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Ecotoxicological information

Short-term toxicity to fish

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Administrative data

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:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report date:
2013

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 850.1075 (Freshwater and Saltwater Fish Acute Toxicity Test)
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Details on test material:
- Purity: 100%

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Samples were collected from each test chamber at the beginning of the test and at 48 and 96 hours (± 1 hour) to measure concentrations of the test substance. Two sets of samples were collected at each interval. All samples were collected at mid-depth from the test chambers, placed in glass scintillation vials, and acidified with 2 drops of 10% phosphoric acid. One set of samples was processed immediately for analysis while the second set was stored refrigerated for possible future analysis.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION:
Individual test solutions were prepared directly in each test chamber. A primary stock solution was prepared by mixing a calculated amount of test substance in 30 L of dilution water at a nominal concentration of 120 mg a.s./L, the highest concentration tested. The primary stock was prepared by mixing 3.6003 grams of test substance with a portion of the pre-measured dilution water in a 2 L glass flask, and sonicating the flask for approximately 5 minutes. The flask was poured into the appropriate aquaria and stirred with the remainder of the dilution water with a top-down electric mixer for approximately 1 hour. The stock solution was adjusted to 100% active ingredient during preparation based on the test substance purity (100%). At the end of mixing, the primary stock solution was clear and colourless with white foam on the surface. Aliquots of the primary stock solution were proportionally diluted with dilution water to prepare four additional test solutions at nominal concentrations of 7.5, 15, 30 and 60 mg a.s./L. The solutions were stirred with a top-down electric mixer for approximately 40 minutes. The 7.5, 15 and 30 mg a.s./L test solutions were clear and colourless with no visible precipitate. The 60 and 120 mg a.s./L test solutions appeared clear and colourless with a glossy surface. The negative control solution was dilution water only.

Test organisms

Test organisms (species):
Pimephales promelas
Details on test organisms:
TEST ORGANISM
- Age at study initiation: juveniles, The average total length of seven fish measured at the end of the test was 3.2 cm, with a range of 3.0 to 3.5 cm.
- Feeding during test: The fish were not fed for at least two days prior to the test or during the test.

ACCLIMATION PERIOD: at least 14 days

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h

Test conditions

Hardness:
Hardness: 144 (mg/L as CaCO3)
Test temperature:
21.9 - 22.3°C
pH:
8.1 - 8.5
Dissolved oxygen:
6.7 - 8.7 mg/L
Salinity:
freshwater
Nominal and measured concentrations:
Nominal concentrations: 0, 7.5, 15, 30, 60 and 120 mg a.s./L
Measured concentrations: 0, 8.9, 19, 35, 72 and 131 mg a.s./L
Details on test conditions:
TEST SYSTEM
- Test vessel: 25-L stainless steel aquaria containing 15 L of test solution
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
Dissolved oxygen and pH were measured in each test chamber at the beginning and end of the test and at approximately 24 hours during the test. The test solutions were not aerated during the exposure period. When 100% mortality occurred in a test chamber, measurements of temperature, dissolved oxygen and pH were taken in that test chamber and then discontinued. Hardness, alkalinity and specific conductance were measured in the dilution water at the beginning of the test.
EFFECT PARAMETERS MEASURED: Observations were made periodically to determine the numbers of dead organisms in each treatment group. Lethality was defined as a lack of visible movement by the test organism. The numbers of individuals exhibiting sublethal effects or abnormal behaviour also were evaluated. Observations were made approximately 4.5, 24, 48, 72, and 96 hours after test initiation.

Results and discussion

Effect concentrations
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
50 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
Fathead minnows in the negative control group appeared normal throughout the test, with no observed mortality or sublethal effects. All fathead minnows in the 8.9, 19 and 35 mg a.s./L treatment groups also appeared normal throughout the test, with no mortality or overt signs of toxicity. Percent mortality at test termination in the 72 and 131 mg a.s./L treatment groups was 100%. The highest mean, measured test concentration causing no mortality at test end was 35 mg a.s./L. The lowest mean, measured test concentration causing 100% mortality at test end was 72 mg a.s./L.
Reported statistics and error estimates:
The mortality data were analysed using the computer program of C. E. Stephan. The program was designed to calculate the LC50 value and the 95% confidence interval by probit analysis, the moving average method, and binomial probability with nonlinear interpolation. In this study, nonlinear interpolation was used to calculate the 24, 48, 72 and 96-hour LC50 values and binominal probability was used to calculate the 95% confidence intervals. Due to the method used to calculate the 96-hour LC50 value, the slope of the dose response curve could not be calculated. The highest mean, measured test concentration causing no mortality at test end and the lowest mean, measured test concentration causing 100% mortality at test end were assessed by visual observation of the mortality and observation data.

Any other information on results incl. tables

Sublethal observations / clinical signs:

Mean, Measured

Concentration (mg a.s./L)

 

Cumulative mortality (No. dead/No. at test start)

 

~4.5 hours

24 hours

48 hours

72 hours

96 hours

Water Control (0.0)

0/7

0/7

0/7

0/7

0/7

8.9

0/7

0/7

0/7

0/7

0/7

19

0/7

0/7

0/7

0/7

0/7

35

0/7

0/7

0/7

0/7

0/7

72

0/7a

4/7b

6/7c

7/7

7/7

131

0/7a

7/7

7/7

7/7

7/7

a7/7 fish lying at the bottom of the test chamber

b3/7 fish lying at the bottom of the test chamber

c1/7 fish lying at the bottom of the test chamber

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
96-h LC50 = 50 mg a.s./L
Executive summary:

The acute toxicity of the test substance to unfed juvenile fathead minnows, Pimephales promelas, a freshwater fish, was determined in an unaerated, static, 96-hour dose response test. The test was conducted in accordance with the OECD Guideline for Testing of Chemicals 203, and U.S. Environmental Protection Agency Series 850 - Ecological Effects Test Guidelines (draft) OPPTS Number 850.1075, Fish Acute Toxicity. Treatments consisted of a dilution water control and five nominal test concentrations of 7.5, 15, 30, 60 and 120 mg a.s./L (equivalent to 8.9, 19, 35, 72 and 131 mg a.s./L based on mean, measured concentrations). The 96-hour LC50 for Pimephales promelas was 50 mg a.s./L based on mean, measured test concentrations and mortality data. The highest mean, measured test concentration causing no mortality at test end was 35 mg a.s./L. The lowest mean, measured test concentration causing 100% mortality at test end was 72 mg a.s./L.