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Ecotoxicological information

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:
2016-02-029 till 2016-04-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
EU Method C.3 (Algal Inhibition test)
Version / remarks:
(2009)
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Version / remarks:
(2006)
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L plus control
- Storage: Routinely, the samples were analysed immediately. Only in exceptional cases, they were stored overnight deep frozen and protected from light.

Sampling schedule:
- Control: at 72 hours
- Test concentration/s: 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L at 0, 24, 48 and 72 hours
Vehicle:
no
Details on test solutions:
Pre-treatment of α-acetyl-ɣ-butyrolacton and preparation of α-acetyl-ɣ-butyrolacton concentrations
Prior to the test the dilution water was adjusted from pH 7.9 to pH 6.0.
A stock solution was prepared to give the desired series of test concentrations. 201.0 mg of α-acetly-ɣbutyrolacton were added to 2 litres of dilution water and stirred for 1 h on a magnetic stirrer.
The pH was measured to be 6.1.
To produce the different α-acetyl-ɣ-butyrolacton concentrations appropriate amounts of the stock solution were diluted with dilution water to a volume of 100 mL and 0.532 mL of the algal inoculum was added to each replicate resulting in a final cell density of 5000 cells/mL. For each α-acetyl-ɣ-butyrolacton concentration and the control 3 replicates were prepared. All flasks were sealed with cotton stoppers.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
Test species
- Name: Desmodesmus subspicatus (formerly Scenedesmus subspicatus) Strain No. 86.81 SAG
- Source: Strain of the test species obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany).
- Maintenance and Acclimatisation: Exponentially growing stock cultures were maintained in the test facility under constant temperature conditions (21-24 °C with a maximum fluctuation of +/- 2 °C) at a light intensity in the range 60 to 120 μE x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter). The growth medium (according to BRINGMANN & KÜHN (1977) was renewed once a week. Cell density measurements were made using a microcell counter, Sysmex F300, Digitana.
- Preparation of pre cultures: Pre cultures were set up three days before the start of a test. They were grown under identical exposure conditions as the stock cultures, except from the use of a different growth medium.
- Test cultures: The algal inocula for the test were taken from an exponentially growing pre culture and were mixed with the growth medium to make up to a final cell density of about 5000 cells per millilitre in the test medium.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
no data
Test temperature:
21 - 24 °C
pH:
Control: 6.5 at 0h and 5.0 at 72 h
Concentrations 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L 6.3 -6.5 at 0 h and 4.4 -4.6 at 72 h
The pH value in α-acetyl-ɣ-butyrolacton slightly increased by more than 1.5 pH units. This increase is not regarded to by relevant to results as all valdity criteria were met.
Dissolved oxygen:
no data
Nominal and measured concentrations:
Concentrations (nominal): 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L plus control
Effective concentrations ranged from 96.9 % to 113.2 % of nominal values at 0 hours, from 42.0 to 76.4 % of nominal values at 24 hours, from 28.8 % to 71.7 % of nominal values at 48 hours and from 17.9 % to 73.7 % of nominal values at 72 hours.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
11.31 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
4.246 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
LOEC
Effect conc.:
8.276 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
The results are expressed in terms of geometric mean measured concentrations.
Effective concentrations ranged from 96.9 % to 113.2 % of nominal values at 0 hours, from 42.0 to 76.4 % of nominal values at 24 hours, from 28.8 % to 71.7 % of nominal values at 48 hours and from 17.9 % to 73.7 % of nominal values at 72 hours.

Reduction of growth rate (ErCx, NOEC [r]) is the preferred endpoint according to OECD 201 and for regulatory purposes in the EU. Results relating to yield (EyCx, NOEC [y]) were calculated to fulfil regulatory requirements in some countries (but not in the EU) and are given in the results section of this report.

Validity criteria fulfilled:
yes
Remarks:
(cell density in control cultures increased by a factor > 16; mean of the replicate coefficient of variation for section-by-section growth <35%; cenfficient of variation of the mean specific growth rate replicates in the control < 7 %)
Conclusions:
The toxicity of α-acetyl-ɣ-butyrolacton to algae (Desmodesmus subspicatus) depending on growth rate were determined at an EC10 of 11.31 mg/L, an EC50 of >100 mg/L after a period of 72 hours. A NOEC of 4.246 mg/L and a LOEC of 8.276 mg/L were calculated.
Executive summary:

The toxicity of α-acetyl-ɣ-butyrolacton to algae (Desmodesmus subspicatus) was determined according to EU method C.3 ‘Freshwater Alga and Cyanobacteria, Growth inhibition test’ (2009) which is equivalent to OECD Guideline for Testing of Chemicals No. 201 (2006). Exponentially growing algal cells were exposed for a period of 72 hours to a range of concentrations, nominally 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L of α-acetyl-ɣ-butyrolacton dissolved in dilution water. The cell densities were measured at 24 hours intervals. Inhibition of the algal population was measured as reduction in growth rate, relative to control cultures grown und identical conditions. The results are expressed in terms of geometric mean measured concentrations. Effective concentrations ranged from 96.9 % to 113.2 % of nominal values at 0 hours, from 42.0 to 76.4 % of nominal values at 24 hours, from 28.8 % to 71.7 % of nominal values at 48 hours and from 17.9 % to 73.7 % of nominal values at 72 hours. An EC10 of 11.31 mg/L and an EC50 of >100 mg/L were measured; a NOEC of 4.246 mg/L and a LOEC of 8.276 mg/L were calculated. This toxicity study is classified as acceptable and satisfies the guideline requirements for the acute algae study.

Description of key information

The toxicity of α-acetyl-ɣ-butyrolacton to algae (Desmodesmus subspicatus) was determined according to EU method C.3 ‘Freshwater Alga and Cyanobacteria, Growth inhibition test’ (2009) which is equivalent to OECD Guideline for Testing of Chemicals No. 201 (2006). Exponentially growing algal cells were exposed for a period of 72 hours to a range of concentrations, nominally 0.88, 1.9, 4.3, 9.4, 21, 45 and 100 mg/L of α-acetyl-ɣ-butyrolacton dissolved in dilution water. The cell densities were measured at 24 hours intervals. Inhibition of the algal population was measured as reduction in growth rate, relative to control cultures grown und identical conditions. The results are expressed in terms of geometric mean measured concentrations. Effective concentrations ranged from 96.9 % to 113.2 % of nominal values at 0 hours, from 42.0 to 76.4 % of nominal values at 24 hours, from 28.8 % to 71.7 % of nominal values at 48 hours and from 17.9 % to 73.7 % of nominal values at 72 hours. An EC10 of 11.31 mg/L and an EC50 of >100 mg/L were measured; a NOEC of 4.246 mg/L and a LOEC of 8.276 mg/L were calculated.

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:
11.31 mg/L

Additional information