Docosyl docosanoate

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:

1 Oct - 4 Oct 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP - Guideline study, tested with the source substance dodecyl oleate (CAS 36078-10-1). According to the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Remarks:

Landesanstalt für Umwelt, Messungen und Naturschutz Baden-Württemberg, Karlsruhe, Germany

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the start (0 h) and at the end of the experimental phase (72 h) samples (10 mL) were taken from the highest, the middle and the lowest concentrations out of each replicate for chemical analysis and pooled in a glass beaker before transferred into brown glass vials (4 mL). At the end additional samples were taken from the test vessels with algae in the same way. Algae were separated by filtration (0.45 µm CA filter) to avoid disturbance during the analytical process.
- Sample storage conditions before analysis: The samples were stored in 4 mL brown glass vials which were filled up to the brim to avoid oxidative processes during storage. Samples were stored for a maximum period of 9 days in the refrigerator at 5 °C ± 3 K before analysis was performed. The cooled samples were sent via over-night express parcel service in an isolated transport box containing cooling elements and arrived at the next morning in good condition at the analytical laboratory.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Due to the low water solubility of the test item, the study was performed with WAF ("water accommodated fraction") prepared with algal growth medium according to OECD 201. The test item is a mobile liquid and therefore difficult to weight. Therefore it was decided to pipette the test item into the 2000 mL beaker containing algal medium. According to the density indication from the sponsor of 0.8529 g/cm³ the amount to be added was converted into volume. The suspension was stirred for 48 hours at 20 - 21 °C in the dark. For stirring a magnetic stirrer with a 2 cm stir bar was used. After stirring was stopped, the suspension was allowed to float and sediment for a period of 70 minutes. After stopping the stirring in all WAFs oily drops were floating on the surface. After one hour sedimentation and flotation period no particles or oily droplets could be observed in suspension which could affect the organisms physically. The WAFs itself were clear and were therefore not filtered to obtain the water soluble fractions (WSF). The test solutions were taken from the middle of the suspension in the beakers using a glass tube and transferred into the test vessels. The loading rates (LR) for the WAFs were chosen 1.11-fold higher than the nominal loading rates intended in the test solutions so as to take into account the dilution by the algal inoculum suspension (10%). 45 mL test solution was transferred into each test vessel (100 mL wide necked Erlenmeyer flask) and 5 mL of the algae inoculum suspension (0.8 E+05 algae mL) were added to obtain an algae starting concentration of 0.8 E+04 algae/mL. For the controls, 5 mL algae inoculum suspension was added to 45 mL algal medium. This medium was stirred in the same way as the WAF. Three replicates have been prepared for the test vessels and six for the control vessels.
- Eluate: no
- Differential loading: yes
- Controls: yes, growth medium control
- Evidence of undissolved material: no

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: CHODAT SAG No. 86.81
- Source (laboratory, culture collection): Obtained from the German Environmental Agency (UBA), Berlin-Marienfelde and cultivated in suspension culture (strain cultured at laboratory since June 2012).
- Method of cultivation: Twice a week the stock suspension is diluted into fresh Holm-Hansen medium under axenic conditions to keep it in exponential growth.

ACCLIMATION
- Culturing media and conditions (same as test or not): Holm-Hansen medium was used for cultivation. Algal growth medium according to OECD 201 was used to prepare the pre-culture three days before initiation of the study and for the study.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

23.0 - 23.2 °C

pH:

7.9 - 8.0

Nominal and measured concentrations:

nominal: control, 3.57, 8.22, 18.90, 43.48 and 100 mg/L
measured: < LOQ (0.005 mg/L) in all test vessels

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks, wide necked
- Type: sealed with a sterile cellulose stopper
- Material, size, headspace, fill volume: glass, 100 mL, headspace: 50 mL, fill volume: 50 mL
- Aeration: shaken
- Initial cells density: 0.07E+05 cells/mL
- Control end cells density: 9.09E+05 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes, according to guideline

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionised water
- Culture medium different from test medium: yes, Holm-Hansen medium was used for cultivation. Algal growth medium according to OECD 201 was used to prepare the pre-culture three days before initiation of the study and for the study.

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous illumination
- Light intensity and quality: 120 µE/m² s ± 5.7% (PAR)

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: To determine the conversion factor between the measured surrogate parameter chlorophyll fluorescence and the biomass, as required according to the OECD guideline, a correlation between fluorescence and cell count was provided. For this purpose six dilutions with the nominal cell counts of 0.05 x 10E+05, 0.1 x 10E+05, 0.5 x 10E+05, 1 x 10E+05, 5 x 10E+05 and 10 x 10E+05 were prepared by diluting the pre-culture used for the test with algal medium. For all dilutions the cell count was determined with the Coulter Counter Z2, and the chlorophyll fluorescence was determined in the same way as in the test. Both values were correlated and the equation describing the curve was calculated. The correlation allows calculation the cell count from the measured surrogate parameter chlorophyll fluorescence.
- Chlorophyll measurement: For measuring the chlorophyll fluorescence of the algae in each test vessel 200 µl test medium was transferred into a 96-well micro-plate. The fluorescence was measured with the microplate reader TECAN infinite F200 at an excitation wave length of 465 nm and an emission wave length of 670 nm. Each measurement was conducted in duplicate. If the variation coefficient was > 10% the measurement was repeated. Algal growth medium was measured as blank. The measured values were corrected for the blank values obtained with the algal medium. The dimensionless values for the fluorescence are a measure for the biomass (OECD 201).

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.3

Reference substance (positive control):

no

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

other: yield

Details on results:

- Exponential growth in the control: yes
- Unusual cell shape: not observed
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Any stimulation of growth found in any treatment: Several loading rates tested resulted in a slight increase of algal growth.

Results with reference substance (positive control):

A quality check takes place at regular intervals using a concentration range of potassium dichromate (Sigma, Steinheim, Germany, Lot No. 074K0617, from November, 22nd, 2005) in order to assess the validity of the test system. The last quality check was performed in June 2012. The ErC50 was 0.94 mg/l (0.84 – 1.05 mg/L 95%-CI) and the EyC50 (Yield) was 0.49 mg/l (0.47 – 0.51 mg/L 95%-CI).

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

Please refer to analogue justification provided in IUCLID section 13.

Reason / purpose for cross-reference:

read-across source

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Remarks on result:

other:

Remarks:

Source: key, RA-A, CAS 36078-10-1, Dako, 2013, D. subspicatus

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Remarks on result:

other: Source: key, RA-A, CAS 36078-10-1, Dako, 2013, D. subspicatus

Description of key information

No effects up to the limit of water solubility based on available data (OECD 201)

Additional information

Since no studies investigating the toxicity to algae of docosyl docosanoate (CAS 17671-27-1) are available for this endpoint, in accordance to Regulation (EC) No. 1907/2006 Annex XI, 1.5 a read across to the structurally related source substance dodecyl oleate (CAS 36078-10-1) was conducted. The read-across substance dodecyl oleate is characterized by a smaller fatty acid chain (C18 unsatd. compared to C22 (target)) and a smaller fatty alcohol chain length (C12 compared to C22 (target)). The smaller alcohol and fatty acid chain length increases the water solubility and therefore the bioavailability in the water. Water solubility is negatively correlated with the C-chain length of the fatty alcohol and fatty acid (Lide, 2005). Therefore, it is possible to extrapolate from a source substance with higher water solubility since it has an increased bioavailability in the water phase (ECHA, 2017).

This read-across is justified in detail in the overall summary (IUCLID section 6.1) and within the analogue justification in IUCLID Section 13. In this case of read-across, the best suited (highest degree of structural similarity, nearest physico-chemical properties) read-across substance was used for the assessment. Nevertheless, as it can be seen in the data matrix of the analogue justification in section 13 and the overall summary, all reliable data support the hazard assessment by showing a consistent pattern of results.

The study with the read-across substance dodecyl oleate (CAS 36078-10-1) was performed according to OECD 201 under GLP conditions (Hafner, 2012). Desmodesmus subspicatus was exposed to the loading rates of five concentrations up to 100 mg/L. Analytical measurements resulted in concentrations below the detection limit (< 0.005 mg/L). The growth of algae was not inhibited resulting in a NOELR (72 h) ≥ 100 mg/L (nominal) and an EL50 (72 h) > 100 mg/L (nominal) based on the growth rate and yield.

Based on the result from one structurally related read-across substance (in accordance to Regulation (EC) No 1907/2006 Annex XI, 1.5) which is characterized by a similar ecotoxicological profile and comparable structure, it can be concluded that docosyl docosanoate (CAS 17671-27-1) will not exhibit effects on algae up to the limit of water solubility.