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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:
2019-04-16 to 2019-07-11
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
2004
Deviations:
no
Qualifier:
according to
Guideline:
other: OECD Series on Testing and Assessment, No. 23, "Guidance Document on Aqueous-phase Aquatic Toxicity Testing of Difficult Test Chemicals", 2nd Ed., February 08, 2019.
Version / remarks:
2019
Deviations:
no
Qualifier:
according to
Guideline:
other: SANCO/3029/99 rev.4 11/07/00: Resiues: Guidance for generating and reporting methods of analysis in support of preregistration data requirements for Annex II (part A; Section 4) an Annex III (part A; Section 5) of directive 91/414
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Control and 100 mg/L.
- Sampling method: Duplicate samples from the freshly prepared media of the only test concentration and the control were taken at the start of the test and at day 1. For the determination of the stability of the test item under the test conditions and of the maintenance of the only test item concentration during the test period, duplicate samples from the aged test media of the only test concentration and the control were collected at day 1 (after 24 hours of exposure) and at the end of the test by pouring togehter the contents of the test beakers of each treatment. All samples were adjusted to pH 3 with HCl, and remained undiluted until analysis.
- Sample storage conditions before analysis: All samples were stored in a freezer (≤-20 °C), protected from light, until analysis was performed. Afterwards the samples were again stored deep frozen (≤ -20 °C) and will be kept stored up to the date of the final report.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION:
The test medium of the only test item concentration of nominal 100 mg/L was prepared by dissolving 69.8 and 80.2 mg test item into 698 and 802 mL test water, respectively by intense stirring for 20 minutes. The solution was sterilized by membrane filtration (cellulose acetate, 0.2 µm). The test media was prepared just before introduction of the daphnids (start of the test) and test medium renewal on Day 1. All glass vessels were sterilized by dry heating (180°C, 3 h) prior to use.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: clone 5
- Age at study initiation: 1.5 to 19.0 hours old
- Method of breeding: The daphnids were bred in the laboratories of ibacon under similar temperature and light conditions as used in the test. The cultivation of the parental daphnids was performed in Elendt M4 medium. The test organisms were not first brood progeny. The daphnids in the stock culture were fed at least on all working days with green algae (Desmodesmus subspicatus) freshly grown in the laboratories of ibacon.
- Source: The daphnids introduced in the test were taken from ibacon's in-house laboratory culture
- Feeding during test : No

ACCLIMATION: Was not necessary, since the test was performed in the same medium as the culturing.
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
2.5 mmol/L (250 mg/L) as CaCO3
Test temperature:
20.5 to 21.9 °C
pH:
7.8 to 8.0
Dissolved oxygen:
8.7 to 9.0 mg/L
Salinity:
Not applicable
Conductivity:
No data
Nominal and measured concentrations:
Nominal: Control and 100 mg test item/L
Measured: The concentrations of the test item were within ±20% of the nominal concentrations during the test. Thus, all reported results refer to nominal values.
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass beaker of 110 mL volume containing approx. 60 mL test medium
- Type: closed . Glass beakers were kept close during the renewal periods with a conical glass stopper to reduce the loss of water due to evaporation and to minimize the entry of dust and bacterial into the solution.
- Volume of solution: 60 mL
- Renewal rate of test solution: Test medium renewal on Day 1
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: According to guideline

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: day 1: 24 h light and day 2: 16 h light - 8 h dark
- Light intensity: 440 to 850 lux

EFFECT PARAMETERS MEASURED: The mobility of the daphnids was determined by visual observation after 24 and 48 hours. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).

VEHICLE CONTROL PERFORMED: no
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
After 48 hours of exposure no immobilisation of the test animals was observed in the control and in the only test concentration of nominal 100 mg test item/L.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Relevant effect levels:
48-h EC50 = 0.672 (95% CI: 0.598-0.757)

Validity criteria of the study:

Control Immobilisation Rate: Was 0% and furthermore no daphnids showed signs of disease or stress; thus the validity criterion was met.

Dissolved Oxygen Concentration: Was 8.8 mg O2/L in all treatment groups at the end of the test; thus validity criterion was met.

Analytical results:

The quantification of the test item in the test samples was performed using liquid chromatography with UV detection. In the freshly prepared test media at the start of the test and at the renewal of the test media 97% of the nominal test concentrations were found. In the aged test media after 24 and 48 hours test duration, 96% of the nominal value was determined. During the test the daphnids were exposed to a mean of 96% of nominal.

Table 1: Summary of analytical results

Nominal

[mg test item/L]

% of nominal1

RSD

[%]

n

Control

n.a.

n.a.

8

100

96

0.5

8

 1mean value of all measured samples per treatment group

RSD: relative standard deviation per treatment group

n: number of analysed samples

n.a.: not applicable

Validity criteria fulfilled:
yes
Conclusions:
The acute immobilisation test with Daphnia magna was performed according to OECD TG 202. Under the conditons given, no immobilisation of the test animals was observed in the control and in the only test item concentration of 100 mg test item/L after 48 hours of exposure. NOEC was determined to be at least 100 mg test item/L.The 48-hour LOEC and the 48-hour EC50 were clearly higher than 100 mg test item/L.
Executive summary:

The acute toxicity of the test item to Daphnia magna was determined in a semi-static Immobilisation test according to OECD TG 202. Young daphnids (<24 hours) were exposed in a semi-static test for 48 hours to the only concentration of nominal 100 mg/L under defined conditions for 48 hours. The recorded effects were the mobility of the daphnids after 24 and 48 hour. After 48 hours of exposure no immobilisation of the test animals was observed in the control and in the only test item concentration of 100 mg test item/L. The 48-hour NOEC was determined to be at least 100 mg test item/L. The NOEC might even be higher than this concentration, but concentrations in excess of nominal 100 mg test item/L have not been tested according to the recommendations of the test guidelines. The 48-hour LOEC and the 48-hour EC50 were clearly higher than 100 mg test item/L. These values could not be quantified due to the absence of toxicity of the test item up to nominal 100 mg test item/L. The initial concentrations and the maintenance of the exposure concentratons during the test were verified analytically. All reported results refer to nominal values since the concentrations of the test item were within ±20% of the nominal concentrations during the test.

Description of key information

The acute immobilisation test with Daphnia magna was performed according to OECD TG 202. Under the conditons given, no immobilisation of the test animals was observed in the control and in the only test item concentration of 100 mg test item/L after 48 hours of exposure. NOEC was determined to be at least 100 mg test item/L.The 48 -hour LOEC and the 48-hour EC50 were clearly higher than 100 mg test item/L (reference 6.1.3-1).

Key value for chemical safety assessment

Additional information

The acute toxicity of the test item to Daphnia magna was determined in a semi-static Immobilisation test according to OECD TG 202. Young daphnids (<24 hours) were exposed in a semi-static test for 48 hours to the only concentration of nominal 100 mg/L under defined conditions for 48 hours. The recorded effects were the mobility of the daphnids after 24 and 48 hour. After 48 hours of exposure no immobilisation of the test animals was observed in the control and in the only test item concentration of 100 mg test item/L. The 48-hour NOEC was determined to be at least 100 mg test item/L. The NOEC might even be higher than this concentration, but concentrations in excess of nominal 100 mg test item/L have not been tested according to the recommendations of the test guidelines. The 48-hour LOEC and the 48-hour EC50 were clearly higher than 100 mg test item/L. These values could not be quantified due to the absence of toxicity of the test item up to nominal 100 mg test item/L. The initial concentrations and the maintenance of the exposure concentratons during the test were verified analytically. All reported results refer to nominal values since the concentrations of the test item were within ±20% of the nominal concentrations during the test (reference 6.1.3-1).