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Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2020-02-04 to 2020-02-14

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

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.0 (NC) and 200 mg/L (A)
- Sampling method: After 0, 24, 48 and 72 h exposure samples were taken from the test solutions: 2 samples of 2 mL per treatment group. The samples were filled from the test vessels directly into 8 mL glass vials without any additional treatment.
- Sample storage conditions before analysis: To each vial, 2 mL acetonitrile was added and samples were stored in the freezer (≤ -18 °C).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 222.4 mg test item (including a factor of 1.11 to take into account the dilution caused by addition of the algal inoculum) to 1000 mL test medium, sonicating (until no larger particles are visible) and shaking for 48 h using an overhead shaker at 22.7–24.1 °C. The pH of the stock solution was adjusted to 9.1 after 24.5 h shaking and adjusted to 8.0 before filtration. The resulting solution was filtered through a fibre glass filter with a retaining range till 0.6 µm. The filter was prepared by rinsing with purified water and preconditioning with ca. 100 mL stock solution (which was discarded) to reduce adsorption of the test item.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): The filtered stock solution was visually clear, without particles.

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
- Source (laboratory, culture collection): Culture Collection of Algae, University of Goettingen
- Age of inoculum (at test initiation):
- Method of cultivation: The strain used for this study has been cultured at Hydrotox GmbH since March 2019. Twice a week, the stock suspension is diluted into fresh Holm-Hansen medium under axenic conditions to keep it in exponential growth.

ACCLIMATION
- Acclimation period: No

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test temperature:

22.0–23.4 °C

pH:

7.5–7.8

Nominal and measured concentrations:

0 and 200 mg/L (nominal)
measured: 0.31–0.58 % of the nominal loading rate

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer glass flasks 100 mL, Schott, Mainz
- Type: open
- Material, size, headspace, fill volume: 50 mL
- Aeration: No
- Initial cells density: 5.0E+04 cells/mL
- Control end cells density: 111.7E+05 cells/mL
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes (OECD TG 201 medium according to OECD 201 (2006))

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuously illuminated
- Light intensity and quality: LEDs (warm white) with a light intensity of 81.3 µE/m²/s ± 7.7 %

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The algal biomass was monitored by measuring the chlorophyll fluorescence at 465 nm after 0, 24, 48, and 72 h.

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 8, 40 and 200 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

potassium dichromate (Honeywell GmbH, Seelze, Germany, Lot: H1030)

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 0.809 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 0.809 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape: no
- Colour differences: no
- Flocculation: No data
- Adherence to test vessels: No data
- Aggregation of algal cells: No data
- Other: No data
- Any stimulation of growth found in any treatment: Yes, approximately 9.5 % growth rate stimulation after 72 h
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: Not applicable, no adverse effects observed.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- ErC50 (72 h): 0.85 mg/L (95 % CI: 0.81–0.89 mg/L)

Reported statistics and error estimates:

As no effect on the test organisms was observed, no statistical data evaluation was performed.

Table 1. Analytically measured test item concentrations after 0 h, 24 h, 48 h and 72 h exposure

Nominal test item loading rate [mg/L]	Sample	Sampling time [h]	Calculated test item concentration [mg/L]	Recovery [%] nominal	Geometric mean test item  concentration [mg/L]
NC	1501-NC1-0h	0	No Peak	--	--
	1501-NC1-24h	24	No Peak
	1501-NC1-48h	48	No Peak	--
	1501-NC1-72h	72	No Peak	--
200	1501-A1-0h	0	1.15	0.58	0.809
	1501-A1-24h	24	0.811	0.41
	1501-A1-48h	48	0.738	0.37
	1501-A1-72h	72	0.621	0.31

 

Criteria of validity

- The biomass (cell concentration) in the control has increased by a factor of ≥ 16 during the test period. 

- The mean coefficient of variation for section-by-section growth rate (day 0–1, 1–2 and 2–) in the control was ≤ 35 %.

- The coefficient of variation of average specific growth rate during the test period in the control replicates was ≤ 7 %.

Validity criteria fulfilled:

yes

Conclusions:

The test item showed no inhibition of the algal growth in comparison to the negative control, no adverse effect on the algal growth was observable. Accordingly, a statistical data evaluation was not necessary for this limit test. As the measured test item concentrations are not within ± 20 % of the nominal concentrations, all results are given in relation to the analytically measured test item concentrations, in line with OECD test guideline 201 (2004) and OECD series on testing and assessment no. 23 (2000),

Executive summary:

In a 72-hour algal growth inhibition test, cultures of Pseudokirchneriella subcapitata (strain no. 61.81 SAG) were exposed to N-(4-((4-(3-phenylureido)phenyl)sulfonyl)phenyl)benzenesulfonamide at nominal concentrations of 0 (control), and 200 mg/L (limit test) under static conditions in accordance with OECD Guideline 201 (23 March 2006), and under GLP. Growth inhibition in the treatment (200 mg/L nominal) was -9.5 % relative to the control, i.e, a stimulating effect. There were no compound related phytotoxic effects.

This toxicity study is classified as acceptable and satisfies the guideline requirements for an algal growth inhibition toxicity study.

As the measured test item concentrations were not within ± 20 % of the nominal concentrations, all results are given as analytically measured test item concentrations.

Results synopsis

Test organism: Pseudokirchneriella subcapitata (strain no. 61.81 SAG)

Test type: Static

72-h LOE_rC: > 0.809 mg/L

72-h NOE_rC: => 0.809 mg/L

Endpoint(s) effected: Growth rate

Description of key information

N-(4-((4-(3-phenylureido)phenyl)sulfonyl)phenyl)benzenesulfonamide is not toxic to algae at maximum water solubility.

LOEC > 0.806 mg/L

NOEC ≥ 0.806 mg/L

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

0.809 mg/L

Additional information

In a 72-hour algal growth inhibition test, cultures of Pseudokirchneriella subcapitata (strain no. 61.81 SAG) were exposed to N-(4-((4-(3-phenylureido)phenyl)sulfonyl)phenyl)benzenesulfonamide at nominal concentrations of 0 (control), and 200 mg/L (limit test) under static conditions in accordance with OECD guideline 201 (23 March 2006), and under GLP. An accompanying substance-specific chemical analysis to determine the test item concentrations in the test vessels was conducted. As the measured test item concentrations are not within ± 20 % of the nominal concentrations, all results are given in as analytically measured test item concentrations, in line with OECD test guideline 201 (2004) and OECD series on testing and assesment no. 23 (2000).