Dilithium peroxide

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 SEP 2021 - 08 OCT 2021

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 23rd March, 2006, Annex 5 corrected: 28th July 2011.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

(published in the Official Journal of the European Union L 54 of 01 March 2016)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Vehicle:

yes

Remarks:

ISO Medium

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 0.01171 g of test item was suspended in 1000 mL of the dilution water (OECD Medium) in order to give the nominal loading rate of 11.71 mg/L of the test item. The further test item concentrations were prepared from this test item stock solution (TISS)
- Controls: untreated control, reference control
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): Reconstituted algal growth medium (OECD medium, according to OECD 201) was used as dilution water in the experiment. Separate stock solutions were first prepared in deionised water (prepared in TOXI-COOP ZRT. by ELIX 3 Int water purification system, Serial No.: F0MA71964D). The growth medium was prepared by adding an appropriate volume of these different stock solutions to deionised water in order to achieve the final concentrations in the prepared algal growth medium.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Common name: Green algae
- Strain: 61.81 SAG (identical strains: CCAP 278/4; UTEX 1648; ATCC 22662)
- Source (laboratory, culture collection): The algae were supplied by the SAG: Collection of Algal Cultures, Inst. Plant Physiology, University of Göttingen, Untere Karspüle 2, D-37073 Göttingen, Germany
- Breeding Conditions: The stock cultures are small algal cultures that are planted on agar regularly. These are transferred to fresh medium at least once every two months under standardized conditions according to the test guidelines.

ACCLIMATION
- Culturing media and conditions (same as test or not): The pre-culture is intended to give a number of algae suitable for the inoculation of test cultures. The pre-culture was prepared with OECD Medium, incubated under the conditions of the test and used.
when still exponentially growing, normally after an incubation period of 2-4 days. (The pre-culture was incubated for four days before this test.) The algal cultures used in this study did not contain deformed or abnormal cells.

Test type:

static

Water media type:

other: OECD medium, according to OECD 201

Limit test:

no

Total exposure duration:

72 h

Test temperature:

22.9 - 23.5°C

pH:

7.88 – 9.78

Nominal and measured concentrations:

0.5, 1.1, 2.42, 5.32 and 11.71 mg/L nominal

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: Volumes of 100 mL algal suspension per replicate were continuously shaken by a laboratory orbital shaker in 250 mL Erlenmeyer flasks
- Initial cells density: 10^4 cells/mL
The test was performed with three replicates per test concentration and six replicates in the control group. Each test unit was uniquely identified with study number, test group and replicate.

GROWTH MEDIUM / TEST MEDIUM
Reconstituted algal growth medium (OECD medium, according to OECD 201) was used as dilution water in the experiment.
Separate stock solutions were first prepared in deionised water (prepared in TOXI-COOP ZRT. by ELIX 3 Int water purification system, Serial No.: F0MA71964D). The growth medium was prepared by adding an appropriate volume of these different stock solutions to deionised water in order to achieve the final concentrations in the prepared algal growth medium.

OTHER TEST CONDITIONS
- Adjustment of pH: The pH of the test solutions was not adjusted during the experiment.
- Photoperiod: The algal culture flasks were continuously illuminated.
- Light intensity and quality: 6998 lux (SD: 46 lux), which was ensured with fluorescent lamps (with a spectral range of 400-700 nm).

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
Based on the results of the preliminary range-finding test five test concentrations in a geometric series with a spacing factor of 2.2 and one control were used in the main test. The following nominal concentrations were tested: 0.5, 1.1, 2.42, 5.32 and 11.71 mg product / L.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7)

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

1.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

2.42 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Key result

Duration:

72 h

Dose descriptor:

EC10

Remarks:

95 % conf. limits

Effect conc.:

1.77 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 1.47 - 2.00

Duration:

72 h

Dose descriptor:

EC10

Remarks:

95 % conf. limits

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Remarks on result:

other: 0.76 - 1.18

Duration:

72 h

Dose descriptor:

EC20

Remarks:

95 % conf. limits

Effect conc.:

2.17 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 1.90 - 2.40

Duration:

72 h

Dose descriptor:

EC20

Remarks:

95 % conf. limits

Effect conc.:

1.23 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Remarks on result:

other: 1.01 - 1.40

Key result

Duration:

72 h

Dose descriptor:

EC50

Remarks:

95 % conf. limits

Effect conc.:

3.24 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 2.98 - 3.52

Duration:

72 h

Dose descriptor:

EC50

Remarks:

95 % conf. limits

Effect conc.:

1.84 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Remarks on result:

other: 1.66 - 2.04

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): No abnormal shape of algal cells was noticed in the control and at the test item concentration of 0.5 mg/L during the experiment. As sign of toxicity swollen cells were found between normal cells in the test group of 1.1mg test item/L. Swollen and thin cells were observed between normal cells at the concentration of 2.42 mg/L during the test, while swollen and thin cells were found in the test groups of 5.32 and 11.71 mg/L.

Analytical results
- Five different test concentrations arranged in a geometric series with a spacing factor of 2.2 and a concurrent control was tested in the main experiment. The concentrations of the test item were analysed at the start and at the end of the test.
In the untreated control group test item concentration was below the quantification limit. The measured test item concentrations were in the range of 94 - 110 % of the nominal concentrations during the experiment, therefore the biological results are based on the nominal test item concentrations.

Results with reference substance (positive control):

For the evaluation of the reliability of the applied test system and the experimental conditions Potassium dichromate is tested at least twice a year. The date of the latest study performed in the laboratory of TOXI-COOP ZRT. (Study number: 392-201-5923) with the reference item Potassium dichromate (K2Cr2O7) was: 06 – 09 April 2021, Final Report issued: 03 May 2021.

The 72h ErC50: 0.62 mg/L, (95 % confidence limits: 0.59 – 0.65 mg/L)
The 72h EyC50: 0.33 mg/L, (95 % confidence limits: 0.32 – 0.34 mg/L)

Validity criteria fulfilled:

yes

Conclusions:

All validity criteria were met and therefore the study is considered as fully valid. Biological results and endpoints are based on nominal test item concentrations.
72h NOEC: 1.1 mg/L
72h LOEC: 2.42 mg/L
72h EC10: growth rate 1.77 mg/L yield 1.00 mg/L
72h EC20: growth rate 2.17 mg/L yield 1.23 mg/L
72h EC50: growth rate 3.24 mg/L yield 1.84 mg/L

Executive summary:

The test was performed according to the OECD Guideline 201 and EU Method C.3 under GLP compliance. Exponentially growing cultures of Raphidocelis subcapitata (also known as Pseudokirchneriella subcapitata) were exposed to nominal concentrations of 0.5, 1.1, 2.42, 5.32 and 11.71 mg/L of the test item for 72hrs under static conditions. No abnormal shape of algal cells was noticed in the control and at the test item concentration of 0.5 mg/L during the experiment. As sign of toxicity swollen cells were found between normal cells in the test group of 1.1 mg test item/L. Swollen and thin cells were observed between normal cells at the concentration of 2.42 mg/L during the test, while swollen and thin cells were found in the test groups of 5.32 and 11.71 mg/L.

All validity criteria were met and therefore the study is considered as fully valid. Biological results and endpoints are based on nominal test item concentrations.

72h NOEC: 1.1 mg/L

72h LOEC: 2.42 mg/L

72h EC10: growth rate 1.77 mg/L; yield 1.00 mg/L

72h EC20: growth rate 2.17 mg/L; yield 1.23 mg/L

72h EC50: growth rate 3.24 mg/L; yield 1.84 mg/L was determined.

Description of key information

The effect of the test item to the unicellular freshwater green algae Pseudokirchneriella subcapitata was determined according to the OECD 201 and EU Method C.3. and in accordance with GLP.

The EC50-values after 72 hours for inhibition of growth rate (ErC50) is 3.24 mg/L and yield (EyC50) is 1.84 mg/L. The 72hours EC-10-values for inhibition of growth rate (ErC10) is 1.77 mg/L and yield (EyC10) is 1.00 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:

3.24 mg/L

EC10 or NOEC for freshwater algae:

1.77 mg/L

Additional information

The test was performed according to the OECD Guideline 201 and EU Method C.3 under GLP compliance. Exponentially growing cultures of Raphidocelis subcapitata (also known as Pseudokirchneriella subcapitata) were exposed to nominal concentrations of 0.5, 1.1, 2.42, 5.32 and 11.71 mg/L of the test item for 72hrs under static conditions. No abnormal shape of algal cells was noticed in the control and at the test item concentration of 0.5 mg/L during the experiment. As sign of toxicity swollen cells were found between normal cells in the test group of 1.1 mg test item/L. Swollen and thin cells were observed between normal cells at the concentration of 2.42 mg/L during the test, while swollen and thin cells were found in the test groups of 5.32 and 11.71 mg/L.

All validity criteria were met and therefore the study is considered as fully valid. Biological results and endpoints are based on nominal test item concentrations.

72h NOEC: 1.1 mg/L

72h LOEC: 2.42 mg/L

72h EC10: growth rate 1.77 mg/L; yield 1.00 mg/L

72h EC20: growth rate 2.17 mg/L; yield 1.23 mg/L

72h EC50: growth rate 3.24 mg/L; yield 1.84 mg/L was determined.