[No public or meaningful name is available]

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:

18 April 2007 to 31 July 2007

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)

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 2.2, 4.6, 10, 22, 46 and 100 mg/L (based on loading rate)
- Sampling method: Just before test start (Day 0) and on the last day of preparation (Day 3) duplicate samples from each test medium and duplicate samples from the control were taken. At the end of the first test medium renewal period (Day 1) and at the end of the last renewal period (Day 4) (stability samples) samples from each test medium and duplicate samples from the control were taken.
However, the samples from the highest loading rate of 100 mg/L were only taken at the start and the end of the first test medium renewal period since all test fish were dead at this test concentration after 24 hours
- Sample storage: The samples were analysed directly after the sampling procedure.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method:
In order to assess the toxicity of the test item to the fish, water accommodated fractions (WAFs) with loading rates of 2.2, 4.6, 10, 22, 46 and 100 mg/L were tested. A control was tested in parallel. The selection of the loading rates for the test was based on the results of a range-finding test (non-GLP).

Prior to the start of the test, individual dispersions with loading rates of 2.2, 4.6, 10, 22, 46 and 100 mg/L were prepared by mixing 11, 23, 50, 110, 230 and 500 mg of the test item, respectively, into 5000 mL of test water. No auxiliary solvent or emulsifier was used. The dispersions were stirred on magnetic stirrers at room temperature in the dark for 3 hours to dissolve a maximal amount of the test item in the test water.

The stirring period of 3 hours was chosen based on the results of a pre-test (non-GLP) in which the maximum concentration of dissolved test item was reached after the stirring period of 3 hours.

After the stirring period of 3 hours, the saturated dispersions filtered through membrane filters just before the start of the test. The filtrates were tested on fish as WAFs.

The test media were prepared just before the fish were transferred to the new test media. The WAFs were characterized by GC analysis and the actual test item concentrations were measured.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
-Common name: Zebra fish
- Source (laboratory, culture collection): The test fish Brachydanio rerio were obtained from laboratories’ in-house breeding programme.

ACCLIMATION
- Acclimation period: The fish were acclimated for one week prior to test start. During holding and acclimatization until one day before the start of the test the fish were fed with commercial fish diet.
- From the acclimated batch of test fish, 10 fish were measured at the start of the test. The mean body length of the fish was 3.1 ± 0.1 cm. The mean body wet weight was 0.61 ± 0.02 g.
- One glass aquarium with 5 liters of test medium was used for each test concentration and the control. At the start of the test 7 fish were introduced into each aquarium in a random area. The loading rate was less than 1 g fish wish wet weight per liter test medium. The test media and control were slightly aerated during the test. The measured temperature through the test was within the range of 21 - 22°C. Each aquarium was illuminated at 50 – 500 Lux with a photoperiod of 16 hours light and 8 hours darkness (with a 30 minute transition period).

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

1.25 mmol/L

Test temperature:

21 – 22°C

pH:

pH 7.3 - 7.4

Nominal and measured concentrations:

Definitive Test: Nominal
2.2, 4.6, 10, 22, 46 and 100 mg/L

Details on test conditions:

Determination of mortality and observed visible abnormalities: Observations were made at 3, 24, 48, 72 and 96 hours. Dead fish were removed at least once daily and discarded.

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

42 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC10

Effect conc.:

22 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:

96 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

22 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

TEST SOLUTIONS
Observations on the test media were carried out during the mixing and testing of the WAFs. No remarkable observations were made concerning the appearance of the test media

TEST ORGANISM
In the control and at the loading rates up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed. At the next highest loading rate of 22 mg/L, all test fish showed symptoms or toxicity after 48 hours. However, all fish survived until the end of the test at this loading rate. At the loading rate of 46 mg/L, all fish showed strong symptoms of toxicity and all four of the seven test fish died by the end of the test. At the highest loading rate of 100 mg/L, all test fish died within 24 hours of test initiation.
Thus, the 96-hour NOEC of the test substance was at the loading rate of 10 mg/L. The 96 hour LOEC and the 96 hour LC10 to Brachydanio rerio was at the loading rate of 22 mg/L.
The 96-hour LC50 of the test substance was calculated to be at the loading rate of 42 mg/L. The 96-hour LC100 was at the loading rate of 100 mg/L.

Validity criteria fulfilled:

yes

Conclusions:

In an OECD 203 study in zebra fish, the 96-hour NOEC (highest concentration tested without observable toxic effects after the exposure period of 96 hours) was the loading rate of 10 mg/L. The 96-hour LOEC (lowest concentration tested with observable toxic effects) and the 96-hour LC0 of test substance to Zebra fish was the loading rate of 22 mg/L.

The 96-hour LC50 of the test substance was calculated to be at the loading rate of 42 mg/L with a 95% confidence interval of 32–57 mg/L. The 96-hour LC100 was the loading rate of 100 mg/L.

Executive summary:

The acute toxicity of the test substance to zebra fish (Brachydanio rerio) was
determined in a 96-hour semi-static test with a daily test medium renewal according to the EU Commission Directive 92/69/EEC, Part C.1 (1992) and the OECD Guideline for Testing of Chemicals No. 203 (1992).

In order to assess the toxicity of the test substance to fish, water accommodated fractions (WAFs) with the loading rates of 2.2, 4.6, 10, 22, 46 and 100 mg/L were tested. Additionally, a control (test water without test item) was tested in parallel. The test method was based on the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures (2000).

For preparation of the WAFs, individual dispersions of the test substance with the loading rates as mentioned above were prepared prior to the start of the test and prior to each test medium renewal. The dispersions were stirred for 3 hours to dissolve a maximum amount of the test item in the dispersion. Thereafter, the dispersions were filtered through membrane filters (0.45 µm) and the undiluted filtrates were tested as WAFs.

At the start of the first and the last test medium renewal periods, the measured test item concentrations (based on a main component of the test item) in the test media were 1.2 and 0.36 mg/L (loading rate 2.2 mg/L), 2.6 and 1.1 mg/L (loading rate 4.6 mg/L), 5.0 and 3.8 mg/L (loading rate 10 mg/L), 8.7 and 6.9 mg/L (loading rate 22 mg/L), 9.9 and 8.1 mg/L (loading rate 46 mg/L) and 26 mg/L (loading rate 100 mg/L, only the first renewal period).

At the end of the test medium renewal periods, the measured concentrations were 0.74 and 0.70 mg/L (loading rate 2.2 mg/L), 2.4 and 1.1 mg/L (loading rate 4.6 mg/L), 5.2 and 1.7 mg/L (loading rate 10 mg/L), 6.9 and 3.8 mg/L (loading rate 22 mg/L), 14 and 12 mg/L (loading rate 46 mg/L) and 32 mg/L (loading rate 100 mg/L, only the first renewal period).

Since water accommodated fractions of the test substance were tested, all reported biological results were based on the loading rates of the test item.

The biological test results (based on the loading rates) were as follows:
– 96-hour LC50: 42 mg/L
(95% confidence interval: 32–57 mg/L)
– 96-hour LC0: 22 mg/L
– 96-hour LC100: 100 mg/L
– 96-hour NOEC: 10 mg/L
– 96-hour LOEC: 22 mg/L

Description of key information

In an OECD 203 study in zebra fish, the 96-hour NOEC (highest concentration tested without observable toxic effects after the exposure period of 96 hours) was the loading rate of 10 mg/L. The 96-hour LOEC (lowest concentration tested with observable toxic effects) and the 96-hour LC0 of test substance to Zebra fish was the loading rate of 22 mg/L.

The 96-hour LC50 of the test substance was calculated to be at the loading rate of 42 mg/L with a 95% confidence interval of 32–57 mg/L. The 96-hour LC100 was the loading rate of 100 mg/L.

 

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

42 mg/L