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EC number: 265-449-9 | CAS number: 65113-55-5
- 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
Long-term toxicity to aquatic invertebrates
Administrative data
Link to relevant study record(s)
- Endpoint:
- long-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- From 2017-01-24 to 2017-11-24
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 211 (Daphnia magna Reproduction Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: container tightly closed, stored in a dry, cool (room temperature) and well-ventilated place
- Stability under test conditions: stable
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: see "Preparation and application of test solution"
- Analytical monitoring:
- yes
- Remarks:
- The method was validated prior to this study according to SANCO 3029/99 rev.4 (2000).
- Details on sampling:
- - Concentrations: all concentration levels and the control
- Sampling method:
At the start of the exposure intervals, samples of the fresh media were taken after preparation of all test item concentrations and analyzed.
At the end of the exposure intervals (48 or 72 hours), samples of the old media were taken directly from the test vessels.
For the longest exposure interval of 72 hours samples were taken at the start (0 hours) and at the end of the exposure interval (72 hours) once within the test period.
- Sample storage conditions before analysis: stored at room temperature until sample preparation and at room temperature until the start of the analysis (in an antosampler), if necessary. - Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:
A saturated solution with a nominal concentration of 1.00 mg/L of the test item was freshly prepared with dilution water, one day prior to the start of the exposure (at -24 hours). The test item was mixed with the dilution water by constant stirring with a magnetic stirrer at approximately 1100 rpm for 24 hours at room temperature. After completion of stirring, 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 were discarded. The filtration was interrupted for 15 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. The following filtrate, i.e. the saturated solution, was for the preparation of five concentration levels. During filtration, the filter was always be kept covered.
5 concentration levels of the test item in a geometric series with a separation factor of √10 were prepared by diluting the saturated solution with dilution water.
- Controls: 10 replicates of dilution water without test item
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): Evidence of undisolved particles only during the preparation of the saturated solution (1mg test item/L), before filtration. Undissolved particles were removed by membrane filtration during the preparation. - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: STRAUS (clone 5)
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), 14195 Berlin, Germany
- Age of parental stock : Less than 24 hours old daphnids. No first brood progeny was used for the test.
- Feeding during test
- Food type: A suspension of Pseudokirchneriella subcapitata and Desmodesmus subspicatus. Origine : Sammlung von Algenkulturen (SAG), Pflanzenphysiologisches Institut der Universität Göttingen, Nikolausberger Weg 18, 37073 Göttingen, Germany
- Amount: 0.2 mg C per Daphnia and day
- Frequency: daily
ACCLIMATION
- Acclimation period: Acclimatization was not necessary, because the composition of the dilution water was equivalent to the culture medium.
- Test type:
- semi-static
- Water media type:
- other: reconstituted water
- Remarks:
- Culture Medium Elendt M4, according to OECD 202, Annex 3(2004)
- Limit test:
- no
- Total exposure duration:
- 21 d
- Hardness:
- 160 to 180 mg CaCO3/L
- Test temperature:
- 20°C + or - 1°C
- pH:
- 6 to 9
- Dissolved oxygen:
- > 3 mg/L
- Nominal and measured concentrations:
- Nominal concentrations :
* 0.100 % of the saturated solution
* 0.316% of the saturated solution
* 1.00 % of the saturated solution
* 3.16% of the saturated solution
* 10.0% of the saturated solution
Geometric mean measured concentrations :
* 0.308 µg/L of the test item
* 0.829 µg/L of the test item
* 2.42 µg/L of the test item
* 9.42 µg/L of the test item
* 42.6 µg/L of the test item - Details on test conditions:
- TEST SYSTEM
- Test vessel: Glass beakers (5 (ID) x 8 (H) cm), 100 mL capacity, with a test volume of 50 mL loosely covered with watch glasses
- Aeration: none
- Renewal rate of test solution (frequency): three times per week (after 48 or 72 hours; i.e Monday, Wednesday and Friday). For this purpose, a second set of test vessels was filled with the freshly prepared test solutions and the daphnids were transferred with a pipette. (50 g test solution per replicate was weighed out into each test vessel. This corresponds to 50 mL. The daphnids were transferred in a small amount of dilution water or test solution by pipette).
- No. of organisms per vessel: 1 daphnid
- No. of vessels per concentration (replicates): 10 replicates
- No. of vessels per control (replicates): 10 replicates
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4 according to OECD 202, Annex 3 (2004), modified to a total hardness of 160 to 180 mg CaCO3/L.
- Intervals of water quality measurement:
* Measurements once within 7 days, in fresh media at the start (0 hours) and in old media at the end (48 or 72 hours) of an exposure interval, in one replicate of the control and the highest test concentration.
* The water quality parameters in fresh media at the start of an exposure interval were measured in an additional replicate without daphnids of the highest test concentration and the control. At the end of an exposure interval, the water quality parameters of the old media were measured in a test vessel of the highest test concentration and the control, which contained daphnids and food algae. The temperature in the incubator was recorded throughout the test period with a thermo-hygrograph.
OTHER TEST CONDITIONS
- Photoperiod: 16/8 hours light/dark cycle
- Light intensity: Max. 1500 lx
TEST CONCENTRATIONS
- Spacing factor for test concentrations: √10
- Range finding study : Yes
- Test concentrations: saturated solution with a nominal concentration of 1.00 mg/L and two further dilution levels prepared by diluting the saturated solution by factor 10 or 100 with dilution water.
- Results used to determine the conditions for the definitive study: yes - Reference substance (positive control):
- not required
- Duration:
- 21 d
- Dose descriptor:
- EC10
- Effect conc.:
- ca. 1.64 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- reproduction
- Remarks on result:
- other:
- Remarks:
- with 95% confidence limits : 0.743 - 3.62 µg/L
- Key result
- Duration:
- 21 d
- Dose descriptor:
- EC50
- Effect conc.:
- ca. 15.2 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- reproduction
- Remarks on result:
- other:
- Remarks:
- with 95% confidence limits: 5.72 - 39.4 µg/L
- Duration:
- 21 d
- Dose descriptor:
- LOEC
- Effect conc.:
- ca. 0.829 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- reproduction
- Key result
- Duration:
- 21 d
- Dose descriptor:
- NOEC
- Effect conc.:
- ca. 0.308 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- reproduction
- Duration:
- 21 d
- Dose descriptor:
- other: LC 20
- Effect conc.:
- ca. 10.7 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Remarks on result:
- other:
- Remarks:
- with confidence limits : 9.42 - 42.6 µg/L
- Duration:
- 21 d
- Dose descriptor:
- LC50
- Effect conc.:
- ca. 12.7 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Remarks on result:
- other:
- Remarks:
- with confidence limits : 9.42 - 42.6 µg/L
- Duration:
- 21 d
- Dose descriptor:
- LOEC
- Effect conc.:
- ca. 42.6 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Duration:
- 21 d
- Dose descriptor:
- NOEC
- Effect conc.:
- ca. 9.42 µg/L
- Nominal / measured:
- meas. (geom. mean)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Details on results:
- 1- Preliminary test (48 hours)
The results of the biological part are presented in Table 1. Analytical results are provided in Table 2.
2- Definitive test (21 days)
- Observations on body length and weight:
The mean values of the body length (excluding the anal spine) of the survived parental daphnids in the tested concentration levels 0.308 to 9.42 µg/L were in the range of 4.11 to 4.20 mm per daphnid and 4.28 mm per daphnid in the control group.
At the highest test concentration level 42.6 µg/L no parent animals survived until the end of the exposure.
The mean dry weight of the survived parental daphnids was in the range of 0.49 to 0.70 mg per daphnid in the concentration levels of 0.308 to 9.42 µg/L and 0.66 mg per daphnid in the control (see table 4).
- Other biological observations:
* number of juveniles per parent : The number of juveniles in all replicates of the control and the test groups was counted and recorded every day. Results concerning the number of living juveniles of all concentration levels and the control for every test day are presented in tables in attached files.
The average number of living juveniles at the end of the test after 21 days per survived parental daphnid was 70.1 in the control group and thus the validity criterion for the reproduction of a mean of ≥ 60 living offspring per survived parent animal in the control was met.
* First appearance of living juvenile:
The first appearance of living juveniles was observed between days 8 and 9 at the introduced parental daphnids of the control and the geometric mean measured concentrations of 0.308 – 0.829 – 2.42 – 9.42 – 42.6 µg/L of the test item. Two parental daphnids of the highest test concentration of 42.6 µg/L died before producing juveniles (see table 5).
* Intrisic rates of natural increase (IR)
The intrinsic rates of natural increase (IR) of the survived parental daphnids accounting for generation time and number of offspring were used for calculation of population growth and maintenance. (see table 6).
* Stillborn Juveniles and aborted eggs
No stillborn juveniles or aborted eggs were observed in the control during the exposure period of 21 days. In all tested concentration levels stillborn juveniles were observed. In the test concentrations of 2.42 and 9.42 µg/L also aborted eggs were observed. For details, see Tables 7 and Table 8. Related to the total number of produced juveniles (dead + alive) per survived daphnid, the percentage of dead juveniles was ≤ 28% in the concentration levels 0.308 to 9.42 µg/L.
* number of dead parents: Accidental and inadvertent mortality as defined in the OECD test guideline and abnormalities were not observed. The test item induced statistically significant adult mortality of 100% after 21 days in the concentration level 42.6 µg/L (Step down Cochran-Armitage Test Procedure, α = 0.05). In the concentration levels of 0.829 and 9.42 µg/L, a mortality rate of 10% of the introduced adults was observed, which is not statistically significant. In the concentration levels of 0.308 and 2.42 µg/L and in the control all parental daphnids survived until the end of the test (21 days) (see table 9). The Cochran-Armitage test procedure revealed a significant increasing trend (p ≤ 0.05) in adult mortality. Therefore, the LC20- and LC50-values were calculated by sigmoidal dose-response regression.
The LC20 for the test item was 10.7 (9.42 – 42.6) mg/L. The LC50 was 12.7 (9.42 – 42.6) mg/L. The LOEC was 42.6 mg/L. The NOEC was 9.42 mg/L. Details are provided in table 9
* Reproductive output:
No parental daphnids were excluded due to accidental or inadvertent mortality from the evaluation of the reproductive output. Since the Cochran-Armitage test procedure revealed a significant increasing trend (p ≤ 0.05) in adult mortality, the evaluation was done based on the cumulative offspring per introduced parent. A statistically significant reduction of the reproductive output in comparison to the reproductive output in the control was determined at the concentration levels 0.829 and 42.6 µg/L (William’s multiple sequential t-test procedure, α = 0.05). The determination of the EC10- and the EC50-value and their 95% confidence limits was carried out by point estimates from the 3 parametric normal CDF.
The EC10 for the test item was 1.64 µg/L (95% confidence limits: 0.743 – 3.62 µg/L). The EC50 was 15.2 µg/L (95% confidence limits: 5.72 – 39.4 µg/L). The LOEC was 0.829 µg/L. The NOEC was 0.308 µg/L (see table 10).
- Mortality of control: No mortality
- Other adverse effects control: No adverse effects
• other measurements throughout the test (dissolved oxygen, pH, hardness, temperature)
The measured water quality parameters (i.e. pH-value, dissolved oxygen concentration, total water hardness and water temperature) were within the acceptable limits. For results, see Tables 11 to 14.
The incubator temperature was in the range of 19 to 21 °C.
• Measured Exposure Concentrations in the Test Vessels during the Definitive Test
The measured concentrations of the test item in fresh media at the start of the respective exposure intervals (0 hours) were in the range of 0.582 to 74.9 µg/L of the test item. At the end of the respective exposure intervals (48 and 72 hours), the measured test item concentrations in the old media were in the range of 0.150 to 41.5 µg/L. The analytical results are presented in Table 15.
The geometric mean measured concentrations were calculated and used for evaluation of the effect-concentrations.
- Abnormal responses:
* Presence of Males : No males were observed in the control or in the test groups during the test.
* Occurence of Ephippia (winter eggs) : No Ephippia were observed in the control or in the test groups during the test. - Reported statistics and error estimates:
- 1-Number of Living Juveniles per Introduced Parental Daphnid (based on the geometric mean measured concentrations of the test item [µg/L])
- Shapiro-Wilk´s Test on Normal Distribution
Number of residuals = 45; Shapiro-Wilk´s W = 0.989; p(W) = 0.938; (p > 0.01) .
- Levene´s Test on Variance Homogeneity (with Residuals), (p > 0.010).
-Trend analysis by Contrasts, The linear trend and the quadratic trend are significants (p ≤ 0.05) The analysis of contrasts revealed a linear trend.
-Williams Multiple Sequential t-test Procedure
A NOEC of 0.308 µg/L is suggested by the program.
2- Calculation of the EC10 and the EC50 for the Reduction of the Reproductive Output of the Introduced Parental Daphnids (based on the geometric mean measured concentrations of the test item [µg/L])
-Estimated parameters of the 3-param. normal CDF
R²: 0.801; adjusted R²: 0.794.Residual standard error: 9.26941.Akaike Criterion (AIC): 332.856.Shapiro Wilk`s test on normal distribution of residuals: p = 0.886.
-Analysis of Variance and Test for Lack of Fit for the 3-param. normal CDF
p(F|Regression) ≤ 0.05, a significant amount of variance is explained by the regression model.
p(F|Lack of Fit) > 0.05, there is no significant lack of fit.
- Point estimates from the 3-param. normal CDF
The confidence limits of the EC10 used as a parameter were computed by means of the standard error of parameter b1; confidence limits for the remaining ECx were estimated by Monte-Carlo simulation using the parameter errors obtained from the inverse Hessian matrix (1000 runs).
3-Adult Mortality
- Qualitative Trend Analysis by Contrasts
The linear trend and quadratic trend are significants (p≤ 0.05)
- Step-down Cochran-Armitage Test Procedure
A NOEC of 9.42 µg/L is suggested by the program.
Calculation of the LC20 and the LC50 for the Adult Mortality
Equation:Y=Bottom + (Top-Bottom)/(1+10^((LogEC50-X)*HillSlope))
Transform: x= log(x) - Validity criteria fulfilled:
- yes
- Conclusions:
- The effects on reproduction were evaluated based on the reproduction per introduced parent animal, since this parameter is the ecologically most relevant response variable and is required by the OECD test guideline 211 (2012), when a significant trend in mortality is detected.
The overall effect threshold for effects of the test item under the test conditions was 0.308 µg/L (NOEC) and 0.829 µg/L (LOEC) based on the geometric mean measured concentrations of the test item. - Executive summary:
A Daphnia magna reproduction test (semi-static, 21 d) of the test item was conducted at the test facility according to OECD 211 (2012) from 2017-01-24 to 2017-11-24, with the definitive exposure phase from 2017-11-03 to 2017-11-24.
Test species was Daphnia magna STRAUS (Clone 5). Ten daphnids, held individually, were used per concentration level and control. At test start the daphnids were 2 to 24 hours old. The study was carried out under semi-static conditions with a medium renewal three times per week (i.e. on Monday, Wednesday and Friday). Aim of the test was to assess the effects on the reproduction capacity and other test item-related effects or parameters such as the intrinsic rate of the natural increase, first appearance of living juveniles, occurrence of aborted eggs and stillborn juveniles, adult mortality, body length and dry weight of the parental daphnids.
The test item is a black powder, which is hardly soluble in water. Five dilution levels of a saturated solution of the test item were selected based on the results of previous tests under GLP conditions as follows: 0.100 - 0.316 - 1.00 - 3.16 - 10.0% of saturated solution.
A stock solution with a nominal concentration of 1.00 mg/L was prepared with dilution water and was used as the highest concentration level and for the preparation of further four concentration levels in the range of 0.100 - 0.316 - 1.00 - 3.16 - 10.0% of the saturated solution.
The test item concentrations were analytically verified via LC-MS/MS in fresh media at the start of the exposure intervals on study days 0, 10 and 17 (0 hours) and in old media at the end of the exposure intervals on study days 3, 12 and 19 (48 or 72 hours, respectively) in all concentration levels and the control.
The measured concentrations of the test item in fresh media at the start of the respective exposure intervals (0 hours) were in the range of 0.582 to 74.9 µg/L of the test item. At the end of the respective exposure intervals (48 and 72 hours), the measured test item concentrations in the old media were in the range of 0.150 to 41.5 µg/L. T
The geometric mean measured concentrations were calculated and used for evaluation of the effect-concentrations. The geometric mean measured concentrations are: 0.308 – 0.829 – 2.42 – 9.42 – 42.6 µg/L.
The environmental conditions were within the acceptable limits. The validity criteria of the test guideline were met
The effects on reproduction were evaluated based on the reproduction per introduced parent animal, since this parameter is the ecologically most relevant response variable and is required by the OECD test guideline 211 (2012), when a significant trend in mortality is detected. A summary of all endpoints based on the geometric mean measured concentrations of the test item is given in Table 1.
Table 1: Endpoints for Reproduction and Mortality (based on the geometric mean measured concentrations of the test item)
Effect values
Test Item
Geometric mean measured concentration of the test item
[µg/L]
EC10 Reproduction
(with 95%confidence limits)
1.64
(Cl: 0.743 – 3.62)
EC50 Reproduction
(with 95%confidence limits
15.2
(Cl: 5.72 – 39.4)
LOECReproduction
0.829
NOECReproduction
0.308
LC20 Adult mortality after 21 days(withconfidence limits)
10.7
(CI: 9.42 – 42.6)
LC50 Adult mortality after 21 days
(withconfidence limits)12.7
(CI: 9.42 – 42.6)
LC100 Adult mortality after 21 days
42.6
LOECAdult mortality after 21 days
42.6
NOECAdult mortality after 21 days
9.42
Reference
Table 1: Immobilization Rates in the non-GLP Preliminary Range Finding Test
(n = 20, divided into 2 replicates with 10 daphnids each)
Dilution level of the saturated solution [%] |
IMMOBILIZATION [%] |
|||||
24 hours |
48 hours |
|||||
Replicates |
Replicates |
|||||
1 |
2 |
MV |
1 |
2 |
MV |
|
100* |
100 |
100 |
100 |
100% mortality after 24 hours |
||
10 |
0 |
0 |
0 |
0 |
0 |
0 |
1 |
0 |
0 |
0 |
0 |
0 |
0 |
Control |
0 |
0 |
0 |
0 |
0 |
0 |
* = saturated solution
Table 2: Measured Concentrations of Solvant Black 46 during the non-GLP preliminary stability test
Sampling date |
2016-08-23 0 hours Start of the exposure |
2016-08-25 48 hours End of the exposure |
|
|
Dilution level of the saturated solution [%] |
Solvant Black 46 1) |
Geometric mean measured test item concentration [mg/L] |
||
Meas. conc. [mg/L] |
Meas. conc. [mg/L] |
% |
||
100* |
0.575 |
0.571 |
99 |
0.573 |
10 |
0.0584 |
0.0604 |
103 |
0.0594 |
Control |
< 0.001 |
< 0.001 |
|
Meas. conc.= measured concentration of the test item, dilution factors taken into account
% = percent of the initially measured concentration of the test item
* = saturated solution
Table 4 : Total Body Length and Dry Weight of the Survived Parental Daphnids
Geometric mean measured concentration |
Total length of the survived parent animals [mm] |
MV |
N |
Dry weight [mg] |
||||||||||
Replicate no. |
|
|
||||||||||||
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
[mm] |
|
S |
MV |
|
42.6 |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
0 |
--- |
--- |
9.42 |
4.25 |
4.00 |
4.25 |
4.00 |
4.00 |
4.50 |
4.00 |
4.00 |
4.00 |
--- |
4.11 |
9 |
4.40 |
0.49 |
2.42 |
4.50 |
4.50 |
4.25 |
4.00 |
4.50 |
3.75 |
4.25 |
4.00 |
4.25 |
4.00 |
4.20 |
10 |
7.00 |
0.70 |
0.829 |
4.00 |
4.00 |
4.00 |
4.50 |
4.00 |
4.25 |
4.00 |
4.00 |
4.25 |
--- |
4.11 |
9 |
6.30 |
0.70 |
0.308 |
4.50 |
4.50 |
4.00 |
4.50 |
4.00 |
4.00 |
4.00 |
4.00 |
4.00 |
4.50 |
4.20 |
10 |
7.00 |
0.70 |
Control |
4.00 |
4.00 |
4.25 |
4.25 |
4.50 |
4.50 |
4.25 |
4.50 |
4.25 |
4.25 |
4.28 |
10 |
6.60 |
0.66 |
N = number of the survived parental daphnids
-- = not applicable, due to the mortality of the parental daphnid(s)
Table 5 : First Appearance of Living Juveniles in the Individual Groups
Geometric mean measured concentration |
Day of first appearance of living juveniles at the introduced parental daphnids in replicate no. |
First |
|||||||||
appearance |
|||||||||||
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
mean day |
|
42.6 |
9 |
8 |
9 |
9 |
8 |
8 |
9 |
9 |
--- |
--- |
8.6 |
9.42 |
9 |
9 |
9 |
9 |
9 |
8 |
8 |
9 |
9 |
9 |
8.8 |
2.42 |
8 |
9 |
8 |
9 |
9 |
9 |
8 |
8 |
9 |
9 |
8.6 |
0.829 |
9 |
8 |
9 |
9 |
9 |
9 |
9 |
9 |
9 |
8 |
8.8 |
0.308 |
9 |
8 |
8 |
9 |
9 |
10 |
8 |
8 |
9 |
9 |
8.7 |
Control |
9 |
9 |
8 |
8 |
9 |
9 |
9 |
9 |
9 |
9 |
8.8 |
-- = not applicable, due to the mortality of the parental daphnid(s)
Table 6 : Intrinsic Rates of Natural Increase
Geometric mean measured concentration |
Intrinsic rate of natural increase in replicate no. |
Mean IR |
CV |
|||||||||
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
MV±SD |
[%] |
|
42.6 |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
--- |
|
9.42 |
0.339 |
0.353 |
0.334 |
0.355 |
0.354 |
0.343 |
0.376 |
0.343 |
0.353 |
--- |
0.350±0.0124 |
3.6 |
2.42 |
0.383 |
0.302 |
0.397 |
0.363 |
0.362 |
0.326 |
0.444 |
0.383 |
0.380 |
0.374 |
0.371±0.0385 |
10.4 |
0.829 |
0.352 |
0.383 |
0.353 |
0.340 |
0.359 |
0.354 |
0.363 |
0.369 |
0.351 |
--- |
0.358±0.0123 |
3.4 |
0.308 |
0.354 |
0.384 |
0.395 |
0.362 |
0.363 |
0.310 |
0.397 |
0.372 |
0.351 |
0.368 |
0.366±0.0251 |
6.9 |
Control |
0.372 |
0.352 |
0.369 |
0.384 |
0.359 |
0.383 |
0.340 |
0.361 |
0.365 |
0.366 |
0.365±0.0133 |
3.6 |
-- = not applicable, due to the mortality of the parental daphnid(s)
Table 7 : Stillborn Juveniles and Aborted Eggs produced by the Survived Parental Daphnids after 21 Days
Geometric mean measured concentration |
Number of |
Total no.
å |
Number of N |
|
stillborn juveniles |
aborted eggs |
|||
42.6 |
9 |
0 |
9 |
0 |
9.42 |
122 |
18 |
140 |
9 |
2.42 |
86 |
6 |
92 |
10 |
0.829 |
97 |
0 |
97 |
9 |
0.308 |
136 |
0 |
136 |
10 |
Control |
0 |
0 |
0 |
10 |
N = number of the survived parental daphnids
Table 8 : Relative Number of Dead (Stillborn + Aborted Eggs) to Total Number of Juveniles per Survived Parental Daphnids
Geometric mean measured concentration |
Number of juveniles |
Percentage of dead juveniles#) [%] |
||
Dead |
Alive |
Total |
||
42.6 |
0 |
0 |
0 |
Not applicable, due to 100% mortality |
9.42 |
140 |
356 |
496 |
28 |
2.42 |
92 |
627 |
719 |
13 |
0.829 |
97 |
574 |
671 |
14 |
0.308 |
136 |
656 |
792 |
17 |
Control |
0 |
701 |
701 |
0 |
Dead = aborted eggs + stillborn juveniles
Total = dead + alive juveniles
#) = related to the total number of juveniles per survived parental daphnid
Table 9 : Mortality (%) of the adult Daphnids after 7, 14 and 21 days of Exposure (n = 10)
Geometric mean measured concentration |
Adult Mortality [%] |
||
7 days |
14 days |
21 days |
|
42.6 |
0 |
70 |
1001) |
9.42 |
0 |
10 |
10 |
2.42 |
0 |
0 |
0 |
0.829 |
0 |
0 |
10 |
0.308 |
0 |
0 |
0 |
Control |
0 |
0 |
0 |
1)= statistically significant (Step‑down Cochran-Armitage Test Procedure,α = 0.05)
Table 10 : Effects on Reproduction for all Introduced and for Surviving Parents
(based on the geometric mean measured concentrations of the test item)
Geometric mean measured concentration of the test item [µg/L] |
Mean number of offspring |
Mean number of offspring |
||||
Mean |
SD |
CV |
Mean |
SD |
CV |
|
42.6 |
19.7 |
11.8 |
60.0 |
--- |
--- |
|
9.42 |
38.9 |
9.34 |
24.0 |
39.6 |
9.66 |
24.4 |
2.42 |
62.7 |
8.83 |
14.1 |
62.7 |
8.83 |
14.1 |
0.829 |
61.6 |
9.54 |
15.5 |
63.8 |
7.00 |
11.0 |
0.308 |
65.6 |
5.78 |
8.81 |
65.6 |
5.78 |
8.81 |
Control |
70.1 |
8.06 |
11.5 |
70.1 |
8.06 |
11.5 |
-- = not applicable, due to the mortality of the parental daphnid(s)
Water Quality Parameters
Comparison of water quality parameters in fresh and old medium once within 7 days.
Table 11 : pH Values
Geometric mean measured concentration |
fresh |
old |
fresh |
old |
fresh |
old |
Day 0 |
Day 3 |
Day 10 |
Day 12 |
Day 17 |
Day 19 |
|
42.6 |
7.49 |
7.97 |
7.79 |
7.47 |
7.53 |
7.33 |
Control |
7.99 |
7.97 |
7.98 |
7.71 |
7.85 |
7.10 |
The pH-values should be in the range of 6 – 9. Significant deviations≥1.5 units between the initial and final pH-values were not observed.
Table 12 : Dissolved Oxygen Concentration [mg/L]
Geometric mean measured concentration |
fresh |
old |
fresh |
old |
fresh |
old |
Day 0 |
Day 3 |
Day 10 |
Day 12 |
Day 17 |
Day 19 |
|
42.6 |
8.74 |
9.66 |
8.64 |
8.31 |
8.84 |
7.95 |
Control |
9.18 |
9.38 |
8.91 |
9.46 |
8.87 |
6.71 |
The dissolved oxygen concentration was above 3 mg/L.
Table 13 : Total Hardness [mg/L] as CaCO3
Geometric mean measured concentration |
fresh |
old |
fresh |
old |
fresh |
old |
Day 0 |
Day 3 |
Day 10 |
Day 12 |
Day 17 |
Day 19 |
|
42.6 |
174 |
167 |
174 |
178 |
175 |
194 |
Control |
178 |
171 |
180 |
178 |
167 |
182 |
The total hardness was above 140 mg/L as CaCO3.
Table 14 : Temperature [°C]
Geometric mean measured concentration |
fresh |
old |
fresh |
old |
fresh |
old |
Day 0 |
Day 3 |
Day 10 |
Day 12 |
Day 17 |
Day 19 |
|
42.6 |
20.6 |
20.4 |
20.6 |
20.8 |
20.7 |
20.6 |
Control |
20.1 |
20.3 |
20.7 |
20.8 |
20.5 |
20.7 |
The temperature was within the range of 20±1°C
Table 15: Measured Concentrations of the Test Item during the Definitive Test
Sampling date |
Day 0 Fresh media |
Day 3 Old media |
Day 10 Fresh media |
Day 12 Old media |
Day 17 Fresh media |
Day 19 Old media |
|
Nominal test item conc. [% of the sat. sol.] |
Solvant Black 46 |
Geometric mean measured conc. [µg/L] |
|||||
Meas. |
Meas. |
Meas. |
Meas. |
Meas. |
Meas. |
||
10.0 |
57.0 |
21.3 |
74.9 |
41.5 |
56.6 |
28.1 |
42.6 |
3.16 |
17.3 |
4.58 |
21.9 |
5.85 |
17.2 |
4.00 |
9.42 |
1.0 |
5.23 |
1.40 |
7.13 |
0.894 |
5.73 |
0.756 |
2.42 |
0.316 |
1.80 |
0.398 |
2.21 |
0.402 |
1.72 |
0.296 |
0.829 |
0.100 |
0.582 |
< LOQ* |
0.699 |
< LOQ* |
0.622 |
< LOQ* |
0.308 |
Control |
< LOQ |
< LOQ |
< LOQ |
< LOQ |
< LOQ |
< LOQ |
|
Meas. conc. = measured concentration of the test item, dilution and weighing factors taken into account
% = percent of the nominal concentration of the test item
Sat. sol. = saturated solution (1 mg test item/L)
LOQ = limit of quantification of the analytical method (0.3 µg test item/L)
* = ½ LOQ (0.150 µg test item/L) was used for calculation of the geometric mean measured concentration
Description of key information
Long term NOEC for reproductive toxicity to daphnids : 0.308 µg/L (OECD TG 211; semi-static)
Key value for chemical safety assessment
Fresh water invertebrates
Fresh water invertebrates
- Effect concentration:
- 0.308 µg/L
Additional information
The reproductive NOEC long-term toxicity was 0.308 µg/L (geometric mean of the measured concentrations)
The mortality NOEC long-term toxicity was 9.42 µg/L (geometric mean of the measured concentrations)
The EC10 reproduction was 1.64 µg/L with 95% C.I. of 0.743 - 3.62
The LC20 mortality was 10.7 µg/L with 95% of 9.42 - 42.6
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