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EC number: 944-232-9 | CAS number: -
- 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:
- From 2016-11-21 to 2016-12-23, with the definitive exposure phase from 2016-12-20 to 2016-12-23 at the test facility.
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 201 (Alga, Growth Inhibition Test)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.3 (Algal Inhibition test)
- Deviations:
- no
- GLP compliance:
- yes
- Analytical monitoring:
- no
- Details on sampling:
- Determination of the test Item
No analytical monitoring was carried out. Due to the low solubility, a specific analytical method is not available and the concentrations are too low for unspecific analysis as the determination of the total organic carbon content. - Vehicle:
- no
- Details on test solutions:
- Preparation of the Saturated solution
A saturated solution with a nominal loading of 5.00 mg/L was prepared once 24 ± 1 hour prior to the start of the exposure. An appropriate amount of the test item was weighed out. The test item was applied onto a glass slide. The glass slide with the test item was inserted into a glass bottle with an appropriate amount of dilution water. The saturated solution was stirred for 24 hours (1100 rpm) with a magnetic stirrer. Undissolved particles were removed by membrane filtration (membrane filter 0.45 µm, RC, MACHEREY-NAGEL). The filter was saturated in order to avoid adsorption during the filtration. The first 25 mL of the filtrate will be 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.
The solution was checked via laser beam (Tyndall effect), which was negative.
Test loading
All terms related to concentration level are given as loading level because partly dissolved compounds and mixtures cannot be related to concentrations. The saturated solution and four dilution levels out of the saturated solution were tested in a geometrical series with a dilution factor of 2: 6.25 - 12.5 - 25.0 - 50.0 - 100% of the saturated solution.
Control
Six replicates (without test item) were incubated under the same conditions as the test loadings. - Test organisms (species):
- Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
- Details on test organisms:
- TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata HINDÁK
- Strain: SAG 61.81
- Source (laboratory, culture collection): SAG, Pflanzenphysiologisches Institut der Universität Göttingen, Nikolausberger Weg 18, D-37073 Göttingen
- Age of inoculum (at test initiation): 4 days
- Method of cultivation: Fresh stocks were prepared every month on Z-Agar. Light intensity amounted to 2567 – 5130 lux corresponding to 35 - 70 µE ∙ m-2 ∙ s-1 for 24 h per day. - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 72 h
- Post exposure observation period:
- No
- Hardness:
- Not specified
- Test temperature:
- min.: 22.0 max.: 23.0 mean value: 22.5
- pH:
- Nominal test item loading pH-values
[mg/L] Start; 0 hours End; 72 hours
5.00 7.75 8.38
2.50 7.78 8.61
1.25 7.80 8.93
0.625 8.04 9.13
0.313 8.14 9.34
Control 8.07 9.26 - Dissolved oxygen:
- Not measured
- Salinity:
- Not measured, freshwater conditions
- Nominal and measured concentrations:
- No analytical monitoring
- Details on test conditions:
- TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type (delete if not applicable): open, cotton wool plugs
- Material, size, headspace, fill volume: sterile 250 mL Erlenmeyer flasks, test volume 100 mL
- Aeration: Test containers were placed on a rotary shaker and oscillated at appr. 70 rpm
- Type of flow-through (e.g. peristaltic or proportional diluter): None
- Renewal rate of test solution (frequency/flow rate): One application at test start
- Initial cells density: Nominal: approximately 2 - 5 x 10^3 cells/mL, Actual: mean 6305 cells/mL
- Control end cells density: Mean 2579163 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6
GROWTH MEDIUM
- Standard medium used: yes
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
Composition of Dilution Water
according to the guideline
Component Concentration [mg/L]
NH4Cl 15
MgCl2 x 6 H2O 12
CaCl2 x 2 H2O 18
MgSO4 x 7 H2O 15
KH2PO4 1.6
FeCl3 x 6 H2O 0.064
Na2EDTA x 2 H2O 0.1
H3BO3 0.185
MnCl2 x 4 H2O 0.415
ZnCl2 3 x 10^-3
Na2MoO4 x 2 H2O 7 x 10^-3
CoCl2 x 6 H2O 1.5 x 10^-3
CuCl2 x 2 H2O 1 x 10^-5
NaHCO3 50
pH 8.1 ± 0.2
OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: 24 h
- Light intensity and quality: Approximately 4440 to 8880 lux, corresponding to 60 to 120
µE ∙ m-2 ∙ s-1, mean value: 5298 lux,
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
-Chlorophyll a-fluorescence
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 found in the definitive test at the saturated solution. - Reference substance (positive control):
- yes
- Remarks:
- (Potassium dichromate)
- Duration:
- 72 h
- Dose descriptor:
- other: NOEL
- Effect conc.:
- 1.25 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- other: LOEL
- Effect conc.:
- 2.5 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Duration:
- 72 h
- Dose descriptor:
- EL10
- Effect conc.:
- 1.86 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: CI
- Remarks:
- 1.66 - 2.03
- Duration:
- 72 h
- Dose descriptor:
- EL20
- Effect conc.:
- 2.27 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: CI
- Remarks:
- 2.16 - 2.34
- Duration:
- 72 h
- Dose descriptor:
- EL50
- Effect conc.:
- 3.08 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- growth rate
- Remarks on result:
- other: CI
- Remarks:
- 2.92 - 3.36
- Duration:
- 72 h
- Dose descriptor:
- other: NOEL
- Effect conc.:
- < 0.313 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Duration:
- 72 h
- Dose descriptor:
- other: LOEL
- Effect conc.:
- 0.313 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Duration:
- 72 h
- Dose descriptor:
- EL10
- Effect conc.:
- 0.836 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Remarks on result:
- other: CI
- Remarks:
- < 0.313 - 1.29
- Duration:
- 72 h
- Dose descriptor:
- EL20
- Effect conc.:
- 1.34 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Remarks on result:
- other: CI
- Remarks:
- 1.15 - 1.60
- Duration:
- 72 h
- Dose descriptor:
- EL50
- Effect conc.:
- 1.88 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- other: yield
- Remarks on result:
- other: CI
- Remarks:
- 1.67 - 2.06
- Details on results:
- - Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test):
- Unusual cell shape: No
- Colour differences: No
- Flocculation: No
- Adherence to test vessels: No
- Aggregation of algal cells: No
- Other:The test media were clear throughout the exposure phase. Microscopic evaluation of the cells at the start and end of exposure revealed no morphological abnormalities. All effect v alues are given based on the nominal test item loadings.
- Effect concentrations exceeding solubility of substance in test medium: No - Results with reference substance (positive control):
- The toxicity of potassium dichromate (SIGMA ALDRICH, batch number MKBV0900V, purity 99.0 %, CAS RN 7778-50-9) to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined over a period of 72 hours from 2016-10-11 to 2016-10-14. The reference item toxicity is in the valid range following test facility SOPS.
EC50-Values of the Reference Item
based on nominal concentrations mg/L, (0-72 hours)
Current Study Valid Range (average ± 3 x SD)
Growth Rate inhibition
ErC50 0.435 0.764 ± 0.513
95% confidence interval 0.414 – 0.458
Yield inhibition
EyC50 0.230 0.402 ± 0.293
95% confidence interval 0.179 – 0.322
SD = Standard deviation - Reported statistics and error estimates:
- ELx-values and statistical Analyses
ELx-values with confidence intervals of growth rate inhibition and for yield inhibition after 72 hours were calculated by sigmoidal dose-response regression.
NOEL, LOEL and statistical analyses
The NOEL and LOEL were determined by calculation of statistically significant differences of growth rates and yield.
The following statistical tests were conducted:
Shapiro-Wilk’s test on normal distribution was calculated with a significance level 0.01.
Levene’s test on variance homogeneity was calculated with a significance level 0.01.
Monotonicity of Concentration/Response was calculated by trend analysis by contrasts (significance level 0.05).
SD Jonckheere-Terpsta test procedure rate was done for growth with a significance level 0.05.
Software
The data for the tables in this report were computer-generated and have been rounded for presentation from the full derived data. Consequently, if calculated manually based on the given data minor deviations may occur from these figures.
Calculations were carried out using software
• Excel, MICROSOFT CORPORATION
• SigmaPlot, SPSS INC.
• GraphPad Prism, GRAPHPAD SOFTWARE, INC.
• ToxRat Version 3.2.1, ToxRat Solutions GmbH - Validity criteria fulfilled:
- yes
- Conclusions:
- In this study, Vat Black 25 was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values (based on nominal test item loadings): The EL50-values with 95 % confidence intervals for inhibition of growth rate (ErL50) after 72 hours were 3.08 (2.92 – 3.36) mg/L, corresponding to 61.6 (58.5 – 67.2) % of the saturated solution, respectively. The EL50-values with 95 % confidence intervals for inhibition of yield (EyL50) after 72 hours were 1.88 (1.67 – 2.06) mg/L, corresponding to 37.5 (33.4 – 41.1) % of the saturated solution, respectively. The NOEL-values for inhibition of growth rate and yield after 72 hours were 1.25 mg/L (corresponding to 25.0 % of the saturated solution) and < 0.313 mg/L (corresponding to < 6.25 % of the saturated solution), respectively.
- Executive summary:
The toxicity of Vat Black 25 (Batch no.:20151007) to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined according to the principles of OECD 201 and Council Regulation (EC) No. 761/2009 C.3 from 2016-11-21 to 2016-12-23, with the definitive exposure phase from 2016-12-20 to 2016-12-23 at the test facility. The aim of the study was the determination of NOEL, LOEL, EL10, EL20- and EL50-values of growth rate and yield over a period of 72 hours.
The study was conducted under static conditions with an initial cell density of 6305 cells/mL. A saturated solution with a nominal loading of 5.00 mg test item/L was prepared with dilution water 24±1 hour prior to the start of the exposure. An appropriate amount of the test item was weighed out. The saturated solution was stirred for 24 ±1hour (1100 rpm) with a magnetic stirrer.Undissolved particles were removed by membrane filtration. The following filtrate, i.e. the saturated solution was used in the test. The solution was checked via laser beam (Tyndall effect), which was negative.
The saturated solution and four dilution levels out of the saturated solution were tested in the definitive test in a geometrical series with a dilution factor of 2:6.25 - 12.5 - 25.0 - 50.0 - 100% of the saturated solution, corresponding to the nominal test item loadings of 0.313 - 0.625 - 1.25 - 2.50 - 5.00 mg/L. Three replicates were tested for each test item concentration and six replicates for the control.The test media were clear throughout the test period.The environmental conditions were within the acceptable limits.
All effect values given are based on the nominal test item loadings of Vat Black 25.
NOEL, LOEL, ELx-values and 95 % Confidence Intervals ofVat Black 25 (0 - 72 hours)
based on the nominal test item loadings
Growth Rate Inhibition
nominal test item loadings[mg/L]
[% saturated solution]
NOEL
1.25
25.0
LOEL
2.50
50.0
ErL10
1.86 (1.66 – 2.03)
37.1 (33.2 – 40.5)
ErL20
2.27 (2.16 – 2.34)
45.4 (43.2 – 46.7)
ErL50
3.08 (2.92 – 3.36)
61.6 (58.5 – 67.2)
Yield Inhibition
nominal test item loadings
[mg/L]
[% saturated solution]
NOEL
< 0.313
< 6.25
LOEL
0.313
6.25
EyL10
0.836 (< 0.313 – 1.29)
16.7 (< 6.25 – 25.7)
EyL20
1.34 (1.15 – 1.60)
26.8 (23.0 – 32.1)
EyL50
1.88 (1.67 – 2.06)
37.5 (33.4 – 41.1)
Reference
Cell Densities
Nominal test item loading |
Replicate |
Cell density [cells/mL] |
|||||||||||
[mg/L] |
[% of the saturated solution] |
No. |
0 hours |
24 hours |
48 hours |
72 hours |
|||||||
5.00 |
100 |
1 |
6305 |
4650 |
3509 |
3317 |
|||||||
2 |
6305 |
5496 |
6426 |
16332 |
|||||||||
3 |
6305 |
3900 |
2756 |
< LOQcell density |
|||||||||
Mean |
6305 |
4682 |
4230 |
n.a. |
|||||||||
2.50 |
50.0 |
1 |
6305 |
16535 |
128052 |
515332 |
|||||||
2 |
6305 |
6939 |
23907 |
489002 |
|||||||||
3 |
6305 |
6897 |
12208 |
471874 |
|||||||||
Mean |
6305 |
10124 |
54722 |
492069 |
|||||||||
1.25 |
25.0 |
1 |
6305 |
30687 |
354576 |
2334737 |
|||||||
2 |
6305 |
28417 |
403429 |
2288675 |
|||||||||
3 |
6305 |
18819 |
258014 |
1803694 |
|||||||||
Mean |
6305 |
25974 |
338673 |
2142369 |
|||||||||
0.625 |
12.5 |
1 |
6305 |
27526 |
312674 |
2338909 |
|||||||
2 |
6305 |
33012 |
402453 |
2555803 |
|||||||||
3 |
6305 |
30765 |
366145 |
2305137 |
|||||||||
Mean |
6305 |
30434 |
360424 |
2399950 |
|||||||||
0.313 |
6.25 |
1 |
6305 |
29677 |
352151 |
2344603 |
|||||||
2 |
6305 |
26159 |
342871 |
2338740 |
|||||||||
3 |
6305 |
29218 |
311637 |
2195140 |
|||||||||
Mean |
6305 |
28351 |
335553 |
2292828 |
|||||||||
Control |
1 |
6305 |
35919 |
418020 |
2578693 |
||||||||
2 |
6305 |
33892 |
418866 |
2682714 |
|||||||||
3 |
6305 |
33531 |
367284 |
2751667 |
|||||||||
4 |
6305 |
30441 |
372702 |
2526316 |
|||||||||
5 |
6305 |
31459 |
363388 |
2358473 |
|||||||||
6 |
6305 |
31989 |
367955 |
2577115 |
|||||||||
Mean |
6305 |
32872 |
384703 |
2579163 |
LOQcell density = Limit of quantification of the cell density (2736 cells/mL).
n.a. = not applicable
Evaluation after 72 hours
Statistically
significant differences of growth rates and yield compared to
control
values are marked (+), not significant differences are marked (-).
Nominal test item loading |
Replicate |
Growth rate |
Inhibition of growth rate |
Yield |
Inhibition of yield |
|||
[mg/L] |
[% of the saturated solution] |
No. |
[d-1] |
[%] |
[cells/mL] |
[%] |
||
5.00 |
100 |
1 |
|
-0.214 |
100 |
|
-2988 |
100 |
2 |
|
0.317 |
84 |
|
10027 |
100 |
||
3 |
|
n.a. |
100 |
|
n.a. |
100 |
||
Mean |
(+) |
n.a. |
95 |
(+) |
n.a. |
100 |
||
2.50 |
50.0 |
1 |
|
1.47 |
27 |
|
509027 |
80 |
2 |
|
1.45 |
28 |
|
482697 |
81 |
||
3 |
|
1.44 |
28 |
|
465569 |
82 |
||
Mean |
(+) |
1.45 |
28 |
(+) |
485764 |
81 |
||
1.25 |
25.0 |
1 |
|
1.97 |
2 |
|
2328432 |
10 |
2 |
|
1.96 |
2 |
|
2282370 |
11 |
||
3 |
|
1.89 |
6 |
|
1797389 |
30 |
||
Mean |
(-) |
1.94 |
3 |
(+) |
2136064 |
17 |
||
0.625 |
12.5 |
1 |
|
1.97 |
2 |
|
2332604 |
9 |
2 |
|
2.00 |
0 |
|
2549498 |
1 |
||
3 |
|
1.97 |
2 |
|
2298832 |
11 |
||
Mean |
(-) |
1.98 |
1 |
(+) |
2393645 |
7 |
||
0.313 |
6.25 |
1 |
|
1.97 |
2 |
|
2338298 |
9 |
2 |
|
1.97 |
2 |
|
2332435 |
9 |
||
3 |
|
1.95 |
3 |
|
2188835 |
15 |
||
Mean |
(-) |
1.97 |
2 |
(+) |
2286523 |
11 |
||
Control |
1 |
|
2.00 |
|
|
2572388 |
|
|
2 |
|
2.02 |
|
|
2676409 |
|
||
3 |
|
2.03 |
|
|
2745362 |
|
||
4 |
|
2.00 |
|
|
2520011 |
|
||
5 |
|
1.97 |
|
|
2352168 |
|
||
6 |
|
2.00 |
|
|
2570810 |
|
||
Mean |
|
2.00 |
|
|
2572858 |
|
n.a. = not applicable
Section-by-Section and Average Specific Growth Ratesof the Control Group (0 - 72 hours)
|
Replicate No. |
Specific growth rate [d-1] |
Mean (0 - 72 hours) |
SD ± |
CV |
Mean CV [%] |
||
section-by-section |
||||||||
0 - 24 hours |
24 - 48 hours |
48 - 72 hours |
||||||
Control |
1 |
1.74 |
2.45 |
1.82 |
2.00 |
0.391 |
19.5 |
20.8 |
2 |
1.68 |
2.51 |
1.86 |
2.02 |
0.439 |
21.8 |
||
3 |
1.67 |
2.39 |
2.01 |
2.03 |
0.361 |
17.8 |
||
4 |
1.57 |
2.50 |
1.91 |
2.00 |
0.471 |
23.6 |
||
5 |
1.61 |
2.45 |
1.87 |
1.97 |
0.429 |
21.7 |
||
6 |
1.62 |
2.44 |
1.95 |
2.00 |
0.412 |
20.6 |
||
|
|
|
Mean |
2.00 |
|
|
||
|
|
|
SD ± |
0.02 |
|
|||
|
|
|
CV [%] |
0.89 |
|
SD = Standard deviation CV = Coefficient of variation
Description of key information
Key value determined in a GLP accredited laboratory study performed in accordance with OECD Guideline 201 (Alga, Growth Inhibition Test).
According to this test, the test item does have toxic effects on alga.
Key value for chemical safety assessment
- EC50 for freshwater algae:
- 3.08 mg/L
- EC10 or NOEC for freshwater algae:
- 1.25 mg/L
Additional information
In a GLP study conducted in accordance with OECD Guideline 201, Vat Black 25 was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values (based on nominal test item loadings): The EL50-values with 95 % confidence intervals for inhibition of growth rate (ErL50) after 72 hours were 3.08 (2.92 – 3.36) mg/L, corresponding to 61.6 (58.5 – 67.2) % of the saturated solution, respectively. The EL50-values with 95 % confidence intervals for inhibition of yield (EyL50) after 72 hours were 1.88 (1.67 – 2.06) mg/L, corresponding to 37.5 (33.4 – 41.1) % of the saturated solution, respectively. The NOEL-values for inhibition of growth rate and yield after 72 hours were 1.25 mg/L (corresponding to 25.0 % of the saturated solution) and < 0.313 mg/L (corresponding to < 6.25 % of the saturated solution), respectively.
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.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.