Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
14 Dec 2018 - 22 May 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report date:
2019

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)
Version / remarks:
Adopted March 23, 2006; Annex 5 corrected 28 July 2011.
Qualifier:
according to guideline
Guideline:
other: Guidance document on aqueous-phase aquatic toxicity testing of difficult test chemicals, OECD series on testing and assessment number 23
Version / remarks:
2000
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Fatty acids, C18-unsatd., dimers, hydrogenated, reaction products with Phytosterol, Isooctadecan-1-ol, Hexadecan-1-ol, Octadecan-1-ol and Docosan-1-ol
EC Number:
817-761-7
Cas Number:
765293-81-0
Molecular formula:
Not applicable
IUPAC Name:
Fatty acids, C18-unsatd., dimers, hydrogenated, reaction products with Phytosterol, Isooctadecan-1-ol, Hexadecan-1-ol, Octadecan-1-ol and Docosan-1-ol
Test material form:
other: waxy paste
Details on test material:
Appearance: White to pale yellow waxy paste
Test item storage: At room temperature
Stable under storage conditions until: 30 May 2019 (retest date)

Sampling and analysis

Analytical monitoring:
yes
Remarks:
UPLC-MS
Details on sampling:
Samples for possible analysis were taken from all test concentrations and the control according to the schedule below.
Frequency: at t=0 h, t=24 h and t=72 h.
Volume: 2.0 mL from the approximate centre of the test vessels.
Storage: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility.

Test solutions

Vehicle:
no
Details on test solutions:
Preparation of Test Solutions
The batch of Plandool-H tested was a white to pale yellow waxy paste UVCB and not completely soluble in test medium at the loading rates initially prepared. No correction was made for the purity/composition of the test item.
Preparation of test solutions started with loading rates individually prepared at 1.0, 10 and 100 mg/L. A two-day period of magnetic stirring at 45°C was applied to ensure maximum dissolution of the test item in medium. The obtained mixtures were allowed to settle overnight (approximately a period of 21 - 24 hours). Thereafter, the aqueous Water Accommodated Fractions (WAFs) were collected by means of siphoning and used as test concentrations. All test solutions were clear and colorless at the end of the preparation procedure.
After preparation, volumes of 50 mL were added to each replicate of the respective test concentration. Subsequently, 1 mL of an algal suspension was added to each replicate providing a cell density of 10000 cells/mL.
Any residual volumes were discarded.

Test organisms

Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
Species: Raphidocelis subcapitata, strain: NIVA CHL 1
Source: In-house laboratory culture.
Reason for selection: This system is a unicellular algal species sensitive to toxic items in the aquatic ecosystem and has been selected as an
internationally accepted species.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
72 h

Test conditions

Test temperature:
22 - 23 °C
pH:
8.1 - 8.2 (control)
8.0 - 8.7 (treatments)
Details on test conditions:
Freshwater Algae Culture
Stock culture:
Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light in a climate room at a temperature of 21-24°C.
Light intensity:
60 to 120 μE/m2/s when measured in the photosynthetically effective wavelength range of 400 to 700 nm.
Stock culture medium: M1.
Pre-culture:
Three days before the start of the test, cells from the algal stock culture were inoculated in culture medium at a cell density of 10000 cells/mL. The pre-culture was maintained under the same conditions as used in the test. The cell density was measured immediately before use.
Pre-culture medium: M2.

Test Concentrations
Plandool-H: WAFs individually prepared at loading rates of 0.32, 1.0, 3.2, 10, 32 and 100 mg/L.
Control(s): Test medium without test item.
Replicates: 3 replicates of each test concentration, 6 replicates of the control, 1 extra replicate of each test group for sampling purposes after 24 hours of exposure, 1 or 2 replicates of each test concentration without algae.

Test Procedure and Conditions
Test vessels: 100 mL, all-glass with aluminium caps, perforated for ventilation, containing 50 mL of test solution
Test Medium: M2
Cell density: An initial cell density of 10000 cells/mL.
Illumination: Continuously using TLD-lamps with a light intensity within the range of 88 to 90 μE.m-2.s-1.
Incubation: Vessels were distributed at random in the incubator and daily repositioned. During incubation the algal cells were kept in suspension by continuous shaking.
Reference substance (positive control):
yes
Remarks:
potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
< 0.32 mg/L
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: The 72h-NOELR for growth rate inhibition was below 0.32 mg/L (loading rate) based on statistical significance
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
0.32 mg/L
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: The 72h-NOELR for growth rate inhibition was 0.32 mg/L (loading rate) based on biological relevance.
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
< 0.32 mg/L
Conc. based on:
test mat.
Basis for effect:
other: Yield
Remarks on result:
other: Effect concentration is indicated using loading rate
Duration:
72 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: Effect concentration is indicated using loading rate
Details on results:
Combined Limit/Range-Finding Test
Inhibition of algal growth rate and yield was found at all test concentrations at the end of the test. At WAFs (Water-accommodated fraction) prepared at loading rates of 1.0, 10 and 100 mg/L, growth rate inhibition of 14, 9.4 and 8.1% was observed, respectively. Yield inhibition of 49, 38 and 34% was observed, respectively.
Based on these results, samples taken from all test concentrations were analysed. The measured concentrations at the start of the test were 0.039, 0.12 and 0.085 mg/L, respectively. After 24 hours of exposure, the concentrations decreased below the limit of detection.

Final Test
Measured Test Item Concentrations
Samples taken from all test concentrations and the control were analysed. The measured concentrations at the start of the test were 0.080, 0.024 and 0.34 mg/L, for WAFs prepared at loading rates of 3.2, 10 and 32 mg/L. Other measured concentrations were below the limit of detection. At the end of the test, all concentrations were below the limit of detection.
Based on these results and by decision of the sponsor, effect parameters were reported and expressed as loading rates.
The concentrations measured in the samples taken from solutions with algae were comparable with the concentrations measured in the samples without algae. However, since measured concentrations were very low and below the limit of detection at the end of the test, it cannot be stated whether the presence of the algae affected the concentration of the test item in test medium throughout the test or not.
Inhibition of Growth Rate and Inhibition of Yield
Inhibition of growth rates was between 6.3 and 20% at the concentrations of Plandool-H tested. Inhibition of yield was between 29 and 67%. No clear dose-response curve was observed for either growth rate or yield inhibition. Statistically significant inhibition was recorded between the values for growth rate or yield at all of the test concentrations when compared to the control group. However, growth rate inhibition was considered to be biologically not relevant at a WAF prepared at a loading rate of 0.32 mg/L, where the observed inhibition was below 10%. The NOELR based on biological relevance was thus set at 0.32 mg/L.
Microscopic observations at the end of the test revealed a normal and healthy appearance of the algal cells exposed to a loading rate of 1.0 mg/L when compared to the control.
Results with reference substance (positive control):
-Results with reference substance valid? Yes
- ErC50 (72 h): 1.11 mg/L potassium dichromate
- Date of reference test: 29 Apr - 02 May 2019

Any other information on results incl. tables

Acceptability of the Test

1. In the control, cell density increased by an average factor of at least 16 within the exposure period (i.e. 241).

2. The mean coefficient of variation for section-by-section specific growth rates in the control cultures did not exceed 35% (i.e. 13%).

3. The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures did not exceed 7% (i.e. 0.76%).

RESULTS

Combined Limit/Range-Finding Test

The mean cell densities measured during the combined limit/range-finding test are presented in Table 1. Table 2 and Table 3 present the percentages growth rate inhibition and yield inhibition per concentration, respectively.

Inhibition of algal growth rate and yield was found at all test concentrations at the end of the test. At WAFs prepared at loading rates of 1.0, 10 and 100 mg/L, growth rate inhibition of 14, 9.4 and 8.1% was observed, respectively. Yield inhibition of 49, 38 and 34% was observed, respectively.

Based on these results, samples taken from all test concentrations were analysed. The measured concentrations at the start of the test were 0.039, 0.12 and 0.085 mg/L, respectively. After 24 hours of exposure, the concentrations decreased below the limit of detection.

Samples taken from the control and the vessel without algae were also analysed. However, results were not approved because of possible instrumental problems. Since a final test was performed, this has no impact on the study results or conclusions.

Table 1 Mean Cell Densities (x104 Cells/mL) during the Combined Limit/Range-Finding Test

Time (h)

Control

Plandool-H Loading rate: 1.0 mg/L

Plandool-H Loading rate: 10 mg/L

Plandool-H Loading rate: 100 mg/L

0

1.0

1.0

1.0

1.0

24

5.3

n.d.

n.d.

5.3

48

31.8

n.d.

n.d.

31.0

72

161.2

83.0

99.8

107.1

n.d. – not determined

Table 2 Percentage Inhibition of Growth Rate during the Combined Limit/Range-Finding Test

Plandool-H Loading rate (mg/L)

Mean

Std. Dev.

n

%Inhibition

Control

1.694

0.0173

6

 

1.0

1.462

0.1065

3

14

10

1.534

0.0106

3

9.4

100

1.557

0.0222

6

8.1

Table 3 Percentage Inhibition of Yield during the Combined Limit/Range-Finding Test

Plandool-H Loading rate (mg/L)

Mean

Std. Dev.

n

%Inhibition

Control

160.2

8.19

6

 

1.0

82.0

26.49

3

49

10

98.8

3.19

3

38

100

106.1

7.35

6

34

Final Test

Measured Test Item Concentrations

Samples taken from all test concentrations and the control were analysed. The measured concentrations at the start of the test were 0.080, 0.024 and 0.34 mg/L, for WAFs prepared at loading rates of 3.2, 10 and 32 mg/L. Other measured concentrations were below the limit of detection. At the end of the test, all concentrations were below the limit of detection.

Based on these results and by decision of the sponsor, effect parameters were reported and expressed as loading rates.

The concentrations measured in the samples taken from solutions with algae were comparable with the concentrations measured in the samples without algae. However, since measured concentrations were very low and below the limit of detection at the end of the test, it cannot be stated whether the presence of the algae affected the concentration of the test item in test medium throughout the test or not.

Inhibition of Growth Rate and Inhibition of Yield

Table 4 shows group mean growth rates and the percentages of growth rate inhibition (total test period) whereas Table 5 shows the values at different time intervals. The group mean yields and the percentages of yield inhibition are summarized in Table 6.

Inhibition of growth rates was between 6.3 and 20% at the concentrations of Plandool-H tested. Inhibition of yield was between 29 and 67%. No clear dose-response curve was observed for either growth rate or yield inhibition. Statistically significant inhibition was recorded between the values for growth rate or yield at all of the test concentrations when compared to the control group. However, growth rate inhibition was considered to be biologically not relevant at a WAF prepared at a loading rate of 0.32 mg/L, where the observed inhibition was below 10%. The NOELR based on biological relevance was thus set at 0.32 mg/L.

Microscopic observations at the end of the test revealed a normal and healthy appearance of the algal cells exposed to a loading rate of 1.0 mg/L when compared to the control.

Table 4 Growth Rate and Percentage Inhibition for the Total Test Period

Plandool-H Loading rate (mg/L)

Mean

Std. Dev.

n

%Inhibition

Control

1.828

0.0139

6

 

0.32

1.713

0.0238

3

6.3#

1.0

1.459

0.0742

3

20*

3.2

1.619

0.0231

3

11*

10

1.638

0.0041

3

10*

32

1.549

0.0083

3

15*

100

1.590

0.0102

3

13*

* - effect was statistically significant, # - effect was statistically significant however biologically not relevant (<10%).

Table 5 Growth Rate and Percentage Inhibition at Different Time Intervals

Plandool-H Loading rate (mg/L)

n

Mean (0-24 h)

%Inhibition (0-24 h)

Mean (24-48 h)

%Inhibition (24-48 h)

Mean (48-72 h)

%Inhibition (48-72 h)

Control

6

2.066

 

1.811

 

1.609

 

0.32

3

2.200

-6.5

1.517

16

1.422

12

1.0

3

2.043

1.1

1.425

21

0.910

43

3.2

3

2.032

1.6

1.624

10

1.202

25

10

3

2.159

-4.5

1.571

13

1.183

26

32

3

2.127

-3.0

1.496

17

1.025

36

100

3

2.132

-3.2

1.530

16

1.108

31

Table 6 Yield and Percentage Inhibition for the Total Test Period

Plandool-H Loading rate (mg/L)

Mean

Std. Dev.

n

%Inhibition

Control

240.3

10.12

6

 

0.32

169.9

12.34

3

29*

1.0

79.9

17.03

3

67*

3.2

128.0

8.75

3

47*

10

135.1

1.67

3

44*

32

103.4

2.59

3

57*

100

116.9

3.62

3

51*

* - effect was statistically significant.

Determination of Effect Concentrations

Table 7 shows the effect parameters based on loading rates.

Table 7 Effect Parameters

Parameter (mg/L)

NOELR*

NOELR#

EL50

Value (Growth rate)

<0.32

0.32

>100

Value (Yield)

<0.32

<0.32

n.d.+

cl – confidence limit, * - based on statistical significance,#- based on biological relevance,+- due to the lack of a dose-response curve this value got not be statistically determined.

Experimental Conditions

Table 8 shows the pH recorded at the beginning and the end of the test. The pH was within the limits prescribed by the study plan (6-9, preferably not varying by more than 1.5 unit).

During the exposure period the temperature measured in the incubator was maintained between 22 and 23°C. Temperature remained within the limits prescribed by the study plan (21-24°C, constant within ±1°C).

Table 8 pH Levels Recorded during the Final Test

Plandool-H Loading rate (mg/L)

pH (t=0 h)

pH (t=72 h)

Control

8.1

8.2

0.32

8.1

8.0

1.0

8.6

8.1

3.2

8.6

8.2

10

8.7

8.2

32

8.6

8.2

100

8.6

8.2

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
For further details please refer to “Any other information on results incl. tables”.