2-benzylheptanol

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:

2016-04-01 to 2016-04-08

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

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Concentrations: 0.316 - 1.00 - 3.16 - 10.0 - 31.6 - 100% of the saturated solution
- Sampling method: All concentration levels and the control were analytically verified via HPLC-DAD after 0 hours (start of exposure) and 72 hours (end of exposure). Separate replicates for the test item analysis at the beginning of exposure were prepared with algae. For the test item analysis after 72 hours samples were taken from the test replicates.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until sample preparation and at room temperature until the start of the analysis

Vehicle:

yes

Remarks:

demineralized water

Details on test solutions:

REPARATION AND APPLICATION OF TEST SOLUTION
- Method: A saturated solution with a nominal loading of 1 mL test item/L was prepared once with demineralized water 96 ± 1 hours prior to the start of the exposure. The test item was placed onto the surface of the demineralized water with a pipette. A slow stirring procedure was applied. Gentle stirring (to avoid formation of an emulsion) was carried out with a magnetic stirrer at 30 ± 2 °C for 72 ± 1 hours and for 24 ± 1 hours at room temperature. After a separation phase of 1 hour at room temperature the saturated solution was taken from the homogeneous phase and was used for testing. The saturated solution was checked after stirring via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion). Then the components of the dilution water were added to the saturated solution. The saturated solution was clear and colorless.
- Controls: Six replicates (without test item) were exposed under the same conditions as the test concentrations. Potassium dichromate was tested as a reference item.
- Concentration of vehicle in test medium: stock solution: 1 mL/L; final test solutions: 0.316 - 1.00 - 3.16 - 10.0 - 31.6 - 100 % of saturated solution
- Evidence of undissolved material: No

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: HINDAK, SAG 61.81
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounted to 35-70 µE * m*exp^-2 * s*exp^-1 for 24 hours per day.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

0.24 mmol Ca+Mg/L

Test temperature:

21 - 24 °C, controlled at ± 2 °C

pH:

8.1 ± 0.2

Dissolved oxygen:

No data was provided.

Salinity:

Freshwater was used.

Nominal and measured concentrations:

Nominal: 0.316 - 1.00 - 3.16 - 10.0 - 31.6 - 100 % of saturated solution
Measured: 0.0695 - 0.21 - 0.673 - 2.16 - 9.01 - 29.5 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks, volume: 250 mL
- Type: sealed with cotton wool plugs
- Fill volume: 100 mL
- Initial cells density: 6305 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- Agitation: Test containers were placed on a rotary shaker and oscillated at approximately 70 rpm.

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: According to guideline

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: 24 h/day
- Light intensity and quality: 68.2-79.6 µE * m^-2 * s^-1; Light homogeneity was within ± 15 % over incubation area.

EFFECT PARAMETERS MEASURED:
- Chlorophyll measurement: yes, Chlorophyll-a-fluorescence daily with excitation at 436 nm, emission at 685 nm
- Microscopic evaluation: Microscopic evaluation of the cells was carried out at the start and end of the incubation. The cells were checked for unusual cell shapes, colour differences, differences in chloroplast morphology, flocculation, adherence of algae to test containers and agglutination of algae cells.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: dilution factor of √10 (~ 3.16)
- Range finding study: yes
- Test concentrations: nominal 0.316 - 1.00 - 3.16 - 10.0 - 31.6 - 100 % of the saturated solution

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.673 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

2.16 mg/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.:

1.89 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 1.53-2.37 95 % Confidence Interval

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

2.8 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 2.37 - 3.30 95 % Confidence Interval

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

5.29 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 4.72 - 5.88 95 % Confidence Interval

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.21 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield Inhibition

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

0.673 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield Inhibition

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.6 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield Inhibition

Remarks on result:

other: < 0.0695-1.19 95 % Confidence Interval

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

1.06 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield Inhibition

Remarks on result:

other: 0.686-1.49 95 % Confidence Interval

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

2.08 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield Inhibition

Remarks on result:

other: 1.70-2.54 95 % Confidence Interval

Details on results:

- Observation of abnormalities: Microscopic evaluation of the cells at the start and end of exposure revealed no morphological abnormalities.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Measured concentrations: The measured concentrations of the test item during the exposure interval were 0.0763 - 0.223 - 0.713 - 2.61 - 9.03 - 29.7 mg/L (0 hours) and in the range of 69 to 100 % (72 hours) of the initially measured values of all concentrations. The geometric mean measured concentrations were determined.

Results with reference substance (positive control):

- Results with reference substance valid: yes
- EC50: Growth rate inhibition: 0.460, Yield inhibition: 0.317

Reported statistics and error estimates:

EC10-, EC20- and EC,50-values with confidence intervals of growth rate and yield inhibition after 72 hours were calculated by sigmoidal dose-response regression.

Evaluation after 72 hours

Statistically significant differences of growth rates and yield compared to control values are marked (+), not significant differences are marked (-).

Geometric mean measured test item concentration [mg/L]	Replicate No.	Growth rate [d^-1]	Inhibition of growth rate [%]	Yield [cells/mL]	Inhibition of yield [%]
29.5	1	< LOQ	100	< LOQ	100
	2	< LOQ	100	< LOQ	100
	3	< LOQ	100	< LOQ	100
	mean	(+) n.a.	100	(+) n.a.	100
9.01	1	0.263	86	7572	100
	2	0.432	77	1611	99
	3	0.604	68	32317	98
	mean	(+) 0.433	77	(+) 18867	99
2.16	1	1.64	14	847177	54
	2	1.68	11	970424	47
	3	1.64	13	852872	54
	mean	(+) 1.65	13	(+) 890158	52
0.673	1	1.82	4	1468710	20
	2	1.87	1	1725803	7
	3	1.86	2	1678669	9
	mean	(-) 1.85	2	(+) 1624394	12
0.210	1	1.80	5	1396544	24
	2	1.93	-2	2032341	-10
	3	1.88	0	189456	3
	mean	(-) 1.87	1	(-) 1739447	6
0.0695	1	1.88	1	1781788	4
	2	1.89	0	18202240	1
	3	1.85	2	1616708	12
	mean	(-) 1.87	1	(-) 1739579	6
Control	1	1.82		1476885
	2	1.93		2025012
	3	1.91		1914676
	4	1.88		1748581
	5	1.91		1955608
	6	1.91		1959047
	mean	1.89		1846635

< LOQ = below the limit of quantification  (2736 cells/mL)        

n.a. = data not applicable

Validity criteria fulfilled:

yes

Conclusions:

The EC50-values with 95 % confidence intervals for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 5.29 (4.72 - 5.88) mg/L and 2.08 (1.70 - 2.54) mg/L, respectively. The NOEC for inhibition of growth rate and yield after 72 h was 0.673 and 0.210 mg/L, respectively.

Executive summary:

The toxicity of the test substance to the unicellular freshwater green alga was determined according to the principles of OECD 201 and Council Regulation (EC) No. 761/2009, C.3. under GLP. Five dilution levels of the saturated solution were tested in a geometrical series with a dilution factor of√10 (approx. 3.16): 0.316 - 1.00 - 3.16 - 10.0 - 31.6 – 100 % of the saturated solution. The measured concentrations in the old media (72 h) containing algae were in the range of 69 to 100 % of the initially measured values of all concentrations, corresponding to the geometric mean measured concentrations: 0.0695 - 0.210 - 0.673 - 2.16 - 9.01 -29.5 mg/L. The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 5.29 mg/L and 2.08 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.89 mg/L and 0.6 mg/L, respectively.

Description of key information

The test revealed an EC50 value of 5.29 mg/L, an EC10 value of 1.89 mg/L and a NOEC of 0.673 mg/L. The values are based on the growth rate.

Key value for chemical safety assessment

EC50 for freshwater algae:

5.29 mg/L

EC10 or NOEC for freshwater algae:

1.89 mg/L

Additional information

The toxicity of the test substance to the unicellular freshwater green alga was determined according to the principles of OECD 201 and Council Regulation (EC) No. 761/2009, C.3. under GLP. Five dilution levels of the saturated solution were tested in a geometrical series with a dilution factor of√10 (approx. 3.16): 0.316 - 1.00 - 3.16 - 10.0 - 31.6 – 100 % of the saturated solution. The measured concentrations in the old media (72 h) containing algae were in the range of 69 to 100 % of the initially measured values of all concentrations, corresponding to the geometric mean measured concentrations: 0.0695 - 0.210 - 0.673 - 2.16 - 9.01 -29.5 mg/L. The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 5.29 mg/L and 2.08 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.89 mg/L and 0.6 mg/L, respectively.