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Toxicity to aquatic algae and cyanobacteria

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Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 6, 2011 - July 7, 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
adopted March 23, 2006
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Version / remarks:
Council Regulation (EC) No. 761/2009 laying down test methods pursuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Remarks:
Because of the low water solubility (0.0028 mg/L), the compound cannot be detected with standard analytical methods and the development of an analytical method with a sufficiently low detection and quantification limit is complex.
Vehicle:
yes
Details on test solutions:
Vehicle:

Nutrient (mg/L)
NH4Cl 15.0
MgCl2 * 6H2O 12.0
CaCl2 * 2H2O 18.0
MgSO4 * 7H2O 15.0
KH2PO4 1.60

FeCl3 * 6H2O 0.0640
Na2EDTA * 2H2O 0.100

H3BO3 0.185
MnCl2 * 4H2O 0.415
ZnCl2 0.00300
CoCl2 * 6H2O 0.00150
CuCl2 * 2H2O 0.00001
Na2MoO4 * 2H2O 0.00700

NaHCO3 50.0

- pH: 7.96
- Total hardness: 24 mg/L as CaCO3 (calculated)

Preparation of the Test Item:
The test medium (reconstituted water and test material) was freshly prepared. Therefore, the calibrated flask with test material and vehicle (reconstituted water) was treated in an ultrasonic device for 1 hour. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Afterwards, the formulation was given through a nutsch filter (pore size 10 - 16 µm) and the filtrate was used for the study.

Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Unicellular green algae
- Strain: Desmodesmus subspicatus
- Source (laboratory, culture collection): The test system used was Desmodesmus subspicatus, strain 86.81 SAG (Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universität Göttingen, Germany). This strain was further cultivated in the laboratories of Non-Clinical Safety.
- Method of cultivation: The permanent algae cultures, the pre-culture and the algae cultures of the study were cultivated in an air-conditioned room with a water temperature of 21.0 – 24.0°C, controlled at ± 2°C. For cultivation, 300 mL Erlenmeyer flasks filled with 100 mL algae suspension were covered with air permeable stoppers (Heinz Herenz Medizinalbedarf GmbH, Hamburg, Germany). The cultures were shaken continuously at about 120 rpm (Universalschüttler SM25, Edmund Bühler GmbH, Hechingen, Germany) and lit between 4440 and 8880 Lux. Fluorescent tubes (Philips Master TL5 – 80 W/840 HO) installed above the flasks served for lighting.


ACCLIMATION
- Acclimation period: 72h
- Culturing media and conditions (same as test or not): same as test
- Any deformed or abnormal cells observed: no
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h
Post exposure observation period:
-
Hardness:
24 mg/L as CaCO3 (calculated)
Test temperature:
22 - 23°C
pH:
7.88 - 8.32
Nominal and measured concentrations:
Nominal concentrations: 100.0 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks
- Type: open
- Material, size, headspace, fill volume: 300 mL Erlenmeyer glas flasks filled with 100 mL cell suspension
- Aeration: no
- Initial cells density: 10 000 cells per ml
- Control end cells density: 1 007 570 cells per 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: reconstituted water according to OECD TG 201
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Light intensity and quality: The light intensity was between 4440 and 8880 Lux


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic particle counter

TEST CONCENTRATIONS
- Test concentrations: 100.0 mg/L
Reference substance (positive control):
no
Remarks:
The accuracy and reliability of the test method is demonstrated periodically as recommended by OECD Guideline No. 201 and the Council Regulation (EC) No. 761/2009. Therefore, potassium dichromate (Art. 104864) was tested as positive control.
Key result
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
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
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Remarks on result:
other: > 0.0028 mg/L (limit of water solubility)
Details on results:
The EC values exceeded the water solubility of 0.0028 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
Results with reference substance (positive control):
The accuracy and reliability of the test method is demonstrated periodically as recommended by OECD Guideline No. 201 and the Council Regulation (EC) No. 761/2009. Therefore, potassium dichromate (Art. 104864) was tested as positive control.

The test item Art. 104864 (potassium dichromate) showed 72h EC values which are within the recommended range (Biomass integral EC50 0.20 – 0.75 mg/L and Growth rate EC50 0.60 – 1.03 mg/L).

Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of the present study, an aqueous solution of 100 mg/L of the test item in an open static system revealed no aquatic toxicity.
The EC50 (0-72h) for Growth rate and Yield were >0.0028 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.
Executive summary:

Objective

The objective of this study was to evaluate the influence of the test itemon the growth and growth rate of the green algae speciesDesmodesmus subspicatus.

Study Design

The study design included one control group and one test material group with six replicates, each containing 100 mL reconstituted water or test medium and about 10000 cells/mL at the start of the experimental phase.

The study was performed as a limit test in an open static test system. The algae were exposed to a filtrate of nominal 100 mg/L.

The growth of the algae was calculated after 24, 48, and 72 hours exposure in the test medium.

Results

The test material concentration in the reconstituted water was not quantified at the start and the end of this study due to the low water solubility (0.0028 mg/L). Because of the low water solubility, the compound cannot be detected with standard analytical methods. The development of an analytical method with a sufficiently low detection and quantification limit is complex.

Due to the absence of any adverse effects at the saturation concentration, the study was performed without analytical concentration verification.

An aqueous preparation of a nominal concentration of 100 mg/L of the test item revealed no toxic effect in Desmodesmus subspicatus.

The following EC values (Growth rate and Yield) were determined:

Parameter (0-72h)

Growth rate

Yield

EC10

95% confidence interval

>0.0028 mg/L (nominal >100 mg/L)

n.d.

>0.0028 mg/L (nominal >100 mg/L)

n.d.

EC20

95% confidence interval

>0.0028 mg/L (nominal >100 mg/L)

n.d.

>0.0028 mg/L (nominal >100 mg/L)

n.d.

EC50

95% confidence interval

>0.0028 mg/L (nominal >100 mg/L)

n.d.

>0.0028 mg/L (nominal >100 mg/L)

n.d.

n.d. = could not be determined

Conclusion

Under the conditions of the present study, an aqueous solution of 100 mg/L of the test item in an open static system revealed no aquatic toxicity.The EC50 (0-72h) for Growth rate and Yield were >0.0028 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
For this endpoint a one-to-one read across was performed to a chemical similar compound of the same chemical class with a comparable phys. chem. profile and similar response in biological assays. The relevant study was performed according to GLP and the methods applied are fully compliant with OECD TG 201. A detailed read across justification is provided in chapter 13 of this dossier.
Reason / purpose:
read-across source
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
Remarks on result:
other: EC50 > water solubility limit
Key result
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: EC50 > water solubility limit

Description of key information

For this endpoint a one-to-one read across was performed to a chemical similar compound of the same chemical class with a comparable phys. chem. profile and similar response in biological assays. The relevant study was performed according to GLP and the methods applied are fully compliant with OECD TG 201. The EC50 and EC10 (growth rate,72 h) exceed the limit concentration of 100 mg/L (nominal) and are thus above the water solubility of the test material.

Key value for chemical safety assessment

Additional information