Disodium 7-[[4,6-bis[(2-hydroxyethyl)amino]-1,3,5-triazin-2-yl]amino]-4-hydroxy-3-[[4-[(4-sulphonatophenyl)azo]phenyl]azo]naphthalene-2-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:

February from 15th to 19th, 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

Adopted April 13, 2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

February 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

The analytical determination of the test substance concentration was performed at the beginning and at the end of the test. The sample for analysis (0 hours) was prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. An analysis of sample was started immediately after delivery.
The sample for analysis at the end of the test (48 hours) was delivered to analytical laboratory immediately after the end of testing. An analysis of sample was started immediately after delivery. All samples were stored at laboratory temperature.

Vehicle:

no

Details on test solutions:

The stock solution of the test substance was prepared in the dilution water. 100 mg of the test substance was weighed into 1000 ml of the dilution water for the limit test.
Stock solution of the test substance: 49.92 mg/500 ml

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Source: Daphnia magna Straus were obtained by aperiodic parthenogenesis from own laboratory healthy stick breeding (at least third generation).
- Culuring: the grow-up gravid females are separated into dilution water. Young daphnia born in 24 hours are segregated to the handling container from which they entered the experiment. At handling a care should be taken of not to penetrate the air under animal’s shell. 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).
- Feeding during test: none.

ACCLIMATION
- Acclimation period: brood daphnids are maintained in dilution water for 48 hours prior to the start of the test.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

2.53 mmol/l

Test temperature:

20 ± 2 °C

pH:

Control: 7.6 - 7.9
Test vessels: 7.6 - 7.8

Dissolved oxygen:

Control: 7.6 - 7.9 mg/l
Test vessels: 7.6 - 7.8 mg/l

Nominal and measured concentrations:

100 mg/l nominal

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 ml glass bekers.
- Type: covered.
- Aeration: none.
- Number of test animals: 20 for concentration and control.
- Numbers of replicates: 4 per concentration.
- Number of test animals per replicate: 5 animals.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: 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 1000 ml of deionized water.
A. 117.6 g CaCl2•2H20 in 1 l of deionized water
B. 49.3 g MgSO4•7H2O in 1 l of deionized water
C. 25.9 g NaHCO3 in 1 l of deionized water
D. 2.3 g KCl in 1 l of deionized water
- Conductivity of deionized water: 1.65 µS/cm
- pH: 7.9
- Oxygen content: 7.9 mg/l
- Content of (Ca+Mg) in dilution water: 2.53 mmol/l

OTHER TEST CONDITIONS
- Adjustment of pH: none.
- Photoperiod: 16-hours light and 8-hours dark cycle.

EFFECT PARAMETERS MEASURED
Immobilisation of daphnia. Those animals that are not able to swim within 15 seconds, after gentle agitation of the test vessels are considered to be immobilized (even if they can still move their antennae). Acute toxicity is expressed as the median effective concentration EC50.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Immobility, any abnormal behaviour and any abnormal appearance was not observed during the limit test.

The analytical results showed that the test substance was stable in dilution water under current test conditions.

Results with reference substance (positive control):

The sensitivity of the test species and correctness of test performance is periodically verified in six-month period by testing with the reference substance, potassium dichromate.
EC50 (48h): 0.83 mg/l (95 % confidence limit: 0.69 – 1.01 mg/l)
EC50 (24h): 0.51 mg/l (95 % confidence limit: 0.44 – 0.58 mg/l)

Determined concentrations at the beginning and at the end of the limit test

Nominal concentration (mg/l)	Determined concentration 0 hours (mg/l)	Determined concentration 48 hours (mg/l)	Change of the test substance concentration in interval 0-48 hrs (% of initial concentration)
100	98.2	97.2	-1

Validity criteria fulfilled:

yes

Remarks:

in the control not more that 10 % of the daphnids have been immobilised and the dissolved oxygen concentration at the end of the test resulted higher than 3 mg/l in control and test vessels

Conclusions:

EC50 (48h) > 100 mg/l (nominal)

Executive summary:

The substance was tested in acute immobilisation test on Daphnia magna; the test was performed according to the OECD guideline 202. The limit test was performed with the test substance nominal concentration 100 mg/l.  

The samples for analytical determination were taken at the beginning and at the end of the test: the test substance was stable in dilution water at conditions of the test (measured concentration was within ± 20 % of the nominal one).

No immobilisation was observed during the test; no abnormal behaviour neither abnormal appearance were observed.

Conclusion

EC50 (48h) > 100 mg/l (nominal)

Description of key information

Not harmful/toxic for aquatic invertebrates after short-term exposures.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

Direct Red 253 was tested in acute immobilisation test on Daphnia magna; the test was performed according to the OECD guideline 202. The limit test was performed with the test substance nominal concentration 100 mg/l. The samples for analytical determination were taken at the beginning and at the end of the test: the test substance was stable in dilution water at conditions of the test (measured concentration was within ± 20 % of the nominal one).

No immobilisation was observed during the test; noabnormal behaviour neither abnormal appearance were observed (Roupcová, 2016).