Bis-biphenyl-4-yl-(9,9‘-spirobifluoren-4-yl)-amine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

November 19, 2014 - March 25, 2015

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)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Batch: OS11008922
Appearance: Pale yellow powder without visible impurities
Water solubility: 1.7e-12 mg/L
Released until: August 15, 2016
Storage: Tightly closed, dark at room temperature (15 to 25°C)

Analytical monitoring:

no

Remarks:

The test item concentration in the reconstituted water was not quantified at the start and the end of this study. Because of the low water solubility (<1.7e-12 mg/L) the compound cannot be detected with standard analytical methods.

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: 7.86
- Total hardness: 24 mg/L as CaCO3 (calculated)

Preparation of the Test Item:
A stock preparation with a test item concentration of 100 mg/L was freshly prepared. For that purpose, the test item was weighed in a calibrated flask and vehicle was added. The preparation was treated in an ultrasonic device for 30 minutes. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Then the formulation was passed through a filter membrane (pore size: 0.2 µm). 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 61.81 SAG (Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universität Göttingen, Germany). This strain was further cultivated in the laboratories of the test facility.
- 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 (Lumilux T5 nws FLH1 HO 80W/840, Osram GmbH, München, Germany) 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:

22.4-23.1°C

pH:

7.68 - 8.62

The pH in the control group did increase more than 1.5 units during the test. This did not influence the outcome of the study.

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
- 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 measured immediately before the start of the study was 6995 Lux ± 5.6% and 7119 Lux ± 8.3% at the end of the study. Thus, the light intensity was in the anticipated range of 4440 – 8880 Lux within ± 15% from the average.


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

TEST CONCENTRATIONS
- Nominal test concentration: 100 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 (Gado, 2014).

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: EC10 >1.7e-12 mg/L

Key result

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: EC20 >1.7e-12 mg/L

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 >1.7e-12 mg/L

Key result

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: EC10 >1.7e-12 mg/L

Key result

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: EC20 >1.7e-12 mg/L

Key result

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: EC50 >1.7e-12 mg/L

Details on results:

The EC values exceeded the water solubility of 1.7e-12 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 was tested as positive control (Gado, 2014).

Under the conditions of the present study, the test item Potassium dichromate showed 72h EC50 of 1.03 mg/L for growth rate and 0.53 mg/L for yield. The results obtained in this study are similar to results of a ring test mentioned in DIN EN ISO 8692.

Objective

The objective of this study was to evaluate the influence of the test item on the growth and growth rate of the green algae species Desmodesmus subspicatus.

Study Design

The study design included one control group and one test item 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 item group and control group.

Results

The test item concentration in the reconstituted water was not quantified at the start and the end of this study. Because of the low water solubility (1.7e-12 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.

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 [mg/L] 95% confidence interval	>1.7e-12 mg/L (nominal >100 mg/L) n.d.	>1.7e-12 mg/L (nominal >100 mg/L) n.d.
EC20 [mg/L] 95% confidence interval	>1.7e-12 mg/L (nominal >100 mg/L) n.d.	>1.7e-12 mg/L (nominal >100 mg/L) n.d.
EC50 [mg/L] 95% confidence interval	>1.7e-12 mg/L (nominal >100 mg/L) n.d.	>1.7e-12 mg/L (nominal >100 mg/L) n.d.

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 >1.7e-12 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.

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 and EC10 value for growth rate was greater than the water solubility of 1.7e-12 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.

Executive summary:

The objective of this study was to evaluate the influence of the test item on the growth and growth rate of the green algae species Desmodesmus subspicatus in a limit test according to OECD Guideline 201. Because of the low water solubility (1.7e-12 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.

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 72 -h EC10 and EC50 value were determined to be greater than the water solubility of 1.7e-12 mg/L (nominal > 100 mg/L).

Reference 2

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:

comparable to 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 for cross-reference:

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

Validity criteria fulfilled:

yes

Conclusions:

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 source compound did not show any evidence for toxicity towards aquatic algae at concentrations above the water solubility.

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 source compound did not show any evidence for toxicity towards aquatic algae at concentrations above the water solubility.

Key value for chemical safety assessment

EC50 for freshwater algae:

100 mg/L

EC10 or NOEC for freshwater algae:

100 mg/L