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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From July 23 to 25, 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Qualifier:
according to
Guideline:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: crystalline
Specific details on test material used for the study:
Test Item name: Dehydrocholic Acid
Batch No.: 2018110457
Appearance: White microcrystalline powder
Expiry date: 30 November 2023
Storage: Room temperature (15 - 25 ºC)

Sampling and analysis

Analytical monitoring:
no

Test solutions

Vehicle:
yes
Remarks:
water
Details on test solutions:
Based on the results of the preliminary test, a single test concentration at 100 mg/L nominal concentration and one control group were tested in a limit test. Biological results and endpoints are based on the nominal concentration.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Twenty animals, divided into four replicates (glass beaker; volume app. 50 mL) of five animals each were exposed to the test concentration or run as control for 48 hours in a static test.
Origin: Laboratory of Hydrobiology (Central Agricultural Office, Directorate of Plant-, and Soil Protection) Hungary
Age of animals: Less than 24 h old at the beginning of the test.
Sex: Female
Animal health: Apparently healthy animals were used in this test with a known history
Acclimatization: Test animals were bred under similar (or the same) conditions as that used during the exposure period (holding water, temperature, background colour etc.), therefore additional acclimatisation before the test was not necessary.
Food and Feeding: The Daphnia were fed with concentrated algal suspension of Raphidocelis subcapitata during the holding. Test animals were not fed during the exposure.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Post exposure observation period:
The immobility or mortality of the Daphnia was determined by visual observation after 24 and 48 hours of exposure.

Test conditions

Hardness:
The reconstituted water (ISO medium) had an approximate theoretical total hardness of 249 mg/L (as CaCO3).
Test temperature:
between 18*C and 22*C with a maximum deviation of * 1*C. As minimum, the ambient temperature was measured continuously with a min/max thermometer. Water temperature was additionally determined in test vessels (at the start and at the end of the test).
pH:
The pH should not vary by more than 1.5 units in any one test and should be within the range 6-9.
The pH was measured in the control and in the test concentration at the start and at the end of the experiment.
Dissolved oxygen:
The oxygen concentration should be * 3 mg/L
The dissolved oxygen concentration was measured in the control and in the test concentration at the start and at the end of the experiment.
Nominal and measured concentrations:
In order to select appropriate test concentrations for use in the definitive test, one non-GLP preliminary range-finding test was conducted to determine the approximate toxicity of the test item.
In the non-GLP preliminary test ten daphnids (divided into 2 replicates) in the test concentrations of 0.1, 1.0, 10.0, 100.0 mg/L and in the control were exposed for 48 hours respectively to check whether the test item had any toxic effects up to the limit concentration.
For preparation of test solutions an amount of 0.0300 g test item was dissolved in 300 mL ISO medium to obtain the 100 mg/L nominal concentration. Thereafter this stock solution was placed into an ultrasonic bath for approximately 10 minutes. The test solutions of subsequent lower test concentrations were prepared by appropriate diluting of this stock solution.
No toxic effects were observed during the preliminary test, therefore, a single concentration of 100 mg/L (limit test concentration) and a concurrent control were tested in the main study.
All biological results are based on the nominal concentration.

Details on test conditions:
The test solution used in the test was prepared by mechanical dispersion without any solubilizing agent. An amount of 0.0406 g of test item was dispersed in 406 mL dilution water (ISO medium; see 5.4) in order to obtain the concentration of 100 mg/L. This solution was placed into an ultrasonic bath for 2x approx. 10 minutes. The test solution was freshly prepared in the testing laboratory just before introduction of test animals.
Twenty animals, divided into four replicates (glass beaker; volume app. 50 mL) of five animals each were exposed to the test concentration or run as control for 48 hours. The test animals were not fed during the test.
The dilution water was not aerated prior to the addition of the test item and test animals. Each test vessel contained approximately 40 mL of test solution and was uniquely identified with study number, test group and replicate.
The immobility or mortality of the Daphnia was determined by visual observation after 24 and 48 hours of exposure. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile.
In addition to immobility, any abnormal behaviour or appearance is reported.

Results and discussion

Effect concentrationsopen allclose all
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:
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:
No immobilization was observed in the control group (< 10 %) and the dissolved oxygen concentration at the end of the test in control and test vessels was more than 3 mg/L in all cases. All validity criteria were within acceptable limits and therefore the study can be considered as valid.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
In this 48-hour acute toxicity test on Daphnia magna, the obtained results showed that the test item Dehydrocholic Acid had no toxic effects on the mobility of the daphnids up to the limit concentration of 100 mg/L.
All validity criteria were met. The results are based on the nominal test item concentration (100 mg/L) and were determined directly from the raw data.
The 48-h NOEC was determined to be 100 mg/L and the 48-h LOEC and the EC50 values were determined to be > 100 mg/L (directly from the raw data).