A mixture of: trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-(4-nitro-2-sulfonatoanilino)phenylazo)phenolato)ferrate(1-); trisodium bis(2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(4-nitro-2-sulfonatophenylazo)phenolato)ferrate(1-); trisodium (2,4(or 2,6 or 4,6)-bis(3,5-dinitro-2-oxidophenylazo)-5-hydroxyphenolato)(2(or 4 or 6)-(3,5-dinitro-2-oxidophenylazo)-5-hydroxy-4(or 2 or 6)-(3-sulfonatophenylazo)phenolato)ferrate(1-); disodium 3,3'-(2,4-dihydroxy-1,3(or 1,5 or 3,5)-phenylenediazo)dibenzenesulfonate

Endpoint summary

Description of key information

Short-term toxicity to fish

LC50 (96 h, Cyprinus carpio) = 3.52 mg/L (nominal)

Long-term toxicity to fish

NOEC (21 d, Oncorhynchus mykiss) =  1.0 mg/L (geometric mean of measured conc.)

Short-term toxicity to aquatic invertebrates

EC50 (48 h, Daphnia magna) =46.52 mg/L (geometric mean of measured conc.)

Toxicity to aquatic algae and cyanobacteria

ErC50 (72 h, Scenedesmus subspicatus)= 21.9 mg/L

Toxicity to microorganisms

IC 50 (3 h) >100 mg/L (nominal)

Additional information

Short-term toxicity to fish

The short-term toxicity to fish (Cyprinus carpio) was evaluated performing a study in accordance with the OECD guideline 203 (1984). Ten fish per concentration were exposed for 96 hours in a semi-static system to a range of nominally 1 to 10 mg/L forming a geometric progression with a factor of 1.8. The test media were renewed after 48 hours. After 24 hours of exposure all fish had died at nominally 10 mg/L and 90 % mortality was recorded at 5.6 mg/l. After 96 hours of exposure the lowest concentration at which 100 % mortality was recorded still was 10 mg/L (LC100), with 90 % mortality at 5.6 mg/L and 40 % mortality at 3.2 mg/L. No mortality of fish was observed at and below nominally 1.8 mg/L (LC0). Effects other than mortality, were observed in fish exposed to 3.2 mg/L and higher. The effects recorded were hypoactivity, loss of equilibrium and immobility. No effects were recorded in the fish exposed to 1.8 mg/L and lower.

Analysis of the samples taken at the start (t=0) of the final test revealed mean actual concentrations of > 95 % relative to the nominal concentrations of 1, 3.2 and 10 mg/L.

The LC50 value calculated at 96 hours was 3.52 mg/L, with a 95 % confidence interval of 3.07 to 4.44.

Long-term toxicity to fish

The toxicity of test substance to fish (Oncorhynchus mykiss) was assessed over a period of 21 days under flow-through conditions. The test was conducted according to the OECD Guideline No. 204 (1984).

This study examined nominal test article concentrations in the range from nominal 0.05 to 4.0 mg/L which were spaced by a geometric progression factor of 3.

At the nominal test concentration of 0.44 mg/L the analysed concentrations of test item in the test medium varied in the range from 67.2 % to 73.9 % of the nominal value, at the next higher test concentration of nominal 1.33 mg/L in the range from 67.1 % to 80.1 %. At the highest test concentration of nominal 4.0 mg/L 86.3 % of nominal was found. At the lower test concentrations the measured test substance concentrations were mostly lower than 80 % of the nominal values, possibly due to adsorption of the test substance onto surfaces (glass walls of the aquaria, test organisms and their feces). Therefore all biological results are based on the mean measured test substance concentrations (calculated as mean of all measurements per test concentration): 0.3 mg/L at a nominal concentration of 0.44 mg/L, 1,0 mg/L at a nominal concentration of 1.33 mg/L, and 3.5 mg/L at a nominal concentration of 4.0 mg/L.

Per concentration, one group of ten fish each was exposed to the test concentrations and untreated test medium (control). No mortality, no significant clinical signs, and no significant influence on fish growth compared to the control were observed at the test concentrations up to and including 1.0 mg/L within 21 days of exposure. At the highest test concentration of 3.5 mg/L (nominal 4.0 mg/L), all fish were dead within the first 4 days of exposure.

Invertebrate acute toxicity

An experimental study for short-term toxicity to Daphnia magna is available on the substance.Daphnia magnawere exposed for 24 hours to a concentration range of 100 to 1000 mg/L forming a geometric progression with a factor of 1.8 according to OECD Guideline 202 (1984) and the EU method C.2 of Directive 84/449. The test was performed in duplicate with 10 daphnia per vessel.

After 24 hours of exposure the total rate of immobilisation of daphnia was 95 % at 1000 mg/L, 70 % at 560 mg/L. 40 % at 320 mg/L, 30 % at 180 mg/L and 15 % at 100 mg/L.

The EC50 (24 h,Daphnia magna) was calculated to be nominally 313 mg/L with 95 % confidence interval ranging from 236 to 415 mg/L (slope = 2.43).

As the experimental phase was only for 24 h, a weight of evidence approach is followed. Specifically, data from Similar Substance 01 is used in order to complete the assessment of aquatic short-term toxicity toDaphnia magnafollowing a read-across approach. Justification for read-across is detailed in attachment to Short-term toxicity to aquatic invertebrates summary.

A static test was performed using Similar Substance 01 according to the OECD Guideline 202 (2004) and the EU method C.2 of the Regulation (EC) 440/2008. 5 concentrations were used ranging from 4.6 to 100 mg/L (nominal concentration). For each test concentration and the blank control, 20 Daphnia were exposed to the test item for 48 hours. After 24 and 48 hours, the immobilised Daphnia were counted.Potassium dichromate K₂Cr₂O₇was used as positive controlin a current reference study to assure that the test conditions are reliable.At the beginning and at the end of the test, the content of the test item in the test solutions was determined using photometer-determination. The concentrations determined at the start of the test were between 45 and 81 % of the nominal concentration. At the end of the test the determined concentrations were between 94 and 108 % of the nominal concentration. Therefore, the determination of the biological results was based on geometric mean of measured concentrations.All validity criteria were met.

Toxicity to aquatic algae and cyanobacteria

The influence of test substance on the growth of the green algaScenedesmus subspicatuswas investigated. The test method was based on the OECD Guideline No. 201 (1984) and the EEC Directive 92/69 Part C.3, and was modified to differentiate between a reduced growth of algae due to direct inhibitory effects of the test article on the algal cells and a reduced growth of algae due to an indirect effect caused by the light absorption of the coloured test solutions.

The growth inhibition effect on Scenedesmus subspicatus was in part caused by the indirect effect, i.e. the light absorption of the coloured test solutions.

Nevertheless, especially at the highest test concentration of 100 mg/L, a direct inhibitory effect of test item on the algae growth cannot be excluded. Therefore, the results of experimental part in which only the direct inhibitory effect of substance is considered should be used in the assessment of the influence of test substance on the growth of Scenedesmus subspicatus.

At the beginning of the test (immediately after treatment), the actual test item mean measured concentrations were found to be in the range from 81 % to 100 % of nominal, and after 72 hours of exposure from 84 % to 95 % of nominal. Based on these results, the biological data are expressed as nominal concentrations.

The ErC50 (72 h, Scenedesmus subspicatus) was found to be 21.9 mg/L (nominal).

Toxicity to microorganisms

The inhibitory effect of test item on aerobic waste-water bacteria was investigated in two respiration tests following the OECD guideline 209 (1984). The two tests were performed in 1989 and 1990.

In the test of 1990 the test item showed slight inhibition (13.5 %) of the respiration rate up to a concentration of 100 mg/L.

Thus, the IC 50 (3 h) was determined to be >100 mg/L.

Even if to the previous study (1989) a reliability is not assignable as no information about the test item is available, the test results support the results obtained in the subsequent study (1990).

JUSTIFICATION FOR CLASSIFICATION OR NON-CLASSIFICATION 

According to the CLP Regulation (EC) no. 1272/2008, Part 4: Environmental Hazards, the substances can be classified for hazardous to the aquatic environment when the following criteria are met:

A )Acute (short-term) aquatic hazard Category Acute 1: 96 hr LC 50 (for fish) and/or 48 hr EC 50 (for crustacea) and/or 72 or 96 hr ErC 50 (for algae or other aquatic plants) ≤ 1 mg/L.

B) Long-term aquatic hazard

(i) Non-rapidly degradable substances for which there are adequate chronic toxicity data available:

Category Chronic 1: Chronic NOEC or ECx (for fish) and/or Chronic NOEC or ECx (for crustacea) and/or Chronic NOEC or ECx (for algae or other aquatic plants) ≤ 0,1 mg/L

Category Chronic 2: Chronic NOEC or ECx (for fish) and/or Chronic NOEC or ECx (for crustacea) and/or Chronic NOEC or ECx (for algae or other aquatic plants) ≤1 mg/L.

(iii) Substances for which adequate chronic toxicity data are not available and the substance is not rapidly degradable and/or the experimentally determined BCF ≥ 500 (or, if absent, the log K ow ≥ 4).

Category Chronic 1: 96 hr LC 50 (for fish) and/or 48 hr EC 50 (for crustacea) and/or 72 or 96 hr ErC 50 (for algae or other aquatic plants) ≤ 1 mg/L

Category Chronic 2: 96 hr LC 50 (for fish) and/or 48 hr EC 50 (for crustacea) and/or 72 or 96 hr ErC 50 (for algae or other aquatic plants)> 1 to ≤10 mg/L

Category Chronic 3:96 hr LC 50 (for fish) and/or 48 hr EC 50 (for crustacea) and/or 72 or 96 hr ErC 50 (for algae or other aquatic plants) > 10 to ≤ 100 mg/L.

 

The substance is not rapidly degradable and the available long-term and short-term toxicity tests to fish fixed the effect levels that meet the criteria related to the hazard categories for Category Chronic 2. On the other hand, short-term data for invertebrates and algae fixed effect levels that meet the criteria for Category Chronic 3.

Therefore, fish is the most sensitive species and the substance is classified for Aquatic toxicity as Category Chronic 2 according to the CLP Regulation (EC) no. 1272/2008.