2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3-yl]benzoxazole-5-sulphonamide

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:

30 December 2016 to 06 March 2017

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:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult items and mixtures, OECD series on testing and assessment number 23

Version / remarks:

2000

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Samples for possible analysis were taken from the test concentration and the control. In addition, the filter used to prepare the saturated solution (SS) was retained for possible analysis of the residue.
- Sampling method: At t = 0, t = 24 and t = 96 h, 3.0 mL was sampled.
- Sample storage conditions before analysis: Samples were stored in a freezer (≤ -15 °C) until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Preparation of test solutions started with a loading rate of 100 mg/L applying three days of magnetic stirring to reach the maximum dissolution of the test material in medium. The obtained aqueous mixture was filtered through a 0.45 μm membrane filter (RC55, Whatman) and the resulting saturated solution (SS) was used as the highest test concentration. Lower test concentrations for the combined limit/range-finding test were prepared by subsequent dilutions of the SS in test medium. All test solutions were clear and colourless at the end of the preparation procedure.

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM
- Common name: Carp
- Strain: Carp (Cyprinus carpio, Teleostei, Cyprinidae) Linnaeus, 1758
- Age at study initiation: Not specified
- Length at study initiation: 3.6 ± 0.2 cm
- Weight at study initiation: 0.58 ± 0.12 g
- Method of breeding: F1 from a single parent-pair bred in UV-treated water

ACCLIMATION
- Acclimation period: At least 12 days after delivery
- Acclimation conditions (same as test or not): Yes
- Type of food during acclimation: Pelleted fish food
- Feeding frequency during acclimation: Daily
- Health during acclimation (any mortality observed): In the batch of fish used for the test, mortality during the seven days prior to the start of the test was less than 5 %.

FEEDING DURING TEST: No

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

180 mg CaCO3/L

Test temperature:

21 to 22 °C

pH:

7.2 to 8.1

Dissolved oxygen:

5.6 to 9.8 mg O2/L

Nominal and measured concentrations:

- Nominal concentration: 100 % of a SS prepared at 100 mg/L
- Measured concentration: 0.018 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 10 L
- Material, size, headspace, fill volume: All-glass vessels containing 9 L of solution
- Aeration: The test media were not aerated during the test
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.45 g fish/L, i.e. 7 fish per 9 litres of test medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Adjusted ISO medium, formulated using RO-water (tap-water purified by reverse osmosis; GEON Waterbehandeling, Berkel-Enschot, The Netherlands) with the following composition: CaCl2.2H2O 211.5 mg/L, MgSO4.7H2O 88.8 mg/L, NaHCO3 46.7 mg/L and KCl 4.2 mg/L
- Culture medium different from test medium: No
- Intervals of water quality measurement: Dissolved oxygen content, pH and temperature were measured daily in all vessels with surviving fish, beginning at the start of the test (day 0).

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hour photoperiod daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
Mortality and other effects were observed at 2.75, 24, 48, 72 and 96 hours following the start of exposure. In addition, fish were observed every afternoon from day 0 to check for any dead or severely distressed fish. Dead fish were removed when observed. At the end of the test the surviving fish were rapidly killed by exposing them to ca. 1.2 % ethylene glycol monophenylether in water.

TEST CONCENTRATIONS
- Range finding study: Yes
- Test concentrations: 1.0, 10 and 100 % of a SS prepared at 100 mg/L
- Results used to determine the conditions for the definitive study: Yes. No mortality or clinical effects were observed at any of the test concentrations during the test period. Therefore, the expected LC50 was above a concentration obtained in 100 % of a SS prepared at 100 mg/L.

Reference substance (positive control):

yes

Remarks:

pentachlorophenol

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 0.018 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

No mortality or clinical effects were observed at the limit concentration and in the control during the test period. The responses recorded in this test allowed for reliable determination of an LC50.
Samples taken from 100 % of the SS prepared at 100 mg/L were analysed. The measured concentration was just above LOD, i.e. 0.021 mg/L at the start of the test. During the exposure period the measured concentration decreased slightly to 73 % of initial at the end of the test. Given this results, the LC50 values were calculated using the TWA concentration, i.e. 0.018 mg/L.

ACCEPTABILITY OF THE TEST
1. No mortality was observed in the control at the end of the test.
2. Test conditions were maintained in a constant manner throughout the test.
3. The dissolved oxygen concentration was at least 60 % of the air saturation value throughout the test (>5 mg/L at 22 °C).
4. Since the measured concentrations deviated by more than 20 % from initial, results were based on the time weighted average exposure concentration.

Results with reference substance (positive control):

The objective was to evaluate Pentachlorophenol (PCP) for its ability to generate acute toxic effects in Cyprinus carpio during an exposure period of 96 hours and to determine the LC50 at all observation times. The reference test was carried out to check the sensitivity of the test system as used by the testing facility. Concentrations tested were 0.10, 0.22 and 0.46 mg/L in ISO-medium.
All carp exposed to a PCP concentration of 0.46 mg/L died within 24 hours, while no mortality occurred at 0.10 and 0.22 mg/L. The 96 h-LC50 for carp exposed to PCP was 0.32 mg/L (95 % confidence interval between 0.22 and 0.46 mg/L). This effect was already reached within 24 hours of exposure. The range of the 96 h-LC50 for carp is generally between 0.10 and 0.46 mg/L based on historical data. Hence, the sensitivity of carp originating from the present batch for PCP falls within the range of sensitivities generally observed.

Reported statistics and error estimates:

No LC50 values could be determined because the observed effects were below 50 %.

Validity criteria fulfilled:

yes

Conclusions:

Due to the low solubility of the test material in water, concentration levels that might be toxic for carp could not be reached. Under the conditions of this study, the 96 h-LC50 was >0.018 mg/L based on the TWA concentration.

Executive summary:

The potential for the test material to cause acute toxicity to fish was investigated in accordance with the standardised guidelines OECD 203, EU Method C.1 and the OECD series on testing and assessment number 23 under GLP conditions.

A limit test was performed under static conditions based on the results of a preceding range-finding test. The test material was not completely soluble in test medium at the loading rate initially prepared. A saturated solution (SS) was prepared at a loading rate of 100 mg/L and used as the test concentration. Seven fish per concentration were exposed to a control and to 100 % of the SS prepared at 100 mg/L. The total exposure period was 96 hours and samples for analytical confirmation of exposure concentrations were taken at the start, after 24 hours of exposure and at the end of the test.

No mortality or clinical effects were observed at the limit concentration and in the control during the test period. The responses recorded in this test allowed for reliable determination of an LC50.

Samples taken from 100 % of the SS prepared at 100 mg/L were analysed. The measured concentration was 0.021 mg/L at the start of the test. During the exposure period the measured concentration decreased slightly to 73 % of initial at the end of the test. Given this result, the LC50 was calculated using the TWA concentration, i.e. 0.018 mg/L.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

Due to the low solubility of the test material in water, concentration levels that might be toxic for carp could not be reached. Under the conditions of this study, the 96 h-LC50 was >0.018 mg/L based on the TWA concentration.

Description of key information

Under the conditions of this study, the 96 h-LC50 was >0.018 mg/L based on the TWA concentration.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

0.018 mg/L

Additional information

The potential for the test material to cause acute toxicity to fish was investigated in accordance with the standardised guidelines OECD 203, EU Method C.1 and the OECD series on testing and assessment number 23 under GLP conditions.

A limit test was performed under static conditions based on the results of a preceding range-finding test. The test material was not completely soluble in test medium at the loading rate initially prepared. A saturated solution (SS) was prepared at a loading rate of 100 mg/L and used as the test concentration. Seven fish per concentration were exposed to a control and to 100 % of the SS prepared at 100 mg/L. The total exposure period was 96 hours and samples for analytical confirmation of exposure concentrations were taken at the start, after 24 hours of exposure and at the end of the test.

No mortality or clinical effects were observed at the limit concentration and in the control during the test period. The responses recorded in this test allowed for reliable determination of an LC50.

Samples taken from 100 % of the SS prepared at 100 mg/L were analysed. The measured concentration was 0.021 mg/L at the start of the test. During the exposure period the measured concentration decreased slightly to 73 % of initial at the end of the test. Given this result, the LC50 was calculated using the TWA concentration, i.e. 0.018 mg/L.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

Due to the low solubility of the test material in water, concentration levels that might be toxic for carp could not be reached. Under the conditions of this study, the 96 h-LC50 was >0.018 mg/L based on the TWA concentration.