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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From January 23th to January 25th 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Sampling method: samples were taken from each test item group and the control at the start and at the end of the test in duplicates. The first series of samples taken (4×5 ml) were sent to the analysis.
- Sample storage conditions before analysis: room temperature
Vehicle:
no
Details on test solutions:
The test solutions used in the test were prepared by mechanical dispersion without using any solubilising agent. After the formulation procedure the test animals were immediately introduced into the test solutions.
A test item solution was first prepared by dissolving an amount of 0.0609 g test item in 609 ml dilution water resulting a nominal concentration of 100 mg/l. This solution was filtrated through a membrane filter (0.22 µm) to separate the possible micelles formed because of the surface activity of test item. The further test item solutions were prepared by the appropriate dilution of this test item stock solution with a dilution factor of 2.0.

nominal conc. mg/l test item stock solution ml ISO medium ml
100 300 –
50 150 q.s. ad 300
25 75 q.s. ad 300
12.5 37.5 q.s. ad 300
6.25 18.75 q.s. ad 300
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Strain/clone: Daphnia magna (Straus)
- Age at study initiation: less than 24 h old at the beginning of the test
- Method of breeding: Daphnia are bred in the laboratory of TOXI-COOP ZRT.
- Source: Laboratory of Hydrobiology (Central Agricultural Office, Directorate of Plant-, and Soil Protection) 2100 Gödöllő, Kotlán S. u. 3. Hungary
- Feeding during test : test animals were not fed during the exposure

ACCLIMATION
- Acclimation period: test animals were bred under similar conditions as that during the exposure period, therefore additional acclimation before the test was not necessary.
- Acclimation conditions: same as test
- Type and amount of food: the daphnia were fed with concentrated algal suspension of Raphidocelis subcapitata during holding
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
The hardness of reconstituted water was measured as 240.3 mg/l (CaCO3) at the start of the study
Test temperature:
The test temperature was in the range of 20.3-20.7 °C measured in the test vessels during the test. The additionally measured ambient temperature within the climate chamber was between 19.9-20.9 °C.
pH:
The pH of the test solution was not adjusted and not varied by more than 1.5 units in any one test. The pH was in the range of 7.32-7.93 during the test.
Dissolved oxygen:
The dissolved oxygen concentration was in the range of 7.06-7.51 mg/l during the test.
Nominal and measured concentrations:
Nominal concentrations: 6.25, 12.5, 25, 50, 100 mg/l
Measured concentrations t 0: 5.85±0.12, 11.8±0.10, 23.9±0.14, 47.7±0.29, 95.5±2.32 mg/l
Measured concentrations t 48h: 6.14±0.08, 12.4±0.46, 25.5±1.23, 51.3±3.11, 103±9.72 mg/l
Details on test conditions:
TEST SYSTEM
- Test vessel: beaker
- Material, size, headspace, fill volume: glass, volume app. 50 ml
- Volume of solution: 40 ml of test solution for each vessel
- Aeration: not aerated
- No. of organisms per vessel: five animals
- No. of vessels per concentration (replicates): four replicates
- No. of vessels per control (replicates): four replicates

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstitued water (ISO medium) was used as dilution water in the experiment. Separate stock solutions of individual substances were first prepared in deionised water. The ISO medium is prepared by adding 25 ml from each of four stock solutions to one litre of deionised water proportionally.

OTHER TEST CONDITIONS
- Adjustment of pH: not adjusted
- Light intensity: the experiment was carried out in complete darkness
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
37.957 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
51.642 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
25 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
50 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
95 % conf. limits
EC10:12.858-44.643 mg/l
EC20: 20.593-48.244 mg/l
EC50: 43.138-65.770 mg/l

Results of the preliminary range-finding test (48h exposure)
nominal conc. mg/l control 0.1 1 10 50 100
no. of treated animals 10 10 10 10 10 10
no. immobilised animals 0 0 0 0 0 0

Nominal concentration
[mg/L]

Number of
Daphnia
tested

Number of immobilised
Daphnia after

% of immobilised
Daphnia after

24 h

48 h

24 h

48 h

Control

20

0

0

0

0

6.25

20

0

0

0

0

12.5

20

0

0

0

0

25

20

0

0

0

0

50

20

0

9

0

45

100

20

11

20

55

100

Validity criteria fulfilled:
yes
Conclusions:
In this 48-hour acute toxicity test on daphnia magna, effect levels were determined:
EC50 (48h) = 51.642 mg/l
NOEC (48h) = 25 mg/l
LOEC (48h) = 50 mg/l
All validity criteria were met. The results are based on nominal test item concentrations.
Executive summary:

Test substance was tested for toxicity to daphnia magna (Straus) in ISO medium. Nominal concentrations of 6.25, 12.5, 25, 50, 100 mg/L were investigated in the main study. Concurrent control ran. Measured test item concentrations remained within ±20% of the nominal over the test period of 48 hours, therefore the biological results are based on the nominal concentrations.

20 animals, divided into 4 replicates of 5 animals each were exposed to test concentrations or run as control for 48 hours of exposure.

For determination of the test item concentrations, samples were taken from each concentration level and control at the start and at the end of the test. Concentrations were determined using HPLC method with UV detection.

Toxic effects were expressed as EC10, EC20, EC50, NOEC and LOEC for 48 hour test period.

For determination of ECx values, Proibit analysis was used, while NOEC and LOEC were determined by Jonckheere-Terpstra Test.

All validity criteria were met and therefore the study can be considered as valid.

EC50 (48h) = 51.642 mg/l

NOEC (48h) = 25 mg/l

LOEC (48h) = 50 mg/l

Description of key information

EC50 (48h) = 51.642 mg/l.

NOEC (48h) = 25 mg/l.

LOEC (48h) = 50 mg/l.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
51.642 mg/L

Additional information

Toxicity to aquatic invertebrates was tested according to OECD guideline 202.

Nominal concentrations of 6.25, 12.5, 25, 50, 100 mg/l were tested in the main study. Concurrent control ran. Measured test item concentrations remained within ±20 % of the nominal over the test period of 48 hours, therefore the biological results are based on the nominal concentrations.

Twenty animals, divided into 4 replicates of 5 animals each were exposed to the test concentrations or run as control for 48 hours of exposure.

Analysis of concentrations was done by HPLC method with UV detection.

Toxic effects in terms of mobility of daphnia were seen.