[No public or meaningful name is available]

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:

2008-04-17 to 2008-05-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

2006

Deviations:

yes

Remarks:

The cell density at test start was 5000 cells/mL and for the control 3 replicates were tested.

GLP compliance:

yes (incl. QA statement)

Remarks:

certified by "Ministerium für Umwelt und Naturschutz, Landwirtschaft und Verbraucherschutz des Landes Nordrhein-Westfalen", Germany

Analytical monitoring:

yes

Details on sampling:

at 0, 24, 48 and 72 hours

Vehicle:

no

Details on test solutions:

Test medium according to guideline

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Strain of the test species obtained from 'The Collection of Algal Cultures' of the Institute of Plant Physiology at the University of Göttingen (Germany).
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: Exponentially-growing stock cultures were maintained in the test facility under constant temperature conditions (23 +/- 2°C) at a light intensity in the range 60 – 120 μE. x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter). The nutrient medium (according to BRINGMANN & KÜHN (1977) was renewed once a week.

ACCLIMATION
- Acclimation period for culturing media: 3 days
- Any deformed or abnormal cells observed: none

Test type:

static

Water media type:

freshwater

Remarks:

Medium according to OECD 201

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

None

Hardness:

22.5 mg/L CaCO3

Test temperature:

21 to 25 °C

pH:

7.9 to 8.3

Dissolved oxygen:

Not specified

Salinity:

Not applicable

Conductivity:

Not specified

Nominal and measured concentrations:

nominal: 0 (control), 50 and 100 mg test item/L
measured: only DOC measurements, please see Table 1 in results section

Details on test conditions:

TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks with cotton stoppers
- Initial cells density: approximately 5000 cells per millilitre
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes, according to OECD 201

OTHER TEST CONDITIONS
- Photoperiod: Continuous uniform illumination was provided in the spectral range 400 to 700 nm
- Light intensity and quality: At the average of the test solutions, a light intensity in the range 60 to 120 μE. x m-2 x s-1, or an equivalent range of 4000 to 8000 lx, was used
- Determination of cell densities: microcell counter

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding test preceded the main test. It provided information about the range of concentrations which were used in the main test.
- Test concentrations (main test): 50 and 100 mg test item/L (nominal)

CULTURING APPARATUS
-Details on culturing apparatus used: Light chamber in which a temperature in the range 21°C to 25°C was maintained at +/- 2°C

Reference substance (positive control):

not specified

Key result

Duration:

72 h

Dose descriptor:

EC10

Remarks:

ErC 10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Remarks:

ErC 10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Key result

Duration:

72 h

Dose descriptor:

EC50

Remarks:

ErC 50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Remarks:

EyC 50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Details on results:

- Exponential growth in the control: yes
- Observation of abnormalities: no abnormailies
- Unusual cell shape: no
- Colour differences: no, not reported
- Flocculation: no, not reported
- Adherence to test vessels: no, not reported
- Aggregation of algal cells: no, not reported
- Any stimulation of growth found in any treatment: no, not reported
- Any observations (e.g. precipitation): no, not reported
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

Not reported

Reported statistics and error estimates:

For the determination of the NOEC/LOEC values, Williams Multiple Sequential t-Test Procedure (α = 0.05, 2-sided) was performed with growth rates and yield. As no effects were found in the test, ECx values were estimated.

Table 1: Results of the chemical analyses (DOC values in mg/L)

Test item concentration [mg/L]	DOC values [mg/L]	DOC values [mg/L]	DOC values [mg/L]	DOC values [mg/L]
	0 h	24 h	48 h	72 h
Control	< 5	< 5	< 5	< 5
50	15.69	13.16	10.52	9.229
100	23.57	20.98	21.49	17.74
100 without algae	23.69	21.27	23.16	22.04

Table 2: Growth Rates and Yield calculated for the entire test duration (day 0-3)

Test item concentration [mg/L]		Growth rate (Day 0 - 3)	% inhibition compared to control	Yield  (Day 0 - 3)	% inhibition compared to control
Control		1.628		65.500
		1.638		67.500
		1.628		65.500
	mean	1.631		66.167
	SD	0.0057		1.1547
	CV	0.4		1.7
50		1.652		70.500
		1.617		63.500
		1.612		62.500
	mean	1.627	0.2	65.500	1.0
	SD	0.0217		4.3589
	CV	1.3		6.7
100		1.633		66.500
		1.628		65.500
		1.647		69.500
	mean	1.636	-0.3	67.167	-1.5
	SD	0.0102		2.0817
	CV	0.6		3.1

VALIDITY OF TEST

The cell concentration in the control cultures should have increased exponentially by a factor of at least 16 within three days (72 hours) (actual value: 133.0).

The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control cultures must not exceed 35 % (actual value: 16.4 %).

The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7 % (actual value: 0.4 %).

Validity criteria fulfilled:

yes

Conclusions:

In an Algae Growth Inhibition Test according to OECD 201, the 72-hour EC10 and EC50 towards Desmodesmus subspicatus based on growth rate was estimated to be > 100 mg/L. The 72-hour EC10 and EC 50 based on yield was estimated to be > 100 mg/L. The 72-hour NOEC both based on the growth rate and yield was ≥ 100 mg/L. All validity criteria of the test guideline were fulfilled. The study is thus regarded valid.

Executive summary:

The growth inhibiting effect of the test item towards the green algae Desmodesmus subspicatus was assessed in a study according to OECD TG 201 and GLP principles. Exponentially growing cells (approx. 5000 cells per mL at start of the test) were exposed for 72 hours to nominal concentrations of 50 and 100 mg/L test item, respectively. Untreated control ran in parallel. For determination of the test item concentrations, samples were taken from each test concentration and from the control at each day of the test. Concentrations were indirectly determined by measurement of carbon. Carbon dioxide was determined by infrared spectrometry. Biological results and endpoints are based on the nominal concentrations. The algal growth in relation to a control culture was determined over a fixed test period of 72 hours and, thus, over several algal generations. The test design included three replicates at each test concentration and for the control. Sealed 300 mL glass Erlenmeyer flasks filled with test liquid were used as test vessels. The algae cell densities were measured in a microcell counter at 24-hour intervals. To determine NOEC/LOEC values, Williams Multiple Sequential t-test Procedure was used. As a result, the 72-hour EC₁₀ and EC₅₀ based on growth rate was estimated to be >100 mg/L. The 72-hour EC₁₀ and EC₅₀ based on yield was estimated to be >100 mg/L. The 72-hour NOEC both based on the growth rate and yield was ≥ 100 mg/L. All validity criteria of the test guideline were fulfilled. The study is thus regarded valid.

Description of key information

In an Algae Growth Inhibition Test according to OECD 201, the 72-hour EC₁₀ and EC₅₀ towards Desmodesmus subspicatus based on growth rate was estimated to be > 100 mg/L. The 72-hour EC₁₀ and EC₅₀ based on yield was estimated to be > 100 mg/L. The 72-hour NOEC both based on the growth rate and yield was ≥ 100 mg/L. All validity criteria of the test guideline were fulfilled. The study is thus regarded valid.

Key value for chemical safety assessment

EC50 for freshwater algae:

100 mg/L

EC10 or NOEC for freshwater algae:

100 mg/L

Additional information

The growth inhibiting effect of the test item towards the green algae Desmodesmus subspicatus was assessed in a study according to OECD TG 201 and GLP principles. Exponentially growing cells (approx. 5000 cells per mL at start of the test) were exposed for 72 hours to nominal concentrations of 50 and 100 mg/L test item, respectively. Untreated control ran in parallel. For determination of the test item concentrations, samples were taken from each test concentration and from the control at each day of the test. Concentrations were indirectly determined by measurement of carbon. Carbon dioxide was determined by infrared spectrometry. Biological results and endpoints are based on the nominal concentrations. The algal growth in relation to a control culture was determined over a fixed test period of 72 hours and, thus, over several algal generations. The test design included three replicates at each test concentration and for the control. Sealed 300 mL glass Erlenmeyer flasks filled with test liquid were used as test vessels. The algae cell densities were measured in a microcell counter at 24-hour intervals. To determine NOEC/LOEC values, Williams Multiple Sequential t-test Procedure was used. As a result, the 72-hour EC₁₀ and EC₅₀ based on growth rate was estimated to be >100 mg/L. The 72-hour EC₁₀ and EC₅₀ based on yield was estimated to be >100 mg/L. The 72-hour NOEC both based on the growth rate and yield was ≥ 100 mg/L. All validity criteria of the test guideline were fulfilled. The study is thus regarded valid.