Butanedioic acid, sulfo-, 4-C12-14 (even numbered)-alkyl esters, disodium salts

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline compliant GPL study with good documentation.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control, 1.0, 3.2, 10, 32 and 100 mg/L
- Sampling method: For the determination of the actual test item concentrations in this semi-static test, duplicate samples were taken from each treatment just before the start of the first and of the last renewal periods (Days 0 and 3). At the end of these renewal periods (Days 1 and 4), additional samples were taken for determination of the stability of the test item during the renewal periods of 24 hours.

- Sample storage conditions before analysis: All samples were taken from the approximate center of each aquarium without mixing the test medium, and were stored deep-frozen (at about -20 °C) immediately after sampling. Based on pre-experiments for investigation of the storage stability, the test item is sufficiently stable in the test water under the storage conditions.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A concentrated stock solution of nominal 100 mg/L was freshly prepared by completely dissolving 600.3 mg of test item in 6 liters of test water or 300 mg of test item (ranged from 300.2 to 301.7 mg) in 3 liters of test water using ultrasonic treatment for 15 minutes and stirring for 15 minutes at room temperature. Adequate volumes of this intensively stirred stock solution were added to the test water in the test vessels and were intensively mixed to prepare the test media with the test concentrations as stated above. The test media were freshly prepared just before introduction of the fish (= start of the exposure) and before each test medium renewal.

- Controls: dilution water only
- Evidence of undissolved material (e.g. precipitate, surface film, etc): No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebra fish
- Strain: not provided
- Source: The fish were obtained from a breeding culture at Harlan Laboratories.
- Age at study initiation (mean and range, SD): not provided
- Length at study initiation (length definition, mean, range and SD): From the acclimated test fish batch, 10 fish were measured at the start of the test. The mean total body length (measured from the mouth to the end of the tail fin) of the fish was 3.0 ± 0.09 cm (Mean ± SD).
- Weight at study initiation (mean and range, SD): the mean body wet weight was 0.20 ± 0.02 g (Mean ± SD).
- Method of breeding: similar to test conditions
- Feeding during test: no

ACCLIMATION
- Acclimation period: Prior to test start, they were acclimated for one week to the test water and temperature.
- Acclimation conditions (same as test or not): similar
- Type and amount of food: During holding and acclimatization until one day before the start of the test, the fish were fed with a commercial fish diet (Tetra Min® Hauptfutter, supplied by TETRA-Werke, 49304 Melle / Germany).
- Feeding frequency: not reported
- Health during acclimation (any mortality observed):During holding and acclimatization, no fish died in the test fish batch and all fish were healthy.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

15 mg/L as CaCO3

Test temperature:

21-22 °C

pH:

7.0 to 7.4

Dissolved oxygen:

The oxygen concentration was at least 8.3 mg/L, corresponding to at least 92% oxygen saturation.

Nominal and measured concentrations:

The following nominal concentrations of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts were tested:1.0, 3.2, 10, 32 and 100 mg test item/L. Additionally, a control (test water without test item) was tested in parallel. In the analyzed biologically relevant test media samples (10 to 100 mg/L) from the start and end of the first and last test medium renewal periods the measured test item concentrations ranged between 100 and 110% of the nominal values. Consequently, the test water renewal ensured constant concentrations of the test item during the test and the reported biological results are related to the nominal test item concentrations.

Details on test conditions:

TEST SYSTEM
- Test vessel: glass vessels filled with 4 L test solution each
- Aeration: not reported, but DO concentration was within the validity range
- Renewal rate of test solution (frequency/flow rate): 24 h renewal interval.
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.34 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water (AAP Medium) prepared according to the test guideline was used for this study.
- Culture medium different from test medium: no
- Intervals of water quality measurement: at every renewal of the freshly prepared and aged solutions

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: A 16 hour light to 8-hour dark photoperiod, with a 30-minute transition period was used
- Light intensity: light intensity during the light period was approximately within the range of 140 to 480 Lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
EC50 96 hours

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Justification for using less concentrations than requested by guideline: NA
- Range finding study
- Test concentrations: not reported
- Results used to determine the conditions for the definitive study: The selection of the test concentrations was based on the results of a range-finding test, pre-experiments to determine the solubility of the test item and the two first main tests. In consultation with the Sponsor the main study had to be repeated three times due to modifications in the test design (change of test media, change of species, semi-static setup).

Reference substance (positive control):

no

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:

LC50

Effect conc.:

18 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% C.I.: 10 - 32 mg test item/L

Details on results:

- Behavioural abnormalities:
- Observations on body length and weight: NA
- Other biological observations: In the control and up to and including the highest test concentration 29.17 mg test item/L (10 mg solid content/L), all fish survived until the end of the test and no visible abnormalities were observed.
- Mortality of control: none
- Other adverse effects control: none
- Abnormal responses: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No remarkable observations were made concerning the appearance of the test media.
- Effect concentrations exceeding solubility of substance in test medium: no

Reported statistics and error estimates:

The LC50 values at different observation times could not be calculated by Probit Analysis or Moving Average Interpolation due to a steep concentration-effect relationship. Instead, the LC50 values were determined as a geometric mean value of the two consecutive test concentrations with 0 and 100% mortality, and the 95% confidence limits for the LC50 as the test concentrations with 0 and 100% mortality. The NOEC, LOEC, LC0 and LC100 were determined directly from the raw data.

Sublethal observations / clinical signs:

In the control and at the test concentrations up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed in the test fish. At the next higher test concentration of 32 mg/L, all test fish showed one or several visible abnormalities. After 48 hours test duration, all seven test fish died at this test concentration. At the highest test concentration of 100 mg/L, all fish were dead already after 24 hours of test duration.

Validity criteria fulfilled:

yes

Executive summary:

In the Klimisch 1 GLP study from Kimmel (2013) the acute toxicity of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts to zebra fish (Danio rerio) was determined in a 96‑hour semi-static test with daily test medium renewal according to the EU Commission Directive 92/69/, Part C.1, the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992). The following nominal concentrations of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts were tested:1.0, 3.2, 10, 32 and 100 mg test item/L. Additionally, a control (test water without test item) was tested in parallel. In the analyzed biologically relevant test media samples (10 to 100 mg/L) from the start and end of the first and last test medium renewal periods the measured test item concentrations ranged between 100 and 110% of the nominal values. Consequently, the test water renewal ensured nearly constant concentration of the test item during the test and the reported biological results were related to the nominal test item concentration. In the control and at the test concentrations up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed in the test fish. At the next higher test concentration of 32 mg/L, all test fish showed one or several visible abnormalities. After 48 hours test duration, all seven test fish died at this test concentration. At the highest test concentration of 100 mg/L, all fish were dead already after 24 hours of test duration. In the control and at the test concentrations up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed in the test fish. At the next higher test concentration of 32 mg/L, all test fish showed one or several visible abnormalities. After 48 hours test duration, all seven test fish died at this test concentration.

The 96‑hour LC₅₀of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts was calculated to be 18 mg/L with a 95% confidence interval of 10-32 mg/L. The 96‑hour LC₁₀₀was 32 mg/L.

 

The results of this study are considered relevant and reliable for the risk assessment.

Description of key information

96 h LC50: 18 mg/L

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

18 mg/L

Additional information

In the Klimisch 1 GLP study from Kimmel (2013) the acute toxicity of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts to zebra fish (Danio rerio) was determined in a 96‑hour semi-static test with daily test medium renewal according to the EU Commission Directive 92/69/, Part C.1, the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992). The following nominal concentrations of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts were tested:1.0, 3.2, 10, 32 and 100 mg test item/L. Additionally, a control (test water without test item) was tested in parallel. In the analyzed biologically relevant test media samples (10 to 100 mg/L) from the start and end of the first and last test medium renewal periods the measured test item concentrations ranged between 100 and 110% of the nominal values. Consequently, the test water renewal ensured nearly constant concentration of the test item during the test and the reported biological results were related to the nominal test item concentration. In the control and at the test concentrations up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed in the test fish. At the next higher test concentration of 32 mg/L, all test fish showed one or several visible abnormalities. After 48 hours test duration, all seven test fish died at this test concentration. At the highest test concentration of 100 mg/L, all fish were dead already after 24 hours of test duration. In the control and at the test concentrations up to and including 10 mg/L, all fish survived until the end of the test and no visible abnormalities were observed in the test fish. At the next higher test concentration of 32 mg/L, all test fish showed one or several visible abnormalities. After 48 hours test duration, all seven test fish died at this test concentration.

The 96‑hour LC₅₀of Butanedioic acid, sulfo-, 4-C12-14-alkyl esters, disodium salts was calculated to be 18 mg/L with a 95% confidence interval of 10-32 mg/L. The 96‑hour LC₁₀₀was 32 mg/L.

 

The results of this study are considered relevant and reliable for the risk assessment.