Dodecanoic acid, ester with 1,2,3-propanetriol

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:

February 28 – March 03, 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Remarks:

Ministerium für Arbeit, Integration und Soziales des Landes Nordrhein-Westfalen

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Samples (at least 2 x about 5 mL) were taken from all five test solutions and controls at the beginning of the test prior to addition of the algae (2 x 6 samples, analysis and retain samples). At the end of the growth test, samples were taken directly from pooled replicates per test concentration and controls (2 x 6 samples). For stabilisation of the samples, all samples were given into 5 mL of methanole.
- Sample storage conditions before analysis: If samples could not be measured on the day of sampling, they were immediately stored at-20°C until analysis. All samples will be discarded after finalisation of the study.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: For the highest test concentration 20.012 mg of the test item were added to sterile growth medium and treated in an ultrasonic bath at about 30 °C for about 6 hours to facilitate the suspension of the test item. Then the suspension was stirred vigorously at about 30 °C using a magnetic stirrer for 17.5 hours. The temperature was thereby slightly increased from room temperature to about 30 °C to promote the solution behavior of the test substance. For 1.5 hours the suspension cooled down to room temperature.
- Differential loading: Individual dilutions from the highest test concentration were prepared.
- Controls: yes, test medium
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): There were no visible residues.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Source (laboratory, culture collection): SAG, Culture Collection of Algae at Pflanzenphysiologisches Institut of the University at Göttingen, Albrecht von Haller Institut, Göttingen, Catalog No 61.81 SAG
- Method of cultivation: The stock cultures are maintained fulfilling the criteria of the OECD guideline (culture medium recommended by Bringmann und Kühn (1980).

ACCLIMATION
- Acclimation period: Prior to testing a pre-culture was established in standard OECD growth medium to obtain exponentially-growing algae for the test. The culture duration of the pre-cultures was 4 days.
- Culturing media and conditions (same as test or not): same as test, sterilised synthetic growth medium according to OECD 201
- Any deformed or abnormal cells observed: no (see table under "Any other information on results")

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

21°C ± 0°C

pH:

controls: 7.45 at test start; 8.23 at test end
test concentrations: 7.30 - 7.54 at test start, 8.09 - 8.55 at test end

Nominal and measured concentrations:

nominal: 0.201, 0.634, 2.00, 6.33 and 20.0 mg/L
measured: 0.058, 0.180, 0.854, 2.58 and 6.83 mg/L (geometric mean)

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL conical glass flasks covered with air-permeable silicone-sponge caps
- Material, size, headspace, fill volume: 100 mL fill volume
- Initial cells density: 10000 cells/mL
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 8

GROWTH MEDIUM
- Standard medium used: yes, sterilised synthetic growth medium according to OECD 201

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: purified water processed using an ELGA „PURELAB Ultra“
- Culture medium different from test medium: yes, culture medium recommended by Bringmann und Kühn (1980)
- Intervals of water quality measurement: The solution pH was measured in one test vessel per test concentration including the control. At the end of the test, the solution pH was measured directly in all individual test vessels. The test temperature was measured in one additional test vessel. The culture vessels were randomly placed and repositioned daily in the incubator.

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Light intensity and quality: 114.93 to 116.66 μE m-2 s-1 (OSRAM Standard “cool white”); the light intensity was measured daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: During the test, the cell concentrations were determined after 24, 48 and 72 h in samples taken (electronic particle counter (CASY 1 Model TT, OMNI Life Science GmbH & Co. KG, Bremen, Germany)
directly from the test vessels.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.16
- Range finding study
- Test concentrations: 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: The highest test concentration was hazy or milky. The dilution of the highest test concentration to 10 mg/L resulted in a clear test media. The growth rate inhibition after 72 hours was 4.6, 58 and 100% compared to controls. The inhibition of yield after 72 hours was 20, 95 and 100% compared to controls.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 6.83 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 6.83 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.:

2.58 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.:

> 6.83 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

3.57 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

2.58 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no (see table under "Any other information on results")

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50: The latest (March 2017) ErC50 value of 2.29 mg/L (nominal; 95% confidence limits: 2.12 – 2.48 mg/L) for inhibition of growth rate is in good agreement with the results of an international ring test with an ErC50 of 3.38 ± 1.30 mg/L.

A slight concentration-dependent inhibiting effect on the growth of the freshwater green algae was observed over the range of the tested concentrations. However, no effects on growth rate were determined > 10% and no effects on yield above 50% were observed.

The measured concentrations at the end of the test were below 80% of the measured initial concentrations. Therefore, the evaluation of the results was based on the geometric mean measured concentrations during exposure.

Table 1: Measured test item concentrations of test solutions.

Nominal test item conc. [mg/L]	Measured conc-day 0 [mg/L]	Measured conc. day 3 [mg/L]	% of nominal at d0	% of nominal at d3	% of initial at d3	Geometric mean measured conc. [mg/L]
Control	<LOQ	<LOQ	-	-	-	-
0.201	0.175	0.019	87.3	9.53	10.9	0.058
0.634	0.570	0.057	89.8	9.02	10.0	0.180
2.00	1.95	0.375	97.3	18.8	19.3	0.854
6.33	5.14	1.30	81.1	20.5	25.2	2.58
20.0	17.5	2.66	87.7	13.3	15.2	6.83

 

Table 2: Inhibition of growth rate and yield compared to controls after 72 hours.

Geometric mean measured conc. [mg/L]	Growth rate		Yield
	Growth rate [1/d]	% Inhibition	Yield [cells x 104/mL]	% Inhibition
Control	1.751	0.0	190.7	0.0
0.058	1.748	0.1 (-)	188.8	1.0 (-)
0.180	1.727	1.3 (-)	177.1	7.1 (-)
0.854	1.736	0.8 (-)	182.1	4.5 (-)
2.58	1.729	1.2 (-)	179.6	5.8 (-)
6.83	1.605	8.3 (+)	123.6	35.2 (+)

(-) not significantly different from controls, (+) significantly different from controls, Williams` Multiple Sequential t-Test, significance level 0.05, one-sided smaller.

Table 3: Appearance of the inoculum culture and appearance of the algae

Date	Arithmetic mean measured conc. [mg/L]	Observations
Test start	Normal appearance of inoculum culture for all test concentrations.
Test end, day 3	Control	Normal appearance of algal cells.
	0.058	Normal appearance of algal cells.
	0.180	Normal appearance of algal cells.
	0.854	Normal appearance of algal cells.
	2.58	Normal appearance of algal cells.
	6.83	Normal appearance of algal cells.

Table 4: Validity criteria for OECD 201.

Criterion from the guideline	Outcome	Validity criterion fulfilled
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period.	The cell number in the control cultures increased by a factor of 191.7 within the 72 hour test period.	yes
The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures must not exceed 35%	The mean of the replicate coefficients of variations in the section-by-section growth rate of controls was 4.6 % during the test period.	yes
The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% in tests with Pseudokirchneriella subcapitata and Desmodesmus subspicatus. For other less frequently tested species, the value should not exceed 10%.	The coefficient of variation of average specific growth rate at test end in replicate control cultures was 1.8 %	yes

 

 

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

ErC50 (72h) > 6.83 mg/L (meas. geom mean) for Pseudokirchneriella subcapitata (OECD 201)

NOEC (72h) = 2.58 mg/L (Meas. geom. mean) for Pseudokirchneriella subcapitata (OECD 201)

Key value for chemical safety assessment

EC50 for freshwater algae:

6.83 mg/L

EC10 or NOEC for freshwater algae:

2.58 mg/L

Additional information

One experimental study is available investigating the toxicity of the substance Dodecanoic acid, ester with 1,2,3-propanetriol (CAS 37318-95-9) to aquatic freshwater algae. The study was conducted according to OECD 201 (GLP) using the freshwater algae Pseudokirchneriella subcapitata. The algae cells were exposed for 72h to five test concentrations of 0.201, 0.634, 2.00, 6.33 and 20.0 mg/L (nominal).The concentrations of the test item in the media were verified by analytical measurement at test start and test termination. At test start the measured concentrations were 0.18, 0.57, 1.95, 5.14 and 17.5 mg test item/L (81.1 – 97.3% of nominal). After 72 hours the measured test item concentrations were 0.019, 0.057, 0.38, 1.30 and 2.67 mg test item/L (9.02 – 20.5% of nominal; 10.0 – 25.2% of initial). Thus, the evaluation of the test was based on the geometric mean measured concentrations (0.058, 0.180, 0.854, 2.58 and 6.83 mg/L).A slight concentration-dependent inhibiting effect on the growth of the freshwater green algae was observed over the range of the tested concentrations. However, no effects above 50% were observed. An EC10/EC50 (72h) > 6.83 mg/L (meas. geom. mean) and a NOEC = 2.58 mg/L (meas. geom. mean) were determined.