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Ecotoxicological information

Short-term toxicity to fish

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Link to relevant study record(s)

Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-08-25 to 2017-08-29
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Version / remarks:
2008
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 4.31 mg/L
- Sampling method: The limit loading level and the control were analytically verified from all freshly prepared and aged test media.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until sample preparation and at room temperature until the start of the analysis (on an autosampler),
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Preparatoin as a WAF, water accomodated fraction.
- Method: An appropriate volume of the test item was introduced in the 2000 mL dilution water by pipette in brown glass flasks. The dispersions were shaken for 24 hours with 20 rpm at room temperature. After completion of shaking and following a separation phase of at least 1 hour of standing, the aqueous phase or WAF was removed from the approximate center of the water body by siphoning.
- Controls: 7 fish in dilution water (without test item) were tested under the same test conditions as the test replicate.
- Chemical name of vehicle: dilution water
- Test concentration: 4.31 mg/L
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
- Common name: zebrafish
- Strain: Danio rerio
- Source: a single brood stock

ACCLIMATION
- Acclimation period: at least 12 days
- Acclimation conditions: same as test
- Type and amount of food during acclimation: Sera Vipan, 4 % of the fish body weight per feeding day.
- Feeding frequency during acclimation: 3 times per week, not fed 24 h before the test start
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Hardness:
57 mg/L as CaCO3
Test temperature:
22.7-23.4°C
pH:
pH at the start of exposure: 7.47-7.60 (4.31 mg/L), 7.32-7.69 (control)
pH at the end of exposure: 7.24-7.32 (4.31 mg/L), 7.17-7.36 (control)
Dissolved oxygen:
9.97 mg/L

Nominal and measured concentrations:
Nominal: 4.31 mg/L
Measured: 3.23-3.66 mg/L (fresh medium)
Details on test conditions:
TEST SYSTEM
- Test vessel: 13 L glass aquaria
- Type: closed, covered with a floating lid
- Material, size, headspace, fill volume: Glass aquaria of 13 L, filled with approximately 8 L of dilution water
- Aeration: No aeration
- Renewal rate of test solution (frequency/flow rate): 24 h
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): One replicate
- No. of vessels per vehicle control (replicates): One replicate
- Biomass loading rate: 0.15 g fish per litre test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap water of local origin
- Chlorine:filtered on activated charcoal and aerated for at least 24 h to remove chlorine.
- Total hardness: 10 - 250 mg CaC03/L
- pH-value: 6.0 - 8.5
- Acidity: 0.1 mmol/L
- Alcalinity: 0.6 mmol/L
- Conductivity: 138 µS/cm

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light / 8 h dark
- Light intensity: 7 - 750 Lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Biological Parameters: observed for visible abnormalities after 3, 24, 48, 72 and 96 h. Measured size and weight of the control fish at the end of the exposure.
- Physical Properties: The pH-value, temperature and oxygen saturation were measured in all test vessels at test start and end and every 24 hours and after renewal of the test media from fresh and old test media. Total hardness was determined at the start of the exposure out of the control. Water temperature was recorded once per hour during the test. Light intensity was measured at the start of the exposure.

TEST CONCENTRATIONS
- Justification for using less concentrations than requested by guideline: Based on the toxicity data of the test item of an acute immobilization test to Daphnia magna (EL50(0-48h) = 4.31 mg/L) and an algae toxicity test (ErL50 (0-72 h) = 6.90 mg/L, EyL50(0 - 72 h)= 4.57 mg/L, based on nominal loading levels of the water accommodated fractions of the test item), a limit test is justified.
- Test concentrations: 4.31 mg/L was tested as a threshold loading level in a limit test
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
LL50
Effect conc.:
> 4.31 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Duration:
96 h
Dose descriptor:
LL0
Effect conc.:
>= 4.31 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Duration:
96 h
Dose descriptor:
LL100
Effect conc.:
> 4.31 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
- Biological Data: No mortality occurred.
- Physicochemical data: The environmental conditions (pH-value, temperature and oxygen-saturation) were determined to be within the acceptable limits. Light intensity was measured at the start of the exposure and ranged from 46 to 62 (mean value 55) Lux. The test solution showed slight turbidity during the period of exposure
- Analytical data: The measured concentrations of the two main constituents of the test item at the end of each exposure interval were in the range of 68 to 75% of the initially measured concentrations.
Sublethal observations / clinical signs:

Biological results:

Nominal loading level of the test item

[mg/L]

Effect

Number of fish effected at observation time [hours]

3 

24

48

72

96

4.31

Quiescence

 

 

7

7

7

7

7

Control

Normal behaviour

7

7

7

7

7

 

Cumulative Mortality [%]:

 

Nominal loading level of the test item [mg/L]

Cumulative mortality [%] after different exposure periods [hours]

3

24

48

72

96

 

4.31

0

0

0

0

0

 

Control

0

0

0

0

0

 

Analytical results:

Measured concentrations of the two main constituents and percent of the initial concentrations:

 

Sampling date

0 hours

(Fresh medium)

24 hours

(Old medium)

24 hours

(Fresh medium)

48 hours

(Old medium)

Nominal concentration

of the test item [mg/L]

Meas. conc.

[mg/L]

Meas.conc.

[mg/L]

%

Meas. conc.

[mg/L]

Meas.conc.

[mg/L]

%

4.31

3.44

2.47

72

3.66

2.48

68

Control

< LOQ

< LOQ

< LOQ

< LOQ

 

Sampling date

48 hours

(Fresh medium)

72 hours

(Old medium)

72 hours

(Fresh medium)

96 hours

(Old medium)

Nominal concentration

of the test item [mg/L]

Meas. conc.

[mg/L]

Meas.conc. [mg/L]

%

Meas. conc.

[mg/L]

Meas.conc.

[mg/L]

%

4.31

3.23

2.41

75

3.61

2.63

73

Control

< LOQ

< LOQ

< LOO

< LOQ

Validity criteria fulfilled:
yes
Conclusions:
The threshold loading of the test item was found to have no lethal effects on zebrafish when tested with the water accommodated fraction with the nominal loading of 4.31 mg/L. In conclusion, the LL0 corresponds to the nominal loading of ≥ 4.31 mg test item/L. The LL50 and LL100 correspond to the nominal loading of > 4.31 mg test item/L.
Executive summary:

The acute toxicity of the test item to fish (zebrafish) was determined according to the OECD Guideline for Testing of Chemicals No. 203 (1992) and EU Council Regulation No. 440/2008/C.1 (2008), in a semi-static limit test in a closed system without headspace with a threshold concentration of 4.31 mg/L. Exposure was assessed for 96 hours. 7 test organisms were exposed each for the test concentration and the control. The water quality parameters pH-value, temperature and oxygen saturation, measured after 0, 24, 48, 72 and 96 h, were determined to be within the acceptable limits.

The concentration of the test item was analytically verified via GC-MS from all freshly prepared media and aged test media during exposure. The measured concentrations of the two main constituents of the test item at the end of each exposure interval were in the range of 68 to 75% of the initially measured concentrations. This means, that the test item was stable during the exposure. All effect values given are based on the nominal test item loading.

The threshold loading of the test item was found to have no lethal effects on zebrafish when tested with the water accommodated fraction with the nominal loading of 4.31 mg/L. In conclusion, the LL0 corresponds to the nominal loading of >= 4.31 mg test item/L. The LL50 and LL100 correspond to the nominal loading of > 4.31 mg test item/L.

Description of key information

One study is available for this endpoint in which the toxicity to fish was assessed according to OECD 203 and EU Method C.1, in a semi-static limit test in a closed system without headspace. The LL0 corresponds to the nominal loading of ≥ 4.31 mg test item/L. The LL50 and LL100 correspond to the nominal loading of > 4.31 mg test item/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Effect concentration:
4.31 mg/L

Additional information

For this endpoint only one test is available. In this acute toxicity test with fish (Danio rerio or zebrafish), the effects of a threshold concentration of 4.31 mg/L of the test item were determined according to OECD 203 (1992).

7 test organisms were exposed each for the test concentration and the control. The water quality parameters pH-value, temperature and oxygen saturation, measured after 0, 24, 48, 72 and 96 h, were determined to be within the acceptable limits.

The concentration of the test item was analytically verified via GC-MS from all freshly prepared media and aged test media during exposure. The measured concentrations of the two main constituents of the test item at the end of each exposure interval were in the range of 68 to 75% of the initially measured concentrations. This means, that the test item was sufficiently stable during the exposure. All effect values given are based on the nominal test item loading.

All validity criteria of the test guideline were met.

The threshold loading of the test item was found to have no lethal effects on zebrafish when tested with the water accommodated fraction with the nominal loading of 4.31 mg/L. In conclusion, the LL0 corresponds to the nominal loading of ≥ 4.31 mg test item/L. The LL50 and LL100 correspond to the nominal loading of > 4.31 mg test item/L.