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Ecotoxicological information

Toxicity to aquatic plants other than algae

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Reference
Endpoint:
toxicity to aquatic plants other than algae
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From November 18th, 2016 to February 14th, 2017
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Justification for type of information:
Details for read across approach are included into the IUCLID section 13.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 221 (Lemna sp. Growth Inhibition Test)
Deviations:
yes
Remarks:
not affecting the integrity or validity of the study
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
Sample of the control and each test group were taken from the freshly prepared bulk preparation on Day 0 and from the pooled replicates on Day 7 for quantitative analysis.
All samples were stored frozen prior to analysis. Duplicate samples were taken at each occasion and stored frozen for further analysis if necessary.
Details on test solutions:
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 from which a series of dilutions was made to give further test concentrations.
Each of the prepared concentrations was inverted several times to ensure adequate mixing and homogeneity.
Test organisms (species):
Lemna minor
Details on test organisms:
TEST ORGANISM
- Source: a culture of Lemna minor was obtained from Canadian Phycological Culture Centre, University of Waterloo, Ontario, Canada.

ACCLIMATION
- Culturing media and conditions: 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:
no
Total exposure duration:
7 d
Test temperature:
24 ± 1 ºC
pH:
Test vessels: 6.7 - 9.5
Control: 6.7 - 10.1
Nominal and measured concentrations:
1.0, 3.2, 10, 32 and 100 mg/l, nominal
Details on test conditions:
TEST SYSTEM
- Test vessel: glass conical flasks were used.
- No. of colonies per vessel: 3 colonies of Lemna minor.
- No. of fronds per colony: total 10 fronds.
- No. of vessels per concentration: 3 replicates.
- No. of vessels per control: 3 replicates.

OTHER TEST CONDITIONS
- Photoperiod: constant illumination.
- Light intensity: approximately 7000 lux

EFFECT MEASURED
The number of fronds present in each test and control culture was recorded on days 0, 3, 5 and 7 along with observations on frond size, appearance, root length and number of colonies present.
In addition the dry weight of the fronds in each control and treatment group was determined on day 7. At the start of the test six replicate samples of fronds identical to those used to inoculate the test vessels were taken and the dry weight determined. At the end of the test the dry weight of colonies from each control and test vessel was determined by drying at 60 °C.

PARAMETERS MEASURED
The pH of each control and test flask was recorded on Day 0 and Day 7. The temperature and light intensity in the incubator were recorded daily.

RANGE-FINDING STUDY
- Test concentrations: 1.0, 10 and 100 mg/l.
- Replicates: 2 replicate flasks were prepared for each control and test concentration.
- Incubation conditions: at 24 ± 1 ºC under continuous illumination (intensity approximately 7000 lux) for 7 days.
- Renewal of test solution: on Days 3 and 5 the test solutions were renewed.
- Observation: recorded on days 0, 3, 5 and 7.

VALIDATION CRITERIA
The results of the test are considered valid if the following performance criterion is met: the doubling time of frond numbers in the controls must be less than 2.5 days (60 h), corresponding to approximately a seven-fold increase in seven days and an average specific growth rate of 0.275 d-1.
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:
both frond number and dry weight
Duration:
7 d
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks:
both frond number and dry weight
Details on results:
Growth Data Based on Frond Number
There were no statistically significant differences between the control, 1.0, 3.2 and 10 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of inhibition of average specific growth rates calculated from frond numbers was 10 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 32 mg/l.
ErC10 (7d): 22 mg/l, based on frond numbers
ErC20 (7d): 97 mg/l, based on frond numbers
ErC50 (7d) > 100 mg/l, based on frond numbers

Yield Based on Frond Number
There were no statistically significant differences between the control, 1.0, 3.2 and 10 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of inhibition of yield calculated from frond numbers was 10 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 32 mg/l.
EyC10 (7d): 3.7 mg/l, based on frond numbers
EyC20 (7d): 16 mg/l, based on frond numbers
EyC50 (7d) > 100 mg/l, based on frond numbers

Growth Data Based on Dry Weight
There were no statistically significant differences between the control, 1.0 and 3.2 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of inhibition of average specific growth rate calculated from dry weight was 3.2 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 10 mg/l.
ErC10 (7d): 7.3 mg/l, based on dry weight
ErC20 (7d): 88 mg/l, based on dry weight
ErC50 (7d) > 100 mg/l, based on dry weight

Yield Based on Dry Weight
There were no statistically significant differences between the control, 1.0 and 3.2 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of inhibition of yield calculated from dry weight was 3.2 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 10 mg/l.
EyC10 (7d): 2.3 mg/l, based on dry weight
EyC20 (7d): 4.4 mg/l, based on dry weight
EyC50 (7d) > 100 mg/l, based on dry weight

OBSERVATIONS
A notable reduction in root length was observed in the 10, 32 and 100 mg/L test cultures.

MEASURED CONCENTRATIONS
Chemical analysis of the test preparations on Day 0 and Day 7 showed near nominal concentrations were obtained for all samples with the exception of the 1.0 mg/l test preparation on Day 7 where a measured concentration of 71 % of nominal was obtained. Given that the mean measured concentration at this level was 83 % of nominal, and that this concentration was below the No Observed Effect Concentration, this decline was considered to have had no adverse effect and as such the results have been calculated based on nominal test concentrations only.

RANGE-FINDING STUDY
The results no significant effect on growth at 1.0 mg/l, however, growth was observed to be reduced at 10 and 100 mg/l.
Chemical analysis of the test preparations on Day 0 (fresh media) and Days 3 and 7 (old media) showed measured concentrations to range from 25 to 95 % of nominal. Examination of the data could find no cause for the low measured concentrations obtained from the 1.0 and 10 mg/l test preparations. What these results did however demonstrate was that the test item was stable over the test duration and hence a static design for the definitive test would be appropriate.
Validity criteria fulfilled:
yes
Remarks:
the doubling time of the control cultures was 1.83 days, in line with the OECD Guideline
Conclusions:
ErC50 (7d) > 100 mg/l (nominal), based on both frond numbers and dry weight
EyC50 (7d) > 100 mg/l (nominal), based on both frond numbers and dry weight
Executive summary:

A study was performed to assess the effect of the test item on the growth of the freshwater plant Lemna minor. The method followed that described in the OECD Guideline No. 221 “Lemna sp. Growth Inhibition Test (March 2006)”.

Following a preliminary range-finding test, Lemna minor was exposed to an aqueous solution of the test item at concentrations of 1.0, 3.2, 10, 32 and 100 mg/l (three replicate flasks per concentration) for a period of 7 days, under constant illumination at a temperature of 24 ± 1 °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 preparations on Day 0 and Day 7 showed near nominal concentrations were obtained for all samples with the exception of the 1.0 mg/l test preparation on Day 7 where a measured concentration of 71 % of nominal was obtained. Given that the mean measured concentration at this level was 83 % of nominal, and that this concentration was below the No Observed Effect Concentration, this decline was considered to have had no adverse effect.

There were no statistically significant differences between the control, 1.0, 3.2 and 10 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of both the inhibition of average specific growth rates and the inhibition of yield calculated from frond numbers was 10 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 32 mg/l.

There were no statistically significant differences between the control, 1.0 and 3.2 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of both the inhibition of average specific growth rates and the inhibition of yield calculated from dry weight was 3.2 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 10 mg/l.

Conclusion

ErC50 (7d) > 100 mg/l (nominal), based on both frond numbers and dry weight

EyC50 (7d) > 100 mg/l (nominal), based on both frond numbers and dry weight

Description of key information

Not harmful/toxic for aquatic plants

Key value for chemical safety assessment

Additional information

There are no information about the potential toxicity to aquatic plants of Direct Violet 051, therefore, the available data on structural analogous Similar Substance 01 have been taken into account. The read across approach can be considered as reliable and adequate for the purpose; details and explanations are detailed in the report attached to the IUCLID section 13.

A study was performed to assess the effect of the Similar Substance 01 on the growth of the freshwater plant Lemna minor. The method followed that described in the OECD Guideline No. 221. Following a preliminary range-finding test, Lemna minor was exposed to an aqueous solution of the test item at concentrations of 1.0, 3.2, 10, 32 and 100 mg/l (three replicate flasks per concentration) for a period of 7 days.

Chemical analysis of the test preparations on Day 0 and Day 7 showed near nominal concentrations were obtained for all samples with the exception of the 1.0 mg/l test preparation on Day 7 where a measured concentration of 71 % of nominal was obtained. Given that the mean measured concentration at this level was 83 % of nominal, and that this concentration was below the No Observed Effect Concentration, this decline was considered to have had no adverse effect.

There were no statistically significant differences between the control, 1.0, 3.2 and 10 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of both the inhibition of average specific growth rates and the inhibition of yield calculated from frond numbers was 10 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 32 mg/l.

There were no statistically significant differences between the control, 1.0 and 3.2 mg/l test concentrations (P ≥ 0.05), however all other test concentrations were significantly different (P < 0.05) and therefore the "No Observed Effect Concentration" (NOEC) in terms of both the inhibition of average specific growth rates and the inhibition of yield calculated from dry weight was 3.2 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) was 10 mg/l.