Registration Dossier

Ecotoxicological information

Short-term toxicity to fish

Currently viewing:

Administrative data

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:
25 Jun 2018 - 17 Aug 2018
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:
other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23
Version / remarks:
2000
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All test concentrations and the control
- Sampling method: 2.0 mL, taken at the start and at the end of the first renewal period (0-24 hour) and at the start and at the end of the last renewal period (0-72 hour).
- Sample storage conditions before analysis: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Preparation of test solutions started with the highest concentration of 10 mg solids/L (26 mg test item/L) applying a 1-hour period of magnetic stirring to accelerate dissolution of the test item in medium. No scattering of laser beam was observed in the highest concentration, indicating complete dissolution of the test item. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure.
- Controls: Test medium without test item or other additives
- Other relevant information: Test item was completely soluble in test medium at the concentrations tested. A correction was made for the purity/composition of the test substance. A correction factor of 2.554 was used. Concentration/doses are expressed as solids.
Test organisms (species):
Cyprinus carpio
Details on test organisms:
TEST ORGANISM
- Common name: Carp
- Strain: Cyprinus carpio Linnaeus, 1758
- Source: Zodiac, proefacc, "De Haar Vissen", Wageningen University and Research Centre, The Netherlands.
- Length at study initiation: 3.4 ± 0.3 cm
- Weight at study initiation: 0.41 ± 0.11 g
- Method of breeding: F1 from a single parent-pair bred in UV-treated water.
- Total fish used: 51

ACCLIMATION
- Acclimation period: at least 12 days after delivery
- Acclimation conditions: Adjusted ISO medium, formulated using tap-water purified by reverse osmosis.
- Type and amount of food during acclimation: Daily with pelleted fish food (Essence (300-500 um), Coppens International bv, Helmond, The Netherlands)
- Validity of batch: In the batch of fish used for the test, mortality during the seven days prior to the start of the test was less than 5%.

FEEDING DURING TEST
- No feeding from 24 hours prior to the test and during the total test period.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
180 mg CaCO3/L
Test temperature:
21 - 22 °C
pH:
7.4 - 7.9
Dissolved oxygen:
New solutions: 8.3 - 8.8 mg/L
Old solutions: 6.0 - 7.8 mg/L
Nominal and measured concentrations:
Nominal: 0.46, 1.0, 2.2, 46 and 10 mg solids/L
Measured: Concentrations measured in freshly prepared solutions were at the level of nominal concentrations (99 – 115%). In spent solutions, measured concentrations were at 95-142% of nominal concentrations. Based on these results, the average exposure concentrations were 0.54, 1.1, 2.5, 5.1 and 11 mg solids/L. See 'Any other information on materials and methods' for calculation of average exposure concentrations and 'Any other information on results' for details on measured concentrations.
Details on test conditions:
TEST SYSTEM
- Test vessel: 6.5 litres, all-glass, containing 5.0 litres of test solution
- Aeration: no
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.57 g fish/Litre
- Introduction of fish: Within 26 minutes after preparation of the test media from a holding tank with comparable water quality parameters and pH and temperature differences between test and holding tank media of less than 1.0 unit and 1.0°C.
- Euthanasia: At the end of the test the surviving fish were rapidly killed by exposing them to ca. 1.2% ethylene glycol monophenylether in water.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Adjusted ISO medium
- Culture medium different from test medium: no
- Intervals of water quality measurement: Dissolved oxygen content, pH and temperature: Daily in all vessels with surviving fish, beginning at the start of the test (day 0).

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h photoperiod daily

EFFECT PARAMETERS MEASURED:
- Mortality and other effects: At 3, 24, 48, 72 and 96 hours following the start of exposure. In addition, every afternoon from day 0 to observe for any dead or severely distressed fish. Dead fish were removed when observed.
- Dissolved oxygen content, pH and temperature: Daily in all vessels with surviving fish, beginning at the start of the test (day 0).

TEST CONCENTRATIONS
- Range finding study test concentrations: 1.0, 10 and 100 mg solids/L.
- Results used to determine the conditions for the definitive study: yes, LC50 expected to be between 1.0 and 10 mg solids/L.
Reference substance (positive control):
yes
Remarks:
Pentachlorophenol (performed Mar 2018)
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
4 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Test substance corrected for water content (specified in report as "solids")
Basis for effect:
mortality
Remarks on result:
other: 95% confidence interval: 3.3 - 4.8 mg solids/L
Details on results:
- No mortality or other effects were observed in the control and the three lowest concentrations tested during the exposure period. All fish exposed to the highest concentration were found dead after the first 24 hours of exposure. Four and six of the fish exposed to 5.1 mg solids/L were found dead after 72 and 96 hour of exposure, respectively.
- Fish exposed to 5.1 mg solids/L were observed to be red coloured from 48 hour of exposure.
- Concentrations measured in freshly prepared solutions were at the level of nominal concentrations (99 – 115%). In spent solutions, measured concentrations were at 95-142% of nominal concentrations. The reason behind the observed increase of concentrations is unknown.
- All environmental parameters remained within the limits prescribed by the study plan throughout the test.
Results with reference substance (positive control):
The 96h-LC50 for carp exposed to PCP was 0.15 mg/L (95 % confidence interval between 0.10 and 0.22 mg/L).
The range of the 96h-LC50 for carp is generally between 0.10 and 0.46 mg/L based on historical data of reference tests performed approximately every 3 months from April 1988 until the end of 2000, and annually since then. Hence, the sensitivity of carp originating from the present batch for PCP falls within the range of sensitivities generally observed during the past years.
Reported statistics and error estimates:
The 96h-LC50 value was calculated using the Spearman-Karber procedure (without trim). ToxRat Professional v 3.2.1 (ToxRat Solutions® GmbH, Germany) was used to perform the analyses.

Table 1: Average Exposure Concentration Versus Nominal Concentration

Nominal concentration 

(mg solids/L)

Measured concentrations (mg solids/L)

Average conc.

(mg solids/L)

t=0h

(new)

t=24h

(old)

t=72h

(new)

t=96h

(old)

0.46

0.52

0.56

0.46

0.65

0.54

1.0

1.0

0.95

1.1

1.2

1.1

2.2

2.3

2.5

2.4

2.8

2.5

4.6

4.7

5.2

5.3

5.4

5.1

10

10

12

n.a.

n.a.

11

n.a. – not applicable, no surviving fish were present in this concentration

Table 2: Incidence of Mortality and Total Mortality During the Final Test


Average exposure conc.
 (mg solids/L)

Initial
Number
of fish

Cumulative mortality

Total
Mortality
(%)

3h

24h

48h

72h

96h

Control

7

0

0

0

0

0

0

0.54

7

0

0

0

0

0

0

1.1

7

0

0

0

0

0

0

2.5

7

0

0

0

0

0

0

5.1

7

0

0

0*

4*

6*

86

11

7

0

7

7

7

7

100

*Fish were observed to be red coloured compared to the control.

Validity criteria fulfilled:
yes
Remarks:
See 'Overall remarks'
Conclusions:
The 96h-LC50 was 4.0 mg solids/L based on average exposure concentrations (95% confidence interval between 3.3 and 4.8 mg solids/L).
Executive summary:

In a study performed in accordance with OECD 203 (1992) and according to GLP principles, the acute toxicity of the test substance to Cyprinus carpio was investigated. The test item consisted of 39.15% solids in water and was completely soluble in test medium at the concentrations tested. A correction factor of 2.554 was used and concentrations/doses are expressed as mg solids/L.

In a semi-static test, carp were exposed for 96 hours to an untreated control and nominal test concentrations of 0.46, 1.0, 2.2, 4.6 and 10 mg solids/L. Measured concentrations were at the level of nominal concentrations (99 -115%) in freshly prepared medium and at 95 - 142% of nominal in spent solutions. Therefore, the average exposure concentrations were calculated to be 0.54, 1.1, 2.5, 5.1 and 11 mg solids/L.

No mortality or other effects were observed in the control and at the three lowest concentrations tested during the exposure period. All fish exposed to the highest concentration were found dead after the first 24 hours of exposure. Six of the 7 fish exposed to 5.1 mg solids/L were found dead after 96 hours of exposure.

The 96h-LC50 was calculated to be 4.0 mg solids/L based on average exposure concentrations (95% confidence interval: 3.3 - 4.8 mg solids/L). The test met all validity criteria and is considered to be reliable without restriction.

Description of key information

Two studies with substance analogues were performed. The 96 -h LC50 values were 4 mg solids/L and 28 mg solids/L for C8 -C18 and C12 Amphoacetates, respectively. The lowest LC50 value is used as a worst-case read-across value for the registered substance.

Key value for chemical safety assessment

LC50 for freshwater fish:
4 mg/L

Additional information

The 96h-LC50 was 4.0 mg solids/L based on average exposure concentrations (95% confidence interval between 3.3 and 4.8 mg solids/L).