Ferrocholinate

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:

22. Apr. 2020 (Study Plan dated); 12. May 2020 (Experimental Starting Date); 15. May 2020 (Experimental Completion Date)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

Adopted 23. Mar. 2006, Annex 5 corrected: 28 July 2011.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

Adopted 07 December 2015.

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

PROPERTIES OF TEST MATERIAL
LogPow: < 0.3 (GLP study 19100101G930 performed at LAUS GmbH).

Analytical monitoring:

yes

Details on sampling:

The cell concentration of each replicate was determined by measuring the cell numbers every 24 hours with an electronic particle counter. Growth rate µ and the yield were determined from the cell number at the respective observation times.

Vehicle:

yes

Details on test solutions:

A stock solution containing 100.1 mg/L test item in algal medium (demineralised water enriched with minerals but without algae) was prepared. The lower treatments (4.6 / 10 / 22 / 46 mg/L) were prepared by dilution of this stock solution with algal medium.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Unicellular freshwater green alga.
Genus: Desmodesmus
Species: subspicatus
SAG Strain Number: 86.81
Taxonomic position: Chlorophyta - Chlorophyceae

The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen). The algae are kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture was prepared. From an aliquot of the perma-nent culture, the pre-culture was prepared.

Water media type:

freshwater

Total exposure duration:

72 h

Test temperature:

20.5 – 22.7 °C

pH:

Blank control: 7.5 (0h) ; 7.5 (72h)
4.6 mg/L nominal concentration: 7.5 (0h) ; 7.5 (72h)
10 mg/L nominal concentration: 7.5 (0h) ; 7.7 (72h)
22 mg/L nominal concentration: 7.5 (0h) ; 7.5 (72h)
46 mg/L nominal concentration: 7.4 (0h) ; 7.4 (72h)
100 mg/L nominal concentration: 7.1 (0h) ; 7.3 (72h)

Nominal and measured concentrations:

4.6 / 10 / 22 / 46 / 100 mg/L nominal concentration
The concentrations tested were based on non GLP pre-tests.

Details on test conditions:

TEST SYSTEM
- Specification
Unicellular freshwater green alga.
Genus: Desmodesmus
Species: subspicatus
SAG Strain Number: 86.81
Taxonomic position: Chlorophyta - Chlorophyceae
- Origin and Culture
The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen). The algae are kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture was prepared. From an aliquot of the perma-nent culture, the pre-culture was prepared.

ANALYTICAL METHOD:
- Sample preparation
At the start of the test, all concentration were measured undiluted. 5 mL of the test item solution was acidified with 50 µL HNO3 (65 % water, Roth) and measured via ICP-OES.
Because the measured concentration lay over the highest calibration level, the concentrations 10 / 22 / 46 and 100 mg/L were diluted as followed to get the values in the calibrated range: 5 mL of the test solution with the concentrations 10 mg/L was diluted with 4.9 mL algal medium (2x); 2 mL of the test solution with the concentration of 22 mg/L was diluted with 7.9 mL algal medium (5x); 1 mL of the test solution with the concentration of 46 mg/L was diluted with 8.9 mL algal medium (10x); 0.5 mL of the test solution with the concentration of 100 mg/L was diluted with 9.4 mL algal medium (20x). Afterwards, 100 µL HNO3 (65 % in water) was added to the dilutions.
For the specificity determination, 1 % HNO3 (65 % water) was added to each sample.

CONDUCT OF THE STUDY:
- Pre-Culture
For the pre-culture an aliquot of the permanent culture was brought into nutrient medium and incubated under continuous lighting for 4 days under test conditions (Lighting: within the specified range of 4440 – 8880 lux; 21.3 – 23.5 °C). The resulting culture grew exponentially.
Before usage, the pre-culture was checked for the absence of cell aggregates and the cell number of the culture was determined.
- Performance of the Study
For each treatment, 200 mL of the respective test item solution was mixed with the necessary amount of algal pre-culture (0.397 mL) to achieve a cell concentration of 1.9 *103 cells/mL (see chapter 12 Devia-tions). For the blank control, 350 mL nutrient medium was used instead of test item solution and mixed with the necessary amount of algal pre-culture (0.695 mL). In this mixture, the pH-value was measured.
The real cell concentration at the beginning of the test was measured with an electronic particle counter in the blank control solution. This measured value was used as start cell concentration for all replicates.
The test vessels were filled with 45 ± 1 mL of the respective test solution and incubated open (covered with perforated plastic foil acting as a stopper) for 72 hours, shaken on an orbital shaker to keep the algae in suspension. Before the start of incubation and every 24 hours, the cell number was determined with an electronic particle counter. After the test the pH value in treatments and blank control was measured again.
At the end of the test, the treatments were examined microscopically in order to assess the appearance of the algae and detect abnormalities (e.g. caused by the exposure to the test item).
Additionally, an extra replicate for each treatment and the control without algae was prepared. To avoid adsorption onto the algae cells, the content of Fe as component of the test item in the test solutions was measured in extra replicates without algae cells at the beginning and at the end of the experiment.
- Experimental conditions
Treatments tested: 4.6 / 10 / 22 / 46 / 100 mg/L nominal concentration
The concentrations to be tested are based on non GLP pre-tests.
Vessels: glass flasks total volume 65 mL
Duration: 72 hours
Lighting: within the specified range (4440 – 8880 Lux)
Temperature: 20.5 – 22.7 °C
Replicates:
Treatments: 3 vessels, filled with 45 ±1 mL test solution and algae
1 vessels, filled with 45 ±1 mL test solution without algae for analytical determination
Blank Control: 6 vessels, each filled with 45 ±1 mL algal medium and algae
1 vessel filled with 45 ±1 mL algal medium without algae for analytical determination

Reference substance (positive control):

yes

Remarks:

Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9)

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

42.1 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

94.8 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

126.94 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

growth rate

Remarks on result:

other:

Remarks:

Extrapolated by the program.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

21 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

42.1 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

30.11 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

138.58 mg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

biomass

Results with reference substance (positive control):

Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9) was used as positive control in a separate reference test.
The 72h-EC50 values of potassium dichromate were determined in a separate reference test. For the estimation of the 72h-EC50 values of the positive control, the fits showed sufficient statistical correspondence of the data with the dose-response-equation. The values were within the range of the laboratory(growth rate 0.65 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

Validity criteria fulfilled:

yes

Remarks:

No observations were made which might cause doubts concerning the validity of the study outcome. The result of the test can be considered valid.

Conclusions:

Based on the values obtained for Acuatic Toxicity (growth rate and yield) of Ferric Choline Citrate in Desmodesmus subspicatus, the classification criteria are not met.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

(Q)SAR

Adequacy of study:

key study

Study period:

05.03.2020

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

1. Software: QSAR Toolbox 3.4.0.17
2. Database version: 3.8.8/3.1.2
3. TPRF v.3.4.1.34101
4. SMILES used as input for the model: CN{+}(C)(C)(CCO).O{-}C(=O)CC1(O)CC(O)O{-}.[Fe]{3+}(.O{-}C(=O)CC(O)(CC(=O)O{-}.[Fe]{3+}2.O{-}C(=O)CC(O)(C(=O)O{-}.N{+}(C)(C)(C)CCO)C(=O)O{-}.2)C(=O)O{-}.N{+}(C)(C)(C)CCO).O{-}C1=O
5. The QSAR Toolbox prediction is attached in the "Attached justification".

Qualifier:

according to guideline

Guideline:

other: "GUIDANCE ON INFORMATION REQUIREMENTS AND CHEMICAL SAFETY ASSESSMENT / CHAPTER R.6: QSARS AND GROUPING OF CHEMICALS"

Version / remarks:

Published by ECHA (May, 2008).

Qualifier:

according to guideline

Guideline:

other: "GUIDANCE DOCUMENT ON THE VALIDATION OF (QUANTITATIVE) STRUCTURE-ACTIVITY RELATIONSHIPS MODELS"

Version / remarks:

Published by OECD (September, 2007).

Specific details on test material used for the study:

- CAS number: 1336-80-7
- Chemical name: 2-HYDROXYETHYL-TRIMETHYL-AMMONIUM,2-HYDROXYPROPANE-1,2,3-TRICARBOXYLATE
- SMILES: CN{+}(C)(C)(CCO).O{-}C(=O)CC1(O)CC(O)O{-}.[Fe]{3+}(.O{-}C(=O)CC(O)(CC(=O)O{-}.[Fe]{3+}2.O{-}C(=O)CC(O)(C(=O)O{-}.N{+}(C)(C)(C)CCO)C(=O)O{-}.2)C(=O)O{-}.N{+}(C)(C)(C)CCO).O{-}C1=O

Key result

Duration:

72 h

Dose descriptor:

EC50

Remarks:

Reproduction (Growth rate) in Pseudokirchneriella subcapitata

Effect conc.:

104 mg/L

Nominal / measured:

not specified

Conc. based on:

not specified

Basis for effect:

growth rate

Remarks on result:

other: QSAR predicted value

Validity criteria fulfilled:

yes

Conclusions:

The value obtained for Acuatic Toxicity EC50 Reproduction (Growth rate) in Pseudokirchneriella subcapitata from the prediction was 104 mg/L (72h).

Description of key information

The following experimental results for Acuatic Toxicity of Ferric Choline Citrate in Desmodesmus subspicatus (OECD 201) were determined:

NOEC (growth rate): 42.10 mg/L

LOEC (growth rate): 94.80 mg/L

EC10 (growth rate): 126.94 mg/L

NOEC (yield): 21.00 mg/L

LOEC (yield): 42.10 mg/L

EC10 (yield): 30.11 mg/L

EC50 (yield): 138.58 mg/L

The value obtained for Acuatic Toxicity EC50 Reproduction (Growth rate) in Pseudokirchneriella subcapitata from the prediction was 104 mg/L (72h).

Key value for chemical safety assessment

EC50 for freshwater algae:

138.58 mg/L

EC10 or NOEC for freshwater algae:

30.11 mg/L