4-methoxybenzyl alcohol

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:

from 2016-04-11 to 2016-04-14

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: 200, 100, 50.0, 25.0, 12.5, 6.25, 0 mg/L (control was analysed undiluted; all other samples were diluted with acetonitrile/HPLC water prior to analysis)
- Sample storage conditions before analysis: Prepared samples were stored at room temperature in the autosampler until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test item was homogenized by melting at approximately 30 °C for 1 hour. Thereafter, the test item was weighed out, added into a measuring flask with the respective volume of dilution water and shaken until the test item was completely dissolved.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Green algae
- Strain: Pseudokirchneriella subcapitata
- Age of inoculum (at test initiation): three days old
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounted to 35-70 µE x m^-2 x s^-1 for 24 hours per day. Culture medium was nutrient medium Z

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

0.24 mmol Ca+Mg/L

Test temperature:

22.5 - 23.5 °C

pH:

7.83 - 8.54

Salinity:

NA

Nominal and measured concentrations:

nominal: 6.25, 12.5, 25.0, 50.0, 100.0 and 200.0 mg/L
measured: 6.60, 12.8, 25.9, 51.9, 106 and 205 mg/L (0 hours); 6.63, 13.0, 25.7, 51.8, 103 and 205 mg/L (72 hours)

Details on test conditions:

TEST SYSTEM
- Test vessel: Sterile Erlenmeyer flasks with cotton wool plugs
- Type: open
- Size, fill volume: 250 mL capacity; 100 mL test volume
- Initial cells density: approx. 5 x 10^3 – 10^4 cells/mL (nominal); 6305 cells/mL (actual)
- Control end cells density: 2039440 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: According to OECD Guideline 201

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Photoperiod: 24 hours/day light
- Light intensity and quality: 63.8 – 79.8 µE x m^-2 x s^-1

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: fluorimeter (determined at 0, 24, 48 and 72 hours)
- Other: Microscopic evaluation of the cells at the start and the end of exposure 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.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study
- Test concentrations: 100, 10 and 1 mg/L
- Results used to determine the conditions for the definitive study: Growth rate inhibition was determined to be 29%, 2% and 1% at 100, 10 and 1 mg/L, respectively.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

141 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CL: 129-155 mg/L

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

53.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CL: 43.7 - 64.9 mg/L

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

64.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Remarks on result:

other: Cl: 58.1-72.7 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

36.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Remarks on result:

other: CL: 26.5-44.1 mg/L

Details on results:

- Exponential growth in the control: yes
- Observation of abnormalities: none
- Any observations that might cause a difference between measured and nominal values: The test media were clear throughout the test period.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50: 0.564 mg/L (95% CL: 0.546 - 0.582) based on growth rate; 0.322 mg/L (95% CL: 0.239 - 0.342) based on yield inhibition
- Other: The toxicity of potassium dichromate to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined over a period of 72 hours from 2015-10-13 to 2015-10-16.

Table 1: Evaluation after 72 hours

Nominal concentration [mg/L]	Replicate	Growth rate (1/d)	Inhibition of growth rate (%)	Yield (cells/mL)	Inhibition of yield (%)
200	1	0.863	55	77675	96
	2	0.638	67	36444	98
	3	0.413	79	15448	99
	Mean	(+) 0.638	67	(+) 43189	98
100	1	1.30	32	305523	85
	2	1.20	38	222836	89
	3	1.28	34	283693	86
	Mean	(+) 1.26	35	(+) 270684	87
50.0	1	1.80	6	1386367	32
	2	1.86	4	1641994	19
	3	1.82	5	1478943	27
	Mean	(+) 1.83	5	(+) 1502435	26
25.0	1	1.92	0	1971423	3
	2	1.93	0	2054865	-1
	3	1.90	1	1882511	7
	Mean	(-) 1.92	0	(-) 1969600	3
12.5	1	1.93	-1	2076064	-2
	2	1.86	4	1642332	19
	3	1.88	2	1793431	12
	Mean	(-) 1.89	2	(-) 1837276	10
6.25	1	1.94	-1	2135488	-5
	2	1.89	2	1844229	9
	3	1.95	-1	2178957	-7
	Mean	(-) 1.93	0	(-) 2052891	-1
0	1	1.92		1968378
	2	1.88		1794164
	3	1.94		2102450
	4	1.86		1669508
	5	1.95		2185497
	6	1.99		2478814
	Mean	1.92		2033135

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

Table 2: Measured Exposure Concentrations during the Definitive Test

	0 h (start of exposure)		72 h (end of exposure)
Nominal concentration [mg/L]	Measured conc. [mg/L]	%	Measured conc. [mg/L]	%
200	205	103	205	102
100	106	106	103	103
50.0	51.9	104	51.8	104
25.0	25.9	103	25.7	103
12.5	12.8	102	13.0	104
6.25	6.60	106	6.63	106
0	< LOQ		< LOQ

Meas. Conc = Measured concentration of the test item (dilution factor taken into account), mean value of two injections

% = Percent of the nominal concentration of the test item

LOQ = Limit of Quantification (0.5 mg/L test item)

Validity criteria fulfilled:

yes

Conclusions:

In this study, the test item was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values (based on nominal test item concentrations): The EC50 and EC10 value was determined to be 141 mg/L and 53.4 mg/L based on growth rate.

Executive summary:

The toxicity of the test item to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined according to OECD 201 and under GLP. The study was conducted under static conditions with an initial cell density of 6305 cells/mL for a period of 72 hours. A stock solution of 200 mg/L was freshly prepared with dilution water and shaken until the test item was completely dissolved. Six concentrations were tested in a geometrical series with a dilution factor of 2 (nominal): 6.25 - 12.5 - 25.0 - 50.0 - 100 - 200 mg/L. The measured concentrations of the test item in the fresh media (0 h) were in the range of 102 to 106% of the nominal values at all tested concentration levels. The measured concentrations in the old media (72 h) were in the range of 102 to 106% of the nominal values of all concentrations. Therefore, all effect values given are based on the nominal test item concentrations of the test item. In this study, the test item was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values: The EC50-values with 95 % confidence interval for inhibition of growth rate (ErC50) was 141.0 mg/L (129 – 155 mg/L). The EC10 value with 95% confidence interval was determined to be 53.4 mg/L (43.7 – 64.9 mg/L) based on growth rate.

Description of key information

In this study, the test item was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values (based on nominal test item concentrations): The EC50 value and EC10 value was determined to be 141 mg/L and 53.4 mg/L based on growth rate, and EC50 and EC10 based on the inhibition of yield were determined to be 64.2 mg/L and 36.4 mg/L, respectively.

Key value for chemical safety assessment

EC50 for freshwater algae:

141 mg/L

EC10 or NOEC for freshwater algae:

53.4 mg/L

Additional information

The toxicity of the test item to the unicellular freshwater green alga Pseudokirchneriella subcapitata was determined according to OECD 201 and under GLP. The study was conducted under static conditions with an initial cell density of 6305 cells/mL for a period of 72 hours. A stock solution of 200 mg/L was freshly prepared with dilution water and shaken until the test item was completely dissolved. Six concentrations were tested in a geometrical series with a dilution factor of 2 (nominal): 6.25 - 12.5 - 25.0 - 50.0 - 100 - 200 mg/L. The measured concentrations of the test item in the fresh media (0 h) were in the range of 102 to 106% of the nominal values at all tested concentration levels. The measured concentrations in the old media (72 h) were in the range of 102 to 106% of the nominal values of all concentrations. Therefore, all effect values given are based on the nominal test item concentrations of the test item. In this study, the test item was found to inhibit the growth of the freshwater green alga Pseudokirchneriella subcapitata after 72 hours with the following effect values: The EC50-values with 95 % confidence interval for inhibition of growth rate (ErC50) was 141.0 mg/L (129 – 155 mg/L) and for inhibition of yield (EyC50) was 64.2 mg/L. The EC10 value with 95% confidence interval was determined to be 53.4 mg/L (43.7 – 64.9 mg/L) based on growth rate (ErC10) and 36.4 mg/L based on yield (EyC10).