A mixture of cis- and trans-cyclohexadec-8-en-1-one

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-10-09 to 2017-10-12

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:

2011

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

GC-MS method

Details on sampling:

- Sampling method: All concentration levels and the control were analytically verified via GC-MS after 0 (start of exposure) and 72 hours (end of the exposure). Separate replicates for each test item concentration and control for the test item analysis at the beginning of the exposure were prepared with algae. The samples for the test item analysis after 72 hours were prepared with algae and incubated under test conditions.
- Sample storage conditions before analysis: all samples were stored at 6 ± 2 °C until sample preparation andat room temperature until the start of the analysis (on an autosampler), if necessary.

Vehicle:

no

Details on test solutions:

A saturated solution with a concentration of 1 mL/L test item was prepared with dilution water. A slow stirring procedure was applied. Gentle stirring (to avoid formation of an emulsion) was carried out with a magnetic stirrer at room temperature for 72 hours. After a separation phase of 15 minutes at room temperature the saturated solution was taken from the bottom of the flask and was used for testing. The solution was checked after stirring via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion). The Tyndall effect was negative.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: Pseudokirchneriella subcapitata HINDÁK, SAG 61.81
- Source (laboratory, culture collection): Sammlung von Algenkulturen (SAG)
Pflanzenphysiologisches Institut der Universität Göttingen Nikolausberger Weg 18, D-37073 Göttingen
- Age of inoculum (at test initiation): A three days old preculture, prepared in dilution water, was used as inoculum.
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounted to 2567 – 5130 lux for 24 hours per day.

ACCLIMATION
- Acclimation period: not needed
- Culturing media and conditions (same as test or not): yes
- Any deformed or abnormal cells observed: no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

Nominal hardness: 0.24 mmol Ca+Mg/L

Test temperature:

21.5-22.5 °C

pH:

Start: 8.02-8.23;
End: 8.32-9.02

Dissolved oxygen:

n.a.

Salinity:

n.a.

Conductivity:

n.a.

Nominal and measured concentrations:

Nominal concentration: 1.00 - 3.16 - 10.0 - 31.6 - 100 % of the saturated solution;
Measured concentrations (geometric mean): 8.69 – 28.4 – 84.8 – 269 – 839 µg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:Sterile headspace flasks, volume: 59 mL, with aluminium tops with PTFE seals
- Type: closed
- Initial cells density: 6839 cells/mL
- Control end cells density: 1051499 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
- Nutrient medium Z according to LÜTTGE et al. (1994)

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: 8.1+/-0.2
- Photoperiod: 24 hours/day light
- Light intensity and quality:Approximately 4440 to 8880 lux, corresponding to 60 to 120 µE*m-2*s-1

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: fluorimeter (Chlorophyll a-fluorescence)


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.16
- Range finding study
- Test concentrations: 1.0-10.0-100% of saturated solution
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (SIGMA-ALDRICH, batch number MKBV0900V, purity 99.0%, CAS RN 7778-50-9)

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

470 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

119 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

28.4 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

122 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

25.3 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

28.4 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Details on results:

See below

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50: 0.535 mg/L(based on the growth rate with headspace); 0.499 mg/L(based on the growth rate without headspace);
- Valid range: 0.754 ± 0.558 mg/L

Reported statistics and error estimates:

Statistical Significance (NOEC/LOEC) – Growth Rate/ Yield
- Shapiro-Wilk´s Test on Normal Distribution
- Levene´s Test on Variance Homogeneity (with Residuals)
- Non-parametric Trend analysis by Contrasts (Monotonicity of Concentration/Response)
- Step-down Jonckheere-Terpstra Test Procedure

Biological Data:

Geometric mean measured test item concentration	Replicate	Growth rate		Inhibition of growth rate	Yield		Inhibition of yield
[µg/L]	No.	[d^-1]		[%]	[cells/mL]		[%]
839	1		-0.223	100		-3339	100
	2		n.a.	100		n.a.	100
	3		-0.285	100		-3933	100
	Mean	(+)	n.a.	100	(+)	n.a.	100
269	1		1.32	22		347053	67
	2		1.29	23		323177	69
	3		1.23	27		262726	75
	Mean	(+)	1.28	24	(+)	310985	70
84.8	1		1.48	12		567907	46
	2		1.50	11		607642	42
	3		1.49	11		595470	43
	Mean	(+)	1.49	11	(+)	590340	43
28.4	1		1.65	2		949633	9
	2		1.65	2		956714	8
	3		1.65	2		949048	9
	Mean	(-)	1.65	2	(-)	951798	9
8.69	1		1.68	0		1037413	1
	2		1.70	-1		1104068	-6
	3		1.71	-2		1149304	-10
	Mean	(-)	1.69	-1	(-)	1096928	-5
Control	1		1.67			1004408
	2		1.66			993172
	3		1.69			1077148
	4		1.71			1143978
	5		1.68			1051751
	6		1.66			997503
	Mean		1.68			1044660

Measured exposure concentrations during the definitive test:

Sampling date	2017-10-09 0 h	2017-10-12 72 h		Geometric mean
Start of analysis	2017-10-09	2017-10-12
Saturated solution [%]	Globanone
	Meas. conc.  [µg/L]	Meas. conc. [µg/L]	[%]	[µg/L]
100	827	852	103	839
31.60	300	242	81	269
10.0	93.5	76.9	82	84.8
3.16	32.8	24.6	75	28.4
1.00	9.55	7.91	83	8.69
Control	< LOQ	< LOQ		< LOQ

LOQ = (1.0 µg test item /L)

Validity criteria fulfilled:

yes

Conclusions:

The growth inhibition effect of the test item on green algae was determined. The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 470 µg/L and 122 µg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 119 µg/L and 25.3 µg/L, respectively. The NOEC-values for both inhibition of growth rate and yield after 72 hours were 28.4 µg/L.

Executive summary:

The growth inhibition effect of the test item on green algae was determined according to OECD TG 201 and GLP. The test was conducted in a closed static system without headspace. Based on the results from the range-finding test,five concentrations with a spacing factor of 3.16 and one control were tested. The test concentrations were verified via GC-MS method. At the start of the exposure the measured concentrations were 9.55 - 32.8 – 93.5 - 300 – 827 µg/L and after 72 hours the amounts were between 75 and 103% of the initial concentrations. The geometric mean measured test item concentrations were calculated to be 8.69 – 28.4 – 84.8 – 269 – 839 µg/L. Therefore, the geometric mean measured concentrations were applied to calculate the biological effect of the test item. Under the test conditions, the EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were determine to be 470 µg/L and 122 µg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 119 µg/L and 25.3 µg/L, respectively. The NOEC-values for both inhibition of growth rate and yield after 72 hours were 28.4 µg/L.

Description of key information

A study on the growth inhibition effect of the item to green algae is available. The test was conducted in a closed static system without headspace according to OECD TG 201 and GLP. Based on the results from the range-finding test,five concentrations with a spacing factor of 3.16 and one control were tested. The test concentrations were verified via GC-MS method. At the start of the exposure the measured concentrations were 9.55 - 32.8 – 93.5 - 300 – 827 µg/L and after 72 hours the amounts were between 75 and 103% of the initial concentrations. The geometric mean measured test item concentrations were calculated to be 8.69 – 28.4 – 84.8 – 269 – 839 µg/L. Therefore, the geometric mean measured concentrations were applied to calculate the biological effect of the test item. Under the test conditions, the EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were determine to be 470 µg/L and 122 µg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 119 µg/L and 25.3 µg/L, respectively. The NOEC-values for both inhibition of growth rate and yield after 72 hours were 28.4 µg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:

470 µg/L

EC10 or NOEC for freshwater algae:

119 µg/L