Trisodium 2-[[6-[(4-amino-6-chloro-1,3,5-triazin-2-yl)methylamino]-1-hydroxy-3-sulphonato-2-naphthyl]azo]naphthalene-1,5-disulphonate

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

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 analyzed on the day of delivery. The samples for analysis at the end of the test (96 hours) were delivered to analytical laboratory immediately after the end of testing. The samples were analyzed 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 under current test conditions.

Vehicle:

no

Details on test solutions:

Standard dilution water was prepared just before the test by admixing 2.5 mL of stored solutions A, B, C and D in 1000 mL of deionized water.A: 117.6 g CaCl2·2H20 in 1L deionized water (Batch No.: PP/2015/01588)B: 49.3 g MgSO4·7H2O in 1L deionized water (Batch No.: PP/2014/03001)C: 25.9 g NaHCO3 in 1L deionized water (Batch No.: PP/2016/04662)D: 2.3 g KCl in 1L deionized water (Batch No.: PP/2015/02174)The dilution water should have a pH 7.8±0.2, value measured in this study was 8.0 in the preliminary test. The content of sum Ca + Mg ions should have the value of 2.50±0.10 mmol/L, value measured in this study was 2.48 mmol/L in the preliminary test. The stock solution of the test substance was prepared in the dilution water. 200 mg of the test substance was weighed into 2000 mL of the dilution water for the preliminary test. The concentrations of solutions used in the preliminary test were obtained by dilution of the stock solution with dilution water.

Test organisms (species):

Poecilia reticulata

Details on test organisms:

TEST ORGANISM- Common name: Poecilia reticulata- Source: certified supplier EMPLA s.r.o., Hradec Králové (Czech Republic) - Age at study initiation: 3 - 4 months- Length at study initiation: 2 - 3 cmACCLIMATION- Acclimation period: 7 days- Acclimation conditions: in dilution water under same conditions as in the test- Type and amount of food during acclimation: “TetraMin” – complete feed for aquarium fish (manufacturer: Tetra, Germany), - Feeding frequency during acclimation: once a day- Health during acclimation (any mortality observed): The mortality of the fish was observed for 12 days from the times of delivery. The recorded mortality was 0.33 % (1 fish from 300 individuals).FEEDING DURING TEST- without feeding, without aeration

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Test temperature:

21.1 - 21.8°C

pH:

7.3 - 8.0

Dissolved oxygen:

6.8 - 8.3 mg/L

Conductivity:

2.48 mmol/L Ca+Mg

Nominal and measured concentrations:

Nominal concentration: 100 mg/L 0 h: 99.7 mg/L (measured) 96 h: 106.2 mg/L (measured)Nominal concentration: 1 mg/L 0 h: 0.98 mg/L (measured) 96 h: 1.03 mg/L (measured)

Details on test conditions:

TEST SYSTEM - Test vessel: glass beaker - Aeration: no - No. of organisms per vessel: 7 - No. of vessels per concentration (replicates): 1 - No. of vessels per control (replicates): 1 OTHER TEST CONDITIONS - Adjustment of pH: no - Photoperiod: daylight TEST CONCENTRATIONS - Test concentrations: 100, 50, 10, 5 and 1 mg/L

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

72 h

Dose descriptor:

LC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC0

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

72 h

Dose descriptor:

LC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

see Table 1

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 the reference substance, potassium dichromate.The results of verification test with K2Cr2O7 are the following:LC50 = 152 mg/L (96 h)From the result of the last interlaboratory test arranged by CSlab spol. s r.o. in 2016 we have obtained data for potassium dichromate 96 hours – LC50 generated at 12 laboratories. The following reference range of values could be calculated from those data:LC50 = 86 – 200 mg/L (96 h)The mortality of test organisms by the reference substance (96 hours, LC50) obtained from our last reference test meets the calculated range from the interaboratory test.

Sublethal observations / clinical signs:

Table 1: Fish mortality

Nominal concentration	mg/L	100	50	10	5	1	Control
Number of fish alive	2 hours	7	7	7	7	7	7
Mortality %	2 hours	0	0	0	0	0	0
Number of fish alive	24 hours	7	7	7	7	7	7
Mortality %	24 hours	0	0	0	0	0	0
Number of fish alive	48 hours	7	7	7	7	7	7
Mortality %	48 hours	0	0	0	0	0	0
Number of fish alive	72 hours	7	7	7	7	7	7
Mortality %	72 hours	0	0	0	0	0	0
Number of fish alive	96 hours	7	7	7	7	7	7
Mortality %	96 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 7 fish, therethrough the conditions of the limit test have been fulfilled. It was demonstrated that the LC50 is higher than this concentration.No mortality was observed in the preliminary test. The exact values of LC50 could not be calculated and the values of LC are given in the form of a range. The LC100 and LC0 values were 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. We choose the nominal concentrations for the evaluation.LC50 > 100 mg/L (96 h) (nominal concentration)LC50 > 100 mg/L (72 h) (nominal concentration)LC50 > 100 mg/L (48 h) (nominal concentration)LC50 > 100 mg/L (24 h) (nominal concentration)LC0 = 100 mg/L (96 h) (nominal concentration)LC0 = 100 mg/L (72 h) (nominal concentration)LC0 = 100 mg/L (48 h) (nominal concentration)LC0 = 100 mg/L (24 h) (nominal concentration)LC100 > 100 mg/L (96 h) (nominal concentration)LC100 > 100 mg/L (72 h) (nominal concentration)LC100 > 100 mg/L (48 h) (nominal concentration)LC100 > 100 mg/L (24 h) (nominal concentration)

Executive summary:

The test substance, Reactive Orange 13, was tested for acute toxicity on fish Poecilia reticulata. The test was performed according to method C.1 – Acute toxicity for fish, 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. 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 7 fish, therethrough the conditions of the limit test have been fulfilled. It was demonstrated that the LC₅₀is higher than this concentration. The test was performed as static.

Samples for the 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 was stable in 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.

For all evaluations and results the nominal concentrations of the test substance were used.

LC₅₀> 100 mg/L (96 h) (nominal concentration)

LC₅₀> 100 mg/L (72 h) (nominal concentration)

LC₅₀> 100 mg/L (48 h) (nominal concentration)

LC₅₀> 100 mg/L (24 h) (nominal concentration)

LC₀= 100 mg/L (96 h) (nominal concentration)

LC₀= 100 mg/L (72 h) (nominal concentration)

LC₀= 100 mg/L (48 h) (nominal concentration)

LC₀= 100 mg/L (24 h) (nominal concentration)

LC₁₀₀> 100 mg/L (96 h) (nominal concentration)

LC₁₀₀> 100 mg/L (72 h) (nominal concentration)

LC₁₀₀> 100 mg/L (48 h) (nominal concentration)

LC₁₀₀> 100 mg/L (24 h) (nominal concentration)

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: meets generally accepted scientific principles, acceptable for assessment

Qualifier:

no guideline available

Principles of method if other than guideline:

The study report contains the limited information on the test conditions because the study has been performed in 1994. However available information is sufficient to conclude on the classification of the substance. Further the test was performed on the vertebrates and use of results from old experimental studies is one of the options to provide information requested by REACH. New experimental studies with vertebrates must only be conducted if there is no adequate existing information.

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

no

Test organisms (species):

Poecilia reticulata

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Remarks on exposure duration:

The exposure time is not explicitly reported. In this laboratory, the exposure time for an acute toxicity test on fish was 96 h.

Nominal and measured concentrations:

1000, 900, 800 and 600 mg/l

Reference substance (positive control):

not specified

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

No death was observed

Conclusions:

According to Regulation (EC) No. 1272/2008 (CLP), the test substance, Reactive Orange 13, need not be classified as Aquatic Acute substance.

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1983

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Justification for type of information:

None

Principles of method if other than guideline:

None

GLP compliance:

no

Specific details on test material used for the study:

None

Analytical monitoring:

not specified

Details on sampling:

None

Vehicle:

yes

Remarks:

water

Details on test solutions:

None

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

None

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Remarks on exposure duration:

None

Post exposure observation period:

None

Hardness:

None

Test temperature:

25

pH:

10

Dissolved oxygen:

None

Salinity:

None

Conductivity:

None

Nominal and measured concentrations:

300 and 1000 mg/l

Details on test conditions:

None

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC100

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

None

Results with reference substance (positive control):

None

Reported statistics and error estimates:

None

Sublethal observations / clinical signs:

None

Validity criteria fulfilled:

no

Conclusions:

The LC50 of FAT 40170 in this fish acute toxicity test was determined to be greater than 1000 mg/L.

Executive summary:

An acute fish toxicity test was performed with Rainbow trout species. Groups of fish were exposed at the test concentrations of 300.0 and 1000.0 mg/L. No mortality was observed throughout the duration of the test. Hence, the LC50 of FAT 40170 was determined to be >1000 mg/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

100 mg/L