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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:
04 July 2012 - 24 December 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Version / remarks:
EC No. 440/2008
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
June 2012
Analytical monitoring:
yes
Details on sampling:
- Concentrations: from control and all test concentrations.
- Sampling method: Two, mid-vessel samples of 10 mL of media taken from control and test vessels at 0 and 72 hrs (fresh media) and at 24 and 96 hours (expired media)
- Sample storage conditions before analysis: One sample analysed, second stored frozen in case of further analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Test glassware was pre-exposed overnight at the test concentration before use in test, to reduce loss of BMI by sorption.
In duplicate, the test substance (7.64 mg, corrected for purity) was added to the dilution medium (20 L) and stirred for approximately 20 minutes. One batch was used as the test medium at a nominal concentration of 360 μg/L and aliquots of the second batch, (9111, 4133, 1878 or 856 mL) were added to dilution medium and adjusted to volume (20 L) to provide test media at nominal concentrations of 164, 74.4, 33.8 and 15.4 μg/L.
- Controls: diluent water
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: Rainbow trout
- Source: commercial fish farm in the UK
- Length at study initiation: mean total length 4.23 cm
- Weight at study initiation: mean wet weight 0.71 g
- Method of breeding: The eggs were obtained from the supplier and were kept in an aerated supply of diluent water under flow-through conditions until use.
- Feeding during test: No
ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions (same as test or not): yes
- Type and amount of food: commercial fish food (TROUW (UK) Ltd; Nutra Fry 02), equivalent to between 3 and 4% of the total wet-weight of fish in the holding tank. No food was given during the 24-hour period immediately before exposure.
- Feeding frequency: daily
- Health during acclimation: No medication was given during the holding period, and mortalities were recorded as <1 % in the 14 days before the definitive test.


Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
154-158 mg/L as CaCO3
Test temperature:
13.1 - 14.2 °C
pH:
8.02 - 8.48
Dissolved oxygen:
89 - 102 % air saturation value
Nominal and measured concentrations:
Nominal concentrations, control, 15.4, 33.8, 74.4, 164, 360 ug/L
Measured concentrations, 0, 6.96, 15.1, 30.9, 59.1, 145 ug/L (geometric mean values; see Table 1 in 'Any other information on results' for details on measured concentrations).
Details on test conditions:
TEST SYSTEM
- Test vessel: Aquarium
- Type: open
- Material, size, headspace, fill volume: glass, 20 L of medium to a depth of 20 cm.
- Aeration: narrow bore glass tubes
- Renewal rate of test solution (frequency/flow rate): Daily batch renewal of the media
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.2489 g bw/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: laboratory tap water, dechlorinated and softened by passage through a water purification system. It was passed through a high grade activated carbon filter to remove chlorine and any organic contaminants. A proportion of the supply then passed through a water softener before final reverse osmosis treatment to produce a highly purified water supply. The two grades of dechlorinated water were then remixed to give a supply with the desired water hardness. This water was then held in an intermediate tank where it was equilibrated to the test temperature and gently aerated before being supplied to the holding and test areas.
- Total organic carbon: 0.65 mg/L
- Particulate matter: <3 mg/L
- Pesticides: <0.17 ug/L
- Chlorine: 0.05 mg/L
- Conductivity: 300 us/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement: Daily records of temperature, pH and dissolved oxygen were kept for each control and test vessel together with measurements of total hardness for selected vessels at 0 hours.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hrs light, 8 hrs dark
- Light intensity: periods of subdued lighting at the beginning and end of each light phase

EFFECT PARAMETERS MEASURED:
- Mortality, at approximately 2, 4, 24, 48, 72 and 96 hours, subjective assessments were also made on the incidence and type of any sub-lethal effects compared with control fish.

TEST CONCENTRATIONS
- Range finding study: A range-finding study using a stated limit of aqueous solubility was conducted.
- Test concentrations: 360 ug/mL
- Results used to determine the conditions for the definitive study: Yes, significant mortality and sub-lethal effects were observed at the nominal test concentration of 360 μg/L.
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
145 µg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Behavioural abnormalities: none
- Mortality of control: none
- Abnormal responses: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no

Measured concentrations of BMI in samples of the freshly prepared test media (on days 0 or 3) ranged between 59 and 111% of their nominal values. In samples of expired media on days 1 and 4 ranged between 14 and 27% of their nominal values (between 14 and 38% of their initial values). Based on a geometric mean, the overall mean measured levels of BMI were 6.96, 15.1, 30.9, 59.1 and 145 μg/L, and these values have been used to determine the study end points.

Study report does not discuss difference in mortality between range-finding study ('significant mortality and sub-lethal effects' at 360 ug/L nominal) and final test ('no mortality or significant treatment-related effects' at 360 ug/L nominal)

No mortality or significant treatment-related effects were noted during the test, therefore the 96-hour LC50 could not be calculated but must be >145 μg/L. The no observed effect concentration (NOEC) was 145 μg/L, and the lowest observed effect concentration must be >145 μg/L.

The measurements of water quality (temperature, pH, concentrations of dissolved oxygen and total hardness) remained within acceptable limits throughout the study. At all concentrations, the test media were colourless.

Table 1: Measured concentrations

Nominal concentration (μg/L)

0 hours [Fresh]

24 hours [Expired]

72 hours [Fresh]

96 hours [Expired]

Overall mean* (μg/L)

Control

nd

nd

nd

nd

nd

15.4

14.1

3.70

11.4

3.95

6.96

33.8

36.6

6.44

24.1

9.25

15.1

74.4

61.9

11.3

78.7

16.6

30.9

164

182

24.8

122

22.2

59.1

360

214

81.7

265

96.5

145

nd: None detected (less than the limit of detection of 2 μg/L

* geometric mean

Validity criteria fulfilled:
yes
Remarks:
See 'Overall remarks' for details on validity criteria.
Conclusions:
No mortality or significant treatment-related effects on the fish were noted during the test; therefore the 96-hour LC50 value could not be calculated but must be >145 μg/L.
The no observed effect concentration (NOEC) was 145 μg/L, and the lowest observed effect concentration must be >145 μg/L.
Executive summary:

A study was performed to assess the acute toxicity of BMI to rainbow trout (Oncorhynchus mykiss) under semi-static conditions, with daily renewal of the media. The study was conducted in accordance with OECD guideline 203 and EC method C1. Groups of seven juvenile fish were exposed to BMI, for 96 hours at nominal concentrations of 15.4, 33.8, 74.4, 164 and 360 μg/L. The test media were prepared by direct addition of the test substance to dilution water; to aid dissolution, stirring was employed. The Measured concentrations of BMI in samples of the freshly prepared test media (on days 0 and 3) ranged between 59 and 111% of their nominal values. In samples of expired media on days 1 and 4 the measured levels ranged between 14 and 27% of their nominal values (between 14 and 38% of their initial values). Based on a geometric mean, the overall mean measured levels of BMI were 6.96, 15.1, 30.9, 59.1 and 145 μg/L, and these values have been used to determine the study end points. Observations of the fish were made after approximately 2, 4, 24, 48, 72 and 96 hours of exposure. No mortality or significant treatment related effects were noted during the test. Therefore, the 96-hour LC50 value could not be calculated but must be >145 μg/L. The no observed effect concentration (NOEC) was 145 μg/L, and the lowest observed effect concentration must be >145 μg/L.

Description of key information

NOEC: 145 ug/L

LC50 and LOEC: >145 ug/L

Key value for chemical safety assessment

LC50 for freshwater fish:
145 µg/L

Additional information

A study was performed to assess the acute toxicity of BMI to rainbow trout (Oncorhynchus mykiss) under semi-static conditions, with daily renewal of the media. The study was conducted in accordance with OECD guideline 203 and EC method C1. Groups of seven juvenile fish were exposed to BMI, for 96 hours at nominal concentrations of 15.4, 33.8, 74.4, 164 and 360 μg/L. The test media were prepared by direct addition of the test substance to dilution water; to aid dissolution, stirring was employed. The Measured concentrations of BMI in samples of the freshly prepared test media (on days 0 and 3) ranged between 59 and 111% of their nominal values. In samples of expired media on days 1 and 4 the measured levels ranged between 14 and 27% of their nominal values (between 14 and 38% of their initial values). Based on a geometric mean, the overall mean measured levels of BMI were 6.96, 15.1, 30.9, 59.1 and 145 μg/L, and these values have been used to determine the study end points. Observations of the fish were made after approximately 2, 4, 24, 48, 72 and 96 hours of exposure. No mortality or significant treatment related effects were noted during the test. Therefore, the 96-hour LC50 value could not be calculated but must be >145 μg/L. The no observed effect concentration (NOEC) was 145 μg/L, and the lowest observed effect concentration must be >145 μg/L.