Reaction mass of 2-octyldodecyl 2-hydroxypropanoate, 2-Octyldodecan-1-ol and 2-octyldodecyl 2-[(2-hydroxypropanoyl)oxy]propanoate

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. July 2017 - 20. Mar. 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2-Octyldodecyl 2-hydroxypropanoate

Molecular formula:

C23H46O3

IUPAC Name:

2-Octyldodecyl 2-hydroxypropanoate

Constituent 2

Reference substance name:

2-octyldodecan-1-ol

EC Number:

226-242-9

EC Name:

2-octyldodecan-1-ol

Cas Number:

5333-42-6

Molecular formula:

C20H42O

IUPAC Name:

2-octyldodecan-1-ol

Constituent 3

Reference substance name:

2-octyldodecyl 2-[(2-hydroxypropanoyl)oxy]propanoate

Molecular formula:

C26H50O5

IUPAC Name:

2-octyldodecyl 2-[(2-hydroxypropanoyl)oxy]propanoate

Test material form:

liquid: viscous

Details on test material:

- State of aggregation: liquid
- Particle size distribution: not relevant

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 6-3-A
- Expiration date of the lot/batch: 03. March 2019
- Purity test date: not stated

RADIOLABELLING INFORMATION (if applicable)
not applicable

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- Stability under test conditions: assumed stable
- Solubility and stability of the test substance in the solvent/vehicle: Water Accomodated fraction used
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not applicable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
none

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The content of DOC in the test vessels was measured at the start and at the end of the test.

Test solutions

Vehicle:

no

Details on test solutions:

The water-accommodated fraction (WAF) was prepared for the test. This was done by mixing the nominal load of 10 g/L resp. 11.2 mL/L test item (based on a density of 0.898 g/mL given by the sponsor) with the corresponding amount of algal medium (demineralised water enriched with minerals but without algae) and stirring vigorously for 7 days. The lower phase of this water-accommodated fraction was used.
The lower treatments were prepared by dilution of this WAF with nutrient medium.

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Specification
Unicellular freshwater green alga.
Genus Desmodesmus
Species subspicatus
SAG Strain Number 86.81
Taxonomic position Chlorophyta - Chlorophyceae

Origin and Culture
The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen). The algae are kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture was prepared. From an aliquot of the permanent culture, the pre-culture was prepared in the first experiment . In the second experiment an aliquot of the stock culture was used for preparing the pre-culture.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

none

Test conditions

Hardness:

The algal medium corresponds to the OECD TG 201 medium.

Test temperature:

20.4 – 24.7 °C

pH:

7.4 - 7.8

Dissolved oxygen:

not stated

Salinity:

not stated

Conductivity:

not stated

Nominal and measured concentrations:

0.1 / 0.32 / 1 / 3.2 / 10 g/L nominal concentration (WAF)
The measured concentrations at the beginning and at the end of the test in the 3 highest concentrated treatments showed clearly the presence of dissolved test item in the test so-lutions. In the 2 lowest concentrated treatments, the measured DOC concentration was in the same range as in the control. Therefore, the determination of the results was based on the geometric mean of the measured concentrations in the 2 highest concentrated treatments. The measured concentration at the nominal treatment 10 g/L can be stated as limit of solubility in test medium.
Geometric Mean of Measured Concentrations
Nominal Concentration Test Item Geometric Mean
mg/L mg/L
Blank control --
100 * --
320 * --
1000 * 1.1
3200 5.9
10000 20.1

Details on test conditions:

Date: 07. August – 10. August 2017 (Exp I)
18. September – 21. September 2017 (Exp II)
Treatments tested: 0.1 / 0.32 / 1 / 3.2 / 10 g/L nominal concentration
The concentrations to be tested were based on non GLP pre-tests.
Number of replicates: 6 replicates for the blank control
3 replicates for each treatment
Vessels: glass flasks, total volume 65 mL
Duration: 72 hours
Lighting: 4500 lux
Temperature: 20.4 – 24.7 °C
Replicates (Blank control): algal medium and algae
Replicates (Treatments): test solution and algae

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

5.9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

5.9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 3

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: limit of solubility

Effect concentrations 4

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: limit of solubility

Effect concentrations 5

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 6

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

>= 5.9 - <= 20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 7

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 8

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

Only in the highest concentration a significant inhibition could be observed, therefore only the two highest concentrations were used for evaluation

Results with reference substance (positive control):

The results of the last study with the positive control K2Cr2O7 are presented in the following table. The study was performed under GLP conditions in February 2017.
Biological Results of K2Cr2O7
Parameter Value 95% confidence interval
72h ErC50 0.94 mg/L 0.90 - 0.98 mg/L
72h EyC50 0.42 mg/L 0.39 - 0.45 mg/L

Reported statistics and error estimates:

Calculation of results was performed with the help of validated software (Microsoft Excel®). The estimation of the biological data (LOEC and NOEC) was accomplished using the software ToxRat® Professional, version 3.2.1.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the presented test conditions, Salacos 13 had an effect on Algae with the following toxicological descriptors:
NOEC (Growth rate) = 5.9 mg/L
LOEC (Growth rate) = 20.1 mg/L
NOEC (Yield) = 5.9 mg/L
LOEC (Yield) = 20.1 mg/L

Executive summary:

Two experiments were performed. In the 1^stexperiment one of the validity criteria was not met. Therefore, the experiment was repeated. The results of the 1^stexperiment are not stated in this report, but will be kept together with the other raw data in the GLP-archive of the test facility.

 

The study was performed using 5 concentrations ranging from 0.1 to 10 g/L nominal concentration. Incubation time (test systemDesmodesmus subspicatus)was 72 hours. The cell concentration of each replicate was determined by measuring the cell numbers every 24 hours with an electronic particle counter. Growth rate µ and the yield[1]were determined from the cell number at the respective observation times.

Significant inhibition of algal growth was observed only at the highest tested concentration 10 g/L (nominal). Therefore, only the two highest concentrated treatments were used for evaluation.

 

At the start and at the end of the test, the content of the test item in the test solutions was estimated by determination of the dissolved organic carbon (DOC) content in the test solutions using a carbon analyser.

The measured concentrations at the beginning and at the end of the test in the 3 highest concentrated treatments showed clearly the presence of dissolved test item in the test solutions. In the 2 lowest concentrated treatments, the measured DOC was in the same range as in the control. Therefore, the determination of the results was based on the geometric mean of the measured concentrations in the 2 highest concentrated treatments. The measured concentration at the nominal treatment 10 g/L can be stated as limit of solubility in test medium.

 

The 72h-EC50s of potassium dichromate (K₂Cr₂O₇, CAS No. 7778-50-9) were determined in a separate reference test. The values lay within the range of the laboratory (growth rate 0.73 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

The following results for the test itemSalacos 13 were determined:

   Results of the test item

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	5.9 mg/L	20.1 mg/L limit of solubility	> 20.1 mg/L	> 20.1 mg/L
Yield	5.9 mg/L	20.1 mg/L limit of solubility	5.9 - 20.1 mg/L	> 20.0 mg/L

 

Note: According to the guideline, NOEC is determined by comparing of the respective treatment with the blank control. Statistically insignificant variation is considered as “no observed effect”. The EC10 which is usually read from the graph toxicity vs. concentration is stated as a range because graphical determination of the ECx values is not possible.

[1]Yield (according to OECD Guideline 201) is defined as the biomass at the end of the exposure period minus the biomass at the start of the exposure period. Calculation see under 8.2 Growth and growth inhibition are quantified from measurements of the algal biomass as a function of time. Algal biomass is defined as the dry weight per volume, e.g. mg algae/L test solution. However, dry weight is difficult to measure and therefore surrogate parameters are used. Of these surrogates, cell counts are most often used.