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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:
16 March 1992 to 20 March 1992
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
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:
Samples of each freshly prepared and old test media were taken throughout the test to determine the concentration of the test material present.
Vehicle:
yes
Remarks:
Analar acetone
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
The test material prepared in analar acetone and dilution water/culture medium to produce a final nominal concentration of 0.2 mg/L. The reported water solubility of the test material lies between the values of <0.14 mg/L and 0.3 mg/L. The limit of detection of the test material in this test was 0.14 mg/L. The concentration of analar acetone present in the test, water/culture medium during the tests was < 100 μL/L.
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: Rainbow trout
- Strain: Walbaum
- Length at study initiation: Mean length of 4.3 cm (range 3.5 to 5.4 cm).
- Weight at study initiation: Mean weight of 0.57 g (range 0.27 to 1.1 g).

ACCLIMATION
- Acclimation period: More than 9 days before use. After a 48 h settling-period fish for testing are acclimated to the test conditions for a minimum of 7 days before the test begins.
- Acclimation conditions: The fish are placed in ~300 L circular self-cleaning fibre-glass tanks at a density appropriate to their size. The tanks receive a continuous flow of temperature controlled water of a suitable and defined quality. The temperature during acclimation was 12 - 14 °C. The water hardness was 268 - 270 mg/L as CaCO3. The pH was 7.0 - 7.2 and the dissolved oxygen was 10.1 - 10.3 mg/L.
- Type and amount of food during acclimation: The fish are fed a maintenance ration of Mainstream Trout Food No. 03 (BP Nutrition Ltd.), the quantities fed depending upon the size and age of the fish and the temperature of the water.
- Health during acclimation: On arrival at the laboratory the fish are given a batch number and a random sample is examined for parasites and bacterial or fungal infections. Fish mortalities in the laboratory are recorded daily. The fish are deemed acceptable for testing if the cumulative mortality in the batch is less than 5 % over a 7 day period preceding the test. If mortality exceeds 5 % or disease is apparent the stock is not used for testing. The O. mykiss used in this experiment were maintained as described above and deemed acceptable for testing.

FEEDING DURING TEST
- Food type: Fish were not fed during the test. The fish were not fed in the 24 h preceding the test.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
242 - 282 mg/L as CaCO3
Test temperature:
16 °C (nominally 11 - 17 °C)
pH:
7.3 - 8.0
Dissolved oxygen:
7.5 - 10.7 mg/L
Nominal and measured concentrations:
Nominal concentration: 0.2 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 14 L spherical glass reaction vessels
- Material, size, headspace, fill volume: Glass, 10 L
- Aeration: The vessels contents were not aerated during the test. Prior to use in tests the water was vigorously aerated for several hours to ensure that no residual chlorine remained.
- Renewal rate of test solution: Daily
- No. of organisms per vessel: 10
- No. of vessels per concentration: One
- No. of vessels per control: One
- No. of vessels per vehicle control: One

TEST MEDIUM / WATER PARAMETERS
During the test temperature of the test media (nominally 11 - 17 °C) was monitored at hourly intervals. The pH and dissolved oxygen concentration of each freshly prepared and old test media were determined at the start and conclusion of the test and at each test medium renewal. The total hardness and residual chlorine concentration of each batch of dechlorinated tap water used in the test was also determined.
- Source/preparation of dilution water: The water used for maintenance of stocks and for the toxicity test with O. mykiss was the laboratory mains supply. The water comes from two pumping stations (Highstead and Eastling) controlled by the Mid Kent Water Company. The water is obtained from bore holes in the chalk of the North Downs. The only chemical treatment prior to its arrival in the laboratory is chlorination to 0.1 mg 1-1.
- Particulate matter: In the laboratory the water was filtered (PALL MDY 1001 Y400) to remove all particles larger than 15 μm (90 % of particles greater than 10 μm) and passed through activated carbon filters (Cano model CT) to remove chlorine and organic contaminants. Both particle and activated carbon filter cartridges' are renewed as recommended by the manufacturers.
- Chlorine: 0.04 mg/L
- Intervals of water quality measurement: Samples of the water, after filtration but before aeration, are taken at approximately 6 month intervals and analysed for a number of parameters. These analyses are carried out by the Mid Kent Scientific Services Limited. The temperature of the water was monitored at hourly intervals by a computer controlled thermocouple system which outputs to a dedicated microcomputer. The pH, concentration of dissolved oxygen, and total hardness were checked weekly. The pH measurements were made with a Corning (240) pH meter calibrated in the range pH 7.0 to pH 9.2 immediately prior to use. The dissolved oxygen measurements were made with a YSI 57 dissolved oxygen meter calibrated in air saturated water immediately prior to use. Total hardness was determined by titration against EDTA in the presence of ammonia buffer and BDH total hardness indicator tablets.
Water quality during tests: The temperature of the water in a vessel adjacent to the test aquaria was monitored as described previously. The pH and concentration of dissolved oxygen in the controls and each test concentration medium were determined immediately prior to and after renewal (i.e. at 24 h intervals). The total hardness and residual chlorine concentration of each batch of water used in the preparation of the test solutions was determined. The residual chlorine concentration was measured using a Lovibond Nesslerizer. All other determinations were made using the methods described previously.

OTHER TEST CONDITIONS
- Temperature adjustment: Heat exchange units. (Stainless steel and perspex) were used to adjust the temperature of the water.

EFFECT PARAMETERS MEASURED
At 24 h intervals the fish were observed and the number of fish exhibiting toxic symptoms was recorded. The symptoms were classified into one of five categories. The live fish were then transferred to freshly prepared test and control media.

TEST CONCENTRATIONS
- Range finding study
Test concentrations: Concentrations of the test material of 0.002, 0.02 and 0.2 mg/L.
Results used to determine the conditions for the definitive study: The concentration tested in the definitive tests was decided upon on the basis of the results of range-finding tests.
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
0.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
The test material at a mean measured concentration of 0.1 mg/L was found not to be toxic to O. mykiss over a 96 hour exposure period.

- Water quality parameters: The water quality parameters were generally in the preferred ranges. However the following should be noted. The total water hardness recorded during the test with O. mvkiss was above the preferred maximum of 250 mg/L (OECD guideline 203, Fish, Acute Toxicity Test). The recorded total hardness range of 242 to 282 mg/L was not regarded sufficient as to jeopardise the outcome of the study.

- Analytical observations: The results of the toxicity tests are expressed below in relation to the mean measured concentration of the test material found in the test media. In the test with O. mykiss the measured concentration of the test material in the freshly prepared test media at the start of each of the 4 exposure periods ranged from 55 to 80 % of the nominal value of 0.2 mg/L. The measured concentration of the test material fell by 52 % between renewals (24 h) of the test media.
No test material was found in any of the control media at the limit of detection of 0.01 mg/L.
Validity criteria fulfilled:
not specified
Conclusions:
Under the conditions of the test, O. mykiss exposed to 0.1 mg/L the test material exhibited no symptoms of toxicity over the duration of the test.
It is concluded that the substance is not acutely toxic to fish at the limit of water solubility.
Executive summary:

The acute toxicity of the test material was assessed in rainbow trout according to EU Test Method C.1. and in compliance with GLP.

The measured concentration of the test material in the freshly prepared test media at the start of each of the 4 exposure periods ranged from 55 to 80 % of the nominal value of 0.2 mg/L. The measured concentration of the test material fell by 52 % between renewals (24 h) of the test media.

The results of the toxicity tests are expressed as the mean measured concentration of the test material present in the test media.

The acute toxicity of the test material to O. mykiss was determined in a 96 hour semi-static toxicity test with daily renewal of the test media.

Under the conditions of the test, O. mykiss exposed to 0.1 mg/L the test material exhibited no symptoms of toxicity over the duration of the test.

Description of key information

Under the conditions of the test, rainbow trout (O. mykiss) exposed to 0.1 mg/L the test material (read-across substance) exhibited no symptoms of toxicity over the duration of the test. It is concluded that the substance is not acutely toxic to fish at the limit of water solubility.

Key value for chemical safety assessment

Additional information

The acute toxicity of the test material was assessed in rainbow trout according to EU Test Method C.1. and in compliance with GLP. The study was awarded a reliability score of 2 in accordance with the criteria set forth by Klimisch et al. (1997).

The measured concentration of the test material in the freshly prepared test media at the start of each of the 4 exposure periods ranged from 55 to 80 % of the nominal value of 0.2 mg/L. The measured concentration of the test material fell by 52 % between renewals (24 h) of the test media.

The results of the toxicity tests are expressed as the mean measured concentration of the test material present in the test media.

The acute toxicity of the test material to O. mykiss was determined in a 96 hour semi-static toxicity test with daily renewal of the test media.

Under the conditions of the test, O. mykiss exposed to 0.1 mg/L the test material exhibited no symptoms of toxicity over the duration of the test.