Dodecanoic acid, monoester with oxybis[propanediol]

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 Feb - 21 Feb 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

March 23, 2006; Annex 5 corrected July 28, 2011

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Dodecanoic acid, monoester with oxybis[propanediol]

EC Number:

619-220-9

Cas Number:

96499-68-2

Molecular formula:

not applicable, the substance is UVCB

IUPAC Name:

Dodecanoic acid, monoester with oxybis[propanediol]

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling: Samples for analysis were taken from all test concentration levels at the start of exposure, at 24 and 48 h and at the end of exposure.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 0.0585 g of the test item and 1300 mL of the test medium were mixed to prepare a stock solution with a nominal loading rate of 45.0 mg/L and stirred for 48 h. The test solutions were prepared by dilution of the stock solution with the appropriate amount of test medium.
- Eluate: no
- Differential loading: no
- Controls: blank control
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Freshwater green algae
- Strain: ATCC22662
- Source: American Type Culture Collection, supplied on 30 June, 1995
- Method of cultivation: sterile conditions; temperature: 21-24 °C; light intensity: nominal 6923 lux; continuous illumination provided by alternative LED light to fluorescent lamp

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not): same as test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Test temperature:

22.1 - 22.5 °C

pH:

7.8 - 7.9

Nominal and measured concentrations:

nominal: Control, 1.00, 3.16, 10.0, 31.6 and 100 mg/L
measured: < detection limit, 0.310, 0.979, 3.02, 10.1 and 39.6 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: sterilised 300 mL Erlenmeyer flasks with gas-permeable Silicosen
- Initial cells density: 0.75 x 10E04 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to guideline
- Culture medium different from test medium: same as test medium
- Intervals of water quality measurement: The pH value was measured at test start and end. The temperature was measured at test start, after 24 and 48 h and at test end.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: continuous light
- Light intensity and quality: 6769-7077 lux in wavelength range 400 - 700 nm; continuous illumination provided by alternative LED light to fluorescent light

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Biomass was measured every 24 hours after test start using chlorophyll fluorescence measurement (spectrofluorometer model RF-6000, Shimadzu).
- Chlorophyll measurement: yes, using a spectrofluorometer
- Other: Cell conditions were evaluated microscopically for each test concentration level at the end of exposure.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Range finding study: Two preliminary tests were performed.
Study 1: Nominal concentrations of 10.0 and 100 mg/L were chosen. Growth rate inhibition was 5.8% at the low concentration and > 100% at the high concentration.
Study 2: Nominal concentrations of 2.00, 16.0 and 50.0 mg/L were chosen. Growth rate inhibition was 1.0% at the lowest concentration, 31% at the intermediate concentration and > 100% at the highest concentration.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

13.6 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 12.4-14.9 mg/L

Effect concentrations 2

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.02 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): At 10.1 and 39.6 mg/L, some bloated cells were observed. Cells of other treatment solutions were the same as the control. For the control cells, the cell condition was normal.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: At test start and end, the test solution of 39.6 mg/L was colourless and clear with some bubbles. All other test solutions were colourless and clear at test start. At test end all other test solutions, except 39.6 mg/L, showed green colour due to algal growth.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- ErC50 (72 h) = 1.3 mg/L

Reported statistics and error estimates:

Estimation of EC50: The percentage inhibition in each exposure level was plotted as a semi-logarithmic graph against the corresponding concentration. Linear regression analysis (least square method) was carried out using the data within the range showing linearity visually (100 and 31.6% levels) to estimate the EC50 and 95% confidence limits, which were estimated from the intercept of the regression line with the parallel drawn to the absciss at 50% inhibition. The EC50 was denoted as ErC50 based on growth rate.

Estimation of NOEC: Regarding growth rate, Bartlett's test was done to determine the homogeneity of variance for the data. Then, one-way analysis of variance (ANOVA) and Dunnett's multiple comparison test was used to determine the significant difference between the control and exposure levels. Concentration levels above the ErC50 were excluded for each analysis.

Any other information on results incl. tables

Table 1: Biomass values

Values are based on the measured value of the pre-culture.

Measured concentration [mg/L]	No.	Chlorophyll fluorescence value (relative value)
		0 h	24 h	48 h	72 h
Control	A	43	200	950	3800
	B	43	180	870	3500
	C	43	190	840	3600
	D	43	180	860	3700
	E	43	190	830	3400
	F	43	190	830	3200
	Mean	43	190	860	3500
	SD	0	7.0	44	230
0.310	A	43	200	980	4100
	B	43	180	900	3500
	C	43	190	850	3300
	Mean	43	190	910	3600
	SD	0	11	64	440
0.979	A	43	210	980	3600
	B	43	200	920	3500
	C	43	190	780	3300
	Mean	43	200	890	3500
	SD	0	12	100	150
3.02	A	43	210	930	3700
	B	43	190	830	3500
	C	43	200	950	3700
	Mean	43	200	900	3600
	SD	0	10	65	130
10.1	A	43	110	400	1500
	B	43	130	430	1500
	C	43	130	460	1700
	Mean	43	120	430	1600
	SD	0	9.9	31	100
39.6	A	43	14	1.2	1.6
	B	43	13	2.0	2.0
	C	43	13	2.7	3.9
	Mean	43	13	2.0	2.5
	SD	0	0.59	0.73	1.3

Table 2: Growth rate and growth inhibition rate

Measured concentration [mg/L]	No.	Growth rate 0-72 h	Growth inhibition rate [%]
Control	A	1.49	-
	B	1.47	-
	C	1.48	-
	D	1.49	-
	E	1.46	-
	F	1.43	-
	Mean	1.47	-
	SD	0.0226	-
0.310	A	1.53	-3.8
	B	1.47	0.16
	C	1.45	1.2
	Mean	1.48	-0.82
	SD	0.0393	2.7
0.979	A	1.48	-0.54
	B	1.47	0.14
	C	1.45	1.4
	Mean	1.46	0.34
	SD	0.0147	1.0
3.02	A	1.49	-1.3
	B	1.47	0.18
	C	1.49	-1.1
	Mean	1.48	-0.74
	SD	0.0118	0.80
10.1	A	1.19	19
	B	1.20	19
	C	1.23	16
	Mean	1.21	18
	SD	0.0209	1.4
39.6	A	1.10	175
	B	-1.03	170
	C	-0.797	154
	Mean	-0.977	166
	SD	0.160	11

Table 3: Validity criteria

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.	factor of 74	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%	4.5%	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%	yes

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

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

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