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Toxicity to aquatic algae and cyanobacteria

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Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
1982-09-17 To 1982-10-01
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Qualifier:
according to
Guideline:
other: Payne AG, Hall RH, 1979, in Aquatic Toxicology, ASTM, STP 667, pp. 171-180; and US EPA, 1978, The Selenastrum capricornutum Printz algal assay. Deviations, reliability, and validity will be evaluated using the USEPA OPPTS 850.5400 (1996) guideline.
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
analytical monitoring was not performed in the study.
Principles of method if other than guideline:
Data is present within the study report to allow for 96 h ErC10, ErC20 and ErC50 endpoint calculations with the emphasis on growth rate. The 96 h modernized endpoints are reported within this summary.
GLP compliance:
not specified
Remarks:
(pre-GLPs)
Analytical monitoring:
no
Details on sampling:
- Concentrations: Not available
- Sampling method: Not available
- Sample storage conditions before analysis: Not available
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A primary stock solution of X0235.01 was prepared by adding a weighed amount of test material to algal growth medium and other stock solutions were prepared by serial dilution. A growth medium control was conducted concurrently and contained no test material. All test concentrations, except 1,000 ppm, were prepared by pipeting 1 milliliter (m2) of the appropriate stock solution to each test flask. The 1,000 ppm test concentration was prepared by adding 50ml of primary stock solution to each test flask.
- Eluate: Not applicable
- Differential loading: Not applicable
- Controls: AAP medium
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Not applicable
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): Not available
- Evidence of undissolved material (e.g. precipitate, surface film, etc): None
Test organisms (species):
Microcystis aeruginosa
Details on test organisms:
TEST ORGANISM
- Common name: Green algae
- Strain: Not available
- Source (laboratory, culture collection): University of Texas at Austin, Austin, Texas
- Age of inoculum (at test initiation): Not available
- Method of cultivation:


ACCLIMATION
- Acclimation period: Not available
- Culturing media and conditions (same as test or not): Same as test
- Any deformed or abnormal cells observed: Not available
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
120 h
Remarks on exposure duration:
96 hour results reported here
Post exposure observation period:
Yes (9 days)
Hardness:
Not available
Test temperature:
Static bioassays were conducted at 24 C (+/-1) degree C in water bath
pH:
Not available
Dissolved oxygen:
Not available
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal laboratory exposure concentrations based on active ingredient were control, 0.01, 0.05, 0.1, 0.5, 1.0 and 1,000mg/L
No measurements of test concentrations.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open / closed: Not available
- Material, size, headspace, fill volume: Test containers were 125-ml flasks with 50 ml of test medium
- Aeration: Not available
- Type of flow-through (e.g. peristaltic or proportional diluter): Not applicable
- Renewal rate of test solution (frequency/flow rate): Not available
- Initial cells density: 1x10(5) cells/ml
- Control end cells density: 18.5x10(5) cells/ml
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3


GROWTH MEDIUM
- Standard medium used: Yes, AAP nutrient medium


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Regular Algal Assay Procedure (AAP) medium prepared with deionized water
- Total organic carbon: Not available
- Particulate matter: Not available
- Metals: Not available
- Pesticides: Not available
- Chlorine: Not available
- Alkalinity: Not available
- Ca/mg ratio: Not available
- Conductivity: Not available
- Culture medium different from test medium: No
- Intervals of water quality measurement: Not available

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: Not applicable
- Photoperiod: Not available
- Light intensity and quality: Approximately 2,000 lux


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Hemacytometer
- Cell counts were made on day 0,2,3,4,5.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2-5 (except for the highest test concentration of 1000 mg/L)
- Justification for using less concentrations than requested by guideline: Not applicable
- Range finding study: no
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
EC10
Effect conc.:
0.89 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC10 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Key result
Duration:
96 h
Dose descriptor:
EC10
Effect conc.:
0.715 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
96 h
Dose descriptor:
EC20
Effect conc.:
2.24 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC20 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
96 h
Dose descriptor:
EC20
Effect conc.:
1.82 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
13.44 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
10.9 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.


- Exponential growth in the control (for algal test): Yes (18.5x growth)
- Observation of abnormalities (for algal test): Not available
- Any stimulation of growth found in any treatment: Not available
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: None
- Effect concentrations exceeding solubility of substance in test medium: None
Results with reference substance (positive control):
Not applicable
Reported statistics and error estimates:
The ErCx values were calculated using Bruce and Versteeg method and NOEC values were calculated by t-test.

Effect of LAS on Microcystis aeruginosa: Cell Countsat 96 hr (study # 29101)

Nominal Conc (mg/L) Algal cell counts (x104/ml) Mean Growth Rate % Inhibition (growth rate) Mean Biomass % Inhibition (biomass) 
0 185 (11) 0.6018 270
0.01 175 (7) 0.5925 1.54  246.5 8.52
0.05 112 (2) 0.5182 13.90  223.0 17.41 
0.1 111 (16) 0.5167 14.15  187  30.74
0.5 111 (17) 0.5167 14.15  189.5  29.63
1 105 (3) 0.5074 15.69  145 46.3 
1000 5 (0) 0 100  7.50 97.22

C10-14 LAS (X0235.01).

Numbers in parenthesis are standard deviation.

 

Validity criteria fulfilled:
yes
Conclusions:
The 96 hour ErC10, ErC20 and ErC50, based on growth rate, are 0.89, 2.24 and 13.44 mg/L, respectively.  These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized ErC10, ErC20 and ErC50 are 0.715, 1.82 and 10.9 mg/L, respectively.
Executive summary:

In a 96 hour algal growth inhibition test, the green alga Microcystis aeruginosa was exposed to Linear Alkylbenzene Sulfonate (C11.9 LAS) in accordance with Payne and Hall, evaluated against USEPA OPPTS 850.5400 (1996). The nominal test concentrations were (test 1) control, 0.01, 0.05, 0.1, 0.5, 1 and 1,000 mg a.i./L,.

The 96 hour ErC10, ErC20 and ErC50, based on growth rate, are 0.89, 2.24 and 13.44 mg/L, respectively.  These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized ErC10, ErC20 and ErC50 are 0.715, 1.82 and 10.9 mg/L, respectively.

 

This study was conducted in accordance with USEPA OPPTS 850.5400. However data is present within the study report to allow for 96 hr ErC10, ErC20 and ErC50 endpoint calculations with the emphasis on growth rate (as recommended by OECD 201). The 96 hr modernized endpoints are reported within this summary. These endpoints differ to those previously reported (96 hr endpoints based on cell count) due to the use of continuous growth rate versus cell count.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1982-08-06 To 1982-08-20
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Qualifier:
according to
Guideline:
other: Payne AG, Hall RH, 1979, in Aquatic Toxicology, ASTM, STP 667, pp. 171-180; and US EPA, 1978, The Selenastrum capricornutum Printz algal assay. Deviations, reliability, and validity evaluated using the subsequent OECD 201 (2006) test guideline.
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
no analytical monitoring was performed in the study
Principles of method if other than guideline:
Data is present within the study report to allow for 72 h EC10 and EC50 endpoint calculations with the emphasis on growth rate (as recommended by OECD 201). The 72 h modernized endpoints are reported within this summary.
GLP compliance:
no
Remarks:
(study conducted prior to GLP)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A primary stock solution of X0235.01 was prepared by adding a weighed amount of test material to algal growth medium, and other stock solutions were prepared by serial dilution. A growth medium control was conducted concurrently and contained no test material. All test concentrations were prepared by pipeting 1.0 milliliter (ml) of the appropriate stock solution to each test flask.
- Controls: AAP medium
- Evidence of undissolved material: No
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Green algae
- Strain: Not available
- Source: University of Texas at Austin, Austin, Texas
- Age of inoculum (at test initiation): Not available
- Method of cultivation: Not available

ACCLIMATION
- Acclimation period: Not available
- Culturing media and conditions: Same as test
- Any deformed or abnormal cells observed: Not available
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
120 h
Remarks on exposure duration:
72 hour results reported here
Post exposure observation period:
Yes (9 days)
Hardness:
Not available
Test temperature:
Static bioassays were conducted at 24 C (+/-1) degree C in water bath
pH:
Not available
Dissolved oxygen:
Not available
Nominal and measured concentrations:
Nominal laboratory exposure concentrations based on active ingredient were 0, 16, 32, 63, 125, 250, 500, and 1,000 mg/L.
No analytical measurements of test concentrations were taken.

Details on test conditions:
TEST SYSTEM
- Test vessel:
- Open / closed: Not available
- Material: 125-ml flasks with 50 ml solution
- Aeration: No
- Initial cells density: 2x10(4) cells/ml
- Control end cells density: Test 1: 206x10(4);
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: Yes, AAP nutrient medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: AAP media prepared with deionized water
- Total organic carbon: Not available
- Particulate matter: Not available
- Metals: Not available
- Pesticides: Not available
- Chlorine: Not available
- Alkalinity: Not available
- Ca/mg ratio: Not available
- Conductivity: Not available
- Culture medium different from test medium: No
- Intervals of water quality measurement: Not available

OTHER TEST CONDITIONS
- Sterile test conditions: Not available
- Adjustment of pH: No
- Photoperiod: Not available
- Light intensity and quality: Approximately 2,000 lux

EFFECT PARAMETERS MEASURED
- Determination of cell concentrations: Hemacytometer
- Cell counts were made on day 0,2,3,4,5.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: no
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
16.15 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC10 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
other: ECr10
Effect conc.:
13.1 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 72-hr ECr10: 13.1 (0 - 99) mg/L
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
289.67 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
other: ECr50
Effect conc.:
235 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 72-hr ECr50: 235 (34.6 - 965) mg/L
Details on results:
These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.

- Exponential growth in the control: Yes, 103X at 72 hrs
- Observation of abnormalities: In all test concentrations >=1.0 mg/L, the algal cells were abnormally large and malformed. Because of this observation, algae previously exposed to 1000 mg/L were tested for their ability to recover. Their growth was similar to the growth of the control during a 9-day recovery period in test material-free medium, and the algal cells had returned to normal appearance.
- Any stimulation of growth found in any treatment:
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No

Reported statistics and error estimates:
The ECx values were calculated using Bruce and Versteeg method and NOEC values were calculated by t-test.

Effect of LAS on Selenastrum capricornutum: Cell Counts at 72 hr (study # 29101)

72 hour cell count

Concentration

cell count

% Inhibition

0

206(11)

 

16

159(10)

22.81553398

32

95(10)

53.88349515

63

62(4)

69.90291262

125

23(2)

88.83495146

250

19(1)

90.77669903

500

18(2)

91.26213592

1000

5(1)

97.57281553

C10-13 LAS(X0235.01).

Numbers in parenthesis are standard deviation.

Validity criteria fulfilled:
yes
Conclusions:
In a 72 hour algae growth inhibition test, the EC10 and EC50 of LAS to Selenastrum capricornutum was 16.15 mg/L and 289.67 mg/L, consecutively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non- normalized EC10 and EC50 of LAS to Selenastrum capricornutum was 13.1 (0 – 99) mg/L and 235 (34.6 – 965) and mg/L, consecutively.

Executive summary:

In a 72 hour algal growth inhibition test, the green alga Selenastrum capricornutum was exposed to Linear Alkylbenzene Sulfonate (C11.9 LAS) in accordance with OECD 201. The nominal test concentrations were (test 1) control, 16, 32, 63, 125, 250, 500, and 1,000 mg a.i./L,.

The 72 hour EC10 and EC50, based on growth rate, are 16.15 mg/L and 289.67 mg/L, consecutively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6.

This algal growth inhibition test satisfies the guideline requirements of the OECD 201 guideline.

 

This study was conducted in accordance with USEPA OPPTS 850.5400. However data is present within the study report to allow for 72 hr EC10 and EC50 endpoint calculations with the emphasis on growth rate (as recommended by OECD 201). The 72 hr modernized endpoints are reported within this summary. These endpoints appear less conservative to those previously reported (96 hr endpoints based on cell count) due to the use of continuous growth rate versus cell count.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From Dec. 06, 1999 to Dec. 24, 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
2 replicates per test concentration against the guideline requirement of 3
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
yes
Remarks:
2 replicates per test concentration against the guideline requirement of 3
GLP compliance:
yes
Remarks:
OECD principles of GLP
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All test concentrations at 0, 48, 96 hr.
- Sampling method: Sampling was performed in the following manner:
(i) At 0 h: 5 mL samples were collected from the test and control media without algae.
(ii) At 48 and 96 h: 5 mL samples were collected from each test flask and two controls (all containing algae).
- Sample storage conditions before analysis: All samples immediately placed into 20 mL polyethylene vials, frozen and stored at a temperature of < -20°C. After the termination of the test, the samples were shipped frozen, packed in dry ice, to Procter & Gamble Eurocor for analysis.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
This test was conducted using two different algal growth media: OECD (ammonia as nitrogen source), and EPA medium (nitrate as nitrogen source).

1. OECD medium: An amount of 253.7 mg test material was dissolved in 5,000 mL OECD medium in order to condition the relevant glass-ware. Further, for the growth inhibition test 516.9 mg of test material was dissolved in 500 mL OECD medium. The pH of the solution was adjusted to 8.0 by addition of 1 M HCl solution.

2. EPA medium: 253.3 mg of test material was dissolved in 5,000 mL EPA medium in order to condition the relevant glass-ware. Further, for the growth inhibition test, 1,089 mg of test material was dissolved in 1,000 mL EPA medium. The pH of the solution was adjusted to 7.4 by addition of 1 M HCl solution.

- Application of test solution: Test solution application was carried out by adding appropriate volumes of the stock solution to the test flasks so as to yield the required test concentrations.
- Controls: Prepared using OECD or EPA medium containing algae only and a single background series containing test substance without algae.
Test organisms (species):
Scenedesmus quadricauda
Details on test organisms:
TEST ORGANISM
- Common name: Green alga
- Strain number: CCAP 276/21
- Class: Chlorophyceae
- Source: CCAP, The Freshwater Biological Association, the Ferry House, Far Sawrey, Ambleside, Cumbria LA22 OLP, England
- Age of inoculum: Not reported
- Method of cultivation: Pre-cultures of algal species in the exponential growth phase were prepared as detailed in OECD and EPA Guideline on the basis of total cell volume, using the OECD and EPA media for ammonia as nitrogen source and nitrate as nitrogen source respectively.
1. In OECD medium: A suspension of algae in OECD medium containing a total cell volume of 0.7 x 10(8) µm3/mL was prepared by dilution of a pre-culture containing 2.7 x10(8) µm3/mL. Addition of 1 mL of this algal suspension to 100 mL of the appropriate solutions of the test substance yielded a mean inoculum cell density of 0.2 x 10(4) cells/mL (measured in the control cultures).
2. In EPA medium: A suspension of algae in EPA medium containing a total cell volume of 0.7 x 10(8) µm3/mL was prepared by dilution of a pre-culture containing 3.1 x 10(8) µm3/mL. Addition of 1 mL of this algal suspension to 100 mL of the appropriate solutions of the test substance yielded a mean inoculum cell density of 0.3 x 10(4) cells/mL (measured in the control cultures).
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
None
Hardness:
24.2 mg equivalent CaCO3/L (in OECD medium)
5.9 mg equivalent CaCO3/L (in EPA medium)
Test temperature:
23±2˚C
The temperature was determined continuously and was within the required limits.
pH:
Overall, in OECD medium, pH ranged 8.1 - 8.2:
8.1 – 8.2 (at test start, without algae)
8.1 – 8.2 (at 72 h, with algae)
8.2 (at 72 h, without algae)

Overall, in EPA medium, pH ranged 7.0 - 9.6:
7.0 – 7.4 (at test start, without algae)
7.4 – 9.6 (at 72 h, with algae)
7.1 – 7.6 (at 72 h, without algae)
Nominal and measured concentrations:
- Nominal test concentrations in the OECD medium were:
0, 1.0, 3.5, 5.7, 10.2, 18.3, 32.5, 102 and 325 mg/L
- Measured test concentrations in OECD medium, 0-48 hr, were:
<0.2, 0.67, 3.96, 5.95, 9.6, 17.4, 30.5, 91.6 and NA mg/L.
- Measured test concentrations in OECD media, 0-48 hr, were 94% of nominal (range, 67-113%). (NA = not measured.)

- Nominal test concentrations in EPA medium were:
0, 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L
- Measured test concentrations in EPA medium, 0-96 hr, were:
<0.2, 1.03, 3.26, 9.98, 33.6, 89.1, NA, NA and NA mg/L.
- Measured test concentrations in EPA media, 0-96 hr, were 93% of nominal (range, 83-98%).

According to EU guideline, the effect values should be based on nominal test concentrations if the measured concentrations were > 80% of nominal value during 48 h incubation in OECD medium and > 80% of nominal during 96 h incubation in the EPA medium. These criteria were met in both test media. Therefore, the results were based on nominal concentration.

Details on measured concentration are provided under ‘Any other information on materials and methods incl. tables’ section.
Details on test conditions:
TEST SYSTEM
- Test vessel: Conical flasks covered with silicon sponge caps
- Type: Open
- Material: Sterile 200 mL conical test flasks containing 1 mL of algal suspension and 100 mL of test material solution.
- Aeration: No
- Agitation: Test containers were placed on a Gallenkamp orbital shaker and oscillated at approx. 100 rpm
- Initial cells density: 0.2 x 10(4) and 0.3 x 10(4) cells/mL in OECD and EPA medium respectively
- Control end cells density (mean of replicates): 13.5 x 10(4) and 47.5 x 10(4) particles/mL (corrected for background) in OECD and EPA medium respectively.
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 4 controls containing algae only
- No. of vessels for background series: 1 containing test substance without algae for the correction of non-algal particles present in the test flasks.
- Pre-treatment of test flasks: The test flasks and the background control were pre-treated with test solutions to prevent a decrease in the available test concentration due to adsorption to the glassware. Solutions in either OECD or EPA medium containing 50 mg/L test material were placed in the test flasks for ≥24 h before the experimental start. Following the equilibration, the content of the flasks were removed, and the flasks were rinsed ten times with ultrapure water.

GROWTH MEDIUM
- Standard medium used: Yes, OECD and EPA media.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared in accordance with standard OECD and EPA guideline. Detailed composition is provided in the study report (Annex A).
- Ca/mg ratio:
OECD medium: CaCl2.2H2O: 18 mg/L/ (∑MgCl2.6H2O: 12 mg/L, MgSO4.7H2O: 15 mg/L)
EPA medium: CaCl2. 2H2O: 4.41 mg/L/ (∑MgCl2.6H2O: 12.16 mg/L, MgSO4.7H2O: 14.7 mg/L)
- Hardness: 24.2 mg equivalent CaCO3/L for OECD medium; 5.9 mg equivalent CaCO3/L for EPA medium
No information on any other water parameters was provided in the study report.
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH was measured in all parent solutions at 0 h and after 72 h in selected cultures. The temperature was registered continuously.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No, however before the test start the pH of OECD and EPA medium was adjusted to 8.0 and 7.4 respectively.
- Photoperiod: Not reported
- Light intensity: 60-120 µmol/sec/m(2) (within the standard range; measured by Bottemanne Weather Instruments Photosynthetic Radiometer RA200 Q)

EFFECT PARAMETERS MEASURED:
- Determination of algal growth: One sample from each flask was taken after 0, 23.5, 47.5, 73 and 96.5 h in OECD medium or after 0, 24, 46.5, 71.0 and 94 h in EPA medium, and the number of algal particles per mL in the samples was counted with Coulter Multisizer IIe. The numbers of algae in the test series were obtained by subtraction of the number of particles in the background control series (without algae) from the number of particles in the test series.
- Determination of algal biomass: Mean total particle volume was calculated from the number of particles and the given size distribution of the particle-counter channels. As the algae have a tendency for synchronous growth, a regular logistic growth curve cannot be obtained from the cell numbers with these species.
- Microscopic evaluation: The morphology of the algae was examined visually with the aid of a microscope also at the start and after 72 h.

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
OECD medium: 1.6 – 3.5
EPA medium: 1.7 – 3.2
- Range finding study: Yes. A range finding study was performed in EPA medium at 0, 0.2, 2.0, 20.2, 202 and 2,020 mg/L. After 3 d of incubation one sample was taken from each flask, and the number of particles per mL in the samples was analysed. Details on range finding study is provided in the study report.
- Results used to determine the conditions for the definitive study: Yes
The range finding study revealed that inhibiting effects could be expected at test material concentrations ≥ 2 mg/L. However, a complete growth inhibition was not achieved at a concentration of 202 mg/L.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate and or 3, 5-dichlorophenol
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
5.7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: In OECD media
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
34.1 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 14.6-19.2 (in OECD media)
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
16.5 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 9.33-14.3 (in OECD media)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
10.7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: In EPA media
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
70.1
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 3.04-70.9 (in EPA media)
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
41.8 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 2.80-43.1 (in EPA media)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
8.2 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: Average of results in OECD and EPA media.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
52.1 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: Average of results in OECD and EPA media.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
29.2 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: Average of results in OECD and EPA media.
Details on results:
- Exponential growth in the control: Yes, after 72 h of exposure, mean of particle counts/mL in the controls was 7.5 x 10(4) and 9.1 x 10(4) in OECD and EPA medium respectively. These represent 38-fold and 30-fold increases over time 0, respectively.
- Microscopic evaluation: Microscopic inspection of the morphology of algal cells in the pre culture at the start of the test revealed normal cells. At the end of the test no abnormal cells were observed in any test concentration.
- Any stimulation of growth found in any treatment: No
- Effect concentrations exceeding solubility of substance in test medium: No
- pH effects: The pH of the OECD medium at the start and end was approx. pH 8. The pH of the EPA medium at the start of the test was approx. 7, but increased to values as high as pH 9.6 after 72 h incubation, due to less buffering capacity. The EPA medium is clearly less buffered than the OECD medium.
Results with reference substance (positive control):
- Results with reference substance valid: Yes, the results were in the valid range. The test system used was checked with reference substances (K2Cr207 and/or 3,5 dichlorophenol) on an approx. yearly basis during an extended period of at least 15 yrs.
Reported statistics and error estimates:
- Results were updated to methods in OECD 201 (2006). Recalculations were performed using data existing within the study report. New 72 h ErC50 and EbC50 endpoint values were derived using the ECx program compiled in SAS v. 9.1.3 based on Bruce and Versteeg (1992).

- The NOEC was estimated by visual comparison of the measured and calculated growth values of the treated algal suspensions with those of the control.
Algal Cell Counts: Effect of MEA on Desmodesmus quadricauda in OECD Medium (study # 51456)
Nominal test conc. (mg/L) Cell Counts (x 1E4/ml) at Time (hr): OECD
0 23.5 47.5 73 96.5
0 0.2 0.4 1.5 7.5 13.5
1 0.2 0.4 1.2 6.7 13.4
3.5 0.2 0.3 1.2 6.0 11.1
5.7 0.2 0.3 1.2 6.6 15.5
10.2 0.2 0.3 1.0 4.8 8.7
18.3 0.2 0.3 0.8 3.4 5.6
32.5 0.2 0.3 0.6 1.2 2.8
102 0.2 0.2 0.2 0.3 0.4
325 0.2 0.2 0.2 0.2 0.1
Cell counts are mean of 4 flasks (control) or 2 flasks (all other test concentrations).
MEA = monoethanolamine (CAS # 141-43-5).
Test conditions: in OECD medium.
Algal Cell Counts: Effect of MEA on Desmodesmus quadricauda in EPA Medium (study # 51456)
Nominal test conc. (mg/L) Cell Counts (x 1E4/ml) at Time (hr): EPA
0 24 46.5 71 94
0 0.3 0.6 1.7 9.2 47.6
1.1 0.3 0.6 1.7 9.2 41.0
3.4 0.3 0.6 1.6 9.0 38.6
10.7 0.3 0.6 1.7 8.1 32.0
34.3 0.3 0.6 1.4 5.0 21.2
107 0.3 0.4 0.8 1.2 4.0
193 0.3 0.4 0.6 0.6 4.2
343 0.3 0.3 0.4 0.4 5.9
1071 0.3 0.3 0.3 0.4 7.5
Cell counts are mean of 4 flasks (control) or 2 flasks (all other test concentrations).
MEA = monoethanolamine (CAS # 141-43-5).
Test conditions: in EPA medium.
Validity criteria fulfilled:
yes
Conclusions:
The 72 h EC50 values of 2-aminoethanolamine (MEA) to Scenedesmus quadricauda for biomass and growth rate were 16.5 and 34.1 mg/L in OECD medium, respectively, and 41.9 and 70.2 mg/L in EPA medium, respectively. The results were based on nominal concentrations; measured concentrations were >80% of nominal.
Executive summary:

The toxicity of 2-aminoethanolamine (MEA) toScenedesmus quadricauda was determined in a 72 h algae growth inhibition test, following the OECD 201 guideline. The test was conducted for 96 hr, but the results are based on 72 hr.

This algal toxicity test was conducted using two different media, OECD medium (ammonia as nitrogen source) and EPA medium (nitrate as nitrogen source).

The nominal test concentrations were:

0, 1.0, 3.5, 5.7, 10.2, 18.3, 32.5, 102 and 325 mg/L (OECD medium)

0, 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L (EPA medium)

Measured concentrationswere > 80% of nominal up to 48 h incubation in the OECD medium, and up to 96 h incubation in the EPA medium. Therefore, the results were based on nominal concentration.

Endpoints evaluated were growth rate and biomass on the basis of mean cell counts over a period of 72 h. There were 2 replicates for each test concentration and 4 for the control.

The No Observed Effect Concentration (NOEC) was 5.7 and 10.7 mg/L in OECD and EPA medium respectively.

The 72 h EC50 for biomass and growth rate were as follows:

ErC50 (OECD medium): 34.1 mg/L

ErC50 (EPA medium): 70.1 mg/L

EbC50 (OECD medium): 16.5 mg/L

EbC50 (EPA medium): 41.9 mg/L.

Averaging the results in OECD and EPA media, the 72 hr results were:

NOEC: 8.2 mg/L

ErC50: 52.1 mg/L

EbC50: 29.2 mg/L

This toxicity test is classified as acceptable and satisfies the guideline requirements for the OECD Guideline 201.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From Dec. 06, 1999 to Dec. 24, 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
2 replicates per test concentration against the guideline requirement of 3
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Deviations:
yes
Remarks:
2 replicates per test concentration against the guideline requirement of 3
GLP compliance:
yes
Remarks:
OECD principles of GLP
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All test concentrations at 0, 48, 96 hr.
- Sampling method:
(i) At 0 h: 5 mL samples were collected from the test and control media without algae
(ii) At 48 and 96 h: 5 mL samples were collected from each test flask and two controls (all containing algae)
- Sample storage conditions before analysis: All samples immediately placed into 20 mL polyethylene vials, frozen and stored at a temperature of < -20°C. After the termination of the test, the samples were shipped frozen, packed in dry ice, to Procter & Gamble Eurocor for analysis.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
This test was conducted with two different algal growth media: OECD (ammonia as nitrogen source), and EPA medium (nitrate as nitrogen source).

1. OECD medium: An amount of 253.7 mg test material was dissolved in 5,000 mL OECD medium in order to condition the relevant glass-ware. Further, for the growth inhibition test 516.9 mg of test material was dissolved in 500 mL OECD medium. The pH of the solution was adjusted to 8.0 by addition of 1 M HCl solution.

2. EPA medium: 253.3 mg of test material was dissolved in 5,000 mL EPA medium in order to condition the relevant glass-ware. Further, for the growth inhibition test, 1,089 mg of test material was dissolved in 1,000 mL EPA medium. The pH of the solution was adjusted to 7.4 by addition of 1 M HCl solution.

- Application of test solution: Test solution application was carried out by adding appropriate volumes of the stock solution to the test flasks so as to yield the required test concentrations.
- Controls: Prepared using OECD or EPA medium containing algae only and a single background series containing test substance without algae.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Green alga
- Strain number: SAG 86.81
- Class: Chlorophyceae
- Source: Collection of Algal Cultures, Institute for Plant Physiology, University of Gottingen, Nikolausberger Weg 18, D-37073 Gottingen, Germany
- Age of inoculum: Not reported
- Method of cultivation: Pre-cultures of algal species in the exponential growth phase were prepared as detailed in OECD and EPA Guideline on the basis of total cell volume, using the OECD and EPA media for ammonia as nitrogen source and nitrate as nitrogen source respectively.
1. In OECD medium: A suspension of algae in OECD medium containing a total cell volume of 0.7 x 10(8) µm3/mL was prepared by dilution of a pre-culture containing 1.7 x 10(8) µm3/mL. Addition of 1 mL of this algal suspension to 100 mL of the appropriate solutions of the test substance yielded a mean inoculum cell density of 0.9 x 10(4) cells/mL (measured in the control cultures).

2. In EPA medium: A suspension of algae in EPA medium containing a total cell volume of 0.7 x 10(8) µm3/mL was prepared by dilution of a pre-culture containing 2.4 x 10(8) µm3/mL. Addition of 1 mL of this algal suspension to 100 mL of the appropriate solutions of the test substance yielded a mean inoculum cell density of 0.9 x 10(4) cells/mL (measured in the control cultures).
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
None
Hardness:
24.2 mg equivalent CaCO3/L (in OECD medium)
5.9 mg equivalent CaCO3/L (in EPA medium)
Test temperature:
23±2˚C
The temperature was determined continuously and was within the required limits.
pH:
Overall, in OECD medium, pH ranged 8.1 - 8.3:
8.1 – 8.2 (at test start, without algae)
8.1 – 8.3 (at 72 h, with algae)
8.2 (at 72 h, without algae)

Overall, in EPA medium, pH ranged 7.0 - 8.0:
7.0 – 7.4 (at test start, without algae)
7.3 - 8.0 (at 72 h, with algae)
7.0 – 7.4 (at 72 h, without algae)

pH values were within the acceptable limits.
Nominal and measured concentrations:
- Nominal test concentrations in OECD medium were:
0, 1.0, 3.5, 5.7, 11.2, 18.3, 32.5, 102 and 325 mg/L.
- Measured test concentrations in OECD medium, 0-48 hr, were:
0, 0.76, 3.23, 4.96, 11.4, 15.6, 31.8, 91.2, and NA mg/L.
- Measured test concentrations in OECD media, 0-48 hr, were 90% of nominal (range, 76-102%). (NA = not measured.)

- Nominal test concentrations in EPA medium were:
0, 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L.
- Measured test concentrations in EPA medium, 0-96 hr, were:
0, 1.01, 3.14, 9.84, 37.8, 102.2, NA, NA, and NA mg/L.
- Measured test concentrations in EPA media, 0-96 hr, were 96% of nominal (range, 92-110%).

According to EU guideline, the effect values should be based on nominal test concentrations if the measured concentrations were > 80% of nominal during 48 h incubation in OECD medium, and > 80% of nominal during 96 h incubation in the EPA medium. These criteria were met in both test media. Therefore, the results were based on nominal test concentration.

Details on measured concentration are provided under ‘Any other information on materials and methods incl. tables’ section.
Details on test conditions:
TEST SYSTEM
- Test vessel: Conical flasks covered with silicon sponge caps
- Type: Open
- Material: Sterile 200 mL conical test flasks containing 1 mL of algal suspension and 100 mL of test material solution.
- Aeration: No
- Agitation: Test containers were placed on a Gallenkamp orbital shaker and oscillated at approx. 100 rpm
- Initial cells density: 0.9 x 10(4) cells/mL in OECD and EPA medium
- Control end cells density (mean of replicates): 69.1 x 10(4) and 124.9 x 10(4) particles/mL (corrected for background) in OECD and EPA medium
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2 controls containing algae only
- No. of vessels for background series: 1 containing test substance without algae for the correction of non-algal particles present in the test flasks.
- Pre-treatment of test flasks: The test flasks and the background control were pre-treated with test solutions to prevent a decrease in the available test concentration due to adsorption to the glassware. Solutions in either OECD or EPA medium containing 50 mg/L test material were placed in the test flasks for ≥24 h before the experimental start. Following the equilibration, the contents of the flasks were removed, and the flasks were rinsed ten times with ultrapure water.

GROWTH MEDIUM
- Standard medium used: Yes, OECD and EPA media.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared in accordance with standard OECD and EPA guideline. Detailed composition is provided in the study report (Annex A).
- Ca/mg ratio:
OECD medium: CaCl2.2H2O: 18 mg/L/ (∑MgCl2.6H2O: 12 mg/L, MgSO4.7H2O: 15 mg/L)
EPA medium: CaCl2. 2H2O: 4.41 mg/L/ (∑MgCl2.6H2O: 12.16 mg/L, MgSO4.7H2O: 14.7 mg/L)
- Hardness: 24.2 mg equivalent CaCO3/L for OECD medium; 5.9 mg equivalent CaCO3/L for EPA medium
No information on any other water parameters was provided in the study report.
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH was measured in all parent solutions at 0 h and after 72 h in selected cultures. The temperature was registered continuously.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No. However before the test start the pH of OECD and EPA medium was adjusted to 8.0 and 7.4 respectively.
- Photoperiod: Not reported
- Light intensity: 60-120 µmol/sec/m(2) (within the standard range; measured by Bottemanne Weather Instruments Photosynthetic Radiometer RA200 Q)

EFFECT PARAMETERS MEASURED:
- Determination of algal growth: One sample from each flask was taken after 0, 22, 45, 67 and 93.5 h in OECD medium or after 0, 23, 45, 69.5 and 91 h in EPA medium, and the number of algal particles per mL in the samples was counted with Coulter Multisizer IIe. The numbers of algae in the test series were obtained by subtraction of the number of particles in the background control series (without algae) from the number of particles in the test series.
- Determination of algal biomass: Mean total particle volume was calculated from the number of particles and the given size distribution of the particle-counter channels. As the algae have a tendency for synchronous growth, a regular logistic growth curve cannot be obtained from the cell numbers with these species.
- Microscopic evaluation: The morphology of the algae was examined visually with the aid of a microscope also at the start and after 72 h.

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
OECD medium: 1.2 – 3.5
EPA medium: 1.7 – 3.2
- Range finding study: Yes. A range finding study was performed with OECD medium at 0, 0.2, 2.1, 20.6, 206 and 2,059 mg/L. After 3 d of incubation one sample was taken from each flask, and the number of particles per mL in the samples was analyzed. Details on range finding study is provided in the study report.
- Results used to determine the conditions for the definitive study: Yes. The range finding study revealed that inhibiting effects could be expected at test material concentrations ≥ 2 mg/L.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate and/or 3, 5-dichlorophenol
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
5.7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: In OECD media
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
16.8 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 14.6-19.2 (in OECD media)
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
11.6 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 9.33-14.3 (in OECD media)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
3.4 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: In EPA media
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
14.7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 3.04-70.9 (in EPA media)
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
11 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 2.80-43.1 (in EPA media)
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
4.6 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Remarks on result:
other: Average of results in OECD and EPA media.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
15.8 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: Average of results in OECD and EPA media.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
11.3 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: Average of results in OECD and EPA media.
Details on results:
- Exponential growth in the control: Yes, after 72 h of exposure, mean of particle counts/ml in the controls was 21.7 x 10(4) and 26.1 x 10(4) in OECD and EPA medium respectively.
- Microscopic evaluation: Microscopic inspection of the morphology of algal cells in the pre culture at the start of the test revealed normal cells. At the end of the test no abnormal cells were observed in any test concentration.
- Any stimulation of growth found in any treatment: No. considerable bacterial growth was observed in the cultures containing high test material concentrations
- Effect concentrations exceeding solubility of substance in test medium: No
- pH effects: no

For details on cell counts, see 'Table 1 and 2' below in the ‘Any other information on results incl. tables’ section.
Results with reference substance (positive control):
- Results with reference substance valid: Yes, the results were in the valid range. The test system used was checked with reference substances (K2Cr207 and/or 3,5 dichlorophenol) on an approx. yearly basis during an extended period of at least 15 yrs.
Reported statistics and error estimates:
- Results were updated to methods in OECD 201 (2006). Recalculations were performed using data existing within the study report. New 72 h ErC50 and EbC50 endpoint values were derived using the ECx program compiled in SAS v. 9.1.3 based on Bruce and Versteeg (1992).
- The NOEC was estimated by visual comparison of the measured and calculated growth values of the treated algal suspensions with those of the control.
Algal Cell Counts: Effect of MEA on Desmodesmus subspicatus in OECD Medium (study # 51456)
Nominal test conc. (mg/L) Cell Counts (x 1E4/ml) at Time (hr): OECD
0 22 45 67 93.5
0 0.9 1.1 4.8 21.7 54.1
1 0.9 1.2 5.2 24.2 97.6
3.5 0.9 1.2 4.8 21.6 88.0
5.7 0.9 1.2 4.9 21.6 83.4
11.2 0.9 1.2 4.0 12.0 32.4
18.3 0.9 1.0 3.2 3.2 5.0
32.5 0.9 1.0 1.6 1.6 3.1
102 0.9 1.0 1.0 -1.0 -6.6
325 0.9 0.9 0.8 0.8 1.2
Cell counts are mean of 4 flasks (control) or 2 flasks (all other test concentrations).
MEA = monoethanolamine (CAS # 141-43-5).
Test conditions: in OECD medium.
Algal Cell Counts: Effect of MEA on Desmodesmus subspicatus in EPA Medium (study # 51456)
Nominal test conc. (mg/L) Cell Counts (x 1E4/ml) at Time (hr): EPA
0 23 45 69.5 91
0 0.9 1.7 5.5 27.9 125.0
1.1 0.9 1.6 5.4 27.6 126.8
3.4 0.9 1.6 5.2 26.8 122.0
10.7 0.9 1.6 3.7 12.6 30.8
34.3 0.9 1.2 2.2 3.0 3.8
107 0.9 0.9 1.1 1.0 1.1
193 0.9 0.8 0.9 0.6 6.0
343 0.9 0.8 0.8 1.0 20.5
1071 0.9 0.8 0.8 1.5 17.6
Cell counts are mean of 4 flasks (control) or 2 flasks (all other test concentrations).
MEA = monoethanolamine (CAS # 141-43-5).
Test conditions: in EPA medium.
Validity criteria fulfilled:
yes
Conclusions:
The 72 h EC50 values of 2-aminoethanol (MEA) to Scenedesmus subspicatus for biomass and growth rate were 11.6 and 16.8 mg/L in OECD medium, respectively, and were 11.0 and 14.7 mg/L in EPA medium, respectively. The results are based on nominal test concentrations; measured concentrations were >80% of nominal.
Executive summary:

The toxicity of 2-aminoethanol (MEA) to Scenedesmus subspicatus was determined in a 72 h algae growth inhibition test, following the OECD 201 guideline. The test was conducted for 96 hr, but results are based on 72 hr.

This algal toxicity test was performed in two different media, OECD medium (ammonia as nitrogen source) and EPA medium (nitrate as nitrogen source). The nominal test concentrations were:

0 (control), 1.0, 3.5, 5.7, 11.2, 18.3, 32.5, 102 and 325 mg/L (OECD medium)

0 (control), 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L (EPA medium)

Measured concentrations were >80% of nominal, up to 48 h incubation in the OECD medium, and up to 96 h incubation in the EPA medium. Therefore, the results were based on nominal test concentration.

Endpoints evaluated were growth rate and yield (biomass) on the basis of mean cell count over a period of 72 h. There were 2 replicates for each test concentration and 4 replicates for the control.

The No Observed Effect Concentration (NOEC) was 5.7 and 3.4 mg/L in OECD and EPA medium respectively.

The 72 h EC50 for biomass and growth rate were as follows:

ErC50 (OECD medium): 16.8 mg/L

ErC50 (EPA medium): 14.7 mg/L

EbC50 (OECD medium): 11.6 mg/L

EbC50 (EPA medium): 11.0 mg/L

Averaging the results in OECD and EPA media, the 72 hr results were:

NOEC: 4.6 mg/L

ErC50: 15.8 mg/L

EbC50: 11.3 mg/L

This toxicity test is classified as acceptable and satisfies the guideline requirements for the OECD Guideline 201.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From Oct. 03, 2011 to Oct. 06, 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
no
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All test concentrations
- Sampling method: Duplicate samples of each test solution were extracted for analysis at 0, 24, 48 and 72 h
- Sample storage conditions before analysis: All samples were collected and immediately preserved with 50% (v/v) methanol to prevent degradation. Test solutions were stored at 4˚C prior to analysis.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solutions were prepared immediately prior to initiating the test at a concentration of 500 mg/L. The dosing solutions were then individually prepared in 100 mL volumetric glass cylinders in fresh 1XAAP media (minimal nutrient media) which was prepared from concentrated stock solutions in Milli-Q filtered water, adjusted to pH 7.51 and sterilized by micropore filtration.
- Controls: Prepared using the 1XAAP media without test material.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: Green alga
- Strain number: Not reported
- Class: Chlorophyceae
- Source: Carolina Biological Supply Company, Unialgal Cultures, 2700 York St. Burlington, NC, 27215
- Age of inoculum: 4 d old preculture
- Method of cultivation: The organisms were cultured using minimal nutrient media (1XAAP Media: 1 mL/L Media Stocks in High Quality water) in 250 mL flasks. The organisms were subcultured regularly on a weekly basis. The density of culture was approx. 4,250,000 cells/mL.
Details on conditions during culture are provided in the study report (Table 1)

- Application of test organism: The preculture was directly pipetted into the test and control replicates to receive an initial cell concentration of approx. 1 x 10(4) cells/mL.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Post exposure observation period:
None
Hardness:
Not reported
Test temperature:
Temperature in media control: 22.0˚C (at test start); 25.0˚C (at test termination)
Temperature in test plates: 21.9˚C (at test start at 60 mg/L); 25.0˚C (at test termination; at both 3.75 and 60 mg/L)
The temperature was determined continuously and was within the required limits of 24±2˚C.
pH:
pH in control: 7.59 (at test start); 8.48 (at test termination)
pH in test plates: 10.01 (at test start at 60 mg/L); 8.21 and 8.24 (at test termination at 3.75 and 60 mg/L respectively)
Nominal and measured concentrations:
Nominal test concentrations were: 0, 3.75, 7.50, 15.0, 30.0 and 60.0 mg/L
Geometric mean measured test concentration: 0, 4.37, 8.70, 17.13, 35.47 and 72.06 mg/L
The geometric mean measured concentrations were 114-120% of the nominal concentration. Results were based on measured concentration.
Details on test conditions:
TEST SYSTEM
- Test vessel: 6-well Falcon polystyrene plates
- Type: Open
- Material, size, headspace, fill volume: Each well of Falcon polystyrene plates received 10 mL of test solution and an aliquot of algae.
- Aeration: No
- Agitation: The plates were incubated in a Percival Scientific Series 101 incubator equipped with constant rotary agitation at 100 rpm.
- Initial cells density: Approx. 1 x 104 cells/mL (348 fluorescence units)
- Control end cells density (mean of replicates): 25,931 fluorescence units
- No. of wells per concentration (replicates): 3
- No. of wells per control (replicates): 3
- Background wells containing media without algae: 2

GROWTH MEDIUM
- Standard medium used: Yes, AAP media.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared in accordance with standard OECD guideline. Detailed composition is provided in the study report (Table 2).
- Ca/mg ratio: CaCl2.2H2O: 4.4 g/L/ (∑MgCl2.6H2O: 12.16 g/L, MgSO4.7H2O: 14.7 g/L)
No information on any other water parameters was provided in the study report.
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH and temperature was measured in high dose group at 0 h and in high and low dose group after 72 and in the media control at start and termination of test.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No, however before the test start the pH of AAP medium was adjusted to 7.51.
- Photoperiod: 24 h light
- Light intensity: 62 µE/m(2)

EFFECT PARAMETERS MEASURED:
- Cell density: The cell density was measured daily by quantifying chlorophyll by in vivo fluorometry in all control and test vessels.
- Microscopic evaluation: The morphology of the algae was examined at the start and after 72 h. The cells were checked for any unusual cell shapes, colour differences, differences in chloroplast morphology, and adherence of algae to test containers or aggregation of algal cells.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: Not conducted. Definitive concentrations were based on a plethora of internal data as well as data from the literature.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
22.6 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 12.82-39.81 mg/L
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
10.6 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 3.66 - 30.9
Details on results:
- Exponential growth in the control: Yes, after 72 h of exposure, mean fluorescence in the controls had increased 74.5-fold.
- % inhibition: The test material showed an initial stimulatory affect before displaying any inhibition.
For details, see 'Table 1' below in the ‘Any other information on results incl. tables’ section.
Reported statistics and error estimates:
- The results of the toxicity test were expressed as a NOEC (no observed effect concentrations), and as an ECx, where x is the percentage of growth reduction the test substance caused.
- Growth inhibition information was summarized using the ECx program compiled in ‘R’ based on Bruce and Versteeg (1992) or ToxCalc 5.0 to generate 72 h ErC10 and ErC50 statistics. ErC-values were defined as the effect concentration with regard to the growth rate during logistic or exponential growth (0-72 h).
- The No Observed Effect Concentration (NOEC) was calculated using JMP Pro 9.

Table 1: Effect of MEA on Scenedesmus subspicatus: 72 h algal growth inhibition test (study # 78305)

Nominal MEA concentration (mg/L)

Geometric mean MEA concentration (mg/L)

Mean Growth Rate

% Inhibition

0

0

1.4652

0

3.75

4.37

1.9883

-35.70

7.50

8.70

2.1584

-47.31

15.00

17.13

0.8658

40.91

30.00

35.47

0.5929

59.53

60.00

72.06

-0.0718

104.90

Test material was MEA (Monoethanolamine), CAS # 141-43-5.

Table 2: Summary of EC calculations based on Bruce and Versteeg (1992) (study # 78305)

Parameter

Estimate (mg/L)

95% Confidence Limits (mg/L)

ErC10

10.63

3.66 – 30.90

ErC50

22.59

12.82 – 39.81

Test material was MEA (Monoethanolamine), CAS # 141-43-5.

Validity criteria fulfilled:
yes
Conclusions:
The 72 h ErC50 of monoethanolamine (MEA) on Scenedesmus subspicatus was 22.6 mg/L, based on geometric mean measured concentration.
Executive summary:

The effect of Monoethanolamine (MEA) on Scenedesmus subspicatus was determined in a 72 h algae growth inhibition test, following the OECD 201 guideline. Reagent-grade MEA from Aldrich was the test substance. Also tested in this study were MEA-LAS (monoethanolamine salt of linear alkylbenzene sulfonate), and sodium LAS.

The nominal test concentrations of MEA were 0, 3.75, 7.50, 15.0, 30.0 and 60.0 mg/L.

Measured concentrations were114-120% of nominal. Results were based on geometric mean measured concentration.

Endpoints evaluated were growth rate and morphology over a period of 72 h. There were 3 replicates for each test concentration and for the control.

The 72 h ErC50 was 22.6 mg/L, based on measured concentrations.

A total of 3 test materials were evaluated in this study. Those results, all based on measured concentrations, were:

    Test Material     72 h ErC50 (mg/L)

           MEA                         22.6

           LAS                             7.39

           MEA-LAS                   1.44

This toxicity test is classified as acceptable and satisfies the guideline requirements for the OECD Guideline 201.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From April 13, 1992 to April 30, 1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
only 2 replicates per test concentration were used against the guideline requirement of 3
GLP compliance:
yes
Remarks:
OECD principles of GLP
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All concentrations at the start and end of the test.
- Sampling method:
(i) At test start: 100 mL samples were collected from the test and control media (without algae)
(ii) At test end: 100 mL samples were collected from all test concentrations (with and without algae)
- Sample storage conditions before analysis: All samples were frozen immediately and send to sponsor for analysis. The results of analysis are not available in this study report.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solutions of the test material were prepared by dissolving 20.0 mg in 1000 mL algal medium.
- Application of test solution: Test solution application was carried out by preparing appropriate dilutions of the stock solution in the test flasks containing algal medium so as to yield the required test concentrations.
- Controls: Prepared using algal medium containing algae only and a single background series containing test substance without algae.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Green alga
- Strain number: ATCC 22662
- Class: Chlorophyceae
- Source: American Type Culture Collection, 12301 Parklawn Drive. Rockville. Maryland 20852, USA
- Age of inoculum: Not reported
- Method of cultivation: A preculture of algae in the exponential growth phase was prepared as detailed in OECD Guideline using the algal medium.
- Preparation of test inoculum: A suspension of algae in the algal medium containing 2 x 10(4) cells/mL was prepared by dilution of a preculture containing 7.8 x 10(5) cells/mL. Addition of 50 mL of this algal suspension to 50 mL of the appropriate solutions of the test material in the test flasks, yielded a mean inoculum cell density measured in the control cultures of 1.1 x 10(4) cells/mL.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
73 h
Post exposure observation period:
None
Hardness:
Not reported. There were 18 mg/L CaCl2(2H2O), based on the composition of the algal NH4-medium.
Test temperature:
23˚C
pH:
Overall, pH ranged 7.9 - 8.7 throughout the test.

7.9 (at test start)
8.2 (at test end)
8.2 – 8.7 (at test end, with algae)
Nominal and measured concentrations:
Nominal concentration: 0, 0.56, 1.0, 1.8, 3.2, 5.6 and 10 mg/L
No details on measured concentration are provided in the study report. The results are based on nominal concentration.
Details on test conditions:
TEST SYSTEM
- Test vessel: Conical flasks covered with silicon sponge caps
- Type: Open
- Material: Sterile 200 mL conical test flasks containing mL of test solution.
- Aeration: No
- Agitation: Test containers were placed on a Gallenkamp orbital shaker and oscillated at 100 rpm
- Initial cells density: 1.0 x 10(4) cells/mL
- Control end cells density (mean of replicates): 136.8 x 10(4) particles/mL (corrected for background)
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 4 controls containing algae only
- No. of vessels for background series: 1 containing test substance without algae for the correction of non-algal particles present in the test flasks.

GROWTH MEDIUM
- Standard medium used: Yes, OECD medium was used with 150 mg/L NaHCO3, not 50 mg/L NaHCO3 as specified in the OECD guideline.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared in accordance with standard OECD guideline. Detailed composition is provided in the study report (Annex D).
- Ca/mg ratio: CaCl2.2H2O: 18 mg/L/MgSO4.7H2O: 15 mg/L
No information on any other water parameters was provided in the study report.
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH was measured in at 0 h (medium without algae) and after 73 h in selected cultures.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No
- Photoperiod: Not reported
- Light intensity: 60-120 µmol/sec/m(2) (within the standard range; radiated by fluorescent lamps and measured by Bottemanne Weather Instruments Photosynthetic Radiometer RA200 Q)

EFFECT PARAMETERS MEASURED:
- Determination of total particle numbers: One sample from each flask was taken after 0, 22.5, 46.5 and 73 h and the number of algal particles per mL in the samples was counted with Coulter Counter model TAII. The numbers of algae in the test series were obtained by subtraction of the number of particles in the background control series (without algae) from the number of particles in the test series.

The effect values were calculated from mean total particle numbers.

- Microscopic evaluation: The morphology of the algae was examined visually with aid of a microscope at the start and the end of the test.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.7- 1.8
- Range finding study: Yes. A range finding study was performed as a simplified version of OECD Guideline at 0, 0.01, 0.10, 1.0, 10, 101 and 1008 mg/L. After 3 d of incubation one sample was taken from each flask, and the number of particles per mL in the samples was analyzed. Details on range finding study is provided in the study report.
- Results used to determine the conditions for the definitive study: Yes. The range-finding test revealed that inhibiting effects could be expected at test concentrations higher than 1.0 mg/L.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
5.1 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 4.2 – 6.1
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
6.8 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 5.6 – 10
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
3.2 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Details on results:
- Exponential growth in the control (mean of replicates): Yes, after 73 h, mean values of particle counts/mL in control group was 136.8 x 10(4), a 140-fold increase compared to time 0.
- Microscopic evaluation: Microscopic evaluation of the cells at start and end of the incubation period revealed no morphological abnormalities, only the algal cells exposed to the two highest concentrations tested (5.6 and 10 mg/L) showed an abnormal shape.
- Any stimulation of growth found in any treatment: No
- Effect concentrations exceeding solubility of substance in test medium: No
- pH effects: The pH of the algal medium at the start of the test was pH 7.9. The pH of the medium containing different test substance concentrations remained constant during the test (pH 8.2 at the end). In the presence of algae the pH of the test solutions was found to give a slight increase with algal cell density at the end of the test to pH 8.2-8.7.

For details of algal cell counts, refer to ‘Table 1’ under ‘Any other information on results incl. tables’ section.
Reported statistics and error estimates:
The EbC50 was calculated by means of a parametric model developed by Kooijman et al. (1983), assuming an error proportional to the number of cells. The EbC10 and EbC90 were calculated the same way.

The ErC50 was calculated by the method given in the OECD guideline (1984). The ErC50 was calculated by linear interpolation of a plot of the percentage reduction in growth against the log concentration of the test substance. The ErC10 and ErC90 were calculated the same way.

The No Observed Effect Concentration (NOEC) was estimated by visual comparison of the measured and calculated growth curves of the treated algal suspensions with those of the controls.

Table 1: Effect of MEA on mean particle counts in 104 particles/mL with green algae (Selenastrum capricornutum) (Study # 35483)

Time (h)

Nominal concentration (mg/L)

0

0.56

1

1.8

3.2

5.6

10

0

1.0

1.0

1.0

1.0

1.0

1.0

1.0

22.5

4.6

4.6

4.7

4.7

4.8

4.4

3.9

46.5

31.6

30.7

32.4

32.0

32.8

24.8

8.1

73.0

136.8

155.6

153.9

156.6

158.6

100.2

16.8

Test material was MEA (Monoethanolamine), CAS # 141-43-5.

Values are the mean of 4 flasks (control) or 2 flasks (all other test concentrations).

Validity criteria fulfilled:
yes
Remarks:
With the exception that the coefficient of variation of growth rate during entire test period was 12.2% (not 7%).
Conclusions:
The 72 h EC50 values of monoethanolamine (MEA) to Selenastrum capricornutum for growth rate and biomass were 5.1 and 6.8 mg/L respectively, based on nominal concentrations
Executive summary:

The toxicity of monoethanolamine (MEA) to Selenastrum capricornutum was determined in a 72 h algae growth inhibition test, following the OECD 201 guideline.

The nominal test concentrations were 0, 0.56, 1.0, 1.8, 3.2, 5.6 and 10 mg/L. Endpoints were growth rate and biomass on the basis of mean total particle number over a period of 73 h. Two replicates was tested for each test concentration and 4 replicates for the control.

The No Observed Effect Concentration (NOEC) was 3.2 mg/L.

The 72 h EC50 for biomass and growth rate were as follows:

ErC50: 5.1 mg/L

EbC50: 6.8 mg/L

This toxicity test is classified as acceptable and satisfies the guideline requirements for the OECD Guideline 201.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
From Dec. 06, 1999 to Dec. 24, 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
only 2 replicates per test concentration were used against the guideline requirement of 3.
GLP compliance:
yes
Remarks:
OECD principles of GLP
Analytical monitoring:
yes
Details on sampling:
- Concentrations: All concentrations at 0 and 169 h
- Sampling method:
(i) At 0 h: 100 mL samples were collected from the test and control media (without algae)
(ii) At 169 h: 100 mL samples were collected from all test concentrations (with and without algae)
- Sample storage conditions before analysis: All samples were stored deep frozen before transfer to sponsor for analysis. The results of analysis are not available in this study report.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solutions of the test material were prepared by dissolving 331.8 mg in 1000 mL algal medium.
- Application of test solution: Test solution application was carried out by preparing appropriate dilutions of the stock solution in the test flasks containing algal medium so as to yield the required test concentrations.
- Controls: Prepared using algal medium containing algae only and a single background series containing test substance without algae.
Test organisms (species):
Microcystis aeruginosa
Details on test organisms:
TEST ORGANISM
- Common name: Blue- green alga (Cyanobacteria)
- Strain number: Not reported
- Source: Dr T. Källqvist of the Norwegian Institute for Water Research (NIVA), P.O. Box 33,
Blindern, N-03130 Oslo, Norway
- Age of inoculum: Not reported
- Method of cultivation: A preculture of algae in the exponential growth phase was prepared as detailed in OECD Guideline using NH4 algal medium (specific to test organism). The concentration of chelators 0.028 mmol and phosporus 4.4 mg/L in the organism specific medium exceeded the guideline medium constituents limits of ≤0.001 mmol and 0.7 mg/L respectively. Microcystis aeruginosa, however, requires the use of this medium.
- Preparation of test inoculum: A suspension of algae in the algal medium containing 2 x 10(4) cells/mL was prepared by dilution of a preculture containing 1.2 x 10(6) cells/mL. Addition of 50 mL of this algal suspension to 50 mL of the appropriate solutions of the test material in the test flasks, yielded a mean inoculum cell density measured in the control cultures of 0.9 x 10(4) cells/mL.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
162 h
Post exposure observation period:
None
Hardness:
Not reported. There were 4 mg/L CaCl2(2H2O), based on the composition of the algal NH4-medium.
Test temperature:
23˚C
pH:
Overall pH range during test: 7.3 - 8.1.

7.9 (at test start)
7.9 – 8.0 (during first 69 h, without algae)
7.8 – 7.6 (after 169 h, without algae)
8.1 – 7.3 (after 169 h, with algae)
Nominal and measured concentrations:
Nominal concentration: 0, 3.2, 10, 56, 100 and 166 mg/L
No details on measured concentration are provided in the study report. The results are based on nominal concentration.
Details on test conditions:
TEST SYSTEM
- Test vessel: Conical flasks covered with silicon sponge caps
- Type: Open
- Materia: Sterile 200 mL conical test flasks containing 100 mL of test solution.
- Aeration: No
- Agitation: Test containers were placed on a Gallenkamp orbital shaker and oscillated at 100 rpm
- Initial cells density: 0.9 x 10(4) cells/mL
- Control end cells density (mean of replicates): 84.8 x 10(4) particles/mL
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 4 controls containing algae only
- No. of vessels for background series: 1 containing test substance without algae for the correction of non-algal particles present in the test flasks.

GROWTH MEDIUM
- Standard medium used: Yes, with the addition of 0.028 mmol of chelators and 4.4 mg/L phosporus in the medium, which exceeded the guideline medium constituents limits of ≤0.001 mmol and 0.7 mg/L respectively. These were added to support the requirement of Microcystis aeruginosa.
- Detailed composition of medium (NH4 medium, per L):
NH4Cl: 38 mg
K2HPO4: 25 mg
MgSO4. 7H2O: 50 mg
CaCl2. 2H2O: 5 mg
NaHCO3: 99 mg
Fe(III) citrate: 5.17 mg
Na2EDTA.2H2O: 4 mg
1 mL trace element solution
Milli Q water to 1 L

Concentrated trace element solution (per L):
H3BO3: 2.88 g
MnCl2. 4H2O: 2.85 g
ZnSO4.7H2O: 0.4 g
CuSO4.5H2O: 0.08 g
Na2MoO4.2H2O: 0.003 g
CoCl2.6H2O: 0.104 g
NaO3V: 0.01 g
In 1L Milli-Q-Water

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared in accordance with standard OECD guideline. Detailed composition is provided in the study report (Annex D). Additional chelator (EDTA) and phosphorus were added, to support the growth of Microcystis aeruginosa.
- Ca/mg ratio: CaCl2.2H2O: 5 mg/L / MgSO4.7H2O: 50 mg/L
No information on any other water parameters was provided in the study report.
- Culture medium different from test medium: No
- Intervals of water quality measurement: pH was measured in at 0 h (medium without algae) and after 69 and 162 h in selected cultures. The pH measurement after 162 h was added because of the low growth rate of the test organism.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No. However before the test start the pH of the algal medium was adjusted to 8.0 using approx. 5 mL of a 2 M HCl solution.
- Photoperiod: Not reported
- Light intensity: 60-120 µmol/sec/m(2) (within the standard range; radiated by fluorescent lamps and measured by Bottemanne Weather Instruments Photosynthetic Radiometer RA200 Q)

EFFECT PARAMETERS MEASURED:
- Determination of total particle numbers: One sample from each flask was taken after 0, 22, 47, 69 and 162 h (the latter due to the relatively low growth rate of the organism) and the number of algal particles per mL in the samples was counted with Coulter Counter model TAII. The numbers of algae in the test series were obtained by subtraction of the number of particles in the background control series (without algae) from the number of particles in the test series.

- Determination of total particle volume: The total volumes in the test series was obtained by correction of the size distribution for the total particle volumes in the background series. The mean total particle volume values calculated were used for further calculations.

The effect values were calculated using total particle volume values instead of particle numbers because the particle numbers were disturbed by other bacterial growth.

- Microscopic evaluation: The morphology of the algae was examined visually with aid of a microscope at the start and the end of the test.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.6 – 3.2
- Range finding study: Yes. A range finding study was performed as a simplified version of OECD Guideline at 0, 0.01, 0.10, 1.0, 10, 101 and 1006 mg/L. After 3 d of incubation one sample was taken from each flask, and the number of particles per mL in the samples was analyzed. Details on range finding study is provided in the study report.
- Results used to determine the conditions for the definitive study: Yes. The range-finding test revealed that inhibiting effects could be expected at test concentrations higher than 1.0 mg/L.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
76.7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Remarks on result:
other: 71.0-82.8
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
47 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks on result:
other: 32-56
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
other: growth rate and biomass
Details on results:
- Exponential growth in the control (mean of replicates): Yes, after 162 h, mean values of particle counts/ mL in control group was 84.8 x 10(4), a 94-fold increase compared to time 0.
- Microscopic evaluation: Microscopic evaluation of the cells at start and end of the incubation period revealed no morphological abnormalities. Bacteria were found in increasing numbers with the test material concentration.
- Any stimulation of growth found in any treatment: No
For details refer to ‘Table 1’ and ‘Table 2’ under ‘Any other information on results incl. tables’ section.
- Effect concentrations exceeding solubility of substance in test medium: No
- pH effects: The pH of the algal medium was found to be pH 7.9 at the start of the test. The pH of the test solutions remained constant during the first 69 h (pH 7.9-8.0). After 169 h the pH showed a small decrease with the test concentrations (pH 7.8-7.6). This tendency was even stronger in presence of algae (pH 8.1-7.3).
Reported statistics and error estimates:
The No Observed Effect Concentration (NOEC) was estimated by visual comparison of the measured and calculated growth curves of the treated algal suspensions with those of the controls.

Table 1: Effect of MEA on mean particle counts in 104particles/mL with blue green algae (Microcystis aeruginosa) (Study # 35482)

Time (h)

Nominal concentration (mg/L)

0

0

3.2

10

32

56

100

166

0

0.9

0.9

0.9

0.9

0.9

0.9

0.9

0.9

22

2.0

1.7

1.8

2.1

2.1

1.8

1.4

1.3

47

4.1

5.2

8.6

25.7

14.3

12.9

5.5

4.6

69

6.3

6.8

7.1

10.5

7.6

7.5

6.3

4.7

162

84.1

85.6

79.4

72.5

492.0

27.9

122.0

13.9

Test material was MEA (Monoethanolamine), CAS # 141-43-5.

Table 2: Effect of MEA on mean total particle volume in 106 µm3/mL with blue green algae (Microcystis aeruginosa) (Study # 35482)

Time (h)

Nominal concentration (mg/L)

0

0

3.2

10

32

56

100

166

0

0.8

0.8

0.8

0.8

0.8

0.8

0.7

0.8

22

1.4

1.4

1.5

1.5

1.4

1.4

1.3

1.1

47

3.7

3.7

4.2

6.2

4.6

1.5

1.3

1.8

69

5.8

6.6

5.7

5.6

5.4

4.4

2.9

1.2

162

77.0

78.1

74.4

68.8

60.0

25.1

0.8

3.4

Test material was MEA (Monoethanolamine), CAS # 141-43-5.

Validity criteria fulfilled:
yes
Conclusions:
The 72 h EC50 values of monoethanolamine (MEA) to Microcystis aeruginosa for growth rate and biomass were 76.7 and 47 mg/L respectively, based on nominal concentrations.
Executive summary:

The toxicity of Monoethanolamine (MEA) to Microcystis aeruginosa was determined in a 162 h algae growth inhibition test, following the OECD 201 guideline. This algae growth inhibition test was run longer (162 h) than the typical test (72 h), because of the relatively slow growth of the test organism.

The nominal test concentrations were 0, 3.2, 10, 56, 100 and 166 mg/L. Endpoints evaluated were growth rate and biomass on the basis of mean total particle volume over a period of 162 h. Two replicates were tested for each test concentration and 4 replicates for control.

The No Observed Effect Concentration (NOEC) was 10.0 mg/L.

The 72 h EC50 for biomass and growth rate were as follows:

ErC50: 76.7 mg/L

EbC50: 47 mg/L.

This toxicity test is classified as acceptable and satisfies the guideline requirements for the OECD Guideline 201.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
(Q)SAR
Adequacy of study:
supporting study
Study period:
28/03/2017
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification
Remarks:
This local QSAR is suitable for predicting the acute toxicity of LAS to algae with chain lengths ranging from 10 to 14 carbons long.
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Version / remarks:
QSAR is built on 72 hr algal toxicity studies following OECD 201 guidelines
Principles of method if other than guideline:
A linear regression was used to generate a QSAR describing the relationship between acute toxicity and chain length of LAS for freshwater algae. QSAR has an r-square of 0.9738. It is suitable for LAS pure homologues and averaged chain lengths based on molecular weight percent. The counter ions of LAS salts are excluded by the model calculations. 72 hr ErC50: log(EC50)= -0.3028x + 5.3915, where x is the chain length of the alkyl carbon backbone.

GLP compliance:
not specified
Specific details on test material used for the study:
72 hr ErC50s were calculated for LAS based on tested materials with average chain lengths of 10.03, 10.95, 11.87, 13.00, and 13.81.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Test type:
static
Water media type:
freshwater
Total exposure duration:
72 h
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
75.69 mg/L
Nominal / measured:
estimated
Conc. based on:
other:
Basis for effect:
not specified
Remarks on result:
other: Predicted
Remarks:
Based on experimental results of LAS homologues with chain lengths C10-14.
Details on results:
The result is predicted for an average LAS chain length of 11.6. Experimental toxicity data was used to generate this QSAR.

 

Average chain length

 

EC50 (mg/L)

 

SD

 10.03  270  8.9
 10.95  111  8.0
 11.87  48  2.7
 13.00  30  1.3
 13.81  18  0.9
Conclusions:
The 72 hour ErC50 to algae (Scenedesmus subspicatus) is predicted to be 75.69 mg/L for MEA-LAS with an average chain length of 11.6.
Executive summary:

The 72 hour ErC50 to algae (Scenedesmus subspicatus) is predicted to be 75.69 mg/L for MEA-LAS with an average chain length of 11.6. The QSAR is based on "pure" LAS homologues tested for algal toxicity. The tested LAS chain lengths are C10, C11, C12, C13, and C14. The QSAR can be used to predict algal toxicity of LAS with pure chain lengths or averaged based on molecular weight percent. log(EC50)= -0.3028x + 5.3915, where x is the chain length of the alkyl carbon backbone.

The model is applicable to LAS C10 -14.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
(Test concentrations were not reported; results were based on the nominal concentrations rather than the measured concentrations)
GLP compliance:
not specified
Analytical monitoring:
not specified
Details on sampling:
Not specified
Vehicle:
not specified
Details on test solutions:
No information on the preparation of test solution were provided. The composition of C10-LAS tested in this study was 0.5%
Test organisms (species):
other: Scenedesmus subspicatus (Desmodesmus subspicatus)
Details on test organisms:
Scenedesmus subspicatus (Desmodesmus subspicatus) ATCC N-11464 French strain (obtained from University of Metz)
Test type:
not specified
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
Not reported
pH:
Not reported
Dissolved oxygen:
Not reported
Salinity:
Not reported
Conductivity:
Not reported
Nominal and measured concentrations:
Not reported
Details on test conditions:
The test was conducted according to OECD guideline 201 and test medium recommended by AFNOR was used. The composition of the test medium was as follows:
i) 40 mg/L Ca(NO3)2.4H2O
ii) 100 mg/L KNO3
iii) 30 mg/L MgSO4.7H2O
iv) 40 mg/L K2HPO4
v) 0.015 mg/L CuSO4.5H2O
vi) 0.03 mg/L (NH4)6 Mo7O24.4H2O
vii) 0.03 mg/L ZnSO4.7H2O
viii) 0.03 mg/L CoCl2.6H2O
ix) 0.03 mg/L Mn(NO3)2.4H2O
x) 0.03 mg/L C6H5O7.H2O
xi) 0.03 mg/L BO3H3
xii) 0.8125 mg/L C6H5FeO7.5H2O
xiii) 0.3125 mg/L FeSO4.7H2O
xiv) 0.3125 mg/L FeCl3.6H2O
In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2.
Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
26.2 mg/L
Basis for effect:
growth rate
Remarks on result:
other: NOEC was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
80 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
88.5 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
270 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
The 72-hr EC50 value of C10-LAS in Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)) was 270 mg/L. The 72-hr NOEC was 80 mg/L.

These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.

Table 1: Non-normalized Toxicity of pure LAS homologues usingScenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus))

 

Test substance

72-hr EC50 value (Std. deviation) (mg/L; based on growth rate)

72-hr NOEC (mg/L; based on growth rate)

Na-C10-LAS

270 (8.9)

80

Na-C11-LAS

111 (8)

40

Na-C12-LAS

48 (2.7)

18

Na-C13-LAS

30 (1.3)

12

Na-C14-LAS

18 (0.9)

7

 

Validity criteria fulfilled:
not specified
Conclusions:
Based on growth rate, the 72-hr EC50 and NOEC values of C10-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 88.5 mg/L and 26.2 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results were 270 mg/L and 80 mg/L.
Executive summary:

The algal toxicity test of C10-linear alkylbenzene sulfonates (C10-LAS) was conducted in accordance with OECD guideline 201, using the test medium recommended by AFNOR.

 

Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C10-LAS tested in this study was0.5% <C10, 96.8% C10, 2.7% C11. The molecular weight of C10-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C10-NaLAS (i.e. 320.7) provided in Table 2 of the publication. Therefore, the molecular weight of C10-LAS was 297.71.

 

In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.

 

Based on growth rate, the 72-hr EC50 and NOEC values of C10-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 88.5 mg/L and 26.2 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized results were 270 mg/L and 80 mg/L.

This toxicity study is classified as acceptable and satisfies the guideline requirements for the OECD 201 freshwater algae and cyanobacteria toxicity test.


Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
(Test concentrations were not reported; results were based on the nominal concentrations rather than the measured concentrations)
GLP compliance:
not specified
Analytical monitoring:
not specified
Details on sampling:
Not reported
Vehicle:
not specified
Details on test solutions:
No information on the preparation of test solution were provided. The composition of C11-LAS tested in this study was 5.5% C10, 93.7% C11 and 0.8% C12. The molecular weight of C11-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C11-NaLAS (i.e. 333.7) provided in Table 2 of the publication. Therefore, the molecular weight of C11-LAS was 310.71.
Test organisms (species):
other: Scenedesmus subspicatus (Desmodesmus subspicatus)
Details on test organisms:
Scenedesmus subspicatus (Desmodesmus subspicatus) ATCC N-11464 French strain (obtained from University of Metz)
Test type:
not specified
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
Not reported
pH:
Not reported
Dissolved oxygen:
Not reported
Salinity:
Not reported
Conductivity:
Not reported
Nominal and measured concentrations:
Not reported
Details on test conditions:
The test was conducted according to OECD guideline 201 and test medium recommended by AFNOR was used. The composition of the test medium was as follows:
i) 40 mg/L Ca(NO3)2.4H2O
ii) 100 mg/L KNO3
iii) 30 mg/L MgSO4.7H2O
iv) 40 mg/L K2HPO4
v) 0.015 mg/L CuSO4.5H2O
vi) 0.03 mg/L (NH4)6 Mo7O24.4H2O
vii) 0.03 mg/L ZnSO4.7H2O
viii) 0.03 mg/L CoCl2.6H2O
ix) 0.03 mg/L Mn(NO3)2.4H2O
x) 0.03 mg/L C6H5O7.H2O
xi) 0.03 mg/L BO3H3
xii) 0.8125 mg/L C6H5FeO7.5H2O
xiii) 0.3125 mg/L FeSO4.7H2O
xiv) 0.3125 mg/L FeCl3.6H2O
In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2.
Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
26.33 mg/L
Basis for effect:
growth rate
Remarks on result:
other: NOEC was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
40 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
73.1 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
111 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
The 72-hr EC50 value of C11-LAS in Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)) was 111 mg/L. The 72-hr NOEC was 40 mg/L.

These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.

Table 1: Non-normalized Toxicity of pure LAS homologues using Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)

 

Test substance

72-hr EC50 value (Std. deviation) (mg/L; based on growth rate)

72-hr NOEC (mg/L; based on growth rate)

Na-C10-LAS

270 (8.9)

80

Na-C11-LAS

111 (8)

40

Na-C12-LAS

48 (2.7)

18

Na-C13-LAS

30 (1.3)

12

Na-C14-LAS

18 (0.9)

7

 

Validity criteria fulfilled:
not specified
Conclusions:
Based on growth rate, the 72-hr EC50 and NOEC values of C11-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 73.1 mg/L and 26.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized were 111 mg/L and 40 mg/L.
Executive summary:

The algal toxicity test of C11-linear alkylbenzene sulfonates (C11-LAS) was conducted in accordance with OECD guideline 201, using the test medium recommended by AFNOR.

 

Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C11-LAS tested in this study was5.5% C10, 93.7% C11 and 0.8% C12. The molecular weight of C11-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C11-NaLAS (i.e. 333.7) provided in Table 2 of the publication. Therefore, the molecular weight of C11-LAS was 310.71.

 

In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.

 

Based on growth rate, the 72-hr EC50 and NOEC values of C11-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 73.1 mg/L and 26.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized were 111 mg/L and 40 mg/L.

This toxicity study is classified as acceptable and satisfies the guideline requirements for the OECD 201 freshwater algae and cyanobacteria toxicity test.


Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
(Test concentrations were not reported; results were based on the nominal concentrations rather than the measured concentrations)
GLP compliance:
not specified
Analytical monitoring:
not specified
Details on sampling:
Not reported
Vehicle:
not specified
Details on test solutions:
No information on the preparation of test solution were provided. The composition of C12-LAS tested in this study was 0.4% C10, 13.9% C11, 84.5% C12 and 1.2% C13. The molecular weight of C12-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C12-NaLAS (i.e. 346.4) provided in Table 2 of the publication. Therefore, the molecular weight of C12-LAS was 323.41.
Test organisms (species):
other: Scenedesmus subspicatus (Desmodesmus subspicatus)
Details on test organisms:
Scenedesmus subspicatus (Desmodesmus subspicatus) ATCC N-11464 French strain (obtained from University of Metz)
Test type:
not specified
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
Not reported
pH:
Not reported
Dissolved oxygen:
Not reported
Salinity:
Not reported
Conductivity:
Not reported
Nominal and measured concentrations:
Not reported
Details on test conditions:
The test was conducted according to OECD guideline 201 and test medium recommended by AFNOR was used. The composition of the test medium was as follows:
i) 40 mg/L Ca(NO3)2.4H2O
ii) 100 mg/L KNO3
iii) 30 mg/L MgSO4.7H2O
iv) 40 mg/L K2HPO4
v) 0.015 mg/L CuSO4.5H2O
vi) 0.03 mg/L (NH4)6 Mo7O24.4H2O
vii) 0.03 mg/L ZnSO4.7H2O
viii) 0.03 mg/L CoCl2.6H2O
ix) 0.03 mg/L Mn(NO3)2.4H2O
x) 0.03 mg/L C6H5O7.H2O
xi) 0.03 mg/L BO3H3
xii) 0.8125 mg/L C6H5FeO7.5H2O
xiii) 0.3125 mg/L FeSO4.7H2O
xiv) 0.3125 mg/L FeCl3.6H2O
In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2.
Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
23.8 mg/L
Basis for effect:
growth rate
Remarks on result:
other: NOEC was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
18 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
63.4 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
48 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
The 72-hr EC50 value of C12-LAS in Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)) was 48 mg/L. The 72-hr NOEC was 18 mg/L.

These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.

Table 1: Non-normalized Toxicity of pure LAS homologues using Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus))

 

Test substance

72-hr EC50 value (Std. deviation) (mg/L; based on growth rate)

72-hr NOEC (mg/L; based on growth rate)

Na-C10-LAS

270 (8.9)

80

Na-C11-LAS

111 (8)

40

Na-C12-LAS

48 (2.7)

18

Na-C13-LAS

30 (1.3)

12

Na-C14-LAS

18 (0.9)

7

 

Validity criteria fulfilled:
not specified
Conclusions:
Based on growth rate, the 72-hr EC50 and NOEC values of C12-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 63.4 mg/L and 23.8 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized results are 48 mg/L and 18 mg/L.
Executive summary:

The algal toxicity test of C12-linear alkylbenzene sulfonates (C12-LAS) was conducted in accordance with OECD guideline 201, using the test medium recommended by AFNOR.

 

Scenedesmus subspicatus ATCC N-11464 French strain obtained from University of Metz was used as the tester strains for this study. The composition of C12-LAS tested in this study was0.4% C10, 13.9% C11, 84.5% C12 and 1.2% C13. The molecular weight of C12-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C12-NaLAS (i.e. 346.4) provided in Table 2 of the publication. Therefore, the molecular weight of C12-LAS was 323.41.

 

In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.

 

Based on growth rate, the 72-hr EC50 and NOEC values of C12-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 63.4 mg/L and 23.8 mg/L, respectively.  These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized results are 48 mg/L and 18 mg/L.

This toxicity study is classified as acceptable and satisfies the guideline requirements for the OECD 201 freshwater algae and cyanobacteria toxicity test.


Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
(Test concentrations were not reported; results were based on the nominal concentrations rather than the measured concentrations)
GLP compliance:
not specified
Analytical monitoring:
not specified
Details on sampling:
Not reported
Vehicle:
not specified
Details on test solutions:
No information on the preparation of test solution were provided. The composition of C13-LAS tested in this study was 0.7% C11, 9.8% C12, 78.3% C13 and 11.2% C14. The molecular weight of C13-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C13-NaLAS (i.e. 362.3) provided in Table 2 of the publication. Therefore, the molecular weight of C13-LAS was 339.31.
Test organisms (species):
other: Scenedesmus subspicatus (Desmodesmus subspicatus)
Details on test organisms:
Scenedesmus subspicatus (Desmodesmus subspicatus) ATCC N-11464 French strain (obtained from University of Metz)
Test type:
not specified
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
Not reported
pH:
Not reported
Dissolved oxygen:
Not reported
Salinity:
Not reported
Conductivity:
Not reported
Nominal and measured concentrations:
Not reported
Details on test conditions:
The test was conducted according to OECD guideline 201 and test medium recommended by AFNOR was used. The composition of the test medium was as follows:
i) 40 mg/L Ca(NO3)2.4H2O
ii) 100 mg/L KNO3
iii) 30 mg/L MgSO4.7H2O
iv) 40 mg/L K2HPO4
v) 0.015 mg/L CuSO4.5H2O
vi) 0.03 mg/L (NH4)6 Mo7O24.4H2O
vii) 0.03 mg/L ZnSO4.7H2O
viii) 0.03 mg/L CoCl2.6H2O
ix) 0.03 mg/L Mn(NO3)2.4H2O
x) 0.03 mg/L C6H5O7.H2O
xi) 0.03 mg/L BO3H3
xii) 0.8125 mg/L C6H5FeO7.5H2O
xiii) 0.3125 mg/L FeSO4.7H2O
xiv) 0.3125 mg/L FeCl3.6H2O
In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2.
Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
31.8 mg/L
Basis for effect:
growth rate
Remarks on result:
other: NOEC was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
12 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
79.6 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
30 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
The 72-hr EC50 value of C13-LAS in Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)) was 30 mg/L. The 72-hr NOEC was 12 mg/L.

These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary.

Table 1: Non-normalized Toxicity of pure LAS homologues using Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)

 

Test substance

72-hr EC50 value (Std. deviation) (mg/L; based on growth rate)

72-hr NOEC (mg/L; based on growth rate)

Na-C10-LAS

270 (8.9)

80

Na-C11-LAS

111 (8)

40

Na-C12-LAS

48 (2.7)

18

Na-C13-LAS

30 (1.3)

12

Na-C14-LAS

18 (0.9)

7

 

Validity criteria fulfilled:
not specified
Conclusions:
Based on growth rate, the 72-hr EC50 and NOEC values of C13-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 79.6 mg/L and 31.8 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results were 30 mg/L and 12 mg/L.
Executive summary:

The algal toxicity test of C13-linear alkylbenzene sulfonates (C13-LAS) was conducted in accordance with OECD guideline 201, using the test medium recommended by AFNOR.

 

Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C13-LAS tested in this study was 0.7% C11, 9.8% C12, 78.3% C13 and 11.2% C14. The molecular weight of C13-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C13-NaLAS (i.e. 362.3) provided in Table 2 of the publication. Therefore, the molecular weight of C13-LAS was 339.31.

 

In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.

 

Based on growth rate, the 72-hr EC50 and NOEC values of C13-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 79.6 mg/L and 31.8 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results were 30 mg/L and 12 mg/L.


Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Justification for type of information:
Refer to the section 13 of the dataset for details on the read across justification.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Deviations:
yes
Remarks:
(Test concentrations were not reported; results were based on the nominal concentrations rather than the measured concentrations)
GLP compliance:
not specified
Analytical monitoring:
not specified
Details on sampling:
Not reported
Vehicle:
not specified
Details on test solutions:
No information on the preparation of test solution were provided. The composition of C14-LAS tested in this study was 0.6% C11, 1% C12, 15.4% C13, 82.1% C14 and 0.9% >C14. The molecular weight of C14-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C14-NaLAS (i.e. 373.7) provided in Table 2 of the publication. Therefore, the molecular weight of C14-LAS was 350.71.
Test organisms (species):
other: Scenedesmus subspicatus (Desmodesmus subspicatus)
Details on test organisms:
Scenedesmus subspicatus (Desmodesmus subspicatus) ATCC N-11464 French strain (obtained from University of Metz)
Test type:
not specified
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
Not reported
pH:
Not reported
Dissolved oxygen:
Not reported
Salinity:
Not reported
Conductivity:
Not reported
Nominal and measured concentrations:
Not reported
Details on test conditions:
The test was conducted according to OECD guideline 201 and test medium recommended by AFNOR was used. The composition of the test medium was as follows:
i) 40 mg/L Ca(NO3)2.4H2O
ii) 100 mg/L KNO3
iii) 30 mg/L MgSO4.7H2O
iv) 40 mg/L K2HPO4
v) 0.015 mg/L CuSO4.5H2O
vi) 0.03 mg/L (NH4)6 Mo7O24.4H2O
vii) 0.03 mg/L ZnSO4.7H2O
viii) 0.03 mg/L CoCl2.6H2O
ix) 0.03 mg/L Mn(NO3)2.4H2O
x) 0.03 mg/L C6H5O7.H2O
xi) 0.03 mg/L BO3H3
xii) 0.8125 mg/L C6H5FeO7.5H2O
xiii) 0.3125 mg/L FeSO4.7H2O
xiv) 0.3125 mg/L FeCl3.6H2O
In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2.
Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
37.3 mg/L
Basis for effect:
growth rate
Remarks on result:
other: NOEC was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
7 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
95.9 mg/L
Basis for effect:
growth rate
Remarks on result:
other: EC50 was normalized to reflect the toxicity of LAS with a carbon chain length 11.6
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
18 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Details on results:
The 72-hr EC50 value of C14-LAS in Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus)) was 18 mg/L. The 72-hr NOEC was 7 mg/L.
These experimental results were normalized to reflect a C11.6 chain length using a highly localized LAS-specific QSAR (Belanger et al, 2016). Toxicity normalization was performed using this basic fo rmula:

ChVc11.6norm = ChVx * (ChVc11.6pred / ChVxpred)

Where ChV is any chronic endpoint result; such as NOEC, LOEC, EC10, etc., ChVc11.6norm is the normalized ChV for the test material,
ChVx is the experimental ChV for the test material,
ChVc11.6pred is the QSAR predicted ChV for the target material with a chain length of C11.6, ChVxpred is the QSAR predicted ChV for a material with the same chain length as the test
material.

A QMRF is available on the QSAR (Belanger et al, 2016) used to provide the predicted results for normalization. More details on this approach are available in the endpoint summary

Table 1: Non-normalized Toxicity of pure LAS homologues using Scenedesmus subspicatus (ATCC N-11464 French strain (Desmodesmus subspicatus))

 

Test substance

72-hr EC50 value (Std. deviation) (mg/L; based on growth rate)

72-hr NOEC (mg/L; based on growth rate)

Na-C10-LAS

270 (8.9)

80

Na-C11-LAS

111 (8)

40

Na-C12-LAS

48 (2.7)

18

Na-C13-LAS

30 (1.3)

12

Na-C14-LAS

18 (0.9)

7

 

Validity criteria fulfilled:
not specified
Conclusions:
Based on growth rate, the 72-hr EC50 and NOEC values of C14-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 95.9 mg/L and 37.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized results were 18 mg/L and 7 mg/L.
Executive summary:

The algal toxicity test of C14-linear alkylbenzene sulfonates (C14-LAS) was conducted in accordance with OECD guideline 201, using the test medium recommended by AFNOR.

 

Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C14-LAS tested in this study was0.6% C11, 1% C12, 15.4% C13, 82.1% C14 and 0.9% >C14. The molecular weight of C14-LAS was adjusted by subtracting the weight of sodium (i.e. 22.99) from the molecular weight of C14-NaLAS (i.e. 373.7) provided in Table 2 of the publication. Therefore, the molecular weight of C14-LAS was 350.71.

 

In the beginning, the cell concentration was adjusted to 10(4) cells/mL, using a Coulter Counter ZM. During the study, the culture flasks were shaken to facilitate the transfer of CO2. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval.

 

Based on growth rate, the 72-hr EC50 and NOEC values of C14-LAS in Scenedesmus subspicatus (French strain (Desmodesmus subspicatus)) were 95.9 mg/L and 37.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-no rmalized results were 18 mg/L and 7 mg/L.

This toxicity study is classified as acceptable and satisfies the guideline requirements for the OECD 201 freshwater algae and cyanobacteria toxicity test.


Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
Dec. 8, 1992 To Dec. 11, 1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
(A dilution factor of 4 was used in study instead of guideline recommendation of ≤3.2)
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
no
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Source (laboratory, culture collection): Institut fur Wasser-, Boden- und Lufthygiene, Berlin

ACCLIMATION
- Acclimation period: 3 days
Test type:
static
Total exposure duration:
72 h
Test temperature:
24 +/- 2 degree C
pH:
7.7-9.0
Nominal and measured concentrations:
0, 0.6, 2.4, 10, 40, and 160 mg/L
Details on test conditions:
TEST SYSTEM
- Initial cells density: 2 x 10^4 cells/ml
- Control end cells density: 95 x 10^4 cells/ml
- No. of vessels per concentration (replicates): 8
- No. of vessels per control (replicates): 8

OTHER TEST CONDITIONS
- Light intensity and quality: 8000 Lux
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
47.3 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
127.9 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
2.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Details on results:
The pH as measured at the end of the experiment was very high (pH=9.0), however, as growth was not affect, the experiment was considered valid.

Cell count ( x 104 cells/mL)

Concentration (mg/L)

0 hrs

24 hrs

48 hrs

72 hrs

Control

2

8

29

95

0.6

2

8

30

89

2.4

2

9

28

85

10

2

9

27

75

40

2

8

20

48

160

2

7

9

8

Validity criteria fulfilled:
yes
Conclusions:
The 72-hr NOEC for algae was 2.4 mg/L. The 72-hr EbC50 was 47.3 mg/L, and the 72-hr ErC50 was 127.9 mg/L for algae.
Executive summary:

Scenedesmus subspicatus was exposed to concentrations of 0, 0.6, 2.4, 10, 40, or 160 mg/L of test substance for 72 hrs. The cell concentration was measured at 24, 48, and 72 hrs. The 72-hr NOEC for algae was 2.4 mg/L. The 72-hr EbC50 was 47.3 mg/L, and the 72-hr ErC50 was 127.9 mg/L for algae. The reported endpoints do not require normalization since the compound tested was C11.6 LAS.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
Jun. 7, 1984 To Jun. 22, 1984
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to
Guideline:
other: EPA-600/9-78-018
Deviations:
yes
Remarks:
Chlorella kessleri test species was used instead of guideline recommendation of Selenastrum capricornutum and test duration was 15 days instead of 96-hr as per EPA guideline.
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
not specified
Details on sampling:
no information
Details on test solutions:
A stock solution of 1,000 mg product/1000 mL DI water was prepared. From this, a second stock solution was prepared with a dilution factor of 20.
Test organisms (species):
other: Chlorella kessleri
Test type:
static
Water media type:
not specified
Total exposure duration:
15 d
Nominal and measured concentrations:
0, 0.01, 0.03, 0.1, 0.3, 1, 3, 10, 30, 100, 300, 1000 mg/L
Details on test conditions:
TEST SYSTEM
- Material, size, headspace, fill volume: 100 mL DI water, and 10 mL mineral medium
- Initial cells density: 10^4 cells/mL
- Control end cells density: 3.7 x 10^15
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionized water, AAP-Medium


OTHER TEST CONDITIONS
- Light intensity and quality: ~2000 lux


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : number of cells was determined on days 5, 7, 12, and 15
Key result
Duration:
15 d
Dose descriptor:
NOEC
Effect conc.:
3.1 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
cell number
Key result
Duration:
15 d
Dose descriptor:
other: FOEC
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
cell number

Number of cells/ml (average of 3 replicates)

Concentration (mg/L)

5 days

7 days

12 days

15 days

0

7.2 x 105

1.5 x 106

3.9 x 106

3.7 x 106

0.01

5.9 x 105

1.4 x 106

2.4 x 106

3.2 x 106

0.03

3.0 x 105

1.4 x 106

3.4 x 106

3.6 x 106

0.1

2.6 x 105

1.3 x 106

2.4 x 106

3.0 x 106

0.3

2.5 x 105

1.3 x 106

3.2 x 106

3.5 x 106

1

2.6 x 105

1.3 x 106

2.5 x 106

2.8 x 106

3

1.6 x 105

1.0 x 106

2.3 x 106

3.2 x 106

10

3.9 x 105

6.3 x 105

1.5 x 106

1.7 x 106

30

1.9 x 105

3.9 x 105

1.1 x 106

1.4 x 106

100

2.9 x 104

4.5 x 104

4.2 x 104

6.1 x 104

300

1.3 x 104

2.7 x 104

1.9 x 104

1.1 x 104

1000

1.6 x 104

2.6 x 104

2.5 x 104

2.0 x 104

Conclusions:
The NOEC for algae was 3.1 mg/L (based on active ingredient), and the FOEC was 10 mg/L (based on active ingredient).
Executive summary:

Chlorella kessleri algae was exposed to concentrations of 0, 0.01, 0.03, 0.1, 0.3, 1, 3, 10, 30, 100, 300, and 1000 mg/L (nominal) of test substance for 15 days. The number of cells were counted to determine the toxicity of the test substance. The NOEC was 3.1 mg/L, based on active ingredient, and the FOEC was 10 mg/L, based on active ingredient. The data are for C11.6 LAS and no normalization is required.

Description of key information

MEA-LAS dissociates into MEA and LAS in liquid media and the acute toxicity algae and aquatic plants of both compounds has been studied. Modelling has also been conducted for MEA-LAS. 

A robust and highly localized LAS (Q)SAR was developed for plant and algal data normalization (Verge and Moreno 1996). The model was built from high quality Desmodesmus subspicatus toxicity data using thoroughly characterized LAS compounds (C10-14). Further external validation was performed using LAS mixtures with different average alkyl chain lengths. Reporting Format (QMRF) documents describing data quality, model development and performance of the models are appended to the Robust study Summaries (RSS).

Study 1 (C10-14 LAS):

In a 96 h algal growth inhibition test, the blue-green alga Microcystis aeruginosa was exposed to C10-14 LAS, sodium salt (average chain length C11.9) in accordance with Payne and Hall, evaluated against US EPA OPPTS 850.5400 (1996). The nominal test concentrations were 0, 0.01, 0.05, 0.1, 0.5, 1 and 1000 mg a.i./L. The 96 h ErC10, ErC20 and ErC50, based on growth rate, were 0.89, 2.24 and 13.44 mg/L, respectively. These results are normalized to reflect the toxicity of a LAS with a carbon chain length of 11.6 (Verge and Moreno, 1996). The non-normalized ErC10, ErC20 and ErC50 are 0.715, 1.82 and 10.9 mg/L, respectively (Ward, 1982).  

Study 2 (MEA-LAS, (Q)SAR):

The 72 h ErC50 to algae (Scenedesmus (renamed Desmosdesmus) subspicatus) was predicted to be 75.69 mg/L for an MEA-LAS with an average chain length of 11.6. The (Q)SAR was based on algal toxicity data using “pure”, thoroughly characterized LAS homologues. This characterization included a detailed analysis of the homologue distribution. The tested LAS chain lengths range from C10 to C14. The (Q)SAR can be used to predict algal toxicity of LAS with pure chain lengths or averaged based on molecular weight percent.

log(EC50)= -0.3028x + 5.3915, where x is the chain length of the alkyl carbon backbone (Verge, 1996).

Study 3 (C10 LAS):

An algal toxicity test with C10 LAS, sodium salt was conducted in accordance with OECD Guideline 201, using the test medium recommended by AFNOR. Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strain. The composition of C10 LAS tested in this study was 0.5% <C10, 96.8% C10, 2.7% C11. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval. Based on growth rate, the 72 h EC50 and NOEC values were 88.5 and 26.2 mg/L, respectively. These results are normalized to reflect the toxicity of a LAS with a carbon chain length of 11.6. The non-normalized results were 270 and 80 mg/L, respectively (Verge, 1996).

Study 4 (C11 LAS):

An algal toxicity test with C11 LAS, sodium salt was conducted in accordance with OECD Guideline 201, using the test medium recommended by AFNOR. Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C11 LAS tested in this study was 5.5% C10, 93.7% C11 and 0.8% C12. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval. Based on growth rate, the 72 h EC50 and NOEC values were 73.1 and 26.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized were 111 and 40 mg/L, respectively (Verge, 1996).

Study 5 (C12 LAS):

An algal toxicity test with C12 LAS, sodium salt was conducted in accordance with OECD Guideline 201, using the test medium recommended by AFNOR. Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C12 LAS tested in this study was 0.4% C10, 13.9% C11, 84.5% C12 and 1.2% C13. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval. Based on growth rate, the 72 h EC50 and NOEC values were 63.4 and 23.8 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results are 48 and 18 mg/L, respectively (Verge, 1996).

Study 6 (C13 LAS):

An algal toxicity test with C13 LAS, sodium salt was conducted in accordance with OECD Guideline 201, using the test medium recommended by AFNOR. Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C13 LAS tested in this study was 0.7% C11, 9.8% C12, 78.3% C13 and 11.2% C14. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval. Based on growth rate, the 72 h EC50 and NOEC values were 79.6 and 31.8 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results were 30 and 12 mg/L, respectively (Verge, 1996).

Study 7 (C14 LAS):

An algal toxicity test with C14 LAS, sodium salt was conducted in accordance with OECD Guideline 201, using the test medium recommended by AFNOR. Scenedesmus subspicatus ATCC N-11464 French strain (Desmodesmus subspicatus) obtained from University of Metz was used as the tester strains for this study. The composition of C14 LAS tested in this study was 0.6% C11, 1% C12, 15.4% C13, 82.1% C14 and 0.9% >C14. Comparison of algal growth rates between the control and test groups were performed to derive the EC50 values. EC5 values were considered as NOEC since no noticeable variation was observed in the 0-5% interval. Based on growth rate, the 72 h EC50 and NOEC values were 95.9 and 37.3 mg/L, respectively. These results are normalized to reflect the toxicity of an LAS with a carbon chain length of 11.6. The non-normalized results were 18 and 7 mg/L ((Verge, 1996). 

Study 8 (C10-13 LAS):

Scenedesmus (renamed Desmodesmus) subspicatus was exposed to concentrations of 0, 0.6, 2.4, 10, 40 or 160 mg/L of C10-13 LAS, sodium salt (average chain length C11.6) for 72 h according to OECD Guideline 201. The cell concentration was measured at 24, 48 and 72 h. The 72 h NOEC was 2.4 mg/L, the 72 h EbC50 was 47.3 mg/L and the 72 h ErC50 was 127.9 mg/L. The reported endpoints do not require normalization since the compound tested was C11.6 LAS (Scholz, 1992).

Study 9 (C10-13 LAS):

Chlorella kessleri algae was exposed to concentrations of 0, 0.01, 0.03, 0.1, 0.3, 1, 3, 10, 30, 100, 300 and 1000 mg/L (nominal) of C10-13 LAS, sodium salt (average chain length C11.6) for 15 d according to EPA Guideline 600/9-78-018. The number of cells were counted to determine the toxicity of the test substance. The NOEC was 3.1 mg/L, based on active ingredient. The data are for C11.6 LAS and no normalization is required (Muehlberg, 1984).

Study 10 (C10-14 LAS):

In a 72 h algal growth inhibition test, the green alga Selenastrum capricornutum (renamed Pseudokirchneriella subcapitata or Raphidocelis subcapitata) was exposed to C10-14 LAS, sodium salt (average chain length C11.9) in accordance with US EPA OPPTS 850.5400 and OECD Guideline 201. The nominal test concentrations were 0, 16, 32, 63, 125, 250, 500 and 1000 mg a.i./L. The 72 h EC10 and EC50, based on growth rate, were 16.15 and 289.67 mg/L, respectively. These results are normalized to reflect the toxicity of a LAS with a carbon chain length of 11.6 (Ward, 1982; Lewis, 1986, Verge, 1996). 

Study 11 (MEA):

The toxicity of monoethanolamine (MEA) to Scenedesmus quadricauda was determined in a 72 h algae growth inhibition test, following OECD Guideline 201. The test was conducted for 96 h, but the results are based on 72 h. This algal toxicity test was conducted using two different media, OECD medium (ammonia as nitrogen source) and EPA medium (nitrate as nitrogen source). The nominal test concentrations were: 0, 1.0, 3.5, 5.7, 10.2, 18.3, 32.5, 102 and 325 mg/L (OECD medium) and 0, 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L (EPA medium). Measured concentrations were >80% of nominal values up to 48 h of incubation in the OECD medium and up to 96 h of incubation in the EPA medium. Therefore, the results were based on nominal concentrations. Endpoints evaluated were growth rate and biomass on the basis of mean cell counts over a period of 72 h. There were 2 replicates for each test concentration and 4 for the control. The NOEC was 5.7 and 10.7 mg/L in OECD and EPA medium, respectively. The 72 h EC50 for biomass and growth rate were as follows: ErC50 (OECD medium): 34.1 mg/L, ErC50 (EPA medium): 70.1 mg/L, EbC50 (OECD medium): 16.5 mg/L and EbC50 (EPA medium): 41.9 mg/L. Averaging the results in OECD and EPA media, the 72 h results were: NOEC: 8.2 mg/L, ErC50: 52.1 mg/L and EbC50: 29.2 mg/L ( Hanstveit, 2000).

Study 12 (MEA):

The toxicity of monoethanolamine (MEA) to Scenedesmus (renamed Desmodesmus) subspicatus was determined in a 72 h algae growth inhibition test, following the OECD Guideline 201. The test was conducted for 96 h, but results are based on 72 h. This algal toxicity test was performed in two different media, OECD medium (ammonia as nitrogen source) and EPA medium (nitrate as nitrogen source).The nominal test concentrations were: 0, 1.0, 3.5, 5.7, 11.2, 18.3, 32.5, 102 and 325 mg/L (OECD medium) and 0, 1.1, 3.4, 10.7, 34.3, 107, 193, 343 and 1071 mg/L (EPA medium). Measured concentrations were >80% of nominal values up to 48 h of incubation in the OECD medium and up to 96 h of incubation in the EPA medium. Therefore, the results were based on nominal test concentrations. Endpoints evaluated were growth rate and yield (biomass) on the basis of mean cell count over a period of 72 h. There were 2 replicates for each test concentration and 4 replicates for the control. The NOEC was 5.7 and 3.4 mg/L in OECD and EPA medium, respectively. The 72 h EC50 for biomass and growth rate were as follows: ErC50 (OECD medium): 16.8 mg/L, ErC50 (EPA medium): 14.7 mg/L, EbC50 (OECD medium): 11.6 mg/L a,d EbC50 (EPA medium): 11.0 mg/L. Averaging the results in OECD and EPA media, the 72 h results were: NOEC: 4.6 mg/L, ErC50: 15.8 mg/L and EbC50: 11.3 mg/L (Hanstveit, 2000).

Study 13 (MEA):

The effect of monoethanolamine (MEA) on Scenedesmus (renamed Desmodesmus) subspicatus was determined in a 72 h algae growth inhibition test, following OECD Guideline 201. Reagent-grade MEA from Aldrich was the test substance. MEA-LAS and Na-LAS were also tested in this study. The nominal test concentrations of MEA were 0, 3.75, 7.50, 15.0, 30.0 and 60.0 mg/L. Measured concentrations were 114-120% of nominal values. Endpoints evaluated were growth rate and morphology over a period of 72 h. There were 3 replicates for each test concentration and for the control. The 72 h ErC50 was 22.6 mg/L, based on geometric mean measured concentrations. The ErC50 for MEA-LAS and Na-LAS were 1.44 and 7.39 mg/L, respectively (Brill, 2011).

Study 14 (MEA):

The toxicity of monoethanolamine (MEA) to Selenastrum capricornutum (renamed Pseudokirchneriella subcapitata) was determined in a 72 h algae growth inhibition test, following OECD Guideline 201. The nominal test concentrations were 0, 0.56, 1.0, 1.8, 3.2, 5.6 and 10 mg/L. Endpoints were growth rate and biomass on the basis of mean total particle number over a period of 73 h. Two replicates was tested for each test concentration and 4 replicates for the control. The NOEC was 3.2 mg/L. The 72 h EC50 for biomass and growth rate were 5.1 and 6.8 mg/L, respectively (Hanstveit, 1992).

Study 15 (MEA):

The toxicity of monoethanolamine (MEA) to Microcystis aeruginosa was determined in a 162 h algae growth inhibition test, following the OECD Guideline 201. This algae growth inhibition test was run longer (162 h) than the typical test (72 h), because of the relatively slow growth of the test organisms. The nominal test concentrations were 0, 3.2, 10, 56, 100 and 166 mg/L. Endpoints evaluated were growth rate and biomass on the basis of mean total particle volume over a period of 162 h. Two replicates were tested for each test concentration and 4 replicates for control. The NOEC was 10.0 mg/L. The 72 h EC50 for biomass and growth rate were as follows: ErC50: 76.7 mg/L and EbC50: 47 mg/L (Hanstveit, 1992).

 

Key value for chemical safety assessment

EC50 for freshwater algae:
10.9 mg/L

Additional information

The point of departure for aquatic PNEC derivation is the 56 d NOEC of 0.268 mg/L from a mesocosm model ecosystem (Belanger, 1997, 2002 and 2004; Lowe, 1996).