7-anilino-4-hydroxy-3-[[6-methoxy-4-[(6-sulpho-2,4-xylyl)azo]-m-tolyl]azo]naphthalene-2-sulphonic acid, sodium salt, compound with 2,2',2''-nitrilotriethanol

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From February 06th to April 08th, 2017

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Justification for type of information:

Details for read across approach are included into the IUCLID section 13.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

april 2004

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

The concentration and stability of the test item in the test preparations was verified by chemical analysis at 0 and 48 hours.
Samples were taken from the control and each test group from the bulk test preparation at 0 hours, and from the pooled replicates at 48 hours for quantitative analysis. All samples were stored frozen prior to analysis. Duplicate samples were taken at 0 and 48 hours, and stored frozen for further analysis, if necessary.
Only the samples for the 50 % and 100 % v/v saturated solution test concentrations were analyzed.

Details on test solutions:

A nominal amount of test item (1100 mg) was dispersed in 11 liters of test water with the aid of propeller stirring at approximately 1500 rpm for 24 hours. After 24 hours, the stirring was stopped and any undissolved test item was removed by filtration through a 0.2 μm Sartorius Sartopore filter (first approximate 2 liters discarded in order to pre-condition the filter) to give a 100 % v/v saturated solution. A series of dilutions was made from this saturated solution to give further test concentrations of 6.25, 12.5, 25 and 50% v/v saturated solution.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Justification for species other than prescribed by test guideline:
- Age of parental stock: gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing.
- Feeding during test: daphnids received no food during exposure.

BREEDING
- Breeding conditions: Elendt M7 medium, in a temperature controlled room maintaining the water temperature at 18 to 22 °C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods.
- Type and amount of food: algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension.
- Feeding frequency: each culture was fed daily.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test temperature:

18 to 22 °C with a maximum deviation of ± 1 °C

pH:

Test vessels: 7.6 - 7.9
Control: 7.8 - 8.0

Dissolved oxygen:

8.6 - 8.8 mg O2/l

Nominal and measured concentrations:

6.25, 12.5, 25, 50 and 100% v/v saturated solution

Details on test conditions:

TEST SYSTEM
- Test vessel: 150 ml glass beakers containing approximately 100 ml of test preparation.
- Type: vessels were then covered to reduce evaporation.
- Aeration: the test vessels were not aerated.
- No. of organisms per vessel: five daphnids were placed in each test and control vessel at random.
- No. of vessels per concentration: 4 replicates.
- No. of vessels per control: 4 replicates.

TEST MEDIUM / WATER PARAMETERS
- Preparation of dilution water: reconstituted water (Elendt M7 medium).

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods.
- Light intensity: between 200 and 1200 Lux.

EFFECT MEASURED
Any immobilization or adverse reactions to exposure were recorded at 24 and 48 hours after the start of exposure. The criterion of effect used was that daphnia were considered to be immobilized if they were unable to swim within 15 seconds after gentle agitation.

PARAMETERS MEASURED
Water temperature was recorded daily throughout the test. Dissolved oxygen concentrations and pH were recorded at the start and termination of the test. The pH and dissolved oxygen concentration were measured using a Hach Flexi handheld meter, whilst the temperature was measured using a Hanna Instruments HI 93510 digital thermometer. The light intensity during the light period was measured using an ATP Instrumentation Lux meter.
The appearance of the test media was recorded daily.

RANGE-FINDING STUDY
- Test concentrations: 0.10, 1.0, 10 and 100 mg/l.
- No. of organisms per vessel: 10 daphnids were placed in each test and control vessel

VALIDATION CRITERIA
The results of the test are considered valid if the following performance criteria are met:
- No more than 10 % of the control daphnids show immobilization, or other signs of disease or stress (e.g. discoloration or unusual behavior such as trapping at the surface water).
- The dissolved oxygen concentration at the end of the test is equal to or greater than 3 mg/l in the control and test vessels.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 98 mg/L

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

The No Observed Effect Concentration after 24 and 48 hours exposure was 98 mg/l.
A sub-lethal effect of exposure was observed in the 6.25 % v/v saturated solution test concentration. The response was trapping at the surface, however, as the effect was only observed in one daphnia after 24 hours this was considered not to be significant.

APPEARANCE OF TEST MEDIUM
At the start and throughout the test, the control was observed to be a clear, colorless solution. The 6.25 v/v saturated solution test group was observed to be a clear, purple solution. The 12.5 % v/v saturated solution test group was observed to be a clear, dark purple solution. The 25, 50 and 100 % v/v saturated solution test groups were observed to be opaque, purple solutions.

MEAASURED CONCENTRATIONS
Analysis of the 50 and 100 % v/v saturated solution test preparations at 0 hours showed measured test concentrations of 48.7 and 98.3 mg/L, respectively. There was no significant change in the measured concentrations at 48 hours, and so the results are based on 0-Hour measured test concentrations only.

RANGE-FINDING STUDY
No immobilization was observed at the test concentrations of 0.10 and 1.0 mg/l; however, immobilization was observed at 10 and 100 mg/l.
Undissolved test item was observed in the 1.0, 10 and 100 mg/l test concentrations. A media preparation trial was undertaken to determine the appropriate test item preparation method under test conditions. Based on this information test concentrations of 6.25, 12.5, 25, 50 and 100 % v/v saturated solution were selected for the definitive test.
Chemical analysis of the 1.0 and 10 mg/l test preparations at 0 hours showed measured test concentrations of 1.08 and 10.9 mg/l, respectively. There was no significant change in the measured concentrations at 48 hours indicating that the test item was stable under test conditions. Analysis of the 100 mg/l test preparation at 0 and 48 hours showed measured test concentrations of 61.0 and 43.7 mg/l, respectively. The results were considered inconclusive due to the presence of undissolved test item in the samples.

Results with reference substance (positive control):

EC50 (48h): 1.2 mg/l (nominal)

Water Quality Measurements

Nominal Concentration (% v/v Saturated Solution)	0 Hours			24 hours	48 Hours
	pH	mg O2/l	T °C	T °C	pH	mg O2/l	T °C
Control	7.8	8.8	21	22	8	8.6	21
6.25	7.7	8.8	21	21	7.9	8.6	21
12.5	7.7	8.8	22	22	7.9	8.6	21
25	7.7	8.8	22	22	7.8	8.6	21
50 (49 mg/l)	7.7	8.8	22	22	7.8	8.6	21
100 (98 mg/l)	7.6	8.8	22	22	7.9	8.7	21

Validity criteria fulfilled:

yes

Remarks:

none of the control daphnids showed immobilization or other signs of disease or stress, and that the oxygen concentration at the end of the test was equal to or greater than 3 mg/l in the control and test vessels

Conclusions:

EC50 (48h) > 98 mg/l (nominal)

Executive summary:

A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (April 2004) No. 202, "Daphnia sp., Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.

Preliminary solubility work conducted indicated that it was not possible to obtain a testable solution of the test item using traditional methods of preparation (e.g. ultrasonication and high shear mixing). A saturated solution method of preparation was identified as the best method to obtain a test solution.

Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to an aqueous solution of the test item at nominal concentrations of 6.25, 12.5, 25, 50 and 100 % v/v saturated solution for 48 hours at a temperature of 21 to 22 °C under static test conditions. The test item solutions were prepared by stirring an excess (100 mg/l) of test item in test water using a propeller stirrer at approximately 1500 rpm for 24 hours. After the stirring period, any undissolved test item was removed by filtration (0.2 μm Sartorius Sartopore filter, first approximate 2 liters discarded in order to pre-condition the filter) to produce a 100 % v/v saturated solution of the test item. This saturated solution was then further diluted as necessary, to provide the remaining test concentrations. Immobilization and any adverse reactions to exposure were recorded after 24 and 48 hours.

Analysis of the 50 and 100 % v/v saturated solution test preparations at 0 hours showed measured test concentrations of 48.7 and 98.3 mg/l, respectively. There was no significant change in the measured concentrations at 48 hours, and so the results are based on 0-Hour measured test concentrations only.

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated and based on the 0-Hour measured test concentrations gave a 48-Hour EC50 value of greater than 98 mg/l. The No Observed Effect Concentration was 98 mg/l.

Conclusion

EC50 (48h) > 98 mg/l (nominal)

Description of key information

Not harmful/toxic for short-term exposure to aquatic invertebrates

Additional information

There are no information about the short-term toxicity to aquatic invertebrates potential of Direct Violet 051, therefore, the available data on structural analogous Similar Substance 01 have been taken into account. The read across approach can be considered as reliable and adequate for the purpose; details and explanations are detailed in the report attached to the IUCLID section 13.

A study was performed to assess the acute toxicity of the Similar substance to Daphnia magna. The method followed was designed to be compatible with the OECD Guideline 202. Following a preliminary range-finding test, twenty daphnids were exposed to the test item at nominal concentrations of 6.25, 12.5, 25, 50 and 100 % v/v saturated solution, for 48 hours, under static conditions. Analysis of the 50 and 100 % v/v saturated solution test preparations at 0 hours showed measured test concentrations of 48.7 and 98.3 mg/l, respectively. There was no significant change in the measured concentrations at 48 hours, and so the results are based on 0-Hour measured test concentrations only. The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated and based on the 0-Hour measured test concentrations gave a 48-Hour EC50 value of greater than 98 mg/l. The No Observed Effect Concentration was 98 mg/l.