Disodium 2,2'-(azodi-p-phenylene)bis[6-methylbenzothiazole-7-sulphonate]

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

Published in O.J. L 142 (2008)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

The analytical determination of the test substance concentrations were performed at the beginning of the test and at the end of the test. The samples for analysis were taken from the highest (100 mg/L) and the lowest test concentration (1 mg/L). The samples for analysis (0 hours) were prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. The samples were analysed on the day of delivery. The samples for analysis at the end of the test (48 hours) were delivered to analytical laboratory immediately after the end of testing. The samples were analysed on the day of delivery. All samples were stored at laboratory temperature. The analytical results showed, that the test substance was stable in dilution water at the conditions of the test.

Vehicle:

no

Details on test solutions:

The holding and culturing of the Daphnia is performed at conditions (light, temperature, medium) similar to those which are used in the test.For holding and culturing of the daphnia the M4 medium was used (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202). For test was used dilution water (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202). Dilution water was prepared just before the test by admixing 2.5 mL of stored solutions A, B, C and D into 1 000 mL of deionized water.As the culturing and test medium was different we included pre-test acclimation period: brood daphnids were maintained in dilution water for 48 hours prior to the start of the test.A: 117.6 g CaCl2·2H20 in 1L deionized waterB: 49.3 g MgSO4·7H2O in 1L deionized waterC: 25.9 g NaHCO3 in 1L deionized waterD: 2.3 g KCl in 1L deionized water The dilution water had a pH 7.8 in the preliminary test. The content of sum Ca + Mg ions were 2.52 mmol/L in the preliminary test. Measured values meet the requirements.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM - Common name: Daphnia magna - Strain/clone: Straus - Source: own laboratory breeding - Feeding during test: no

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

2.52 mmol/L (Ca+Mg ions)

Test temperature:

20±2°C, with a variation of ± 1°C at each individual test

pH:

7.8

Dissolved oxygen:

8.1 - 8.8 mg/L

Nominal and measured concentrations:

Nominal concentration: 1 mg/L Measured concentration: 0.86 mg/L (0 hours) Measured concentration: 1.02 mg/L (48 hours)Nominal concentration: 100 mg/L Measured concentration: 99.3 mg/L (0 hours) Measured concentration: 119.5 mg/L (48 hours)

Details on test conditions:

TEST SYSTEM - Test vessel: glass beaker - Aeration: without aeration - No. of organisms per vessel: 10 - No. of vessels per concentration (replicates): 2 - No. of vessels per control (replicates): 1 OTHER TEST CONDITIONS - Adjustment of pH: no - Photoperiod: daylight RANGE-FINDING STUDY - Test concentrations: 100, 50, 10, 5 and 1 mg/L - Results used to determine the conditions for the definitive study: The experiment with the concentration of 100 mg/L included in the preliminary test has been performed using 20 daphnia, therethrough the conditions of the limit test given in guideline have been fulfilled.

Reference substance (positive control):

yes

Remarks:

potassium dichromate (CAS: 7778-50-9)

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Results with reference substance (positive control):

REFERENCE SUBSTANCEReference substance: K2Cr2O7EC50 = 1.34 mg/L (24 h)EC50 = 0.74 mg/L (48 h)Interlaboratory test: EC50 = 0.46 – 1.08 mg/L (48 h, K2Cr2O7)

Table 1: Immobolization of daphnia

Nominal concentration	mg/L	100	50	10	5	1	Control
Number of mobile animals	24 hours/series 1	10	10	10	10	10	10
Number of mobile animals	24 hours/series 2	10	10	10	10	10	10
Number of mobile animals	24 hours/total	20	20	20	20	20	20
Immobilisation %	24 hours	0	0	0	0	0	0
Number of mobile animals	48 hours/series 1	10	10	10	10	10	10
Number of mobile animals	48 hours/series 2	10	10	10	10	10	10
Number of mobile animals	48 hours/total	20	20	20	20	20	20
Immobilisation %	48 hours	0	0	0	0	0	0

Validity criteria fulfilled:

yes

Conclusions:

The experiment with the concentration of 100 mg/L included in the preliminary test has been performed using 20 daphnia, therethrough the conditions of the limit test given in guidelines have been fulfilled. It was demonstrated that the EC50 is higher than this concentration.No immobilisation was observed in the preliminary test. Therefore exact values of EC50 could not be calculated and the values of EC are given in the form of a range. The EC0 and EC100 values are identified by direct observation.There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 percent of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values. Because concentrations of the test substance have been satisfactorily maintained within ± 20 percent of the measured initial concentration throughout the test, the nominal concentrations were used for all evaluations and results.EC50 > 100 mg/L (24 h, Daphnia magna) (nominal concentration)EC50 > 100 mg/L (48 h, Daphnia magna) (nominal concentration)EC0 = 100 mg/L (24 h, Daphnia magna) (nominal concentration)EC0 = 100 mg/L (48 h, Daphnia magna) (nominal concentration)EC100 > 100 mg/L (24 h, Daphnia magna) (nominal concentration)EC100 > 100 mg/L (48 h, Daphnia magna) (nominal concentration)

Executive summary:

The test substance, Direct Yellow 28, was tested in acute immobilisation test on Daphnia magna. The test was performed according to method C.2 – Daphnia sp. Acute Immobilisation Test, Council Regulation (EC) No. 440/2008, published in O.J. L 142, 2008.

The preliminary test was performed in a range of the test substance nominal concentrations 1 – 100 mg/L. The test was performed as static. Based on no toxicity of the test substance found in the preliminary test, the study was terminated. The experiment with the concentration of 100 mg/L included in the preliminary test has been performed using 20 daphnia, therethrough the conditions of the limit test given in guideline have been fulfilled. It was demonstrated that the EC50 is higher than this concentration.

The samples for analytical determination were taken at the beginning and at the end of the test. The analytical results from the preliminary test showed, that the test substance, Direct Yellow 28, was stable in the dilution water at conditions of the test.

There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 percent of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values.

The nominal concentrations were used for all evaluation and results.

EC50 > 100 mg/L (24 h, Daphnia magna) (nominal concentration)

EC50 > 100 mg/L (48 h, Daphnia magna) (nominal concentration)

EC0 = 100 mg/L (24 h, Daphnia magna) (nominal concentration)

EC0 = 100 mg/L (48 h, Daphnia magna) (nominal concentration)

EC100 > 100 mg/L (24 h, Daphnia magna) (nominal concentration)

EC100 > 100 mg/L (48 h, Daphnia magna) (nominal concentration)

Description of key information

The effective concentration EC50 > 100 mg/L was determined based on immobilisation of daphnia.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L