Dipotassium dihydrogen 4-[5-[3-carboxylato-5-hydroxy-1-(4-sulphonatophenyl)-1H-pyrazol-4-yl]penta-2,4-dienylidene]-4,5-dihydro-5-oxo-1-(4-sulphonatophenyl)-1H-pyrazole-3-carboxylate

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 - 30 Nov 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hessisches Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Test material

Test material information

Constituent 1

Reference substance name:

Dipotassium dihydrogen 4-[5-[3-carboxylato-5-hydroxy-1-(4-sulphonatophenyl)-1H-pyrazol-4-yl]penta-2,4-dienylidene]-4,5-dihydro-5-oxo-1-(4-sulphonatophenyl)-1H-pyrazole-3-carboxylate

EC Number:

257-479-6

EC Name:

Dipotassium dihydrogen 4-[5-[3-carboxylato-5-hydroxy-1-(4-sulphonatophenyl)-1H-pyrazol-4-yl]penta-2,4-dienylidene]-4,5-dihydro-5-oxo-1-(4-sulphonatophenyl)-1H-pyrazole-3-carboxylate

Cas Number:

51858-17-4

Molecular formula:

C25H16N4O12S2.2K

IUPAC Name:

dipotassium dihydrogen 4-{5-[3-carboxylato-5-oxo-1-(4-sulfonatophenyl)-1,5-dihydro-4H-pyrazol-4-ylidene]penta-1,3-dien-1-yl}-5-hydroxy-1-(4-sulfonatophenyl)-1H-pyrazole-3-carboxylate

Test material form:

solid: crystalline

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All test item concentrations and the control
- Sampling method: Duplicate samples were taken at test start and test end.
- Sample storage conditions before analysis: All samples were stored in a freezer (≤ - 20 °C), protected from light until analysis was performed.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test medium of the highest test concentration of nominal 100 mg test item/L was prepared by dissolving 85.7 mg test item into 857 mL test water by intense stirring for 15 min. Adequate volumes of this test medium were diluted with test water to prepare the test media of the other desired test concentrations.
- Controls: Yes (test medium only)
- Observations: The test medium showed blue coloration in all concentrations.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Strain/clone: Clone 5
- Source: in-house laboratory culture
- Age at test start: From 2.25 to 19.25 h old, not first brood progeny.
- Feeding during test: No feeding during the test.

ACCLIMATION
- Acclimation conditions (same as test or not): Same as test
- Type and amount of food: Green algae (Desmodesmus subspicatus)
- Feeding frequency: At least on all working days.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

2.5 mmol/L (= 250 mg/L) as CaCO3

Test temperature:

19.7 - 20.0 (Control)
19.6 - 20.2 (Test item concentrations)

pH:

7.9

Dissolved oxygen:

8.3 - 8.8 mg/L (Control)
8.5 - 8.9 mg/L (Test item concentration)

Conductivity:

< 5 μS/cm

Nominal and measured concentrations:

Control, 6.3, 12.5, 25, 50 and 100 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
Test vessel:
- Type: open (covered with glass plate)
- Material, size, headspace, fill volume: 100 mL glass beakers, containing approx 60 mL, 40 mL headspace
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt medium (M4) was used according to the guideline
- Alkalinity: 0.9 mmol/L
- Conductivity: < 5 μS/cm
- Culture medium different from test medium: No
- Intervals of water quality measurement: The water temperature, pH-values and dissolved oxygen concentrations were determined at test start and test end in each treatment group.

OTHER TEST CONDITIONS
- Photoperiod: 16 h light : 8 h dark
- Light intensity: 260 to 360 lux

EFFECT PARAMETERS MEASURED: The mobility of the daphnids was determined by visual observation after 24 and 48 h. Those animals that are not able to swim within 15 sec after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).

RANGE-FINDING STUDY: non-GLP pre-experiment
- Results used to determine the conditions for the definitive study: The test item concentrations of the main test were based on the results of a non GLP pre-experiment.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (tested at least twice a year)

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

93.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: Statistical evaluation was based on two values only:

Remarks:

50 and 100 mg/L

Effect concentrations 2

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Mortality of control: no mortality in the controls
- Others: Colouration caused by the test item was observed in all test item concentrations but especially in the highest two test item concentration.

Results with reference substance (positive control):

The EC50 after 24 h was determined to be 1.58 mg test item/L, indicating that the sensitivity of the daphnids was consistent with the level proposed by the OECD 202 guideline (EC50 (24 h) between 0.6 and 2.1 mg potassium dichromate/L).

Reported statistics and error estimates:

The software used to perform the statistical analysis was ToxRat Professional, Version 3.2.1, ToxRat® Solutions GmbH.

Any other information on results incl. tables

Analytical Results:

At the start of the test 102% of the nominal test concentrations were found (average of all test concentrations). After 48 h test duration, 102% of the nominal value was determined (average of all test concentrations). During the test the daphnids were exposed to a mean of 102% of nominal. Thus, the effect concentrations were based on the nominal values.

Table 1: Summary of Analytical Results

Sample Description	% of	RSD
[mg test item/L]	nominal 1)	[%]	n
Control	n.a.	n.a.	4
6.3	109	2	4
12.5	104	1	4
25	100	1	4
50	98	1	4
100	99	0	4

1) mean value of all measured samples per treatment group RSD: relative standard deviation per treatment group n: number of analysed samples n.a.: not applicable

Table 2: Biological Results

Nominal Concentration	No. of Daphnia tested	No. of immobilized Daphnia after		% of immobilized Daphnia after
[mg test item/L]		24 h	48 h	24 h	48 h
Control	20	0	0	0	0
6.3	20	0	0	0	0
12.5	20	0	0	0	0
25	20	0	0	0	0
50	20	0	0	0	0
100	20	0	14	0	70