N-(4-chloro-2,5-dimethoxyphenyl)-3-hydroxy-4-[(E)-2-[2-methoxy-5-(phenylcarbamoyl)phenyl]diazen-1-yl]naphthalene-2-carboxamide; N-(5-chloro-2-methylphenyl)-3-hydroxy-4-[(E)-2-[2-methoxy-5-(phenylcarbamoyl)phenyl]diazen-1-yl]naphthalene-2-carboxamide

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:

1982-02-15 - 1982-02-19

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Pre-guideline study performed similar to OECD 203 with GLP compliance. The results are plausible and well presented. No analytical confirmation of test item concentration, however test item assumed to be stable.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes

Remarks:

Principles of Good Laboratory Practice (GLP), German Federal Register, Vol. 43, No. 247, December 22, 1978

Analytical monitoring:

no

Details on sampling:

Not applicable

Vehicle:

yes

Details on test solutions:

The test substance-dispersion is miscible with water in every ratio. Starting with a stock solution in dilution water (10% stock solution), the amounts corresponding to the respective test concentrations had been pipetted into the test basins and mixed with a glass bar.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Species : zebrafish, Danio rerio (HAMILTON-BUCHANAN)
Origin : West Aquarium, Bad Lauterberg
Date of hatching: 1981-10-09
Date of receipt: 1982-01-20

Until day 7 before start of the experiment, the fish were kept in dechlorinated tap water and further on until transfer into the test basins in dilution water:
Temperature : 23 ± 1 °C
Oxygen content : >= 80 % of the saturation value
Duration of light period : 12 hours daily
Stocking rate: <= 1 g fish/L water
Feeding : twice daily ad libitum
Food : Tetra Min, Tetra Werke, Meile (Germany)

Immediately before start of the experiment the body length of 10 fish representative for the used fish batch were measured. These animals were not used for the experiment:
Batch No.: 11/82 (1982-02-15); n=10; Body length (Variation range): 2.5 to 3.2 cm; mean: 2.8 cm; s: ± 0.24

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

Not applicable

Hardness:

The total hardness of the dilution water: 2.5 mmol Ca^2+ + Mg^2+/L during the study.
The molar ration of Ca++ to Mg++ was 4:1, the molar ratio of Na+ to K+ was 10:1.

Test temperature:

Temperature (°C)
Concentration groups and control group: 22.0 - 22.9

pH:

Concentration groups: 7.5 - 7.9
Control group: 7.6 - 7.8

Dissolved oxygen:

Oxygen content (mg/L):
Concentration groups: 6.2 to 8.6
Control group: 6.4 to 8.6

Salinity:

Water quality was according to OECD 203

Nominal and measured concentrations:

5 test groups with 10 fishes, each, had been evaluated:
Nominal test item concentration (42.8% pigment) [mg/L]
0 / 1 / 10 / 100 / 500

Details on test conditions:

The study was conducted in a static system. The test chambers, which were calibrated to 10 liters, were made of glass (length 29 cm, width 21 cm, height 23 cm) and stood in a water bath made of Hostalit Z® with a Plexiglas viewing panel. The temperature of the water bath was regulated by a thermostat to 22 ± 1 °C. The chambers were illuminated from above from 06.00 a.m. to 06.00 p.m. The light intensity directly over the chambers was approximately 700 lux.
The test chambers were not aerated during the course of the study.
The test substance-dispersion is miscible with water in every ratio. Starting with a stock solution in dilution water, the amounts corresponding to the respective test concentrations had been pipetted into the test basins and mixed with a glass bar.

Study groups
There were 10 fish in each group. Concentration of test item (42.8% pigment) [mg/L]
0 / 1 / 10 / 100 / 500

Test procedure
After the test concentration had been prepared and water parameters recorded, 10 fish were assigned to each test and control chamber stochastically. The fish were not fed during the entire study period. Inspection of the fish took place after 2-4, 24, 48, 72 and 96 hours and involved recording the mortality and visible alterations in appearance and behavior. The water parameters were measured and recorded at the same time points.

At the end of the study, surviving fish were killed with 0.05% MS 222 (3-Aminobenzoic acid ethyl ester, methane sulfonate, Sandoz).

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

LC0

Effect conc.:

500 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: corresponding to 214 mg/L in relation to the submission substance (42.8%)

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

500 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: corresponding to 214 mg/L in relation to the submission substance (42.8%)

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 500 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: corresponding to 214 mg/L in relation to the submission substance (42.8%)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 500 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: corresponding to 214 mg/L in relation to the submission substance (42.8%)

Details on results:

Observations
With the exposure groups 10, 100 and 500 mg/L , the heavy coloration of the water by the test substance compromized the observation of symptoms. However, fish of all test groups showed no difference to the control in terms of behaviour.
Fish from exposure groups 100 and 500 mg/L showed a light red coloring at the end of the test.

Mortality
In this 96-hour acute toxicity study of the submission substance in zebrafish (Danio rerio) mortality occurred neither in the control group nor in the treatment groups.

Results with reference substance (positive control):

not applicable

Reported statistics and error estimates:

not applicable

Validity criteria fulfilled:

yes

Conclusions:

The test was performed with GLP compliance and similar to OECD-guideline 203. Using a dispersion of the submission substance as test item (42.8% of submission substance), high concentrations far above water solubility could be tested avoiding formation of any precipitate.
Under the conditions of this test the LC50 of the test item after 96 hours was > 500 mg/L (214 mg/L submission substance) and the LC0 equal to 500 mg/L (214 mg/L submission substance), based on nominal concentrations, each.
As the test item contains a relevant amount of the submission substance, the test results are considered adequate to fulfill the endpoint requirements.

Executive summary:

The test item (42.8% submission substance) was tested in zebrafish (Danio rerio) for 96 hours in a static system. The test item is a dispersion of the submission substance and readily miscible with water. Such, concentrations of the submission substance far above the level of water solubility could be tested avoiding formation of any precipitate. The nominal concentrations tested were 0 / 1 / 10 / 100 / 500 mg/L with 10 fish for each concentration group. Concentration groups 10, 100 and 500 mg/L showed heavy coloration of the water. Analytical monitoring of the concentration of the test substance in the test water was not conducted as the test item is stable in water. In this 96-hour acute toxicity study, no effects in comparison to the control could be observed at any concentration of the test item (1, 10, 100 and 500 mg/L). Neither in the control nor in the concentration groups any mortalities or behavioural abnormalities occurred. Under the conditions of this test the LC50 of the test item after 96 hours was > 500 mg/L and the LC0(96h) = 500 mg/L nominal concentration (corresponding to 214 mg/L submission substance). As the test item contains a relevant amount submission substance, the test results are considered adequate to fulfill the endpoint requirements.

Description of key information

The test was performed with GLP compliance and similar to OECD-guideline 203. Using a dispersion of the submission substance as test item (42.8% of submission substance), high concentrations far above water solubility could be tested avoiding formation of any precipitate.
Under the conditions of this test the LC50 of the test item after 96 hours was > 500 mg/L (214 mg/L submission substance) and the LC0 equal to 500 mg/L (214 mg/L submission substance), based on nominal concentrations, each.
As the test item contains a relevant amount of the submission substance, the test results are considered adequate to fulfill the endpoint requirements.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

> 214 mg/L