C.I. Reactive Yellow 2

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:

27 February 2020 - 2 March 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

17 July 1992

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: The test material was stored in a dark storage place at room temperature.
- Stability under storage conditions: stable.
- Solubility in dimethylsulfoxide: ≤ 439 g/L
- Solubility in acetone: ≤ 0.03 g/L
- Water solubility: ≤ 310 g/L

Analytical monitoring:

yes

Details on sampling:

- Frequency of measurement: at the start and end of exposure.
- Sample for measurement: the test solution was taken out from the middle layer of the test vessel.
- Volume of sample: approximately 10 mL (all test levels).
- Analytical condition: Calibration curve, LC-MS.

Vehicle:

no

Details on test solutions:

PRETREATMENT OF TEST SOLUTION TO DETERMINE TEST MATERIAL CONCENTRATION
- The test material (69.1 mg) was dissolved and filled up to 50 mL with ultrapure water to obtain 1000 mg/L solution of the test material. This was diluted with dechlorinated tap water to prepare a 10.0 mg/L standard solution.

PREPARATION OF TEST SOLUTION FOR DEFINITIVE STUDY
- The test material (0.387 g) was weighed and dissolved in dilution water (2.8 L) to prepare a 100 mg/L test solution.

Test organisms (species):

Oryzias latipes

Details on test organisms:

TEST ORGANISM
- Common name: Medaka.
- Source: Test facility
- Age at study initiation: 2 months.
- Size: total length = 2.0 ± 1.0 cm

ACCLIMATION
- Acclimation period: 28 days.
- Water: dechlorinated tap water.
- Acclimation type: flow-through regime.
- Dissolved oxygen concentration: at least 80 % of air saturation value.
- Temperature: 24 ± 1 °C
- Lighting condition: room light, 16-hour light/8-hour dark.
- Feed: Newly hatched Artemia nauplii.
- Feeding amount and frequency: saturation volume every day.
- Mortality of test group: 0 % (during 7 days before the start of exposure).
- Feed withdrawal: 24 hours before the start of exposure.

FEEDING DURING TEST
- No feeding.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

40 CaCO3 mg/L

Test temperature:

23.1 - 23.7 °C

pH:

7.7 - 7.9

Dissolved oxygen:

8.5 - 9.0 mg/L (at least 60% of saturate concentration at the water temperature)

Nominal and measured concentrations:

Nominal: 100 mg/L
Measured: At the start = 101 mg/L, at the end = 107 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 3 L glass tank (diameter: 16 cm, depth: 17 cm).
- Cover on test vessel: transparent plastic lid.
- Water bath: plastic tank, warming/cooling unit: RX-401HPN-Z (IWAKI).
- Aeration: conduct gently.
- No. of organisms per vessel: 7 fish/test level
- Replicates: 1 replicate/test level
- Volume of test solution: approximately 2.8 L/test level

DILUTION WATER
- Dechlorinated tap water, aerated.
- It was confirmed that the concentration of residual chlorine was < 0.02 mg/L at use.

OTHER TEST CONDITIONS
- Adjustment of pH: no adjustment.
- Photoperiod: 16-hour light/8-hour dark.
- Light intensity: room light.

EFFECT PARAMETERS MEASURED
- Mortality and visible abnormality was observed at 3, 24, 48, 72 and 96 hours after the start of exposure. A fish was considered dead when the observable motion (motion of mouth and opercula etc) was not observed and touching of the caudal peduncle with a glass rod caused no reaction.
- Total length and body of the test organism were compared to test organisms in the control group.
- Appearance of test solution was observed at the start and end of exposure.

CONDITION OF TEST SOLUTION
- Item of measurement: dissolved oxygen concentration, pH and temperature.
- Frequency of measurement: at the start of exposure and every 24 hours.
- Sample for measurement: the test solution was taken out from the test vessel.
- Instrument: dissolved oxygen meter HQ30d (HACH), pH meter HM-21P (DKK-TOA) and glass thermometer.

TEST CONCENTRATIONS
- Range finding study: yes (4 fish were exposed to a nominal concentration of test material of 100 mg/L and mortality assessed after 3, 24, 48, 72, and 96 hours)
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

copper(II) sulphate

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)

Remarks on result:

other: The results are presented in terms of nominal concentrations since the measured concentrations of the test material in the test solutions were maintained within the range of 80-120% of the nominal during exposure.

Details on results:

BIOLOGICAL RESULTS
- Number of dead fish in the exposure level during exposure was not confirmed. Number of dead fish in the control at the end of exposure was 0, which met the criterion for the validity of the test.
- No abnormal response was observed during exposure or in the control.
- Size of test organism: total length = 2.0 ± 0.1 cm, bodyweight: 0.076 ± 0.011 g

TEST SOLUTIONS
- The test solution in exposure level was yellow and clear at the start and end of exposure.
- The test solution in the control was colourless and clear at the start and end of exposure.
- The measured values of dissolved oxygen concentration, pH and temperature during exposure were 8.5 - 9.0 mg/L, pH 7.7-7.9 and 23.1-23.7 °C, respectively.
- The measured values of dissolved oxygen concentration met the criterion for the validity of the test (at least 60% of saturate concentration at the water temperature).
- The measured concentrations of the test material in the test solutions at the start of exposure was 101 mg/L (101% of the nominal concentrations), and those at the end of exposure was 107 mg/L (107% of the nominal concentrations). The measured concentrations of test material were maintained within the range of 80 - 120 % of the nominal concentrations.
- The LC50 was measured every 24 hours ( > 100 mg/L).

Results with reference substance (positive control):

- Reference substance: copper (II) sulphate pentahydrate.
- 96-hour LC50: 0.37 mg/L (this value was in the stipulated range (mean ± 2SD of 0.13 - 0.89 mg/L, n = 67) to background data in the Test Facility. (Values converted into copper(II) sulphate).

Sublethal observations / clinical signs:

LC50 to Medaka

Exposure duration (hour)	LC50 (mg/L)
24	> 100
48	> 100
72	> 100
96	> 100

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the study the 96–hour LC50 to Medaka was > 100 mg/L.

Executive summary:

The acute toxicity of the test material on Medaka was investigated in accordance with the standardised guideline OECD 203, and under GLP conditions.

This study was conducted as a limit test in order to confirm the effect of the test material on the test organisms, Medaka, at the upper limit concentration of the test method. The test material (0.387 g) was weighed and dissolved in dilution water (2.8 L, dechlorinated tap water) to prepare the 100 mg/L of test solution. Mortality and visible abnormality were observed at 24 hour intervals after the start of the exposure. A fish was considered as dead when the observable motion was not observed and touching of the caudal peduncle with a glass rod cased no reaction.

Under the conditions of the study, no adverse effect on the test organisms was found in the definitive study. The test material concentration was maintained within 80–120 % of the nominal concentration. The environmental conditions were within the suitable range. Therefore, it was concluded that this study complied with the applied test guidelines.

The 96–hour LC50 to Medaka was > 100 mg/L.

Description of key information

Under the conditions of the study the 96-hour LC50 to Medaka was > 100 mg/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

100 mg/L

Additional information

The acute toxicity of the test material on Medaka was investigated in accordance with the standardised guideline OECD 203, and under GLP conditions. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

This study was conducted as a limit test in order to confirm the effect of the test material on the test organisms, Medaka, at the upper limit concentration of the test method. The test material (0.387 g) was weighed and dissolved in dilution water (2.8 L, dechlorinated tap water) to prepare the 100 mg/L of test solution. Mortality and visible abnormality were observed at 24 hour intervals after the start of the exposure. A fish was considered as dead when the observable motion was not observed and touching of the caudal peduncle with a glass rod cased no reaction.

Under the conditions of the study, no adverse effect on the test organisms was found in the definitive study. The test material concentration was maintained within 80–120 % of the nominal concentration. The environmental conditions were within the suitable range. Therefore, it was concluded that this study complied with the applied test guidelines.

The 96–hour LC50 to Medaka was > 100 mg/L.