Farnesol

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:

06 December 2016 - 09 December 2016

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: All concentration levels and the control
- Sampling method: Samples were taken at the start of the test (0 h, without algae) and at the end of exposure (72 h, with algae)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A saturated solution with a nominal loading of 1000 mg/L test item was prepared once with demineralised water at 96 ± 1 hour prior to the start of the exposure. The test item was pipetted on to the surface of the demineralised water and gentle stirring (to avoid formation of an emulsion) was carried out with a magnetic stirrer at room temperature for 96 ± 1 hour. After a separation phase of 1 hour at room temperature the saturated solution was taken from the bottom and was used for testing. Then the components of the dilution water were added to the saturated solution.
- Controls: Dilution water
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): The saturated solution was checked after stirring via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion), which was negative.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Green algae
- Strain: HINDÄK, SAG 61.81
- Age of inoculum (at test initiation): 4 days old
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Cultures were incubated under continuous light (2590 - 5180 lux).

ACCLIMATION
- Culturing media and conditions (same as test or not): No, cells cultured in nutrient medium Z
- Any deformed or abnormal cells observed: No

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

0.24 mmol Ca+Mg/L

Test temperature:

22.0 - 22.5°C

pH:

Start: 7.35 - 7.86
End: 8.13 - 8.95

Dissolved oxygen:

Not reported

Nominal and measured concentrations:

Nominal concentrations: 1.00, 3.16, 10.0, 31.6 and 100% of the saturated solution, corresponding to the geometric mean measured test item concentration of 0.0299, 0.0472, 0.0832, 1.42 and 6.37 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks, sealed with cotton wool plugs
- Type: Closed
- Material, size, headspace, fill volume: 100 mL test volume
- Aeration: Not reported
- Initial cells density: 6305 cells/mL
- Control end cells density: 2366235 cells/mL after 72 hours
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: Yes (OECD TG 201 medium)

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: Yes, nutrient medium Z used for culture medium and OECD TG 201 medium used for test medium

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: No
- Photoperiod: Continuous light
- Light intensity and quality: Approximately 4440 to 8880 lux

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: Fluorimeter
- Chlorophyll measurement: The cell density was measured daily via Chlorophyll a-fluorescence, excitation at 436 nm, emission at 685 nm.
- Other: Microscopic evaluation of the cells at the start and the end of the incubation period was carried out. 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.

The pH-values at the start of exposure were measured in one additional replicate of each test item concentration and the control. At the end of exposure, the pH-values were measured from pooled samples of each test item concentration and the control. The room temperature was measured continuously. Light intensity was measured prior to the start of exposure.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: Square root of 10
- Range finding study: Yes, 72 hours exposure
- Test concentrations: 1, 10 and 100% saturated solution, corresponding to measured concentrations of <0.05, 0.480 and 6.30 mg/L, respectively at the test start (0h), and at the end of exposure (72h) were <0.05, 0.416 and 6.19 mg/L without algae, respectively, and <0.05, 0.250 and 6.12 mg/L with algae, respectively.
- Results used to determine the conditions for the definitive study: Yes, at 1, 10 and 100% saturated solution, the growth rate inhibitions were 1, 4 and 100%, respectively, and the yield inhibitions were 1, 21 and 100%, respectively

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

1.49 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CL: 0.503-4.46 mg/L

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

1.17 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CL: 0.413-4.15 mg/L

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.083 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.:

0.334 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

cell number

Remarks on result:

other: 95% CL: 0.240-0.461 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.079 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

cell number

Remarks on result:

other: 95% CL: 0.0606-0.121 mg/L

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.047 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

cell number

Details on results:

- Exponential growth in the control: Yes

Microscopic evaluation of the cells at the start and end of exposure revealed no morphological abnormalities.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50: ErC50 = 0.435 mg/L (95% CL: 0.414-0.458 mg/L), EyC50 = 0.230 (95% CL: 0.179-0.322 mg/L)
- Other: ErC50 valid range: 0.764 mg/L (3xSD = 0.513 mg/L), EyC50 valid range: 0.402 (3xSD = 0.293 mg/L)

Reported statistics and error estimates:

ECx-values with confidence intervals of growth rate inhibition and for yield inhibition after 72 hours were calculated by sigmoidal dose-response regression. Shapiro-Wilk’s test on normal distribution was calculated with a significance level 0.01. Levene’s test on variance homogeneity was calculated with a significance level 0.01. Monotonicity of Concentration/Response was calculated by trend analysis by contrasts (significance level 0.05). The NOEC and LOEC were determined for growth rate using SD Jonckheere-Terpsta test procedure rate with a significance level 0.05 and for yield using Wiliams Multiple t-test with a significance level 0.05.

Table 1. Mean growth rate and yield after 72 hours

Geometric mean measured test item concentration (mg/L)	Mean growth rate (1/d)	Inhibition of growth rate (%)	Mean yield (cells/mL)	Inhibition of yield (%)
Control	1.98	-	2359930	-
0.0299	(-) 1.97	0	(-) 2312805	2
0.0472	(-) 1.96	1	(-) 2250017	5
0.0832	(+) 1.94*	2	(+) 2104462	11
1.42	(+) 1.20	39	(+) 228807	90
6.37	(+) n.a.	100	(+) n.a.	100

n.a. = not applicable

* A statistically significant difference was found but an inhibition of 2 % is considered to be not biologically significant.

Validity criteria fulfilled:

yes

Remarks:

For the control cultures, the increase of cell growth was 375-fold (≥16-fold) , the mean CV for section-by-section specific growth rates was 27.3% (≤35%) and the CV of average specific growth rates during the whole test period was 0.55% (≤7%).

Conclusions:

The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 1.49 and 0.334 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.17 and 0.0785 mg/L, respectively. The NOEC-values of growth rate and yield after 72 hours were 0.0832 and 0.0472 mg/L, respectively.

Executive summary:

The algal toxicity of the test item was assessed in a 72 -hour static test with green alga Pseudokirchneriella subcapitata. Algae were exposed to 1.00, 3.16, 10.0, 31.6 and 100% of a saturated solution, corresponding to the geometric mean measured test item concentrations of 0.0299, 0.0472, 0.0832, 1.42 and 6.37 mg/L, alongside an untreated control and positive control. The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 1.49 and 0.334 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.17 and 0.0785 mg/L, respectively. The NOEC-values of growth rate and yield after 72 hours were 0.0832 and 0.0472 mg/L, respectively. This study is considered to be reliable without restriction (Klimisch 1) as it is GLP-compliant and was conducted according to OECD 201 and EU C.3.

Description of key information

The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 1.49 and 0.334 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.17 and 0.0785 mg/L, respectively. The NOEC-values of growth rate and yield after 72 hours were 0.0832 and 0.0472 mg/L, respectively.

Key value for chemical safety assessment

EC50 for freshwater algae:

1.49 mg/L

EC10 or NOEC for freshwater algae:

1.17 mg/L

Additional information

The algal toxicity of the test item was assessed in a 72 -hour static test with green alga Pseudokirchneriella subcapitata (2017). Algae were exposed to 1.00, 3.16, 10.0, 31.6 and 100% of a saturated solution, corresponding to the geometric mean measured test item concentrations of 0.0299, 0.0472, 0.0832, 1.42 and 6.37 mg/L, alongside an untreated control and positive control. The EC50-values for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 1.49 and 0.334 mg/L, respectively. The EC10-values for inhibition of growth rate (ErC10) and yield (EyC10) after 72 hours were 1.17 and 0.0785 mg/L, respectively. The NOEC-values of growth rate and yield after 72 hours were 0.0832 and 0.0472 mg/L, respectively. This study is considered to be reliable without restriction (Klimisch 1) as it is GLP-compliant and was conducted according to OECD 201 and EU C.3.