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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:
From 11 November 2002 to 15 November 2002
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
GLP compliance:
yes (incl. certificate)
Remarks:
(according to OECD, EC German principles of GLP)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0, 0.625, 1.25, 2.50, 5.0, 10.0 mg/L
- Sampling method: The test substance concentration levels 2.5, 5.0 and 10.0 mg/L and the control were analytically verified via analysis of dissolved organic carbon (DOC) at test initiation.
Vehicle:
yes
Details on test solutions:
PREPARATION TEST SOLUTION: Test concentrations were prepared by dilution of a stock solution at 62.5 mg/L.
- Dispersion treatment: 1 min at the instrument Ultraturrax at 20000 rpm
- Vehicle: Dilution water
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
- Common name: Zebrafish
- Source: Aquarium AM Aegi, Feldstr. 7, D-30159 Hannover
- Feeding during test: No

ACCLIMATION
- Acclimation period: At least 12 d
- Temperature: 23 ± 2°C
- Light intensity: Diffuse light (0.1 - 10 µmol x m (2) x s (1), natural photoperiod).
- Food: Trouvit 40/2; Milkivit, D-86666, Bürgheim. The amount of food was 4% of the fish body weight per feeding day.
- Feeding frequency: 3 times per week
- Water (maintenance): Tap water. The water was filtered on activated charcoal and aerated for at least 24 h to remove chlorine. The water was analysed once a year according to German tap water regulation. Nominal water parameters were:
Total hardness: 40 - 180 mg CaCO3/L
pH-value: 6.0 - 8.5
Dissolved oxygen: >80% of the air saturation value
The water was changed at least once per week.
- Health: No disease treatments were administered throughout holding.




Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
86 mg/L as CaCO3
Test temperature:
21.0 - 22.7 °C
pH:
7.14-7.64
Dissolved oxygen:
92-100%
Nominal and measured concentrations:
Nominal concentration: 0, 0.625, 1.25, 2.50, 5.0 and 10.0 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass aquaria loosely covered by glass tops
- Test volume: 10 L/vessel.
- Aeration: Gentle aeration was provided.
- No. of organisms per vessel: 7
- No. of vessels per concentration: 1
- No. of vessels per control: 1
- Fish density in the vessel: < 1 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Same tap water was used for holding and testing. Detail of holding water is provided under ‘Details on test organisms’ section above.
- Intervals of water quality measurement: pH value, temperature and oxygen saturation were measured in all vessels at the beginning of test and every 24 h. Total hardness of the water was determined at the beginning of test from the control vessel. During the test the room temperature was recorded continuously with a thermo-hygrograph.

OTHER TEST CONDITIONS
- Photoperiod: A natural photoperiod was maintained during the test.
- Light intensity: 0.1 – 10.0 µmol x m (-2) x s (-1).

EFFECT PARAMETERS MEASURED: The test fish were observed after 24, 48, 72 and 96 h for the following endpoints:
1. Mortality (evident by no visible movement and if touching of the caudal peduncle produced no reaction)
2. Visible abnormalities (e.g. loss of equilibrium, swimming behavior, respiratory function, pigmentation etc).
3. Body weight of fish
4. Fish size

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: A 72 h non-GLP range finding test was carried out with five fish/concentration.
- Test concentrations: 0, 1.0 and 10.0 mg/L
- Results used to determine the conditions for the definitive study: Yes. In the range finding study, 100% mortality was observed at 10.0 mg/L dose level within 24 h of exposure. No mortality was observed at the remaining test concentration throughout the range finding study.
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
5.1 mg/L
Nominal / measured:
nominal
Basis for effect:
mortality
Remarks on result:
other: 4.5-5.7 mg/L
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
1.25 mg/L
Nominal / measured:
nominal
Basis for effect:
mortality
Details on results:
- Behavioural abnormalities: Slowed escape reflex, lying on its side, lethargy, staggering, hyperventilation were observed in animals at concentrations above 1.25 mg/L.
- Effect concentrations exceeding solubility of substance in test medium: The test substance was clearly dissolved at all concentration levels. Slight formation of foam was observed at 10 mg/L on Days 0-2.

Table 1: Cumulative mortality data in the range-finding study

Nominal concentration (mg/L)

Cumulative Mortality (n=5)

24 h

48 h

72 h

Control

0

0

0

1.0

0

0

0

10

100

100

100

 

Table 2: Cumulative mortality data in the definitive study

Nominal concentration (mg/L)

Cumulative Mortality [ %]

24 h

48 h

72 h

96 h

Control

0

0

0

0

0.625

0

0

0

0

1.25

0

0

0

0

2.5

0

0

0

0

5.0

0

14

14

43

10

57

100

100

100

 

Table 3: Results based on the mortality data in the definitive study

Time (h)

LC50 (mg/L)

95% Confidence Limits (mg/L)

24

9.7

8.5->10

48

5.8

5.3-6.5

72

5.8

5.3-6.5

96

5.1

4.5-5.7

Validity criteria fulfilled:
yes
Remarks:
(O2 saturation was ≥ 60%, mortality in control was 0%)
Conclusions:
Under the conditions of the study, the 96 h LC50 and NOEC of the test substance were calculated to be 5.1 mg/L (c.i.- 4.5-5.7) and 1.25 mg/L, respectively.
Executive summary:

A study was conducted to determine the acute toxicity of the test substance to Zebra fish (Danio rerio) according to OECD Guideline 203, in compliance with GLP. Seven test organisms in single replicates were exposed to the substance at nominal concentrations of 0, 0.625, 1.25, 2.50, 5.00 and 10.0 mg/L for 96 h under static conditions. Mortality was observed at 24, 48, 72 and 96 h. The dose levels of 2.5, 5.0 and 10.0 mg/L and the control were analytically verified via analysis of dissolved organic carbon (DOC) at test initiation. Under the conditions of the study, the 96 h LC50 and NOEC were calculated to be 5.1 (c.i.-4.5-5.7) and 1.25 mg/L (nominal), respectively (Scheerbaum, 2002).

Description of key information

Key value for chemical safety assessment

LC50 for freshwater fish:
5.1 mg/L

Additional information

A study was conducted to determine the acute toxicity of the test substance to Zebra fish (Danio rerio) according to OECD Guideline 203, in compliance with GLP. Seven test organisms in single replicates were exposed to the substance at nominal concentrations of 0, 0.625, 1.25, 2.50, 5.00 and 10.0 mg/L for 96 h under static conditions. Mortality was observed at 24, 48, 72 and 96 h. The dose levels of 2.5, 5.0 and 10.0 mg/L and the control were analytically verified via analysis of dissolved organic carbon (DOC) at test initiation. Under the conditions of the study, the 96 h LC50 and NOEC were calculated to be 5.1 (c.i.-4.5-5.7) and 1.25 mg/L (nominal), respectively (Scheerbaum, 2002).