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Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
06 February 2018 - 30 April 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
other: OECD Guideline No. 221 “Lemna sp. Growth Inhibition Test (March 2006)”
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Source Client, Batch no 021340
- Expiration date of the lot/batch: 01 July 2022
- Purity test date: not stated

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: Poorly soluble, WAF's prepared
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.10, 0.32, 1.0, 3.2, 10 and 32 mg/L loading rate WAF
- Sampling method: Samples were taken from the control and each loading rate WAF test group from the freshly prepared bulk test preparation on Days 0, 2 and 5 and from the corresponding old or expired pooled replicates on Days 2, 5 and 7 for quantitative analysis.
- Sample storage conditions before analysis: All samples were stored frozen prior to analysis. Duplicate samples were taken at each occasion and stored frozen for further analysis if necessary.
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Nominal amounts of test item (20 and 200 mg) were each separately added to the surface of 2 liters of culture medium to give the 10 and 100 mg/L loading rates respectively. After the addition of the test item, the culture medium was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixtures allowed to stand for 1-Hour.

Visual observations made on the WAFs indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAFs by filtering through a glass wool plug.
Test organisms (species):
other: Lemna Gibba
Details on test organisms:
TEST ORGANISM
A culture of Lemna gibba was supplied by Bayer CropScience AG, 40789 Monheim, Germany in 2007. Cultures were maintained in the laboratory by the periodic replenishment of culture medium. The culture was maintained in the laboratory at a temperature of 24 ± 1 ºC under continuous illumination (intensity approximately 7000 lux) for at least 7 days prior to the start of the test.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
7 d
Remarks on exposure duration:
not specified
Post exposure observation period:
not specified
Hardness:
not specified
Test temperature:
24 ± 1 °C
pH:
5.4 - 6.6
Dissolved oxygen:
not specified
Salinity:
not specified
Conductivity:
not specified
Nominal and measured concentrations:
Nominal: 0.10, 0.32, 1.0, 3.2, 10 and 32 mg/L
Measured: Chemical analysis of the freshly prepared test preparations on Days 0, 3 and 5 showed measured test concentrations to range from less than the limit of quantification (LOQ) of the analytical method, determined to be 0.0020 mg/L, to 0.18 mg/L. Analysis of the corresponding old or expired test preparations on Day 2, 5 and 7 showed measured test concentrations to range from less than the LOQ to 0.12 mg/L.
Details on test conditions:
TEST SYSTEM
As in the range-finding tests 500 ml glass conical flasks were used. Three flasks each containing 250 mL of solution were prepared for the control and each treatment group.
The control group was maintained under identical conditions but not exposed to the test item.
Each control and test flask was inoculated with 3 colonies of Lemna gibba (total 12 fronds). The flasks were then incubated at 24 ± 1 ºC under constant illumination (intensity approximately 7000 lux) on a black, non-reflective surface for 7 days.
On Days 2 and 5 the test solutions were renewed.

TEST CONCENTRATIONS
- Range finding study: 10 and 100 mg/L
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test: 0.10, 0.32, 1.0, 3.2, 10 and 32 mg/L loading rate WAF.
Reference substance (positive control):
yes
Key result
Duration:
7 d
Dose descriptor:
other: See table 1
Effect conc.:
ca. 0.32 - ca. 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Table 1
Details on results:
Details on results
- Exponential growth in the control (for algal test): n/a
- Observation of abnormalities (for algal test): none
- Unusual cell shape: slight gibbosity above 3.2 mg/L
- Colour differences: some colourosis above 0.32 mg/L
Results with reference substance (positive control):
- Results with reference substance valid? yes

- EC50:
8.7 mg/L Frond number for growth rate
8.9 mg/L Dry weight for growth rate
6.4 mg/L Frond number for yield
6.0 mg/L Dry weight for yield

NOEC:
1.0 mg/L Frond number for growth rate
4.0 mg/L Dry weight for growth rate
1.0 mg/L Frond number for yield
1.0 mg/L Dry weight for yield

LOEC:
2.0 mg/L Frond number for growth rate
8.0 mg/L Dry weight for growth rate
2.0 mg/L Frond number for yield
2.0 mg/L Dry weight for yield
Reported statistics and error estimates:
Frond number

Average Specific Growth Rate:There were no statistically significant differences between the control, 0.10 and 0.32 mg/L loading rate WAFs (P≥0.05), however all other loading rates were significantly different (P<0.05) and therefore the NOEL in terms of inhibition of average specific growth rates calculated from frond numbers was 0.32 mg/L loading rate WAF. Correspondingly the LOEL was 1.0 mg/L loading rate WAF.

Yield: There were no statistically significant differences between the control and 0.10 mg/L loading rate WAF (P≥0.05), however all other loading rates were significantly different (P<0.05) and therefore the NOEL in terms of inhibition of average specific growth rates calculated from yield was 0.10mg/L loading rate WAF. Correspondingly the LOEL was 0.32 mg/L loading rate WAF.


Dry weight

Average Specific Growth Rate: There were no statistically significant differences between the control and 0.10 mg/L loading rate WAF (P≥0.05), however all other loading rates were significantly different (P<0.05) and therefore the NOEL in terms of inhibition of average specific growth rate calculated from dry weight was 0.10 mg/L loading rate WAF. Correspondingly the LOEL was determined to be 0.32 mg/L loading rate WAF.

Yield: There were no statistically significant differences between the control and 0.10 mg/L loading rate WAF (P≥0.05), however all other loading rates were significantly different (P<0.05) and therefore the NOEL in terms of inhibition of yield calculated from dry weight was 0.10 mg/L loading rate WAF. Correspondingly the LOEL was determined to be 0.32 mg/L loading rate WAF.

Table 1. Effect of PR81:5 on Lemna gibba.

Response Variable

Measurement Variable

EL50(mg/L)

No Observed Effect Loading Rate (NOEL) (mg/L)

Lowest Observed Effect Loading Rate (LOEL) (mg/L)

Average Specific Growth Rate

 

Frond number

38

0.32

1.0

Dry weight

 

>100

0.1

0.32

Yield

Frond number

4.4

0.1

0.32

Dry weight

 

2.1

0.1

0.32

Validity criteria fulfilled:
yes
Conclusions:
The effect of the test item on the growth of Lemna gibba has been investigated over a 7-Day period and gave the following results:
- EL50:
38 mg/L Frond number for growth rate
>100 mg/L Dry weight for growth rate
4.4 mg/L Frond number for yield
2.1 mg/L Dry weight for yield

NOEL:
0.32 mg/L Frond number for growth rate
0.1 mg/L Dry weight for growth rate
0.1 mg/L Frond number for yield
0.1 mg/L Dry weight for yield

LOEL:
1.0 mg/L Frond number for growth rate
0.32 mg/L Dry weight for growth rate
0.32 mg/L Frond number for yield
0.32 mg/L Dry weight for yield

Description of key information

Key value for chemical safety assessment

EC50 for freshwater algae:
38 mg/L
EC10 or NOEC for freshwater algae:
0.32 mg/L

Additional information