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Ecotoxicological information

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:
Experimental starting date: 19 March 2012 Experimental completion date: 14 May 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study.
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:
Preliminary test: a sample of each test concentration was taken for chemical analysis at 0 and 24 h in order to determine the stability of the test item under test conditions. All samples were stored at approximately -20 °C prior to analysis. Only concentrations within the range to be used for the definitive test were analysed.
Definitive test: the concentration and stability of the test item in the test preparations were verified by chemical analysis at 0 (fresh media), 24 and 96 h (old media). Water samples were taken from the control and 100 mg/L test vessel at 0 (fresh water), 24 and 96 h (old media) for quantitative analysis. The samples were stored at approximately -20 °C prior to analysis. Duplicate samples, and samples at 24 (fresh media), 48 and 72 h (old media), were taken and stored at approximately -20 °C for further analysis if necessary.

Vehicle:
no
Details on test solutions:
Preliminary test: An amount of the test item was dissolved in dechlorinated tap water, with the aid of high shear mixing at approximately 1500 rpm for approximately 5 min, and the volume adjusted to 2 L to give 2000 mg/L stock solution. Aliquots of the 2000 mg/L stock solution were wach separately diluted in a final volume of 20 L of dechlorinated tap water, and stirred using a flat bladed mixer for approximately 1 min, to give the 10 and 100 mg/L test concentrations respectively. The stock solution was inverted several times to ensure adequate mixing and homogeneity. Three fish were added to each 20 L test and control vessel. The control group was maintained under identical conditions but not exposed to the test item. Each vessel was covered to reduce evaporation. After 3, 6, 24, 48, 72 and 96 h any mortalities or sub-lethal effects of exposure were determined by visual inspection of the test fish.

Definitive test: the test item was dissolved directly in dechlorinated tap water. An amount of test item was dissolved in dechorinated tap water, with the aid of high shear mixing at approximately 1500 rpm for approximately 5 min, and the volume adjusted to 2 L to give a 2000 mg/L stock solution. The entire volume of this stock solution was diluted in a final volume of 20 L of dechlorinated tap water, and stirred using a flat bladed mixer for approximately 1 min, to give the 1000 mg/L test concentration. The stock solution was inverted several times to ensure adequate mixing and homogeneity.
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: rainbow trout
- Source: Brow Well Fisheries Limited, Hebden, near Skipton, Yorkshire, UK and maintained in-house since 7 March 2012
- Age at study initiation (mean and range, SD): juvenile
- Length: mean standard length of 5.1 cm (sd= 0.1) at the end of the definitive test
- Weight: mean weight of 2.00 g (sd= 0.14) at the end of the definitive test
- Feeding during test: feeding was discontinued approximately 24 h prior to the start of the definitive test
- Food type: commercial trout pellets
- Frequency: fish received no food during exposure

ACCLIMATION
- Acclimation period: 11 April 2012 to 23 April 2012
- Type and amount of food: commercial trout pellets
- Health during acclimation (any mortality observed): there was zero mortality in the 7 d prior to the start of the test

OTHER
- Fish were maintained in a glass fiber tank with a "single pass" water renewal system.
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Post exposure observation period:
Mortalities and sub-lethal effects of exposure were recorded at 3, 6, 24, 48, 72 and 96 h after the start of exposure. The criteria of death were taken to be the absence of both respiratory movement and response to physical stimulation.
Hardness:
140 mg/L as CaCO3
Test temperature:
The water temperature was controlled at approximately 14 °C
pH:
Approximately 7
Dissolved oxygen:
≥ 9.8 mg O2/L.
Nominal and measured concentrations:
Nominal: 100 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 20 L glass expsoure vessels
- Type: closed. The test vessels were covered to reduce evaporation
- Aeration: test vessles were aerated via narrow bore glass tubes
- Type of flow-through (e.g. peristaltic or proportional diluter): semi-static
- Renewal rate of test solution (frequency/flow rate): the test preparations were renewed daily to ensure that the concentrations of the test item remained near nominal and to prevent the build up of nitrogenous waste products
- No. of organisms per vessel: 7
- Biomass loading rate: 0.70 g bw/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test water used for both the range-finding and definitive tests was the same as that used to maintain the stock fish. Laboratory tap water was dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex Water Softener). After dechlorination and softening the water was passed through a series of computer controlled plate heat exchangers to achieve the required temperature.
- Other: the diet and diluent water were considered not to contain any contaminant that would affect the integrity and outcome of the study.

OTHER TEST CONDITIONS
- Photoperiod: the lighting cycle was controlled to give 16 hr light and 8 fr darkness cycle with 20 min dawn and dusk transition periods.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: Three fish were added to each 20 L test and control vessel. The control group was maintained under identical conditions but not exposed to the test item. Each vessel was covered to reduce evaporation. After 3, 6, 24, 48, 72 and 96 h any mortalities or sub-lethal effects of exposure were determined by visual inspection of the test fish. There were no mortalities at the test concentrations of 10 and 100 mg/L. Therefore, a single test concentration of 100 mg/L was selected for the definitive test.
Reference substance (positive control):
no
Remarks:
The control group was maintained under identical conditions but not exposed to the test item.
Duration:
96 h
Dose descriptor:
LC50
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.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Remarks:
and sub-lethal effects
Details on results:
Range-finding test
There were no sub-lethal effects of exposure during the range-finding test. The results showed no mortalities at the test concentrations of 10 and 100 mg/L.

Chemical analysis of the 100 mg/L test preparation at 0 and 24 h showed measured concentrations of 100 % and 97 % of nominal respectively indicating that the test item was stable under test conditions.

Definitive Test
- Mortality: there were no mortalities in 7 fish exposed to a test concentration of 100 mg/L for a period od 96 h. The results of the definitive test showed the highest test concentration resulting in 0 % mortality to be ≥ 100 mg/L, the lowest concentration resulting in 100 % mortality to be > 100 mg/L. The No Observed Effect Concentration (NOEC) was 100 mg/L. It was not considered unnecessary and unrealistic to test at concentrations in excess of 100 mg/L.
- Sub-lethal effects: there were no sub-lethal effects of exposure observed in 7 fish to a test concentration of 100 mg/L for a period of 96 h
- Observations on test item solubility: the test preparations were observed to be clear colourless solutions throughout the duration of the test
Sublethal observations / clinical signs:

Physico-chemical measurements

Temperature was maintained at approximately 14 °C throughout the test, while there were no treatment related differences for oxygen concentration of pH.

The oxygen concentration in some of the test vessels was observed to have an air saturation value (ASV) in excess of 100 %. This was considered to be due to the presence of microscopic air bubbles in the media super-saturating the diluents and was considered not to have had an impact on the outcome or integrity of the test as no adverse effects were observed.

Verification of test concentrations

Analysis of the test preparations at 0, 24 and 96 h showed measured test concentrations to range from 97 % to 102 % of nominal and so it was considered justifiable to estimate the LC50values in terms of the nominal test concentrations only. 

Validity criteria fulfilled:
yes
Conclusions:
The acute toxicity of the test item to rainbow trout was assessed according to OECD guideline 203. The 96-Hour LC50 based on nominal test concentrations was > 100 mg/L. The NOEC was 100 mg/L.
Executive summary:

Introduction

A study was performed to assess the acute toxicity of the test item to rainbow trout (Oncorhynchus mykiss). The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No 203, “Fish, Acute Toxicity Test” referenced as Method C.1 of Commission Regulation (EC) No. 440/2008.

Methods

Following a preliminary range-finding test, 7 fish were exposed to an aqueous solution of the test item, at a single concentration of 100 mg/L for a period of 96 h at a temperature of approximately 14 °C under semi-static test conditions. The number of mortalities and any sub-lethal effects of exposure in each test and control vessel were determined 3 and 6 hours after the start of exposure and then daily throughout the test until termination after 96 h.

Results

The 96-Hour LC50based on nominal test concentrations was greater than 100 mg/L. The No Observed Effect Concentration was 100 mg/L.

It was considered unnecessary an unrealistic to test at concentrations in excess of 100 mg/L.

Analysis of the test preparations at 0, 25 and 96 h showed measured test concentrations to range from 97 % to 102 % of nominal and so the results are based on nominal test concentrations only.

Description of key information

The acute toxicity of the test item to rainbow trout was assessed according to OECD guideline 203. The 96-Hour LC50 based on nominal test concentrations was >100 mg/L. The NOEC was 100 mg/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Effect concentration:
100 mg/L

Additional information

Introduction

A study was performed to assess the acute toxicity of the test item to rainbow trout (Oncorhynchus mykiss). The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No 203, “Fish, Acute Toxicity Test” referenced as Method C.1 of Commission Regulation (EC) No. 440/2008.

Methods

Following a preliminary range-finding test, 7 fish were exposed to an aqueous solution of the test item, at a single concentration of 100 mg/L for a period of 96 h at a temperature of approximately 14 °C under semi-static test conditions. The number of mortalities and any sub-lethal effects of exposure in each test and control vessel were determined 3 and 6 hours after the start of exposure and then daily throughout the test until termination after 96 h.

Results

The 96-Hour LC50based on nominal test concentrations was greater than 100 mg/L. The No Observed Effect Concentration was 100 mg/L.

It was considered unnecessary an unrealistic to test at concentrations in excess of 100 mg/L.

Analysis of the test preparations at 0, 25 and 96 h showed measured test concentrations to range from 97 % to 102 % of nominal and so the results are based on nominal test concentrations only.