Ruthenium, dichloro[1,3-dihydro-4,5-dimethyl-1,3-bis(2,4,6-trimethylphenyl)-2H-imidazol-2-ylidene](2-thienylmethylene)(tricyclohexylphosphine)-, (SP-5-41)-

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:

2016-06-22 to 2016-10-20

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

slowly decomposition in water

Analytical monitoring:

yes

Details on sampling:

At the start of the test (oh), the control and following test solutions: 6.25; 25; and 100 mg/l were sampled ( each group: 3 samples of 10 ml)
After 24 and 48 h exposure, only the 2 additional vessels (control; 6.25; 25; 100 mg/l) for chemical analysis were sampled (each group: 2 samples of 10 ml)
At the end of the test (72h), one replicate of the test vessels and two additional vessels from control; 6.25; 25; and 100 mg/l were sampled (each group: 3 samples of 10 ml)
The samples were filtered (0.45µm syring filter), filled in plastic centrifuge tubes and stored (<2weeks) in the fridge (5°C +-3°C). Of each treatement, one of the samples from 0h; 24 h; 48; and 72 h was sent to the analytical laboratory and analyzed. The remaining samples were stored as retain samples in the fridge until finalization of the study

Vehicle:

no

Details on test solutions:

The stock solution was prepared by adding 111.0 mg test item (including a factor of 1.11 to take into account the dilution caused by addition of the algal inoculum) to 1000 mll OECD TG 201 medium and shaking for 24 h using an overhead shaker at 21.9-22.4 °C in the dark. As the test substance is a very poorly soluble solid, the stock solution was filtered through a fibre glass filter with retaining range till 0.6 µm. The filter was prepared by rinsing with purified water and preconditioning with ca 250 ml stock solution (which was discarded), to reduce adsorption of the test item. The negative control (OECD 201 medium) was treated in the same way as the test substance solution. The corresponding volume of the test solution and algal inoculum (resulting in a methodical dilution by a factor of 1.11 and a nominal start concentration of 7X10³ cells/ml) were transfered to the test vessels. Four additional vessels without test organisms were prepared for treatment group, control; 6.25; 25; and 100 mg/l, for the accompanying chemical analysis.

Test organisms (species):

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

Details on test organisms:

The test organism Pseudokirchneriella subcapitata, strain No. 61.81 SAG, originates from the Culture Collection of Algaes at the University of Goettingen and was cultivated in suspension culture. The strain used for the study has been cultured at the Lab, twice a week, the stock suspension is inoculated into fresh Holm-Hansen medium under axenic conditions to keep it in exponential growth.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

OECD 201 medium were used

Test temperature:

during exposure: 23.0 - 23.5 °C

pH:

control: 7.8 - 7.9
test medium: 7.8 - 8.0

Salinity:

not applicable

Nominal and measured concentrations:

nominal concentrations: 0; 6.25; 12.5; 25; 50; and 100 mg/l

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer glass flasks 100 ml
- Type (delete if not applicable): open
- Initial cells density: a nominal start concentration of 7 x 10³ cells/ml
- Control end cells density: 570 x 10³ cells/ml
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
GROWTH MEDIUM
- Standard medium used: yes, OECD 202 medium
- Culture medium different from test medium: no
OTHER TEST CONDITIONS
- Sterile test conditions: yes/no
- Adjustment of pH: no
- Photoperiod:continuously
- Light intensity and quality: mean light intensity was 74.7µE/m/s +-4.8%, fluorescent tubes, each 58 W
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : 0h; 24; 48h; and 72h
- Determination of cell concentrations: fluorimeter
- Chlorophyll measurement: exitation wavelength of 465 nm, emision wavelength of 670 nm

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.11
- Range finding study: yes
- Test concentrations: 6.25; 12.5; 25; 50; 100 mg/l

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

50.13 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EL10

Effect conc.:

49.98 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- The morphology of the algal cells in the test loading rates as well as in the control showed no obvious abnormality
- The measured Ru-concentrations in the test substance treatments were below the limit of quantification (< 1µg/l) up to 1.6 µg/l. The Ru-concentrations in the control were below the limit of quantification

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: ErC50(72h) = 0.831 mg/l (CL 95%: 0.790-0.869 mg/l)

Reported statistics and error estimates:

statistical evaluation with ToxRat Professional 3.2.1

Table 1 Inhibition of growth rate after 24 h; 48 h and 72 h

Nominal test item Loading rate [mg/l]	Inhibition of growth rate [%]
	24 h	48 h	72 h
Control	-	-	-
6.25	39.4	28.6	16.9
12.5	8.3	13.9	9.6
25	45.9	27.3	17.5
50	39.4	30.6	21.1
100	275.9	222.1	164.1

Table 2 Inhibition of yield after 24 h; 48 h, 72 h

Nominal test item Loading rate [mg/l]	Inhibition of yield [%]
	24 h	48 h	72 h
Control	-	-	-
6.25	48.2	56.6	51.0
12.5	13.0	35.0	34.3
25	59.9	57.8	54.1
50	52.2	60.8	60.7
100	122.4	105.6	101.2

 

Validity criteria fulfilled:

yes

Conclusions:

The test substance is classified as harmful to aquatic life with long lasting effects.

Executive summary:

A valid static algae inhibition test with the test substance according to OECD Guideline under GLP was performed. The EL50 loading rate of 50.13 mg/l after 72 hours was determined.

Description of key information

The test substance is classified as harmful to aquatic life with long lasting effects.

Key value for chemical safety assessment

EC50 for freshwater algae:

50.13 mg/L

EC10 or NOEC for freshwater algae:

49.98 mg/L