Registration Dossier

Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2019
Reliability:
1 (reliable without restriction)

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
23 March 2006, Annex 5 corrected 28 July 2011)
Deviations:
no
Principles of method if other than guideline:
Test item: Acetylglycyrrhetinic acid
Test species: Raphidocelis subcapitata (formerly known as Pseudokirchneriella subcapitata and Selenastrum capricornutum)); Strain number: 61.81 SAG. Exponentially-growing cultures; preculture was incubated for four days before the test
Dilution water: OECD medium, prepared in the laboratory of TOXI-COOP ZRT
Concentrations: Based on the preliminary test nominal concentrations of 6.25, 12.5, 25.0, 50.0 and 100.0 mg/L were investigated in the main study. Concurrent control ran. Biological results and endpoints are based on the nominal concentrations.
Test design: Exponentially-growing cultures of Raphidocelis subcapitata were exposed to the test item under defined conditions. The algal growth in relation to a control culture was determined over a fixed test period of 72 hours and, thus, over several algal generations. The test design included three replicates at each test concentration and six replicates for the untreated control. The alga cell concentration was approximately 104 cells/mL at the start of the test in all of the test cultures. Glass flasks with total capacity of 250 mL were used as test vessels. The volume of the test liquid in the vessels was 100 mL. The alga cell concentration was determined by manual cell counting by microscope in each testing flask during the 72-hour test, in 24-hour intervals
Analyticals: According to the Sponsor’s request there was no analytical measurements conducted
Endpoints: EC10, EC20, EC50, NOEC and LOEC for both response variables (i.e. average specific growth rate and yield).
Statistics: For the determination of the LOEC and NOEC statistical analysis was not necessary the values were determined directly from the raw data, the ECx values were also given directly from the raw data.
Validity: All validity criteria were met and therefore the study can be considered as valid
Results: All biological results are based on the nominal concentrations
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid

Test solutions

Vehicle:
yes
Details on test solutions:
Reconstituted algal growth medium (OECD medium, according to OECD 201) was used as dilution water in the experiment

Test organisms

Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)

Study design

Water media type:
other: OECD medium, according to OECD 201
Limit test:
no
Total exposure duration:
72 h

Test conditions

Test temperature:
Culture temperature was checked at the beginning of the study and each day thereafter in a flask filled with water, in the climatic chamber. In addition temperature was continuously measured (with a min/max thermometer) within the climate chamber. The temperature was between 22.7 and 23.2°C measured in the flask and in the range of 21.8– 23.8°C measured within the climate chamber.
pH:
The pH was measured in each test group at the start (before test solutions had been distributed into the test vessels) and in each test vessel at the end of the test. The pH of the control medium was not increased by more than 1.5 units during the test. The range of the pH was 7.66 – 9.94 during the experiment.
Nominal and measured concentrations:
The following nominal concentrations were tested: 6.25, 12.5, 25.0, 50.0 and 100.0 mg/L.

Details on test conditions:
Exponentially-growing cultures of Raphidocelis subcapitata were exposed to the test item under defined conditions. The algal growth in relation to a control culture was determined over a fixed test period of 72 hours and, thus, over several algal generations. The test design included three replicates at each test concentration and six replicates for the untreated control. The alga cell concentration was approximately 104 cells/mL at the start of the test in all of the test cultures. Glass flasks with total capacity of 250 mL were used as test vessels. The volume of the test liquid in the vessels was 100 mL. The alga cell concentration was determined by manual cell counting by microscope in each testing flask during the 72-hour test, in 24-hour intervals.

Validity of the Test

The cell density in the control cultures increased by a factor of 56.67 within 72 hours. This corresponds to a specific growth rate of 1.34 day-1.
The mean coefficient of variation (CV) for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72 h-tests) in the control cultures did not exceed 35 %
- CV for section-by-section growth rate day 0-1: 31.15 %
- CV for section-by-section growth rate day 1-2: 24.02 %
- CV for section-by-section growth rate day 2-3: 5.93 %
The mean coefficient of variation for section-by-section specific growth rates: 20.37 %.
The coefficient of variation of average specific growth rates during the whole test period (day 0-3) in the control cultures was 5.20 %
All validity criteria were met, therefore the study can be considered as valid.
Reference substance (positive control):
yes

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Key result
Duration:
72 h
Dose descriptor:
EC20
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Key result
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield
Details on results:
Abnormal appearance of the algal cells was not observed in the control and in the test item treated group during the experiment.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
All validity criteria were met and therefore the study can be considered as valid. Biological endpoints of the study are summarised in the Result