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Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 December 2017 to ****
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
CAS Number: 15337-18-5
Purity: 96.6%
Physical state/Appearance: Yellow viscous liquid
Analytical monitoring:
yes
Details on sampling:
Water samples were collected from the control and the 100% v/v saturated solution test group at 0, 24, 48 and 72 hours from fresh media and at 24, 48, 72 and 96 hours from old media for quantitative analysis. Duplicate samples at 0 and 72 hours (fresh media) and 24 and 96 hours (old media) were taken and stored frozen for further analysis if necessary.

Preparation of Test Samples
The test samples were thawed with the aid of sonication before being filtered through 0.45 µm cellulose acetate filters.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
Definitive Test
In accordance with the recommendations of REACH, the test was conducted according to the threshold approach recommended by ECHA. As the EC50 values obtained for both the Algal Growth Inhibition and the Acute Toxicity to Daphnia magna tests (Envigo study numbers VP09MY and TL23VK, respectively) were greater than 100% v/v saturated solution, this test was conducted at a single concentration of 100% v/v saturated solution to assess fish toxicity at this concentration.

Test Solution Preparation
A nominal amount of test item (550 mg) was dispersed in 11 liters of test water with the aid of propeller stirring at approximately 1500 rpm for 48 hours. After 48 hours the stirring was stopped and any undissolved test item was removed by filtration through a 0.2 µm Sartorius Sartopore filter (first approximate 2 liters discarded in order to pre condition the filter) to give a 100% v/v saturated solution. The test solution was mixed with a flat bladed stirrer for approximately 1 minute to ensure adequate mixing and homogeneity. The concentration and stability of the test item in the test preparations were verified by chemical analysis of the fresh media at 0 and 72 hours and in the old media at 24 and 96 hours.
Test organisms (species):
Pimephales promelas
Details on test organisms:
Test System and Supporting Information
The test was carried out using juvenile fathead minnow (Pimephales promelas). Fish were bred in house and maintained since 15 January 2018.

There was no mortality in the 7 days prior to the start of the test and the fish had a mean standard length of 2.2 cm (standard deviation = 0.23) and a mean body weight of 0.10 g (standard deviation = 0.032) at the end of the definitive test. Based on the mean body weight value this gave a loading rate of 0.16 g body weight/liter. The stock fish were fed commercial GM 300 which was discontinued approximately 26 hours prior to the start of the definitive test. The diet and diluent water are considered not to contain any contaminant that would affect the integrity and outcome of the study.

The test water used for the definitive test was the same as that used to maintain the stock fish.
Test type:
semi-static
Water media type:
other: Laboratory tap water
Limit test:
yes
Total exposure duration:
96 h
Hardness:
140 mg/L as CaCO3
Test temperature:
19 to 21 ºC
pH:
7.8 to 9.3
Dissolved oxygen:
8.4 to 11.2 mg O2/L
Nominal and measured concentrations:
Nominal: 100% v/v Saturated Solution
Measured: Less than the LOQ of the analytical method employed, which was determined to be 0.010 mg/l as zinc (equivalent to 0.093 mg/L as test item).
Details on test conditions:
Experimental Preparation
The concentration of the test item in the test preparations were verified by chemical analysis at 0 and 48 hours (fresh media) and at 24 and 72 hours (old media).

Exposure Conditions
In the definitive test, a 5 liter glass exposure vessels containing 4.5 liters of test media were used for each control and test concentration. At the start of the test, seven fish were placed in each test vessel at random. The test vessels were then covered to reduce evaporation and maintained at approximately 19 C to 21 ºC in a temperature controlled room with a photoperiod of 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods for a period of 96 hours. The test vessels were aerated via narrow bore glass tubes. The fish were not individually identified and received no food during exposure.
The control group was maintained concurrently under identical conditions, but were not exposed to the test item.
A semi static test regime was employed in the test involving a daily renewal of the test preparations to aid in maintaining exposure levels and prevent the build up of nitrogenous waste products.
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 100 other: v/v saturated solution
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
100 other: v/v saturated solution
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
Measured concentrations in the definitive test were less than the limit of quantification (LOQ) of the analytical method employed were obtained, which was determined to be 0.010 mg/l as zinc (equivalent to 0.093 mg/L as test item). Given that the toxicity in this study cannot be attributed to a single elemental component, but to the dissolved fraction of the total test item and free zinc, which was below the quantifiable limit of the analytical method, the results were based on nominal concentrations as % v/v saturated solution. No toxic effects were observed at the limit of saturation in the test medium.
Validity criteria fulfilled:
yes
Conclusions:
Exposure of fathead minnow (Pimephales promelas) to the test item resulted in an LC50 value of greater than 100% v/v saturated solution, based on the nominal test concentration. The No Observed Effect Concentration (NOEC) was 100% v/v saturated solution.
Executive summary:

A study was performed to assess the acute toxicity of the test item to the freshwater fish, fathead minnow (Pimephales promelas). The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No 203, "Fish, Acute Toxicity Test" and Method C.1, “Acute Toxicity for Fish”, of Commission Regulation (EC) No. 440/2008.

 

Preliminary solubility work conducted indicated that it was not possible to obtain a testable solution of the test item using traditional methods of preparation. A preliminary media preparation trial indicated that a saturated solution method of preparation was the most appropriate for this test item. In accordance with the recommendations of REACH, the test was conducted according to the threshold approach recommended by ECHA.  As the EC50 values obtained for both the Algal Growth Inhibition and Acute Toxicity to Daphnia magna tests (Envigo study numbers VP09MY and TL23VK, respectively) were greater than 100% v/v saturated solution, the test was conducted at a single concentration of 100% v/v saturated solution to ensure that toxicity was not observed at this concentration. 

 

Seven fish were exposed to an aqueous solution of the test item, at a single concentration of 100% v/v saturated solution for a period of 96 hours at a temperature of 19 to 21 ºC under semi static test conditions.  The number of mortalities and any sub lethal effects of exposure in each test and control vessel were recorded 1, 3 and 6 hours after the start of exposure and then daily throughout the test until termination at 96 hours.

 

Test solutions were collected at 0 and 72 hours (fresh media) and 24 and 96 hours (old media) and were analyzed by inductively coupled plasma with mass spectrometric detection (ICP-MS) to confirm test item concentrations based on the quantitation of elemental zinc. Measured concentrations in the definitive test were less than the limit of quantification (LOQ) of the analytical method employed were obtained, which was determined to be 0.010 mg/l as zinc (equivalent to 0.093 mg/L as test item). Given that the toxicity in this study cannot be attributed to a single elemental component, but to the dissolved fraction of the total test item and free zinc, which was below the quantifiable limit of the analytical method, the results were based on nominal concentrations as % v/v saturated solution. Exposure of fathead minnow (Pimephales promelas) to the test item resulted in an LC50 value of greater than 100% v/v saturated solution, based on the nominal test concentration. The No Observed Effect Concentration (NOEC) was 100% v/v saturated solution. No toxic effects were observed at the limit of saturation in the test medium.

Description of key information

Exposure of fathead minnow (Pimephales promelas) to the test item resulted in an LC50 value of greater than 100% v/v saturated solution, based on the nominal test concentration.  The No Observed Effect Concentration (NOEC) was 100% v/v saturated solution.

Key value for chemical safety assessment

Additional information