(2S)-2-amino-3-[3,5-diiodo-4-(4-methoxyphenoxy)phenyl]propanoic acid

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 March 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Name: O-METHYL-L-DIIODOTHYRONIN
Batch No.: B488840
Appearance: white, powder
Manufacturing date: September 25, 2018
Expiry date: September 24, 2020
Storage condition: at room temperature, protected from light

Analytical monitoring:

yes

Details on sampling:

The actual concentration of the test substance was determined at the start and at the end of a 72 hours incubation period in filtered samples of the cultures exposed to the test substance and also in one blank containing 100.0 mg test substance per L test medium (nominal concentration), but no algae.
Sampling and analysis date:
February 05, 2019 Start of experiment
February 08, 2019 End of experiment
Six replicate samples were analysed from the test solutions at the start and at the end of the experiment.
Six replicate samples were analysed from the control.

Vehicle:

no

Details on test solutions:

The test solution used in the test was prepared by mechanical dispersion without using of any solubilising agent.
The test item is poorly water soluble material. For preparation of the test solution a supersaturated solution (100 mg/L nominal loading) was first prepared by dispersing/dissolving an amount of 0.0806 g test item was dissolved in 806 mL dilution water (OECD Medium). This solution was agitated by orbital shaker (~300-350 rpm) for a period of at least 2 days (approx. 4 days in this test) and then the non-dissolved test material was separated by filtration through a membrane filter (0.45 μm) in order to obtain the saturated test solution. Untreated control ran parallel in the test.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

Species: Raphidocelis subcapitata (formerly known as Pseudokirchneriella subcapitata)
Strain number: 61.81 SAG (identical strains: CCAP 278/4; UTEX 1648; ATCC 22662)
Origin: The algae were supplied by the SAG: Collection of Algal Cultures, Inst. Plant Physiology, University of Göttingen, Untere Karspüle 2, D-37073 Göttingen, Germany
Breeding Conditions: The stock cultures are small algal cultures that are planted on agar regularly. These are transferred to fresh medium at least once every two months under standardized conditions according to the test guidelines.
Pre-culturing: The pre-culture is intended to give an amount of algae suitable for the inoculation of test cultures. The pre-culture was prepared with OECD Medium, incubated under the conditions of the test and used when still exponentially growing, normally after an incubation period of 2 - 4 days. (The pre-culture was incubated for four days at this test.) The algal cultures used in this study did not contain deformed or abnormal cells.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Hardness:

OECD medium, prepared in the laboratory of TOXI-COOP ZRT.

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 22.5 and 22.9 °C measured in the flask and in the range of 21.4 – 23.6 °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 range of the pH was 7.71 – 9.08 during the experiment. The pH of the control medium was not increased by more than 1.5 units during the test.

Nominal and measured concentrations:

The test solution used in the test was prepared by mechanical dispersion without using of any solubilising agent.
The test item is poorly water soluble material. For preparation of the test solution a supersaturated solution (100 mg/L nominal loading) was first prepared by dispersing/dissolving an amount of 0.0806 g test item was dissolved in 806 mL dilution water (OECD Medium). This solution was agitated by orbital shaker (~300-350 rpm) for a period of at least 2 days (approx. 4 days in this test) and then the non-dissolved test material was separated by filtration through a membrane filter (0.45 μm) in order to obtain the saturated test solution. Untreated control ran parallel in the test.

Details on test conditions:

Dilution water: OECD medium, prepared in the laboratory of TOXI-COOP ZRT.
Concentrations: Limit test; using a single concentration at saturation (corresponding to 100 mg/L nominal concentration); concurrent control ran.
The exposure concentration of the saturated test solution was analytically determined to be 0.57 mg/L (calculated as the geometric mean of start and end 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 six replicates in the treatment group and 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.
Analytics: Concentration of the test item were determined using HPLC-UV method at the start and at the end of the test.
Endpoints: EC10, EC20, EC50, NOEC and LOEC for both response variables (i.e. average specific growth rate and yield).
Statistics: Statistical comparisons of average specific growth rates and yield in control and in treated group were carried out using Independent Samples T-Test (2-tailed; α = 0.05) by SPSS software.
Validity:All validity criteria were met and therefore, the study can be considered as valid

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 0.57 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat. (dissolved fraction)

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.:

> 0.57 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr. (dissolved fraction)

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

Details on results:

The growth inhibition calculated for the 72 hours exposure in treatment group compared to the control was 1.9 % for the average specific growth rate and 7.9 % for the yield.
Based on the results of the statistical evaluation (T-Test; 2-tailed; α = 0.05) neither the average specific growth rate nor the yield showed significant difference compared to the control.
Accordingly, the 72-h NOEC based on growth rates and yield was determined as the saturation concentration (equivalent to 100 mg/L nominal concentration; 0.57 mg/L measured).

Validity criteria fulfilled:

yes

Conclusions:

Endpoints (0-72 h) Growth rate (r) Yield (y)
EC10 > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal) > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)
EC20 > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal) > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)
EC50 > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal) > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)
NOEC 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal) 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)
LOEC > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal) > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

Executive summary:

In this 72-h algal growth inhibition test with Raphidocelis subcapitata, the obtained results showed that the test item O-METHYL-L-DIIODOTHYRONIN had no toxic effect at aquatic saturation (measured value: 0.57 mg/L; equivalent to 100 mg/L nominal concentration). Accordingly, the 72-h EC50 value was determined to be > 0.57 mg/L (equivalent to 100 mg/L nominal concentration). The 72-h NOEC was determined to be 0.57 mg/L (equivalent to 100 mg/L nominal concentration).

All validity criteria were met.

Description of key information

Endpoints (0-72 h)       Growth rate (r)                            Yield (y)

EC10              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

EC20              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

EC50              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

NOEC              0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)              0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

LOEC              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)              > 0.57 mg/L (saturation) (equivalent to > 100 mg/L nominal)

Key value for chemical safety assessment

EC50 for freshwater algae:

100 mg/L