4,5-dihydro-5-oxo-4-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]-1-(4-sulphophenyl)-1H-pyrazole-3-carboxylic acid, sodium salt

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

Study period:

2017-04-08 to 2017-05-02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: control and 100 mg/L
- Sampling method: taken in 24 h intervals from fresh and aged test solutions. For each sampling also a retain sample was taken.
- Sample storage conditions before analysis: stored deep frozen (10 mL with 0.05 mL formic acid)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: stock solution S1: weighed into a volumetric flask and added test medium up to the bench mark then homogenised by shaking.
- Evidence of undissolved material: the solution was clear and yellow.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Source: own stock breeding of the lab, originally purchased from the Karlsruhe Institute of Technology
- Method of breeding: fed with commercial fish flake food (TetraMin) and protein-enriched pellet food (Caviar) at least once per day
- Maintenance of the brood fish: continuously reared in the lab

FEEDING DURING TEST
- Frequency: none

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

232 mg CaCO3/L

Test temperature:

21.7 – 22.6 °C

pH:

7.13 – 7.88

Dissolved oxygen:

98 - 104 % of air saturation

Conductivity:

698 µS/cm

Nominal and measured concentrations:

Nominal: 100 mg/L
Measured (geometric mean): 79.4 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 2 L glass beaker
- Fill volume: filled with 2 L of test solution
- Aeration: continuous aeration of the test tanks with a membrane pump using a Pasteur pipette
- Renewal of test solution: every 24 hours
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: max. loading of 0.19 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted test water consisting of analytical grade salts dissolved in purified water with a ratio of Ca:Mg and Na:K of 4:1 and 10:1, respectively

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Biological Assessment: effects and mortality assessed at 0, 4, 24, 48, 72 and 96 hours after test start; all fish were euthanized, weighed and measured at the end of test.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0.1, 1.0, 10.0, 100 mg/L
- Results used to determine the conditions for the definitive study: No mortality occurred up to 100 mg/L.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 79.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

79.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Mortality: No mortality was observed
- Clinical signs: No sublethal effects were observed
- Length and weight: The average weight of the test organisms was 0.0546 ± 0.0191 g; the average length was 18.2 ± 2.3 mm.
- Analytical Results: The measured content of test in fresh test solutions was between 94 % and 103 % of nominal and 52 -77 % of nominal in aged test solutions

Reported statistics and error estimates:

All toxicological endpoints were evaluated using nominal and measured concentrations.

Sublethal observations / clinical signs:

Biological results:

% mortality of fish in the main test:

Test item concentration nominal/ measured [mg/L]	Control	Nominal: 100 mg/L Measured: 79.4 mg/L
Time [h]	Mortality (%)
0	0	0
4	0	0
24	0	0
48	0	0
72	0	0
96	0	0

Analytical results:

Determined concentrations of test item during the test:

Test item nominal [mg/L]	Sampling	Test item		Geometric mean	Test item concentration measured
		[mg/L]	[%]	[%]	[mg/L]
0	0 h fresh	n.d.	-	-	-
	24 h aged	n.d.	-
	24 h fresh	n.d.	-
	48 h aged	n.d.	-
	48 h fresh	n.d.	-
	72 h aged	n.d.	-
	72 h fresh	n.d.	-
	96 h aged	n.d.	-
100	0 h fresh	100	100	79.4	79.4
	24 h aged	56.6	57
	24 h fresh	102	102
	48 h aged	77.2	77
	48 h fresh	94	94
	72 h aged	51.7	52
	72 h fresh	103	103
	96 h aged	66.1	66

Validity criteria fulfilled:

yes

Conclusions:

The LC50 (96 h) of the test item was determined to be > 100 mg/L (test item nominal) and > 79.4 mg/L (test item measured). The corresponding NOEC (mortality) (96 h) was 100 mg/L test item (nominal) and 79.4 mg/L test item (measured), respectively. No sublethal effects were observed in the control and at the limit test concentration of 100 mg/L (nominal) and 79.4 mg/L (measured) after 96 h.

Executive summary:

The effects of the test item on the mortality of the Zebrafish Danio rerio were determined under worst-case exposure conditions according to OECD 203. The test item was evaluated in a semi-static limit test with daily water renewal,with one test item concentration (100 mg/L, nominal) and one control with untreated test medium. The test concentration and control contained seven fish. The test duration was 96 hours. Assessments on effects and mortality after 0, 4, 24, 48, 72 and 96 hours were conducted.

The test concentration was analytically verified. The measured content of test in fresh test solutions was between 94 % and 103 % of nominal and  52 -77 % of nominal in aged test solutions. Therefore, the effect concentrations were also evaluated based on measured concentrations, more precisely, geometric mean of the measured concentrations.

The LC50 (96 h) of the test item was determined to be > 100 mg/L (test item nominal) and > 79.4 mg/L (test item measured, geometric mean). The corresponding NOEC (mortality) (96 h) was 100 mg/L test item (nominal) and 79.4 mg/L test item (measured, geometric mean), respectively. No sublethal effects were observed in the control and at the limit test concentration of 100 mg/L (nominal) and 79.4 mg/L (measured) after 96 h.

Description of key information

For this endpoint only one test is available. The effects of the test item on the mortality of the Zebra fish Danio rerio were determined according to OECD 203. The test item was evaluated in a semi-static limit test with daily water renewal. One test item concentration (100 mg/L, nominal) and one control with untreated test medium were used.

The LC50 (96 h) of the test item was determined to be > 100 mg/L (test item nominal), which corresponds to > 79.4 mg/L measured, geometric mean. The corresponding NOEC (mortality) (96 h) was 100 mg/L test item (nominal) and 79.4 mg/L test item (measured, geometric mean), respectively.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

79.4 mg/L

Additional information

For this endpoint, one test were the mortality of the Zebrafish Danio rerio was determined according to OECD 203 is available.The test item was evaluated in a semi-static limit test with daily water renewal. One test item concentration (100 mg/L, nominal) and one control with untreated test medium was used. The test concentration and control contained seven fish each. The test duration was 96 hours. Assessments of effects and mortality after 0, 4, 24, 48, 72 and 96 hours were conducted.

The LC50 (96 h) of the test item was determined to be > 100 mg/L (test item nominal), which corresponds to > 79.4 mg/L measured, geometric mean. The corresponding NOEC (mortality) (96 h) was 100 mg/L test item (nominal) and 79.4 mg/L test item (measured), respectively. No sublethal effects were observed in the control and at the limit test concentration of 100 mg/L (nominal) and 79.4 mg/L (measured, geometric mean) after 96 h.