SUMILIZER GP

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Please refer to Test guideline in 'Materials and methods'

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

yes

Remarks:

Measured concentrations were below loading rate, effect concentrations refer to loading rate. Test item dispersion was filtered before testing

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

1992

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss Federal Office of Public Health, Switzerland

Analytical monitoring:

yes

Details on sampling:

- Concentrations: stock solutions, control, solvent control and 1 mg/L
- Sampling method: Duplicate samples were taken from the approximate center of the aquaria without mixing of the test medium and from freshly prepared test soutions, control and solvent control at the start of the test (Day 0) and at the last test medium preparation on Day 3.
- Sample storage conditions before analysis: deep-frozen (at about -20 °C) immediately after sampling

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solutions were prepared at the start of the test and at each test medium renewal by diluting 300 mg of the test item in 3 mL N,N-dimethylformamide (DMF) and ultrasonicating for 15 minutes. 250 mL were dosed into 2500 mL test water to obtain a loading rate of 10 mg/L. The dispersion was then stirred at room temperature over 96 hours. After stirring, the dispersion was filtered through a membrane filter (0.45 µm) just before test start and medium renewal.
- Controls: control: dilution water only, solvent control: dilution water and DMF
- Chemical name of vehicle: DMF
- Concentration of vehicle in test medium: stock solution: 100%, final test solutions: 100 µL/L, control: 0%, solvent control: 100 µL/L

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebra fish
- Source: obtained from Zoohaus Schaub, Liestal, Switzerland
- Length at study initiation: mean: 3.3 cm (SD: 0.2 cm)
- Weight at study initiation: mean: 0.37 g (SD: 0.11 g)

ACCLIMATION
- Acclimation period: 1 week
- Acclimation conditions: same as test
- Type and amount of food during acclimation: Tetra Min® Hauptfutter (TETRA-Werke, Germany)
- Feeding frequency during acclimation: not reported, fish were fed until one day before the start of the test
- Health during acclimation: no mortality observed

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

250 mg/L as CaCO3

Test temperature:

21 - 22 °C

pH:

7.8 - 7.9

Dissolved oxygen:

8.3 - 9.5 mg/L

Nominal and measured concentrations:

Nominal loading rate: 0 (control), 0 (solvent control), 10 mg/L
Measured concentrations: please refer to 'Any other information on results incl. tables'

Details on test conditions:

TEST SYSTEM
- Test vessel: aquarium
- Material, fill volume: glass, 5 L
- Renewal rate of test solution: daily
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- No. of vessels per vehicle control (replicates): 1
- Biomass loading rate: 0.51 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water, composition: CaCI2 x 2H2O (294 mg/L), MgSO4 x 7H2O (123 mg/L), NaHCO3 (65 mg/L), KCl (65 mg/L)
- Ca/Mg ratio: 4:1
- Intervals of water quality measurement: appearance of medium, pH, dissolved oxygen and temperature were measured daily in freshly prepared and old medium before water renewal

OTHER TEST CONDITIONS
- Photoperiod: 16/8 h light/dark with a 30-minute transition period
- Light intensity: 280 - 500 lux

EFFECT PARAMETERS MEASURED: mortality and visible abnormalities after 3, 24, 48, 72 and 96 hours

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

loading rate

Basis for effect:

mortality (fish)

Remarks on result:

other: No toxic effect up to its solubility limit in the test water

Details on results:

- Mortality of control: 0%
- Effect concentrations exceeding solubility of substance in test medium: clear test medium during the whole test

Sublethal observations / clinical signs:

Table 1: Number of affected fish during the test

Concentration (mg/L)	Number of affected fish* / number of dead fish
	3 h	24 h	48 h	72 h	96 h
Control	0/0	0/0	0/0	0/0	0/0
Solvent control	0/0	0/0	0/0	0/0	0/0
10	0/0	0/0	0/0	0/0	0/0

Table 2: Measured test item concentrations

Nominal concentration of the test item [µg/L]	Sampling day / Age of sample (h)	Measured concentration   [µg/L]	% of nominal concentration
Stock solution
10000	0 / 0	91660	92
	3 / 0	83835	84
Treatment samples
Undiluted filtrate 10 mg/L loading rate	0 / 0	1.67	n.a.
	0 / 0	2.26	n.a.
	1 / 24	0.785	n.a.
	1 / 24	n.d.	n.a.
	3 / 0	n.d.	n.a.
	3 / 0	n.d.	n.a.
	4 / 24	n.d.	n.a.
	4 / 24	n.d.	n.a.
Biological control samples
	0 / 0	n.d.	n.a.
	1 / 24	n.d.	n.a.
	3 / 0	n.d.	n.a.
	4 / 24	n.d.	n.a.
Solvent control
	0 / 0	n.d.	n.a.
	1 / 24	n.d.	n.a.
	3 / 0	n.d.	n.a.
	4 / 24	n.d.	n.a.
Spiked water samples
50.3	0 / 0	41.0	82
	0 / 0	48.4	96

n.a. = not applicable

n.d. = no test item detected, respectively < LOD of 0.2 µg/L

LOQ = 0.6 µg/L

Table 3: Validity criteria of OECD 203

Criterion from the guideline	Outcome	Criterion fulfilled
The mortality in the control(s) should not exceed 10% (or one fish if less than ten are used) at the end of the test	0	yes
The dissolved oxygen concentration must have been at least 60 per cent of the air saturation value throughout the test	8.3 - 9.5 mg/L	yes
Analytical measurement of test concentrations is compulsory	Test item concentrations were measured	yes

Validity criteria fulfilled:

yes

Description of key information

No adverse effect up to its solubility limit in the test water

LC50 (96 h): >10 mg/L (Danio rerio, OECD 203, loading rate)

Additional information

A GLP study according to OECD guideline 203 on the acute toxicity to the zebrafish Danio rerio was performed in 96-hour semi-static test with a daily test medium renewal.

Due to the very low water solubility of the test item, an undiluted filtrate (0.45 µm filter pore size) of a supersaturated dispersion with the maximum concentration of dissolved test item was used as the only test concentration. The limit test was performed with the test substance loading rate of 10 mg/L. Additionally, a control and a solvent control (100 µL/L N,N-dimethylformamide) were tested in parallel.

The analytically determined test item concentrations in the freshly prepared test medium were in the range of 2.3 µg/L to < LOD (limit of detection: 0.2 µg/L). At the end of the test medium renewal periods of one day, the test item concentrations were in the range of 0.79 µg/L to < LOD. In the control, the solvent control and at the loading rate of 10 mg/L no mortality or other visible abnormalities were determined up to the solubility limits of the test item.

The LC50 (96 h) was therefore determined to be at a loading rate of > 10 mg/L.