Dodecanoic acid, 2,2-dimethyl-3-oxopropyl ester

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2004-08-18 to 2005-08-12

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

adopted 2002

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

adopted 1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Sampling preparation:
Aqueous samples (10.0 mL) were membrane filtrated (Nylon, 0.45 μm). The filtrate was acidified with 2M-HCl to pH 2 – 3. 10.0 mL of the filtrate were given on the top of a column with 5.0 g Extrelute. After 30 minutes, elution was slowly performed with ethyl acetate (50 mL, 30 minutes), the eluate was rotated down to dryness and 500 μL ISTD-solution were added. At last, 400 μL of the solution were mixed with 50 resp. 100 μL SIL -Mix in vials, and silylation was performed at 80 °C for 60 minutes.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
1. Experiment:
- Method: As solubility lies below 100 mg/L, the water-accommodated fraction was prepared for the test. This was done by weighing the nominal load of 100 mg/L, adding the corresponding amount of deionized water and shaking vigorously for 24 hours. The resulting solution was filtrated through 0.45 μm filters.
- Differential loading: 100 mg/L
- Controls: six replicates (deionized water with nutrient medium and algae)
- Replicates: six replicates
2. Experiment:
- Method: The water-accommodated fraction was prepared for the test. This was done by weighing the nominal load of 100 mg/L, adding the corresponding amount of deionized water and stirring slowly for 2 hours. The solution was left to stand for 15 minutes. Then the solution was filtrated.
- Differential loading: 100 mg/L
- Controls: six replicates (deionized water with nutrient medium and algae)
- Replicates: three replicates
3. Experiment:
- Method: A stock solution containing 1 g/L in acetone was prepared. This solution was used to prepare the treatment. 100 μL of the acetonic stock solution per litre were used.
- Differential loading: 1 mg/L
- Controls: six replicates (deionized water with 100 μL/L acetone, nutrient medium and algae)
- Replicates: six replicates for the treatment, additional three replicates of control and treatment for analytical determination after 4 hours
4. Experiment:
- Method: A stock solution containing 1 g/L in acetone was prepared. This solution was used to prepare the treatments to be tested. 100 μL of the acetonic stock solution per litre were used at the most.
- Differential loading: 1 mg/L
- Controls: six replicates (deionized water with 100 μL/L acetone, nutrient medium and algae)
- Replicates: three replicates for each treatment, additional three replicates of control and each treatment for analytical determination after 4 hours

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: Desmodesmus subspicatus
- Strain: CHODAT
- Source: Institut für Pflanzenphysiologie of Universität Göttingen
- Method of cultivation: Four days before the start of each test, an aliquot of the stock culture containing a few cells was brought into 50 mL pre-culture medium and incubated for 96 hours. The resulting culture is growing exponentially. After adjustment to a cell concentration of about 5E04/mL by photometric measurement and addition of pre-culture medium, the culture was usable for the test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

1. Experiment: 24 - 25 °C
2. Experiment: 23 - 25 °C
3. Experiment: 24 °C

pH:

1. Experiment: 8.9 to 9.6
2. Experiment: 8.5 to 9.4
3. Experiment: 8.6 to 9.4

Nominal and measured concentrations:

1. Experiment 100 mg/L
2. Experiment 100 mg/L
3. Experiment 1 mg/L
4. Experiment 0.1 mg/L; 0.18 mg/L; 0.32 mg/L, 0.56 mg/L and 1 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: screw cap cuvettes d=6mm
- Initial cells density: 5E04 cells/mL
- No. of vessels per concentration (replicates): six replicates
- No. of vessels per control (replicates): three to six replicates

GROWTH MEDIUM
- in accordance to the guideline

TEST MEDIUM / WATER PARAMETERS
- in accordance to the guideline

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Light intensity and quality: 8000 ± 2000 Lux

EFFECT PARAMETERS MEASURED
- Determination of cell concentrations: photometer

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.18 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

0.32 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Details on results:

1. Experiment: The test solution went flocculent during the test, probably caused by undissolved parts of the test item. Therefore the WAF for the second experiment was prepared by stirring instead of shaking.
2. Experiment: Because of the test item’s physico-chemical properties causing film-formation at the top of the water phase, the preparation of the WAF is not very reproducible. Therefore for the third experiment, a stock solution in acetone was prepared and the test solutions were prepared by spiking the necessary amount of deionized water with this stock solution.
3. Experiment: As the algal growth was inhibited, a further experiment using five concentrations of the test item in a geometric series was performed.
4. Experiment: Only a slight inhibition could be observed. As a dose-response correlation is visible, this test was used to determine the results.

Reported statistics and error estimates:

Determination of the NOEC
The difference between the treatment 0.18 mg/L and the control can be considered as not significant (level of significance: 97.5 %) as all calculated t-values were smaller than the limit of significance. Therefore the concentration 0.18 mg/L is stated as NOEC.
Determination of the LOEC
As the difference between the treatment 0.32 mg/L and the control can be considered as significant, the treatment 0.32 mg/L can be stated as LOEC.

3. Experiment:

At the beginning and 4 hours after the start of the test, the content of the test item in each test solution was determined using GC. The measured concentration of the fresh treatment at the start of the test was 32 % of the nominal concentration. Sample preparation and clean-up requires about one hour. During this time, the test item already undergoes hydrolysis which probably caused this low recovery. The measured concentration after 4 hours was below the limit of quantification. The test item is not stable in water (as has been demonstrated in the respective study “determination of water solubility”). Due to the rapid degradation, referring to determined values is not meaningful and only the nominal values were used for the determination of the biological results.

4. Experiment:

At the begin and 4 hours after the start of the test, the content of the test item in each test solution was determined using GC. The measured concentrations of the freshly prepared test solutions (at the start of the test) were between 27 and 30 % of the nominal concentrations. The three lower treatments lay below the limit of quantification. Sample preparation and clean-up requires about one hour. During this time, the test item already undergoes hydrolysis which probably caused these low recoveries. After 4 hours, the test item could be detected only in the highest treatment. The measured concentration was 8 % of the nominal concentration. The test item is not stable in water (as has been demonstrated in the respective study “water solubility determination”). Due to the fast degradation referring to determined values is not meaningful and only the nominal values were used.

Validity criteria fulfilled:

yes

Conclusions:

The 72 h EC50 was determined to be > 1 mg/L and the 72 h NOEC was 0.18 mg/L.

Executive summary:

The study was performed in order to evaluate the toxic potential of 2,2-Dimethyl-3-lauroyloxy-propanal the toxic potential of test item towards Desmodesmus subspicatus CHODAT, an unicellular freshwater green alga. The incubation period of 72 hours corresponds to a subchronic test, as during this period, about five cell divisions take place. Four experiments were performed. As the test item is poorly water soluble, the “water-accommodated fraction” was tested in the first experiment. This was done by weighing the nominal load 100 mg/L, adding the corresponding amount of dilution water and shaking vigorously for 24 hours. The resulting solution was filtrated through 0.45μm filters. The treatment was used to incubate the unicellular freshwater green alga Desmodesmus subspicatus for a period of 72 hours. The cell concentration of each replicate was determined by measuring the absorption of the cuvettes at 440 nm every 24 hours with a spectral photometer. With these measured values, the number of cells was calculated (linear correlation between cell concentration and absorption given). Then the growth rate, the area under the growth curve (AUC1) and the yield2 were determined. The test solution went flocculent during the test probably caused by undissolved parts of the test item. The second experiment was performed likewise, but with stirring over 2 hours instead of shaking. The resulting solution was left to stand for about 15 minutes, then the solution was filtrated. No inhibition was observed. The test item forms a surface film, thus the preparation of the WAF is not very reproducible. Therefore for the third experiment, a stock solution in acetone was prepared. The treatment concentration of 1 mg/L was prepared by spiking the corresponding amount of deionized water with this stock solution. The treatment concentration of 1 mg/L is considered to be well above maximal water solubility of 0.227 mg/L. 25 % inhibition of growth rate, 72 % inhibition of AUC, and 73 % inhibition of yield were observed. At the beginning and after 4 hours of the test, the content of the test item and its oxidation product (carbonic acid) in each test solution was determined using GC. Based on these experiments, the fourth experiment was performed using five concentrations between 0.1 and 1 mg/L. The test solutions were prepared by spiking with a stock solution in acetone. No inhibition occurred in the two lower concentrations. The three higher concentrations inhibited the algal growth only slightly. At the beginning and 4 hours after the start of the test, the content of the test item in each test solution was determined using GC. The measured concentrations of the freshly prepared test solutions (at the start of the test) were between 27 and 30 % of the nominal concentrations. The three lower treatments were below the limit of quantification. Sample preparation and clean-up requires about one hour. During this time, the test item already undergoes hydrolysis which caused these low recoveries. After 4 hours remaining test item was only detected in the highest concentration. The measured concentration was 8 % of the nominal concentration. The test item is not stable in water (as has been demonstrated in the respective study “water solubility determination”). Due to the fast degradation referring to determined values is not meaningful and only the nominal values were used. The 72 h EC50 was determined to be > 1 mg/L and the 72 h NOEC was 0.18 mg/L.These data were already submitted in a NONS dossier under Directive 92/32/EEC (notification number 05-04-1922-00 from 2005-10-25) and considered valid and uncritical from the German Competent Authority (BAUA).

Description of key information

Aldehyde L was assessed in a toxicity to algae study according to EU-method C.3 and OECD guideline 201. The 72 h EC50 was determined to be > 1 mg/L and the 72 h NOEC was 0.18 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:

1 mg/L

EC10 or NOEC for freshwater algae:

0.18 mg/L

Additional information

The study was performed in order to evaluate the toxic potential of 2,2-Dimethyl-3-lauroyloxy-propanal the toxic potential of test item towards Desmodesmus subspicatus CHODAT, an unicellular freshwater green alga. The incubation period of 72 hours corresponds to a subchronic test, as during this period, about five cell divisions take place. Four experiments were performed. As the test item is poorly water soluble, the “water-accommodated fraction” was tested in the first experiment. This was done by weighing the nominal load 100 mg/L, adding the corresponding amount of dilution water and shaking vigorously for 24 hours. The resulting solution was filtrated through 0.45μm filters. The treatment was used to incubate the unicellular freshwater green alga Desmodesmus subspicatus for a period of 72 hours. The cell concentration of each replicate was determined by measuring the absorption of the cuvettes at 440 nm every 24 hours with a spectral photometer. With these measured values, the number of cells was calculated (linear correlation between cell concentration and absorption given). Then the growth rate, the area under the growth curve (AUC1) and the yield2 were determined. The test solution went flocculent during the test probably caused by undissolved parts of the test item. The second experiment was performed likewise, but with stirring over 2 hours instead of shaking. The resulting solution was left to stand for about 15 minutes, then the solution was filtrated. No inhibition was observed. The test item forms a surface film, thus the preparation of the WAF is not very reproducible. Therefore for the third experiment, a stock solution in acetone was prepared. The treatment concentration of 1 mg/L was prepared by spiking the corresponding amount of deionized water with this stock solution. The treatment concentration of 1 mg/L is considered to be well above maximal water solubility of 0.227 mg/L. 25 % inhibition of growth rate, 72 % inhibition of AUC, and 73 % inhibition of yield were observed. At the beginning and after 4 hours of the test, the content of the test item and its oxidation product (carbonic acid) in each test solution was determined using GC. Based on these experiments, the fourth experiment was performed using five concentrations between 0.1 and 1 mg/L. The test solutions were prepared by spiking with a stock solution in acetone. No inhibition occurred in the two lower concentrations. The three higher concentrations inhibited the algal growth only slightly. At the beginning and 4 hours after the start of the test, the content of the test item in each test solution was determined using GC. The measured concentrations of the freshly prepared test solutions (at the start of the test) were between 27 and 30 % of the nominal concentrations. The three lower treatments were below the limit of quantification. Sample preparation and clean-up requires about one hour. During this time, the test item already undergoes hydrolysis which caused these low recoveries. After 4 hours remaining test item was only detected in the highest concentration. The measured concentration was 8 % of the nominal concentration. The test item is not stable in water (as has been demonstrated in the respective study “water solubility determination”). Due to the fast degradation referring to determined values is not meaningful and only the nominal values were used. The 72 h EC50 was determined to be > 1 mg/L and the 72 h NOEC was 0.18 mg/L.These data were already submitted in a NONS dossier under Directive 92/32/EEC (notification number 05-04-1922-00 from 2005-10-25) and considered valid and uncritical from the German Competent Authority (BAUA).