[No public or meaningful name is available]

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2020-05-01 to 2020-05-07, with the definitive exposure from 2020-05-04 to 2020-05-07

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

ISO 10253 (Water quality - Marine Algal Growth Inhibition Test with Skeletonema costatum and Phaeodactylum tricornutum)

Principles of method if other than guideline:

None

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Determination of
Samples were analyzed with an LC-MS/MS method, implemented
the test item under non-GLP and documented finally in the GLP raw data. The method was validated under GLP based on the criteria set forth by SANCO/3029/99 rev.4 11/07/00.

Sampling schedule
All concentration levels and the control were analytically verified via LC-MS/MS at the start (0 hours) and at the end of the exposure (72 hours) with algae.

Sampling and pre-treatment
At the start, samples were taken from one additional replicate of the test item concentration and the control and at the end samples were taken from pooled replicates.

Vehicle:

no

Details on test solutions:

Preparation of the saturated solution
A saturated solution with a nominal loading rate of 100 mg test item/L was prepared once 72 ± 1 hour prior to the start of the exposure. An appropriate amount of the test item was pipetted onto the surface of the dilution water. For this purpose, the density was taken into account (0.98 g/cm3). A slow stirring procedure was applied for 72 ± 1 hour at room temperature in the dark. After a separation phase of 1 hour (at room temperature), the saturated solution was removed by siphoning (from the approximate bottom of the glass flask).


Preparation of the test item solutions
Out of the saturated solution 5 concentrations were tested in a geometric series with a dilution factor of 2:
6.25 – 12.5 – 25 – 50 – 100% of the saturated solution. The dilution levels are based on the results of a preliminary range finding test (non-GLP).


Control
Six replicates (without test item) were tested under the same test conditions as the test item replicates.

Test organisms (species):

Skeletonema costatum

Details on test organisms:

TEST ORGANISM
- Common name: Skeletonema costatum (GREVILLE) Cleve 1873, CCAP 1077/1C
- Strain: CCAP 1077/1C
- Source (laboratory, culture collection):
Culture Collection of Algae and Protozoa (CCAP)
SAMS Research Services Ltd
Dunstaffnage Marine Laboratory
Dunbeg, OBAN; Argyll PA37 1QA; Scotland, UK


- Age of inoculum (at test initiation): 4 days
- Method of cultivation: Fresh stocks were prepared every month in dilution water. Light intensity amounted to 2567 – 5130 lux corresponding to 35 - 70 µE ∙ m-2 ∙ s-1 for 24 h per day.

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

No

Hardness:

Not specified

Test temperature:

min.: 18.5 max.: 19.5 mean value: 19.0

pH:

Geometric mean measured test item concentration Nominal test item loading pH-value
[µg/L] [mg/L] Start; 0 hours End; 72 hours
165 100 8.21 7.99
53.2 50.0 8.27 8.06
22.7 25.0 8.32 8.14
10.4 12.5 8.34 8.14
4.52 6.25 8.35 8.14
Control 8.37 8.14

Dissolved oxygen:

Not measured

Salinity:

Not measured

Nominal and measured concentrations:

Nominal test item concentrations 6.25 - 12.5 - 25.0 - 50.0 - 100 % of the saturated solution, corresponding to nominal loadings of 6.25 to 100 mg/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type (delete if not applicable): open, cotton wool plugs
- Material, size, headspace, fill volume: sterile 250 mL Erlenmeyer flasks, test volume 100 mL
- Aeration: Test containers were placed on a rotary shaker and oscillated at appr. 40 rpm
- Type of flow-through (e.g. peristaltic or proportional diluter): None
- Renewal rate of test solution (frequency/flow rate): One application at test start
- Initial cells density: Nominal: approximately 1 x 10^4 cells/mL, Actual: mean 9439 cells/mL
- Control end cells density: Mean 161374 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6


GROWTH MEDIUM
- Standard medium used: yes


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:

According to the CCAP Administration double concentrated medium (F/2 + Si) with natural seawater (salinity of 30 +/- 5 S):
Composition of Dilution Water

Component Concentration [mg/L]
NaNO3 150
NaH2PO4  H2O 10
Na2SiO3  9 H2O 60
Titriplex III 8.32
FeCl3  6 H2O 6.30
CuSO4  5 H2O 0.02
ZnSO4  7 H2O 0.044
CoCl2  6 H2O 0.02
MnCl2  4 H2O 0.36
Na2MoO4  4 H2O 0.012
Vitamin B1 (Thiamin hydrochloride) 0.2
Vitamin H (Biotin) 0.1 µg/L
Vitamin B12 (Cyanocobalamine) 0.1 µg/L
pH 8.0 +/- 0.2

The pH of the test medium has to be 8.0 +/- 0.2 and was adjusted prior to testing with addition of 1 N NaOH and HCl, if necessary.

Characterization Data of Natural Seawater
Location Alfred-Wegener-Institut,
Biologische Anstalt Helgoland,
Kurpromenade 201, D-27498 Helgoland Latitude:54.11°, Longitude: 7.53°
Arrival Date 2019-09-24
pH 7.83
Salinity [S] 32.7
Pretreatment
The natural seawater was filtrated (membrane filter, 0.2 µm RC), adjusted to a salinity of 30 S with demineralized water and sterilized (autoclave d at 121 °C) until addition of the F/2 + Si components
Storage conditions Room temperature



OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: 24 h
- Light intensity and quality: Approximately 2567 – 3300 lux, measured: 2578 - 3046 lux with a mean of 2775 lux



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
-Chlorophyll a-fluorescence
The cell density was measured daily via Chlorophyll a-fluorescence, excitation at 436 nm, emission at 685 nm. Dilution water was used as a background signal. No self-fluorescence was observed in the preliminary range finding test at the saturated solution.

Reference substance (positive control):

yes

Remarks:

(Potassium dichromate)

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

22.7 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

22.7 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

53.2 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

53.2 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

38.9 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CI: 25.3 - 48.4

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

45.5 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CI: 30.2 - 51.1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

63.4 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CI: 55.9 - 99.5

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

32.5 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield inhibition

Remarks on result:

other: CI: 26.2 - 39.2

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

36.1 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield inhibition

Remarks on result:

other: CI: 29.4 - 41.8

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

46.5 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield inhibition

Remarks on result:

other: CI: 40.0 - 50.5

Duration:

72 h

Dose descriptor:

other: NOEL

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

other: LOEL

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

other: NOEL

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

other: LOEL

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

other: EL10

Effect conc.:

38.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Cl: 27.3 - 46.2

Duration:

72 h

Dose descriptor:

other: EL20

Effect conc.:

44.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Cl: 31.7 - 48.4

Duration:

72 h

Dose descriptor:

other: EL50

Effect conc.:

57.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Cl: 52.1 - 77.7

Duration:

72 h

Dose descriptor:

other: EL10

Effect conc.:

33.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: Cl: 28.1 - 38.9

Duration:

72 h

Dose descriptor:

other: EL20

Effect conc.:

36.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: Cl: 30.9 - 41.0

Duration:

72 h

Dose descriptor:

other: EL50

Effect conc.:

44.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: Cl: 39.8 - 47.9

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test):
- Unusual cell shape: No
- Colour differences: No
- Flocculation: No
- Adherence to test vessels: No
- Aggregation of algal cells: No
- Other: The test media were clear throughout exposure period (possible turbidity related to algae growth not taken into account).
- Effect concentrations exceeding solubility of substance in test medium: No

Results with reference substance (positive control):

The toxicity of 3,5-Dichlorophenol (SIGMA-ALDRICH, batch number MKBZ0947V, purity 100.0%, CAS RN 591-35-5) to the marine diatom alga Skeletonema costatum was determined over a period of 72 hours from 2020-03-09 to 2020-03-12. The reference item toxicity is in the valid range which was established by calculation of the average of the historic reference data since 2011, and the limits were set using the threefold standard deviation of these values.

EC50-Values of the Reference Item
based on nominal concentrations [mg/L], (0-72 hours)
Current Study Valid Range (average ± 3 x SD)
Growth Rate inhibition
ErC50 1.62 1.70 ± 0.910
95% confidence interval 1.58 – 1.65
Yield inhibition
EyC50 1.17 1.40 ± 0.914
95% confidence interval 1.07 – 1.27
SD = Standard deviation

Reported statistics and error estimates:

EC/EL-values and statistical analyses
EC/EL10, EC/EL20 and EC/EL50-values with confidence intervals of growth rate and yield inhibition after 72 hours were calculated by sigmoidal dose-response regression with the software GraphPad Prism.


NOECNOEL, LOEC/LOEL and statistical analyses
The NOEC/NOEL and LOEC/LOEL were determined by calculation of statistically significant differences of growth rate and yield using Dunnett’s Method. A Shapiro-Wilk’s Normality test and a Brown-Forsythe’s Equal Variance test were done first. P-values for both Normality and Equal Variance tests are 0.05. The -value (acceptable probability of incorrectly concluding that there is a difference) is  = 0.01. Data were transformed for growth rate (Y = Y squareroot).


Software
The data for the tables in this report were computer-generated and have been rounded for presentation from the full derived data. Consequently, if calculated manually based on the given data minor deviations may occur from these figures.
Calculations were carried out using software
•Excel, MICROSOFT CORPORATION
•SigmaPlot, SPSS INC.
•GraphPad Prism, GRAPHPAD SOFTWARE, INC.

Microscopic evaluation of the cells at the start and the end of exposure revealed no morphological abnormalities. All effect values given are based on the geometric mean measured test item concentrations and the nominal loadings of Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol, respectively.

Cell Densities

Geometric mean measured test item concentration       Nominal test item loading       Replicate       Cell density [cells/mL]

[µg/L]       [mg/L]       No.       0 hours       24 hours       48 hours       72 hours

165       100       1       9439       < LOQcell density       < LOQcell density       5065

             2       9439       < LOQcell density       6871       18329

             3       9439       2947       5282       19764

             Mean       9439       n.a.       n.a.       14386

53.2       50.0       1       9439       4501       23699       58610

             2       9439       5612       20903       63855

             3       9439       5134       32766       64994

             Mean       9439       5082       25789       62486

22.7       25.0       1       9439       17851       65637       159684

             2       9439       18836       62961       183615

             3       9439       20425       65056       174583

             Mean       9439       19037       64551       172627

10.4       12.5       1       9439       27481       71663       185847

             2       9439       22532       72460       178478

             3       9439       19525       84932       194652

             Mean       9439       23179       76352       186326

4.52       6.25       1       9439       24525       68826       172419

             2       9439       22071       67767       162111

             3       9439       22515       71446       193695

             Mean       9439       23037       69346       176075

Control       1       9439       26461       72745       152771

      2       9439       21962       80034       151142

      3       9439       24713       70398       185893

      4       9439       24161       65899       153648

      5       9439       22122       69054       155060

      6       9439       24183       73189       169730

      Mean       9439       23934       71887       161374

n.a. = not applicable

LOQcell density = 2712

 

Evaluation after 72 hours

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

Geometric mean measured test item concentration       Nominal test item loading       Repl.       Growth rate       Inhibition of growth rate       Yield       Inhibition of yield

[µg/L]       [mg/L]       No.       [d-1]       [%]       [cells/mL]       [%]

165       100       1              -0.21       100              -4374       100

             2              0.22       77              8890       94

             3              0.25       74              10325       93

             Mean       (+)       0.09       84       (+)       4947       96

53.2       50.0       1              0.61       36              49171       68

             2              0.64       33              54416       64

             3              0.64       32              55555       63

             Mean       (+)       0.63       33       (+)       53047       65

22.7       25.0       1              0.94       0              150245       1

             2              0.99       -5              174176       -15

             3              0.97       -3              165144       -9

             Mean       (-)       0.97       -2       (-)       163188       -7

10.4       12.5       1              0.99       -5              176408       -16

             2              0.98       -4              169039       -11

             3              1.01       -7              185213       -22

             Mean       (-)       0.99       -5       (+)*       176887       -16

4.52       6.25       1              0.97       -2              162980       -7

             2              0.95       0              152672       0

             3              1.01       -7              184256       -21

             Mean       (-)       0.97       -3       (-)       166636       -10

Control       1              0.93                     143332       

      2              0.92                     141703       

      3              0.99                     176454       

      4              0.93                     144209       

      5              0.93                     145621       

      6              0.96                     160291       

      Mean              0.95                     151935       

*       = statistical significance is caused by growth stimulation

 

Section-by-Section and Average Specific Growth Rates of the Control Group

      (0 - 72 hours)

      Replicate No.       Specific growth rate [d-1]       Mean

(0 - 72 hours)       SD

±       CV

[%]       Mean CV [%]

             section-by-section                            

             0 - 24 hours       24 - 48 hours       48 - 72 hours                            

Control       1       1.03       1.01       0.74       0.93       0.161       17.4       16.7

      2       0.84       1.29       0.64       0.92       0.336       36.3       

      3       0.96       1.05       0.97       0.99       0.046       4.67       

      4       0.94       1.00       0.85       0.93       0.079       8.48       

      5       0.85       1.14       0.81       0.93       0.179       19.2       

      6       0.94       1.11       0.84       0.96       0.135       14.0       

                    Mean       0.945       

                    SD ±       0.03       

                    CV [%]       2.90       

SD = Standard deviation       CV = Coefficient of variation

Validity criteria fulfilled:

yes

Conclusions:

In this study, Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol was found to inhibit the growth of the marine diatom alga Skeletonema costatum after 72 hours with the following effect values (based on geometric mean measured concentrations and nominal test item loadings): The EC50-values with 95% confidence intervals for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 63.4 (55.9 – 99.5) and 46.5 (40.0 – 50.5) µg test item/L, respectively. The EL50-values with 95% confidence intervals for inhibition of growth rate (ErL50) and yield (EyL50) after 72 hours were 57.5 (52.1 – 77.7) and 44.8 (39.8 – 47.9) mg test item/L, respectively. The NOEC/NOEL-values for both inhibition of growth rate and yield after 72 hours were 22.7 µg test item /L, corresponding to 25.0 mg test item/L.

Executive summary:

The toxicity of Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol (Batch no.: ESD0033040) to the marine diatom alga Skeletonema costatum was determined according to the principles of DIN EN ISO-Guideline 10253 from 2020-05-01 to 2020-05-07, with the definitive exposure from 2020-05-04 to 2020-05-07 at the test facility. The aim of the study was the determination of NOEC/NOEL, LOEC/LOEL, EC/EL10, EC/EL20 and EC/EL50-values of growth rate and yield over a period of 72 hours.

The study was conducted under static conditions with an initial cell density of 9439 cells/mL. A saturated solution of 100 mg test item/L was prepared once 72 ± 1 hour prior to the start of the exposure. An appropriate amount of the test item was pipetted onto the surface of the dilution water. For this purpose, the density was taken into account (0.98 g/cm3). A slow stirring procedure was applied for 72 ± 1 hour at room temperature in the dark. After a separation phase of 1 hour (at room temperature), the saturated solution was removed by siphoning (from the approximate bottom of the glass flask). Five concentrations were tested in a geometrical series with a dilution factor of 2 (nominal): 6.25 - 12.5 - 25.0 - 50.0 - 100 % of the saturated solution, corresponding to nominal loadings of 6.25 to 100 mg/L. Three replicates were tested for each test item concentration and six replicates for the control. The environmental conditions were within the acceptable limits.

The concentrations of the test item, Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol, were analytically verified by LC-MS/MS at the start and the end of the exposure of all test concentrations and the control. The analytical method was validated according to SANCO/3029/99 rev.4. The measured concentrations of the test item, Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol, at the start of the exposure (0 hours) were in the range of 24.6 to 713 µg/L. The measured concentrations of the test item at the end of the exposure (72 hours) were in the range of 0.828 to 38.2 µg/L, corresponding to 3 to 5% of the initially measured values.

NOEC, LOEC and ECx-values and 95% Confidence Intervals of Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol (0 - 72 hours)

based on the geometric mean measured test item concentration [mg/L]

      Growth Rate Inhibition                                                 

Geometric mean measured test item concentration [µg/L]       Growth Rate Inhibition

Nominal test item loading [mg/L]

NOEC/NOEL       22.7       25.0

LOEC/LOEL       53.2       50.0

ErC10 / ErL10       38.9 (25.3 – 48.4)       38.8 (27.3 – 46.2)

ErC20 / ErL20       45.5 (30.2 – 51.1)       44.1 (31.7 – 48.4)

ErC50 / ErL50       63.4 (55.9 – 99.5)       57.5 (52.1 – 77.7)

      Yield Inhibition

Geometric mean measured test item concentration [µg/L]       Yield Inhibition

Nominal test item loading [mg/L]

NOEC/NOEL       22.7       25.0

LOEC/LOEL       53.2       50.0

EyC10 / EyL10       32.5 (26.2 – 39.2)       33.5 (28.1 – 38.9)

EyC20 / EyL20       36.1 (29.4 – 41.8)       36.5 (30.9 – 41.0)

EyC50 / EyL50       46.5 (40.0 – 50.5)       44.8 (39.8 – 47.9)

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

08.03.07 - 16.03.07

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

ISO 10253 (Water quality - Marine Algal Growth Inhibition Test with Skeletonema costatum and Phaeodactylum tricornutum)

Version / remarks:

2006

GLP compliance:

yes

Specific details on test material used for the study:

Test item: Dodicor V 5654
Batch: 07-YS-019
Date test item received: 12.02.07

Analytical monitoring:

yes

Vehicle:

no

Test organisms (species):

Skeletonema costatum

Details on test organisms:

The strain of Skeletonema costatum (Greville) used in these tests is NIVA BAC 1. The culture mediUill
is prepared from natural seawater obtained from a depth of ca. 130 m. The salinity is adjusted to 30 ±5
%0. The culture medium is prepared from natural seawater by addition of 15 ml ISO nutrient stock
solution 1, 0.5 ml nutrient stock solution 2, and 1 ml nutrient stock solution 3 to 1 litres of seawater
(the compositions of the ISO nutrient media are given in Table 1 on page 12 of this report). The preculture
is maintained under the same conditions as used in the test (see Summary of Test Conditions).
Culture is semi-continuous under continuous even fluorescent lighting at an intensity of 60 - 90 )lmol
m-2
S-l, with continuous agitation on an orbital shaker.

Temperature is maintained at 20±2.0°C. Algal cultures are used for tests when they are detennined to
be in a phase of exponential growth. Cell density in the pre-culture is measured immediately before a
test in order to calculate the required inoculum volume.

Test temperature:

20 +- 2 °C

pH:

start 6.13 - 8.09
end 7.40 - 8.37

Salinity:

Natural seawater salinity 34 promille

Nominal and measured concentrations:

55.9 - 100 -321 - 559 - 1000 mg/L and a control

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

412 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

321 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Validity criteria fulfilled:

yes

Conclusions:

A growth inhibition test was performed with Skeletonema costatum using Dodicor V 5654 at
nominal concentrations of 55.9- 1000 mg r l
. The following results were obtained:
72-h EC50: 412 mg/L
24-h EC50 : 382 mg/L
48-h EC50 : 400 mg/L
72-hNOEC: 321 mg/L

Executive summary:

A growth inhibition test was performed withSkeletonema costatumusing Dodicor V 5654 at

nominal concentrations of 55.9- 1000 mg rl

.The following results were obtained:

72-hEC50: 412 mg/L

24-hEC50 : 382 mg/L

48-hEC50 : 400 mg/L

72-hNOEC: 321 mg/L

 

Comments: Stock solution of 1054 mg!l of the test item was prepared according to procedure D, as

described in the OSPAR Guidelines for Toxicity Testing of Substances and Preparations

Used and Discharged Offshore. See pages10-11of this report. Test solutions were

prepared from the Water Accommodated Fraction (WAF).

Note:test results obtained when the test item is prepared according to procedure A refer to

the nominal concentrations of the test item as stated. Test results obtained when the test

item is prepared according to procedures B, C or D refer only to Water-Accommodated

Fractions of the test item prepared at the nominal concentrations of the test item stated.

ECso (48 h) value has been calculated manually from the dose-response curve (see

comments to Table 3 on Page 17 of this report).

Optimal pH range for the algae test is 7.8 - 8.2. The two highest test concentrations of

Dodicor V 5654 showed pH-values below this range at the start of the test.

72-h NOEC refers to the highest test concentration showing no statistically significant

Description of key information

Two studies according ISO 10253 Guidline with Skeletonema costatum are available. 

In one study, Reaction products of DL-methionine and C18 unsaturated fatty acid chloride and isopropanol was found to inhibit the growth of the marine diatom alga Skeletonema costatum after 72 hours with the following effect values (based on geometric mean measured concentrations and nominal test item loadings): The EC50-values with 95% confidence intervals for inhibition of growth rate (ErC50) and yield (EyC50) after 72 hours were 63.4 (55.9 – 99.5) and 46.5 (40.0 – 50.5) µg test item/L, respectively. The EL50-values with 95% confidence intervals for inhibition of growth rate (ErL50) and yield (EyL50) after 72 hours were 57.5 (52.1 – 77.7) and 44.8 (39.8 – 47.9) mg test item/L, respectively. The NOEC/NOEL-values for both inhibition of growth rate and yield after 72 hours were 22.7 µg test item /L, corresponding to 25.0 mg test item/L.

A second growth inhibition test was performed with Skeletonema costatum using Dodicor V 5654 at

nominal concentrations of 55.9- 1000 mg r l

. The following results were obtained:

72-h EC50: 412 mg/L

24-h EC50 : 382 mg/L

48-h EC50 : 400 mg/L

72-hNOEC: 321 mg/L

Key value for chemical safety assessment

EC50 for marine water algae:

63.4 µg/L

EC10 or NOEC for marine water algae:

38.9 µg/L