LUMIPOD

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31 July 2020

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

OECD Guideline 201 "Water Quality - Freshwater Algae (Unicellular Green Algae) Growth Inhibition Test" (May 2012).

Deviations:

no

GLP compliance:

no

Remarks:

OECD 201 GLP test is in progress.

Analytical monitoring:

yes

Remarks:

An organic carbon measurement was performed for the stock solution and two test concentrations in order to estimate the amount of test element present in the test solutions and its stability throughout the period of time exposure.

Details on sampling:

The test element considered to be slightly soluble in water, solubilization tests were carried out in order to optimize the dissolution.
These tests were carried out in different media (ultrapure water, algae test media) and the solubilization rate was estimated on the amount of organic carbon measured in the aqueous phase.
The various tests carried out to solubilize the test item showed very similar results.
Indeed, whatever the stirring time, the test medium used, the percentage of solubilized organic carbon was between 75 and 78% compared to the carbon content measured in the test item.
At the highest concentration tested, 500 mg/L, the percentage of solubilized organic carbon was identical to the other test solutions. However, at this concentration, the texture of the prepared solution was not suitable for the ecotoxicity tests. Thus, the maximum concentration tested was limited to 200 mg/L.
The TOX20022 being composed of 80 to 100% lipoamino acid, the follow-up of the solubilized organic carbon content shows that a significant part of the element is present in the test solutions but does not allow to distinguish the part of lipoamino acid dissolved from to the other constituents potentially present in the tested product (which may represent up to 20% of the product).
In addition, it seemed that a part of the test item was absorbed, among other things, on the bar magnet used to perform the magnetic stirring.
In the remainder of the report, the test concentrations will therefore be presented in nominal concentrations of TOX20022.

Vehicle:

no

Details on test solutions:

Range of nominal concentrations (in mg/L): 2.90 - 6.40 - 14.1 - 31.0 - 68.2 - 150
Control: 6 vials containing 25 mL of growth medium and algal inoculum (0.8 * 104 cells/mL)
Replicates: 3 vials per concentration, containing 25 mL of test solution and algal inoculum (0.8 * 104 cells/mL)
Negative controls: 1 vial per concentration, containing 25 mL of test solution without algal inoculum and 1 vial containing 25 mL of growth medium without algal inoculum (Blank)

Test organisms (species):

other: Raphidocelis subcapitata (formerly called Pseudokirchneriella subcapitata), strain CCAP 278/4, in exponential growth phase

Details on test organisms:

Raphidocelis subcapitata (formerly called Pseudokirchneriella subcapitata), strain CCAP 278/4, in exponential growth phase

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

Temperature, 22 ± 2°C

pH:

7.7-7.8

Dissolved oxygen:

8.8-8.9 mg/L

Conductivity:

675-676 µS/cm

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass vials containing 25 mL of test solution
- Aeration: None within the test period
- Renewal rate of test solution (frequency/flow rate): None
- Initial cells density: 0.8 x 10^4/ mL
- Control end cells density: 59.96 x 10^4/ mL
- Negative controls: 1 vial per concentration, containing 25 mL of test solution without algal inoculum and 1 vial containing 25 mL of growth medium without algal inoculum (Blank)
- No. of vessels per concentration (replicates): 3 vials per concentration, containing 25 mL of test solution and algal inoculum (0.8 * 10^4cells/mL)
- No. of vessels per control (replicates): 6 vials containing 25 mL of growth medium and algal inoculum (0.8 * 10^4 cells/mL)

GROWTH MEDIUM
- Standard medium used: yes OECD growth medium

OTHER TEST CONDITIONS
Temperature, 22 ± 2°C; continuous illumination (5190 - 5490 Lux), constant agitation (120 rpm).

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Daily: algae cell counting with the particle counter

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
- Test concentrations: 2.90 - 6.40 - 14.1 - 31.0 - 68.2 - 150 mg/L (nominal)

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 150 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

A measurement of the organic carbon was performed on the stock solution and two test concentrations to verify the stability of the sample (based on the organic carbon content present in the test solutions).
To follow-up the organic carbon in the test solutions, additional vessels, without algae, were prepared and placed under the test conditions.
The results obtained during this test are presented in Table below.

Table: Measurement of organic carbon (mg O.C./L) in test solutions

Nominal test concentration (mg/L)	Theoretical O.C. (mg/L)	Solubilized O.C (mg O.C./L)/ % Day 0	Solubilized O.C (mg O.C./L)/ % Day 1	Solubilized O.C (mg O.C./L)/ % Day 2	Solubilized O.C (mg O.C./L)/ % Day 3	O.C. decrease at the end of the test (%)
31.0	18.0	14.2/ 78.6	13.6/ 75.5	13.7/ 76.2	13.6/ 75.5	4.2
150	87.2	75.4/ 86.5	72.0/ 82.7	71.7/ 82.3	68.2/ 78.3	9.5
200	116.2	99.0/ 85.2	91.8/ 79.0	92.3/ 79.4	85.2 73.4	13.9

The amount of solubilized organic carbon measured in the test solutions is between 78.6 and 86.5% and the O.C. remain relatively stable over 72 hours under test conditions (greater than 73%).

Table: Growth rate and inhibition of algae growth after 72 hours

Nominal test concentration (mg/L)	Growth rate (days-1)	Growth rate inhibition (%)
Control	1.438	/
2.90	1.422	1.1
6.40	1.435	0.2
14.1	1.447	- 0.6
31.0	1.443	- 0.3
68.2	1.439	- 0.1
150.0	1.406	2.2

No significant inhibition of growth on Raphidocelis subcapitata was observed during this test at the
nominal concentration of 150 mg/L in TOX20022 (maximum concentration tested).

 

Validity criteria fulfilled:

yes

Conclusions:

No significant inhibition of growth on Raphidocelis subcapitata was observed during this test at the
nominal concentration of 150 mg/L in the registered substance (maximum concentration tested). ErC50 is higher than 150 mg/L (nominal concentation).

Description of key information

No significant inhibition of growth on Raphidocelis subcapitata was observed during this test at the nominal concentration of 150 mg/L in the registered substance (maximum concentration tested). ErC50 is higher than 150 mg/L (nominal concentation).

Key value for chemical safety assessment

EC50 for freshwater algae:

150 mg/L