 4: Section-by-Section and Average Specific Growth Rates of the Control Group (0 - 72 hours)
|
Replicate No. |
Specific growth rate [d_1] section-by-section |
Mean (0 – 72 hours) |
SD ± |
CV [%] |
Mean CV [%] |
||
0 - 24 hours |
24-48 hours |
48 - 72 hours |
||||||
Control |
1 |
1.84 |
1.99 |
1.97 |
1.93 |
0.081 |
4.2 |
7.74 |
2 |
1.81 |
2.07 |
1.96 |
1.95 |
0.134 |
6.9 |
||
3 |
1.73 |
2.10 |
2.06 |
1.96 |
0.205 |
10.4 |
||
4 |
1.78 |
2.01 |
2.03 |
1.94 |
0.141 |
7.3 |
||
5 |
1.86 |
2.03 |
1.91 |
1.93 |
0.091 |
4.7 |
||
6 |
1.66 |
2.14 |
2.04 |
1.94 |
0.252 |
12.9 |
||
|
Mean |
1.94 |
|
|||||
SD ± |
0.01 |
|
||||||
CV [%] |
0.60 |
|
Table 5: Environmental Conditions
Geometric mean measured test item concentration [mg/L] |
pH-values
|
||
Start; 0 hours |
End; 72 hours |
||
1.14 |
7.64 |
8.73 |
|
0.808 |
7.68 |
9.17 |
|
0.385 |
7.72 |
9.47 |
|
0.204 |
7.77 |
9.63 |
|
0.117 |
7.86 |
9.65 |
|
Control |
7.93 |
9.38 |
|
Room temperature [°C]: |
min.: 22.0 |
max.: 24.0 |
mean value: 23.0 |
Light intensity [lux] |
n = 24 |
mean value: 5778 range of the measured values: 5040 to 6172 equalling -12.8 to 6.82% |
CV [%]: 5.21% |
CV = Coefficient of variation
n = number of measuring points
Table 6: Measured Concentration of the test item in the Definitive Test with Algae
Sampling date |
Fresh Media, 0 hours |
Old Media, 72 hours |
|
|||
Nominal test item concentration [mg/L] |
Test Item |
Geometric mean measured concentration of the test item [mg/L] |
||||
Meas. cone. [mg/L] |
% |
Meas. cone. [mg/L] |
% |
|||
10.0 |
1.57 |
2 |
1.27 |
1 |
1.41 |
|
3.16 |
0.756 |
2 |
0.864 |
2 |
0.808 |
|
1.00 |
0.388 |
2 |
0.382 |
2 |
0.385 |
|
0.316 |
0.196 |
2 |
0.212 |
2 |
0.204 |
|
0.100 |
0.121 |
2 |
0.113 |
2 |
0.117 |
|
Control |
< LOQ |
< LOQ |
|
|||
Meas. conc. = measured concentration of the test item, dilution factors taken into account
% = percent of the nominal measured concentration of the test item
LOQ = limit of quantification (0.0700 mg/L of the test item)
Validity Criteria
The study meets the validity criteria of the guideline:
Table 7: Validity Criteria
Validity Criterion |
Required |
This study |
Increase of the cell growth in the control cultures |
Exponentially, ≥ 16-fold corresponding to a specific growth rate of 0.92 /day |
341-fold (specific growth rate 1.94 /day) |
Mean coefficients of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control cultures |
< 35 % |
7.74 % |
Coefficient of variation of average specific growth rates during the whole test period in replicate control cultures |
< 7 % |
0.60 % |
Description of key information
The test item was tested to Pseudokirchneriella subcapitata in an open static test system according to OECD 201 (2011). Under the test conditions, the EC50-vaiues for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were > 1.41 mg/L and 0.870 (C.I: 0.764 - 0.999) mg/L, respectively. The EC10 value for growth rate was 0.785 mg/L (reference 6.1.5-1).
Key value for chemical safety assessment
- EC10 or NOEC for freshwater algae:
- 0.785 mg/L
Additional information
The toxicity of the test item to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined according to the principles of OECD 201 and Council Regulation (EC) No. 266/2016/Method C.3. The aim of the study was the determination of the effects on growth rate and yield over a period of 72 hours. The study was conducted under static conditions with an initial cell density of 6442 cells/mL. A saturated solution with a nominal concentration of 100 mg/L was prepared with dilution water. Out of the saturated solution five concentration levels were tested in a geometrical series with a dilution factor of 2: 6.25 - 12-5 - 25.0 - 50.0 - 100 % (1.41, 0.808, 0.385, 0.204 and 0.117 mg/L). Three replicates were tested for each test item concentration level and six replicates for the control. The environmental conditions were within the acceptable limits. The measured concentrations in the test solutions at the start of the exposure were 2% of the nominal concentrations. At the end of exposure after 72 hours, the measured concentrations were in the range of 1 to 2% of the nominal concentrations. All validity criteria were met. Under the test conditions, the EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were > 1.41 mg/L and 0.870 (C.I: 0.764 - 0.999) mg/L, respectively. The EC10 value for growth rate was 0.785 mg/L.
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