Octahydro-4,7-methano-1H-inden-5-yl acetate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

12 March 2010 and 20 May 2010.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to OECD TG 201 in compliance with GLP, without deviations that influence the quality of the results.

Justification for type of information:

The information of Cyclacet is used for read across to Cyclacet Dihydro (In the Endpoint summary further details are provided)

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations:
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test: 1.0, 3.2, 10, 32 and 100 mg/l.

- Sampling method & Sample storage conditions before analysis:

Verification of test concentrations
Samples were taken from the control (replicates R1 - R6 pooled) and each test group (replicates R1 - R3 pooled) at 0 and 72 hours for quantitative analysis. Duplicate samples were taken at 0 and 72 hours and stored at approximately 20ºC for further analysis if necessary.
The method of analysis, stability, recovery and test preparation analyses are described in the attached Appendix 4.

Vehicle:

no

Details on test solutions:

Experimental Preparation
For the purpose of the definitive test, the test item was dissolved directly in culture medium.
An amount of test item (100 mg) was dissolved in culture medium with the aid of ultrasonication for approximately 45 minutes and the volume adjusted to 1 litre to give a 100 mg/l stock solution. A series of dilutions was made from this stock solution to give further stock solutions of 32, 10, 3.2 and 1.0 mg/l. An aliquot (500 ml) of each of the stock solutions was separately inoculated with 5.4 ml of algal suspension to give the required test concentrations of 1.0, 3.2, 10, 32 and 100 mg/l.
The stock solutions and each of the prepared concentrations were inverted several times to ensure adequate mixing and homogeneity.
The concentration and stability of the test item in the test preparations were verified by chemical analysis at 0 and 72 hours (see Appendix 4).

Exposure conditions
As in the range-finding test 250 ml glass conical flasks were used. Six flasks each containing 100 ml of test preparation were used for the control and three flasks each containing 100 ml were used for each treatment group.
The control group was maintained under identical conditions but not exposed to the test item.
Pre-culture conditions gave an algal suspension in log phase growth characterised by a cell density of 3.67 x 10^5 cells per ml. Inoculation of 500 ml of test medium with 5.4 ml of this algal suspension gave an initial nominal cell density of 4 x10^3 cells per ml and had no significant dilution effect on the final test concentration.
The flasks were plugged with polyurethane foam bungs and incubated (INFORS Multitron- Version 2 incubator) at 24 ± 1°C under continuous illumination (intensity approximately 7000 lux) provided by warm white lighting (380 – 730 nm) and constantly shaken at approximately 150 rpm for 72 hours.
Samples were taken at 0, 24, 48 and 72 hours and the cell densities determined using a Coulter® Multisizer Particle Counter.

- Eluate: not applicable

- Controls:

A positive control (Harlan Laboratories Ltd Project Number: 0039/1127) used potassium dichromate as the reference item. Details of the positive control are given in Appendix 2. The positive control was conducted between 12 January 2010 and 15 January 2010.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Test Species
The test was carried out using Desmodesmus subspicatus strain CCAP 276/20. Liquid cultures of Desmodesmus subspicatus were obtained from the Culture Collection of Algae and Protozoa (CCAP), Dunstaffnage Marine Laboratory, Oban, Argyll, Scotland. Master cultures were maintained in the laboratory by the periodic replenishment of culture medium (Section 3.2). The master cultures were maintained in the laboratory under constant aeration and constant illumination at 21 ± 1°C.
Prior to the start of the test sufficient master culture was added to approximately 100 ml volumes of culture media contained in conical flasks to give an initial cell density of approximately 10^3 cells/ml. The flasks were plugged with polyurethane foam stoppers and kept under constant agitation by orbital shaker (100 – 150 rpm) and constant illumination at 24 ± 1°C until the algal cell density was approximately 10^4 - 10^5 cells/ml.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

Not applicable

Hardness:

Not recorded.

Test temperature:

Temperature was maintained at 24 ± 1ºC throughout the test. The temperature within the incubator was recorded daily.

pH:

The pH values of each test and control flask are given in Table 2 see in any other information in results section.

The pH values of the control cultures (see Table 2 in any other information in results section) were observed to increase from pH 7.2 at 0 hours to pH 7.6 – 7.7 at 72 hours. The pH deviation in the control cultures was less than 1.5 pH units after 72 hours and therefore was within the limits given in the Test Guidelines.

The pH of each control and test flask was determined at initiation of the test and after 72 hours exposure. The pH was measured using a WTW pH 320pH meter.

Dissolved oxygen:

Not recorded.

Salinity:

freshwater used

Nominal and measured concentrations:

The range-finding test was conducted by exposing Desmodesmus subspicatus cells to a series of nominal test concentrations of 0.10, 1.0, 10 and 100 mg/l for a period of 72 hours.
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test: 1.0, 3.2, 10, 32 and 100 mg/l.

Details on test conditions:

Range-finding test
The test concentrations to be used in the definitive test were determined by a preliminary range-finding test. The range-finding test was conducted by exposing Desmodesmus subspicatus cells to a series of nominal test concentrations of 0.10, 1.0, 10 and 100 mg/l for a period of 72 hours.
The test was conducted in 250 ml glass conical flasks each containing 100 ml of test preparation and plugged with polyurethane foam bungs to reduce evaporation. Two replicate flasks were used for each control and test concentration. The test item was dissolved directly in culture medium.
An amount of test item (50 mg) was dissolved in culture medium with the aid of ultrasonication for approximately 8 minutes and the volume adjusted to 500 ml to give a 100 mg/l stock solution. A series of dilutions was made from this stock solution to give further stock solutions of 10, 1.0 and 0.10 mg/l. An aliquot (200 ml) of each of the stock solutions was separately inoculated with 2.9 ml of algal suspension to give the required test concentrations of 0.10, 1.0, 10 and 100 mg/l.

The stock solutions and each of the prepared concentrations were inverted several times to ensure adequate mixing and homogeneity.
The control group was maintained under identical conditions but not exposed to the test item.
At the start of the range-finding test a sample of each test and control culture was removed and the cell density determined using a Coulter® Multisizer Particle Counter. The flasks were then plugged with polyurethane foam bungs and incubated (INFORS Multitron- Version 2 incubator) at 24 ± 1ºC under continuous illumination (intensity approximately 7000 lux) provided by warm white lighting (380 – 730 nm) and constantly shaken at approximately 150 rpm for 72 hours.
After 72 hours the cell density of each flask was determined using a Coulter® Multisizer Particle Counter.

Culture Medium
The culture medium used for both the range-finding and definitive tests was the same as that used to maintain the stock culture.
The culture medium is defined in the attached Appendix 3.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CL not stated

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Remarks on result:

other: 95% CL not stated

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

9.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Remarks on result:

other: 95% CL 7.6-11mg/l

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CL 21-29mg/l

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CL not stated

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

3.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Remarks on result:

other: 95%CL not stated

Details on results:

Range-finding Test
The cell densities and percentage inhibition of growth values from the exposure of Desmodesmus subspicatus to the test item during the range-finding test are given in Table 1 in any other information in results section.
The results showed no effect on growth at the test concentrations of 0.10 and 1.0 mg/l. However, growth was observed to be reduced at 10 and 100 mg/l.
Based on this information test concentrations of 1.0, 3.2, 10, 32 and 100 mg/l were selected for the definitive test.

Definitive Test
Cell density values determined at each sampling time and pH values at 0 and 72 hours are given in Table 2 in any other information in results section. Daily specific growth rates for the control cultures are given in Table 3 in any other information in results section. Growth rate and yield values for the control and test cultures after 72 hours and percentage inhibition values are given in Table 4 in any other information in results section.
The mean cell densities versus time for the definitive test are presented in Figure 1 see attached section. Percentage inhibition values are plotted against test concentration in Figure 2 and Figure 3 see both in attached section.

Validation criteria
The following data show that the cell concentration of the control cultures increased by a factor of 67 after 72 hours. This increase was in line with the OECD Guideline that states the enhancement must be at least by a factor of 16 after 72 hours.
Mean cell density of control at 0 hours : 4.53 x 10^3 cells per ml
Mean cell density of control at 72 hours : 3.03 x 10^5 cells per ml
The mean coefficient of variation for section by section specific growth rate for the control cultures was 26% and hence satisfied the validation criterion given in the OECD Guideline which states the mean must not exceed 35%.
The coefficient of variation for average specific growth rate for the control cultures over the test period (0 – 72 h) was 0% and hence satisfied the validation criterion given in the OECD Guideline which states that this must not exceed 7%.

Growth data
From the data given in Tables 2 and 4 in any other information in results section, it is clear that the growth rate (r) and yield (y) of Desmodesmus subspicatus (CCAP 276/20) were affected by the presence of the test item over the 72-Hour exposure period.
Accordingly the following results were determined from the data based on the nominal test concentrations:

Inhibition of growth rate
ErC10 (0 - 72 h) : 11 mg/l
ErC20 (0 - 72 h) : 15 mg/l
ErC50 (0 - 72 h) : 25 mg/l; 95% confidence limits 21 - 29 mg/l

where ErCx is the test concentration that reduced growth rate by x%.
Statistical analysis of the growth rate data was carried out for the control and all test concentrations using one way analysis of variance incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf 1981) and Dunnett's multiple comparison procedure for comparing several treatments with a control (Dunnett 1955). There were no statistically significant differences between the control, 1.0 and 3.2 mg/l test concentrations (P>0.05), however all other test concentrations were significantly different (P<0.05) and, therefore the "No Observed Effect Concentration" (NOEC) based on growth rate was 3.2 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) based on growth rate was 10 mg/l.

Inhibition of yield
EyC10 (0 - 72 h) : 3.6 mg/l
EyC20 (0 - 72 h) : 5.1 mg/l
EyC50 (0 - 72 h) : 9.2 mg/l; 95% confidence limits 7.6 - 11 mg/l
where EyCx is the test concentration that reduced yield by x%.

Statistical analysis of the yield data was carried out as in reported statistics and error estimates. There were no statistically significant differences between the control and 1.0 mg/l test concentration (P>0.05), however all other test concentrations were significantly different (P<0.05) and, therefore the "No Observed Effect Concentration" (NOEC) based on yield was 1.0 mg/l. Correspondingly the "Lowest Observed Effect Concentration" (LOEC) based on yield was 3.2 mg/l.

Observations on cultures
All test and control cultures were inspected microscopically at 72 hours. After 72 hours there were no abnormalities detected in the control or test cultures at 1.0, 3.2, 10 and 32 mg/l, however no intact cells were observed to be present in the test cultures at 100 mg/l.

Observations on test item solubility
At the start of the test all control and test cultures were observed to be clear colourless solutions. After the 72-Hour test period all control, 1.0, 3.2 and 10 mg/l test cultures were observed to be green dispersions. The 32 mg/l test cultures were observed to be extremely pale green dispersions whilst the 100 mg/l test cultures were observed to be clear colourless solutions.

Physico-chemical measurements
The pH values of each test and control flask are given in Table 2 in any other information in results section. Temperature was maintained at 24 ± 1ºC throughout the test.
The pH values of the control cultures (see Table 2 in any other information in results section) were observed to increase from pH 7.2 at 0 hours to pH 7.6 – 7.7 at 72 hours. The pH deviation in the control cultures was less than 1.5 pH units after 72 hours and therefore was within the limits given in the Test Guidelines.

Verification of test concentrations
Analysis of the test preparations at 0 hours (see Appendix 4) showed measured test concentrations to range from 101% to 109% of nominal. A decline in measured test concentrations was observed at 72 hours in the range of 3% to 19% of nominal. This decline was considered to be due to both slight instability of the test item in culture medium as was observed in the preliminary stability analyses conducted and also adsorption of the test item to the algal cells present. Whilst the preliminary recovery analyses conducted in the presence of algal cells indicated that no immediate adsorption occurred this does not preclude long term adsorption over the test period. Adsorption was not a factor in the preliminary stability analyses conducted as no algal cells were present.

Current regulatory advice is that in cases where a decline in measured concentrations is observed, geometric mean measured concentrations should be used for calculating EC50 values. It was therefore considered justifiable to base the results on the geometric mean measured test concentrations in order to give a “worst case” analysis of the data.

The geometric mean measured test concentrations were determined to be:
Nominal Test Concentration (mg/l) Geometric Mean Measured Test Concentration (mg/l) Expressed as a % of the Nominal Test Concentration
1.0 0.32 32
3.2 0.54 17
10 1.6 16
32 9.2 29
100 44 44
It is however considered that as the decline in measured concentrations was attributable to adsorption to biological matter rather than instability, the algal cells were exposed to nominal test concentrations throughout the duration of the test. As such it is considered appropriate to base the results on the nominal measured test concentrations only.

Results with reference substance (positive control):

Appendix 2      Positive Control

A positive control (Harlan Laboratories Ltd Project No: 0039/1127) used potassium dichromate as the reference item at concentrations of 0.0625, 0.125, 0.25, 0.50 and 1.0 mg/l.
Exposure conditions and data evaluation for the positive control were similar to those in the definitive test.

Exposure of Desmodesmus subspicatus (CCAP 276/20) to the reference item gave the following results:
ErC50(0 – 72 h)        :          0.49 mg/l*
EyC50(0 – 72 h)       :          0.18 mg/l, 95% confidence limits 0.16 – 0.21 mg/l

No Observed Effect Concentration (NOEC) based on growth rate:              0.0625 mg/l
No Observed Effect Concentration (NOEC) based on yield:                          0.0625 mg/l
Lowest Observed Effect Concentration (LOEC) based on growth rate:        0.125 mg/l
Lowest Observed Effect Concentration (LOEC) based on yield:                   0.125 mg/l
The results from the positive control with potassium dichromate were within the normal ranges for this reference item.

*It was not possible to calculate 95% confidence limits for the ErC50 value as the data generated did not fit the models available for the calculation of confidence limits.

Reported statistics and error estimates:

Statistical analysis
One way analysis of variance incorporating Bartlett's test for homogeneity of variance (Sokal and Rohlf 1981) and Dunnett's multiple comparison procedure for comparing several treatments with a control (Dunnett 1955) was carried out on the growth rate and yield data after 72 hours for the control and all test concentrations to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package (SAS 1999 - 2001).

Determination of ECx values
For each individual test vessel (mean values for yield), percentage inhibition (arithmetic axis) was plotted against test concentration (logarithmic axis) and a line fitted by computerised interpolation using the Xlfit software package (IDBS). ECx values were then determined from the equation for the fitted line.
Where appropriate 95% confidence limits for the EC50 values were calculated, using the simplified method of evaluating dose-effect experiments of Litchfield and Wilcoxon (1949).

Table 1              Cell Densities and Percentage Inhibition of Growth from the Range-finding Test

Nominal Concentration (mg/l)		Cell Densities*(cells per ml)		Inhibition Values (%)
		0 Hours	72 Hours	Growth Rate	Yield
Control	R1	4.41E+03	2.22E+05
	R2	3.74E+03	1.83E+05	-	-
	Mean	4.08E+03	2.03E+05
0.10	R1	4.65E+03	2.28E+05
	R2	4.53E+03	1.56E+05	4	6
	Mean	4.59E+03	1.92E+05
1.0	R1	4.86E+03	1.73E+05
	R2	5.67E+03	1.93E+05	9	11
	Mean	5.26E+03	1.83E+05
10	R1	6.18E+03	1.41E+05
	R2	5.06E+03	1.43E+05	17	31
	Mean	5.62E+03	1.42E+05
100	R1	4.83E+03	4.30E+03
	R2	5.09E+03	2.29E+03	111	101
	Mean	4.96E+03	3.30E+03

*Cell densities represent thean number of cells per ml calculated from thean of the cell counts from 3 counts for each of the replicate flasks.

R₁and R₂= Replicates 1 and 2

Table 2              Cell Densities and pH Values in the DefinitiveTest

Nominal Concentration (mg/l)		pH	Cell Densities*(cells per ml)				pH
		0 h	0 h	24 h	48 h	72 h	72 h
Control	R1	7.2	4.78E+03	1.56E+04	7.17E+04	2.89E+05	7.6
	R2	7.2	4.46E+03	1.03E+04	6.55E+04	2.99E+05	7.7
	R3	7.2	4.32E+03	9.66E+03	6.50E+04	3.11E+05	7.7
	R4	7.2	4.28E+03	1.15E+04	6.93E+04	3.09E+05	7.7
	R5	7.2	4.77E+03	1.07E+04	7.35E+04	3.01E+05	7.6
	R6	7.2	4.58E+03	1.23E+04	6.93E+04	3.09E+05	7.6
	Mean		4.53E+03	1.17E+04	6.90E+04	3.03E+05
1.0	R1	7.2	3.56E+03	1.07E+04	5.38E+04	3.12E+05	7.6
	R2	7.1	3.87E+03	1.67E+04	5.21E+04	3.14E+05	7.6
	R3	7.2	4.26E+03	1.39E+04	5.33E+04	3.10E+05	7.6
	Mean		3.90E+03	1.38E+04	5.31E+04	3.12E+05
3.2	R1	7.1	4.01E+03	1.45E+04	5.49E+04	2.89E+05	7.6
	R2	7.2	4.49E+03	1.03E+04	5.77E+04	2.79E+05	7.6
	R3	7.2	4.51E+03	8.20E+03	5.71E+04	2.73E+05	7.6
	Mean		4.34E+03	1.10E+04	5.66E+04	2.80E+05
10	R1	7.1	4.36E+03	6.99E+03	3.45E+04	1.38E+05	7.5
	R2	7.2	3.78E+03	9.12E+03	4.06E+04	1.38E+05	7.5
	R3	7.2	4.66E+03	9.98E+03	3.92E+04	1.43E+05	7.5
	Mean		4.27E+03	8.70E+03	3.81E+04	1.40E+05
32	R1	7.1	4.50E+03	1.05E+04	2.27E+04	3.01E+04	7.4
	R2	7.2	4.18E+03	8.39E+03	1.11E+04	1.24E+04	7.4
	R3	7.2	4.24E+03	7.70E+03	1.97E+04	2.75E+04	7.4
	Mean		4.31E+03	8.87E+03	1.78E+04	2.33E+04
100	R1	7.1	4.12E+03	1.58E+03	1.08E+03	1.25E+03	7.4
	R2	7.2	4.42E+03	1.33E+03	1.82E+03	9.52E+02	7.3
	R3	7.2	4.71E+03	1.33E+03	1.33E+03	9.68E+02	7.3
	Mean		4.42E+03	1.41E+03	1.41E+03	1.06E+03

 

*Cell densities represent thean number of cells per ml calculated from thean of the cell counts from 3 counts for each of the replicate flasks.

R₁- R₆= Replicates 1 to 6

Table 3              Daily Specific Growth Rates for the Control Cultures in the Definitive Test

		Daily Specific Growth Rate (cells/ml/hour)
		Day 0 - 1	Day 1 - 2	Day 2 - 3
Control	R1	0.057	0.064	0.058
	R2	0.039	0.077	0.063
	R3	0.037	0.079	0.065
	R4	0.044	0.075	0.062
	R5	0.041	0.080	0.059
	R6	0.047	0.072	0.062
	Mean	0.044	0.075	0.062

 

R₁- R₆= Replicates 1 to 6

Table 4              Inhibition of Growth Rate and Yield in the Definitive Test

Nominal Concentration (mg/l)		Growth Rate (cells/ml/hour)		Yield (cells/ml)
		0 – 72 h	% Inhibition	0 – 72 h	% Inhibition*
Control	R1	0.059		2.85E+05
	R2	0.060		2.94E+05
	R3	0.060		3.07E+05
	R4	0.060	-	3.04E+05	-
	R5	0.060		2.96E+05
	R6	0.060		3.05E+05
	Mean	0.060		2.99E+05
	SD	0.000		8.54E+03
1.0	R1	0.060	0	3.08E+05
	R2	0.061	[2]	3.10E+05
	R3	0.060	0	3.05E+05
	Mean	0.060	[1]	3.08E+05	[3]
	SD	0.001		2.43E+03
3.2	R1	0.059	2	2.85E+05
	R2	0.059	2	2.75E+05
	R3	0.059	2	2.68E+05
	Mean	0.059	2	2.76E+05	8
	SD	0.000		8.18E+03
10	R1	0.049	18	1.33E+05
	R2	0.049	18	1.35E+05
	R3	0.050	17	1.38E+05
	Mean	0.049	18	1.35E+05	55
	SD	0.001		2.54E+03
32	R1	0.028	53	2.56E+04
	R2	0.016	73	8.18E+03
	R3	0.027	55	2.33E+04
	Mean	0.024	60	1.90E+04	94
	SD	0.007		9.46E+03
100	R1	-0.016	127	-2.87E+03
	R2	-0.020	133	-3.46E+03
	R3	-0.020	133	-3.74E+03
	Mean	-0.019	131	-3.36E+03	101
	SD	0.002		4.45E+02

[ ]

*In accordance with the OECD test guideline only thean value for yield for each test concentration is calculated

R₁– R₆= Replicates 1 to 6

SD= Standard Deviation

[Increase in growth as compared to controls]

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test item on the growth of Desmodesmus subspicatus has been investigated over a 72-Hour period and gave the following results based on nominal test concentrations:

Response Variable EC50 (mg/l) 95% Confidence No Observed Lowest Observed
Limits (mg/l) Effect Effect
Concentration Concentration
(NOEC) (mg/l) (LOEC) (mg/l)

Growth Rate 25 21-29 3.2 10
Yield 9.2 7.6 - 11 1.0 3.2

Executive summary:

The acute toxicity of Cyclacet towards Desmodesmus subspicatus was investigated according to OECD guideline 201 under GLP. Algal cells were exposed to nominal concentrations of 1.0, 3.2, 10, 32 and 100 mg/l and observed for 72 hours. The 72h-EC50 and 72h-NOEC (both based on growth rate) were found tob e 25 and 3.2 mg/l respectively based on nominal concentrations.

Analysis of the test preparations at 0 hours showed measured test concentrations to range from 101% to 109% of nominal. A decline in measured test concentrations was observed at 72 hours in the range of 3% to 19% of nominal. This decline was considered to be due to both slight instability of the test item in culture medium as was observed in the preliminary stability analyses conducted and also adsorption of the test item to the algal cells present. Whilst the preliminary recovery analyses conducted in the presence of algal cells indicated that no immediate adsorption occurred this does not preclude long term adsorption over the test period. Adsorption was not a factor in the preliminary stability analyses conducted as no algal cells were present. It is considered that as the decline in measured concentrations was attributable to adsorption to biological matter rather than instability, the algal cells were exposed to nominal test concentrations throughout the duration of the test. As such it is considered appropriate to base the results on the nominal measured test concentrations only.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Information is based on read across

Justification for type of information:

The full read across justification is presented in the Endpoint summary, the accompanying file is also attached there.

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

25 mg/L

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

3.2 mg/L

Conc. based on:

test mat.

Basis for effect:

growth rate

Validity criteria fulfilled:

yes

Description of key information

The acute, ErC50 and chronic toxicity, EC10, for algae are 25 and 3.2 mg/l, respectively. The information is retrieved from Cyclacet tested in an OECD TG 201.

Key value for chemical safety assessment

EC50 for freshwater algae:

25 mg/L

EC10 or NOEC for freshwater algae:

3.2 mg/L