Reaction products of 1,3-dihydroxybenzene, 4-amino-5-hydroxynaphthalene-2,7-disulphonic acid, 4-methoxy-1-aminobenzene, 2-(4-aminoanilino)-5-nitrobenzenesulphonic acid, sodium salt

Short-term toxicity to fish

Link to relevant study record(s)

Reference

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:

yes

Remarks:

See Any othfer information on materials and methods

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Test substance name: Acid Brown 235
- Other name: Korostan Brown DGR
- Source and lot/batch No. of test material: 7012/2007
- Expiration date of the lot/batch: May 2018
- Appearance: dark brown powder

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 (10 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.

Vehicle:

no

Details on test solutions:

The stock solution of the test substance was prepared in the dilution water. 400 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 cm:

ACCLIMATION
- Acclimation period: 7 days
- Acclimation conditions: same as test
- Type and amount of food during acclimation: TetraMin – complete feed for aquarium fish (manufacturer: Tetra GmbH, Germany)
- Feeding frequency during acclimation: once a day
- Health during acclimation: mortality 0.33 % (1 fish from 300 individuals)

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Test temperature:

22 ± 2°C, with a variation of ± 1°C at each individual test

pH:

7.5 - 8.1

Dissolved oxygen:

6.8 - 8.6 mg/L

Nominal and measured concentrations:

nominal concentration 10 mg/L:
9.60 mg/L (0 h)
9.54 mg/L (96 h)
nominal concentration 100 mg/L:
89.8 mg/L (0 h)
99.3 mg/L (96 h)

Details on test conditions:

TEST SYSTEM
- Aeration: without aeration
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
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
B: 49.3 g MgSO4·7H2O in 1L deionized water
C: 25.9 g NaHCO3 in 1L deionized water
D: 2.3 g KCl in 1L deionized water
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 2.55 mmol/L in the preliminary test.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: daily light

TEST CONCENTRATIONS
- Test concentrations: 100, 80, 40, 20 and 10 mg/L

Reference substance (positive control):

yes

Remarks:

potassium dichromate (CAS: 7778-50-9)

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:

no animals died
see Table 1

Results with reference substance (positive control):

- Positive control substance: potassium dichromate
- LC50 = 110 mg/L (Poecilia reticulata, 96 h)
- Interlaboratory test data: LC50 = 86 - 200 mg/L (Poecilia reticulata, 96 h)

Sublethal observations / clinical signs:

Table 1:Fish mortality

Nominal concentration	mg/L	100	80	40	20	10	Control
Number of fish alive	2 hours	7	7	7	7	7	7
Mortality %		0	0	0	0	0	0
Number of fish alive	24 hours	7	7	7	7	7	7
Mortality %		0	0	0	0	0	0
Number of fish alive	48 hours	7	7	7	7	7	7
Mortality %		0	0	0	0	0	0
Number of fish alive	72 hours	7	7	7	7	7	7
Mortality %		0	0	0	0	0	0
Number of fish alive	96 hours	7	7	7	7	7	7
Mortality %		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 (Poecilia reticulata, 96 h) (nominal concentration)
LC50 > 100 mg/L (Poecilia reticulata, 72 h) (nominal concentration)
LC50 > 100 mg/L (Poecilia reticulata, 48 h) (nominal concentration)
LC50 > 100 mg/L (Poecilia reticulata, 24 h) (nominal concentration)

LC0 = 100 mg/L (Poecilia reticulata, 96 h) (nominal concentration)
LC0 = 100 mg/L (Poecilia reticulata, 72 h) (nominal concentration)
LC0 = 100 mg/L (Poecilia reticulata, 48 h) (nominal concentration)
LC0 = 100 mg/L (Poecilia reticulata, 24 h) (nominal concentration)

LC100 > 100 mg/L (Poecilia reticulata, 96 h) (nominal concentration)
LC100 > 100 mg/L (Poecilia reticulata, 72 h) (nominal concentration)
LC100 > 100 mg/L (Poecilia reticulata, 48 h) (nominal concentration)
LC100 > 100 mg/L (Poecilia reticulata, 24 h) (nominal concentration)

Executive summary:

The test substance, Acid Brown 235, 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 10 – 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.

Test results:

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

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

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

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

 

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

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

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

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

 

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

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

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

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

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

100 mg/L