Isooctadecanoic acid, monoester with propane-1,2-diol

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:

16 - 19 Jan 2018

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)

Version / remarks:

Adopted 23 March 2006

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

29 December 1992

GLP compliance:

yes (incl. QA statement)

Remarks:

The department of health of the government of the United Kingdom, Brixham, UK

Analytical monitoring:

yes

Remarks:

LC-MS/MS

Details on sampling:

- Concentrations: culture medium control, solvent control and nominal concentrations of 0.0625, 0.125, 0.25, 0.50 and 1.0 mg/L
- Sampling method: At study start, samples were taken from excess test solutions and at study end from the test vessels of the dilution water control, solvent control and each test concentration. At 0-h, an aliquot was removed from each of the excess dosing solutions (three aliquots for 0.125 mg/L) and an equivalent volume of THF was added (resulting in a ×2 dilution). The sample was mixed and an aliquot transferred to an LCMS vial for analysis. At 72-h sampling occasion, an aliquot was removed from a single test vessel for each concentration (three test vessels for 0.125 mg/L) and an equivalent volume of THF was added (resulting in a ×2 dilution). The sample was mixed and an aliquot transferred to an LCMS vial for analysis.

Vehicle:

yes

Remarks:

Dimethylformamide (DMF)

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A primary stock concentrate of the substance, with a nominal concentration of 10 g/L, was prepared by adding a nominal 0.100 g of test substance (actual weight 0.10056 g) and making up to 10 mL with the solvent Dimethylformamide (DMF) in a volumetric flask. The stock was shaken to mix and observed to be clear and colorless. The stock was used to prepare the test solutions by direct addition of the appropriate amount, using a microliter syringe, to stirring dilution water in a volumetric flask.
- Differential loading: yes
- Controls: The solvent control was prepared in the same way as the test concentrations using solvent only
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Dimethylformamide (DMF)
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): 0.1 mL/L
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: CCAP 278/4
- Source (laboratory, culture collection): laboratory culture
- Age of inoculum (at test initiation): 4-day old culture, in the exponential growth phase
- Method of cultivation: same as test

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

21.7 - 22.2 °C

pH:

7.29 - 7.83

Nominal and measured concentrations:

Nominal: control, solvent control, 0.0625, 0.125, 0.25, 0.50 and 1.0 mg/L
Mean measured:

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL glass conical flasks
- Type (delete if not applicable): closed with foam bungs
- Fill volume: 100 mL
- Initial cells density: 0.5 × 10E+04 cells/mL
- Control end cells density: Control: 38.93 × 10E+04 cells/mL, Solvent control: 39.62 × 10E+04 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): 6
- Other: One blank vessel (without algal inoculum) was incubated concurrently for each control and test concentration sampling occasion.

GROWTH MEDIUM
- Standard medium used: yes, AAP-medium according to guideline

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to guideline
- Culture medium different from test medium: no
- Intervals of water quality measurement: The pH of the control and test solutions was measured at the start of the test, using the excess remaining after filling the test vessels. At the end of the test the pH of one replicate vessel (containing algae) from each test concentration was determined. The temperature of the incubator was measured daily. The light intensity was measured once during the study, in each of four representative positions, using a photometer.

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: "cool-white" illumination, 6380 lx
- Other: orbital shaking at 160 rpm

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Algal cell densities of the inoculum and test cultures were determined after 24, 48, and 72 h by electronic particle counting, using a Coulter counter and counting between a lower and upper threshold equivalent spherical diameter of approx. 2.3 and 5.0 μm, respectively.
- Other: At the end of the test microscopic observations were made on samples taken from a single replicate of the control and each test substance concentration.

TEST CONCENTRATIONS
- Range finding study: yes, non-GLP
- Test concentrations: culture medium control, solvent control and nominal test concentrations of 0.0625, 0.125, 0.25, 0.50 and 1.0 mg/L (without analytical confirmation of the test concentrations)
- Results used to determine the conditions for the definitive study: No toxicity effects were observed in any concentration or control

Reference substance (positive control):

not specified

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 0.59 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 0.59 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes, 79.4 and 80.9 fold biomass increase over the 72 h for the control and solvent control respectively.
- Observation of abnormalities (for algal test): The microscopic observations, made at the end of the test, showed that compared to the control the algal cells sampled from the solvent control and each test concentration appeared normal.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: The stock was observed to be clear and colorless.
- Effect concentrations exceeding solubility of substance in test medium: No effects observed.

BIOLOGICAL RESULTS:

Table 1: Algal cell particle density.

Nominal concentration (mg/L)	Measured concentration (mg/L)	Replicate	Algal cell particle density (10E+4 cells/mL)
			24 h	48 h	72 h
Culture medium control	0	A	2.47	9.59	37.9
		B	2.25	9.6	34.3
		C	2.85	10.6	39.9
		D	2.74	10.4	40.5
		E	2.89	10.3	40.5
		F	2.81	10.3	40.5
		Mean	2.67	10.13	38.93
Solvent control	0	A	2.8	10.6	40.1
		B	2.78	10.2	35.9
		C	2.85	10.3	40.2
		D	3.05	10.6	41.4
		E	2.78	10.2	40.9
		F	2.66	9.99	39.2
		Mean	2.82	10.32	39.62
0.0625	0.036	A	2.81	10.6	41
		B	3.02	10.6	39.6
		C	2.82	10	39
		Mean	2.88	10.4	39.87
0.125	0.068	A	2.87	10.8	39.2
		B	2.94	10.2	37.9
		C	2.93	9.93	36.8
		Mean	2.91	10.31	37.97
0.25	0.15	A	2.97	10.3	38
		B	2.95	10.1	38.8
		C	3.06	10.5	36.1
		Mean	2.99	10.3	37.63
0.5	0.29	A	3	10.5	38.2
		B	2.91	11.1	39.3
		C	2.93	10.1	35.2
		Mean	2.95	10.57	37.57
1	0.59	A	3.02	10.2	39.4
		B	3.01	10.1	39.9
		C	2.62	11	37
		Mean	2.88	10.43	38.77

Table 2: Mean growth rates over the test period.

Measured concentration (mg/L)	Mean growth rate/day (0-72 hours)	Mean growth rate/day 95% Cl	Percentage of solvent control (%)
0	1.457	1.434 - 1.481	100
0	1.464	1.446 - 1.482	-
0.036	1.466	1.445 - 1.487	100
0.068	1.45	1.423 - 1.476	99
0.15	1.447	1.416 - 1.477	99
0.29	1.446	1.399 - 1.493	99
0.59	1.457	1.423 - 1.490	100

Further effect values reported in the study:

- LOEC > 0.59

- ErC20 > 0.59

- EyC50 > 0.59

- EyC20 > 0.59

- EyC10 > 0.59

ANALYTICAL RESULTS:

Table 3: Analytical results.

Nominal concentration (mg/L)	Measured concentration				Mean measured concentration (mg/L)	Mean measured concentration (%)
	0 h		72 h
	(mg/L)	% of nominal	(mg/L)	% of nominal
Control	<LOQ	-	<LOQ	-	0	-
Solvent Control	<LOQ	-	<LOQ	-	0	-
0.0625	0.72	115	<LOQ	-	0.036	58
0.125	0.14^a	109	<LOQ^b	-	0.068	55
0.25	0.29	118	<LOQ	-	0.15	59
0.5	0.59	117	<LOQ	-	0.29	59
1	1.2	118	<LOQ	-	0.59	59

a Mean of triplicate analyses: 0.14, 0.13, 0.14 mg/L

b Mean of triplicate analyses: <LOQ, <LOQ, <LOQ mg/L

The limit of quantification (LOQ) in this study was 0.04 mg/L for all test solutions. The instrument LOQ was 0.02 mg/L but during analysis, samples from the control and each test concentration were diluted ×2, doubling the LOQ.

VALIDITY CRITERIA:

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.	79.4 and 80.9 fold biomass increase over the 72 hours for the control and solvent control respectively.	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%	14% for the control and 17% for the solvent control.	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%.	1.5% for the control and 1.2% for the solvent control.	yes

Validity criteria fulfilled:

yes

Remarks:

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

Description of key information

ErC50 (72 h) = 0.59 mg/L (arithm. mean meas.) for Pseudokirchneriella subcapitata (OECD 201)

Additional information

One study is available testing the toxicity of the substance on algae. The study (2018) was conducted according to the OECD guideline 201 and under GLP standards. Pseudokirchneriella subcapitata was exposed for 72 h to the nominal test item concentrations of 0.0625, 0.125, 0.25, 0.50 and 1.0 mg/L (nominal). Since the substance is poorly soluble in water a non-GLP solubility trial was performed prior to test start with different preparation methods of the stock solution. On this basis, dimethylformamide (DMF) was used as a solvent in a final concentration of 0.1 mL/L for the preparation of the final test solutions. Therefore, a dilution water and a solvent control were included in the test. All solutions were clear and colourless at test start. An analytical monitoring took place and all tested concentrations including the control and the solvent control were measured via LC-MS/MS at the beginning and at the end of the test.

 

Since the recoveries of the concentrations were < 80% the results were expressed in terms of mean measured concentrations. The study did not result any effects on the growth rate or yield of the green algae. Thus, an ErC50 (72 h) > 0.59 mg/L and an ErC10 (72 h) > 0.59 mg/L (arithm. mean meas.) were derived. All validity criteria given by the guideline were fulfilled.