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Toxicity to aquatic plants other than algae

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Reference
Endpoint:
toxicity to aquatic plants other than algae
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 221 (Lemna sp. Growth Inhibition Test)
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: MH-12/2014 (china)
- Expiration date of the lot/batch: 13 November 2019

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Frozen in the dark
Analytical monitoring:
not specified
Details on sampling:
- Concentrations:
Range-finding Test: 0.10, 1.0, 10 and 100 mg/L
Definitive Test: 100 mg/L
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
Range finding test:
The test concentration to be used in the definitive test was determined by a preliminary range-finding test. The range-finding test was conducted by exposing Lemna minor to a series of nominal test concentrations of 0.10, 1.0, 10 and 100 mg/L for a period of 7 days. The test was conducted in glass conical flasks (250 mL). Two replicate flasks were prepared for each control and test concentration. The test item was dissolved directly in culture medium. A nominal amount of test item (100 mg) was dissolved in culture medium and the volume adjusted to 1 liter to give a 100 mg/L test concentration from which a series of dilutions were made to give further test concentrations of 10, 1.0 and 0.10 mg/L. Each of the prepared concentrations was inverted several times to ensure adequate mixing and homogeneity. The control group was maintained under identical conditions but not exposed to the test item. At the start of the range-finding test the number of fronds present in each test and control culture was recorded along with observations on frond size, appearance, root length and number of colonies present. The flasks were then incubated at 24 ± 2 ºC under continuous illumination (intensity approximately 7000 lux) on a black, non-reflective surface for 7 days. On Days 2 and 5 the test solutions were renewed, and observations on the test organisms were recorded on Days 2, 5 and 7. In order to determine the stability of the test item under test conditions a sample of each test concentration was taken for chemical analysis on Day 0 (fresh media) and Day 5 (old media). An additional sample of each test concentration was prepared on Day 0 and incubated alongside the test until Day 7 in order to determine stability over the entire test duration. All Day 0 and Day 5 samples were stored frozen prior to analysis.

Definitive Test:
Based on the result of the range-finding test a “limit test” was conducted at a concentration of 100 mg/L to confirm that at the maximum concentration given in the OECD Test Guidelines, no effect on growth was observed.

Experimental Preparation
A nominal amount of test item (200 mg) was dissolved in culture medium and the volume adjusted to 2 liters to give the 100 mg/L test concentration. The prepared concentration was inverted several times to ensure adequate mixing and homogeneity. The concentration and stability of the test item in the test preparations were verified by chemical analysis on Day 0 and Day 7.

Exposure Conditions:
As in the range-finding test glass conical flasks were used. Six flasks each containing 250 mL of solution were prepared for the control and 100 mg/L 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 minor (total 11 fronds). The flasks were then incubated at 24 ± 2 ºC under constant illumination (intensity approximately 7000 lux) on a black, non-reflective surface for 7 days. A static testing regime was employed.

Test organisms (species):
Lemna minor
Details on test organisms:
TEST ORGANISM
- Source (laboratory, culture collection): A culture of Lemna minor was obtained from Canadian Phycological Culture Centre, University of Waterloo, Ontario, Canada. 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:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
7 d
Test temperature:
24 ± 1 ºC
pH:
6.5 - 7.6
Nominal and measured concentrations:
Range-finding Test : 0.10, 1.0, 10 and 100 mg/L
Definitive Test : 100 mg/L
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Duration:
7 d
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
Average Specific Growth Rate
Duration:
7 d
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield (Frond Number, Dry Weight)
Duration:
7 d
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
Average Specific Growth Rate
Duration:
7 d
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Yield(Frond Number, Dry Weight )
Results with reference substance (positive control):
- Results with reference substance valid: yes
- Relevant effect levels:
EC50: 3.4 mg/L (Average specific growth rate - frond number and dry weight)
EC50: 2.6 mg/L( Yield: Frond Number)
EC50: 2.7 mg/L( Yield: Dry Weight)

No Observed Effect Concentration (NOEC) (mg/L):
2.5 (Average specific growth rate - frond number)
1.25 (Average specific growth rate - dry weight)
2.5 (Yield: Frond Number)
1.25 (Yield: Dry Weight)

Lowest Observed Effect Concentration (LOEC) (mg/L):
5.0 (Average specific growth rate - frond number)
2.5 (Average specific growth rate - dry weight)
5.0 (Yield: Frond Number)
2.5 (Yield: Dry Weight)
Validity criteria fulfilled:
yes
Conclusions:
Exposure of Lemna minor to the test item based on nominal concentrations gave the following results:
EC50: >100 mg/L (Average specific growth rate - frond number and dry weight)
EC50: >100 mg/L (Yield: Frond Number, dry weight)

No Observed Effect Concentration (NOEC):
100 mg/L (Average specific growth rate - frond number)
100 mg/L (Yield: Frond Number, dry weight)
Executive summary:

A key study was performed to assess the effect of the test item on the growth of the freshwater plant Lemna minor according to the OECD Guideline 221. Following a preliminary range-finding test, Lemna minor was exposed to an aqueous solution of the test item at a concentration of 100 mg/L (six replicate flasks) for a period of 7 days, under constant illumination at a temperature of 24 ± 2 °C. The number of fronds in each control and treatment group was recorded on Days 0, 3, 5 and 7 along with observations on plant development. Chemical analysis of the test preparation on Day 0 (fresh media) and Day 7 (old media) showed measured test concentrations to range from 99 % to 102 % of nominal and so the results are based on nominal test concentrations only.

 

Exposure of Lemna minor to the test item based on nominal concentrations gave the following results:

EC50: >100 mg/L (Average specific growth rate - frond number and dry weight)

EC50: >100 mg/L( Yield::Frond Number, dry weight)

No Observed Effect Concentration (NOEC):

100 mg/L (Average specific growth rate - frond number)

100 mg/L (Yield: Frond Number, dry weight)

Description of key information

The EC50 and NOEC to Lemna were >100 mg/L and 100 mg/L respectively.

Key value for chemical safety assessment

EC50 for freshwater plants:
100 mg/L
EC10 or NOEC for freshwater plants:
100 mg/L

Additional information

A study was performed to assess the effect of the test item on the growth of the freshwater plant Lemna minor according to OECD Guideline 221. Following a preliminary range-finding test, Lemna minor was exposed to an aqueous solution of the test item at a concentration of 100 mg/L (six replicate flasks) for a period of 7 days, under constant illumination at a temperature of 24 ± 2 °C. The number of fronds in each control and treatment group was recorded on Days 0, 3, 5 and 7 along with observations on plant development. Chemical analysis of the test preparation on Day 0 (fresh media) and Day 7 (old media) showed measured test concentrations to range from 99 % to 102 % of nominal and so the results are based on nominal test concentrations only.

 

Exposure of Lemna minor to the test item based on nominal concentrations gave the following results:

EC50 : >100 mg/L (Average specific growth rate - frond number and dry weight)

EC50 : >100 mg/L( Yield: Frond Number, dry weight)

No Observed Effect Concentration (NOEC):

100 mg/L (Average specific growth rate - frond number)

100 mg/L (Yield: Frond Number, dry weight)