Phosphoric acid, C8-16-alkyl esters, reaction products with (Z)-N-9-octadecenyl-1,3-propanediamine

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:

2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

The temperature in the experiment was in the range of 21 .8 - 25.5 °C and therefore higher than stated in the study plan. Because normal growth was observed, this can be stated as uncritical.
Due to a technical accident only 182 mL instead of 200 mL test solution was used for preparation of treatment 32 mg/L. As the amount of algae added to the solution was adjusted to the amount of test solution and the extinction was in the expected range at the start of the test, this can be stated as uncritical.
Considering that these deviation are uncritical, the study is considered without significant deviation.

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Phosphoric acid, C8-16-alkyl esters, reaction products with (Z)-N-9-octadecenyl-1,3-propanediamine

EC Number:

291-909-3

EC Name:

Phosphoric acid, C8-16-alkyl esters, reaction products with (Z)-N-9-octadecenyl-1,3-propanediamine

Cas Number:

90506-47-1

Molecular formula:

complex substance

IUPAC Name:

Phosphoric acid, C8-16-alkyl esters, reaction products with (Z)-N-9-octadecenyl-1,3-propanediamine

Test material form:

liquid: viscous

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and batch No.of test material: ARCOT 3135 from PCAS, Batch N° 80555C079
- production date: May 2015
- date of receipt: 08 June 2015
- Expiration date of the batch: May 2016
- Purity test date: 28 May 2015

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature at room temperature (19.2 - 24.0 °C), in proper conditions in a closed vessel.

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The water-accommodated fractions (WAFs) were prepared for the test. This was done mixing the nominal loads with the corresponding amount of nutrient medium (demineralised water enriched with minerals but without algae) and shaking vigorously for 24 hours. The resulting solutions were filtrated through 0.45 µm nylon filters.
- Controls: Positive control used: potassium dichromate K2Cr207 (CAS No. 7778-50-9) was used as positive control in a separate reference test. Blank control: deionised water with nutrient medium and algae

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: freshwater green alga
- Genus: Desmodesmus
- Species: subspicatus
- SAG Strain Number: 86.81
- Source: The culture of Desmodesmus subspicatus was obtained in February 2014 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen).
- Method of cultivation: kept as stock culture on solid agar at 2-8 oc. From the stock culture, a permanent culture was prepared. From an aliquot of the permanent culture, the pre-culture was prepared.

ACCLIMATION
- Acclimation period: 96 hours
- Culturing media and conditions: Four days before the start of each test, an aliquot of the permanent culture was brought into pre-culture medium and incubated under continuous lighting for 96 hours. The resulting culture grew exponentially. Before usage, the pre-culture was checked for the absence of cell aggregates and the cell number of culture was determined via photometric measurement.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

As recommended in the OECD Guideline.

Test conditions

Test temperature:

21.8 - 25.5 °C

pH:

7.7 - 7.8

Nominal and measured concentrations:

1.0 / 3.2 / 10 / 32 / 100 mg/L nominal concentration

Details on test conditions:

TEST SYSTEM
- Test vessel: glass flasks total volume 65 mL
- Type: open
- Material, size, headspace, fill volume:
- Initial cells density: cell concentration of approximately 5*10^3 cells/mL
- No. of vessels per concentration (replicates): three replicates for each treatment
- No. of vessels per control (replicates): six replicates for the control

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Adjustment of pH: yes, varying between 7.7 - 7.8
- Light intensity and quality: 5300 lux during all the study.

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: The cell numbers were determined by photometric measurement of optical density.

TEST CONCENTRATIONS
- Test concentrations: 1.0 / 3.2 / 10 / 32 / 100 mg/L nominal concentration

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

3.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 3

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

2.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 4

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

17 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 5

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

1.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

Yield

Effect concentrations 6

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

5.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

Yield

Reported statistics and error estimates:

The estimation of the biological data was accomplished using the software ToxRat@ Professional, version 3.1 The details of calculation are stated in the annex, page 29 of the complete study report attached to the Iuclid file. Biological results were stated using two significant digits.

Any other information on results incl. tables

From the cell numbers, the Growth Rate and the Yield were calculated. The means and standard deviations at the end of the test are given in the following table:

Nominal Concentration in mg/L		Growth Rate (0-72h) [day^-1]	Yield (0-72h) [Cell Concentration/mL]
Blank control	Mean	1.38	332743
	SD	0.04	37397
1.0 mg/L	Mean	1.39	351428
	SD	0.06	57882
3.2 mg/L	Mean	1.26	231051
	SD	0.03	18337
10 mg/L	Mean	0.89	74382
	SD	0.08	18389
32 mg/L	Mean	0.37	11499
	SD	0.10	4861
100 mg/L	Mean	0.25	7187
	SD	0.19	7178

SD = Standard Deviation

Applicant's summary and conclusion

Conclusions:

The 72h ErC50 is 17 mg/L (95% confidence interval 14 - 20 mg/L)
The 72 EyC50 is 5.1 mg/L (95% confidence interval 4.4 - 5.8 mg/L)

Executive summary:

A study was conduct for the Determination of the toxicity of ARCOT 3135 in Desmodesmus subspicatus according to OECD 201 resp.EU C.3. The study was performed using 5 concentrations ranging from 1 .0 to 100 mg/L nominal concentration. Incubation time (test system Desmodesmus subspicatus) was 72 hours. The cell concentration of each replicate was determined by measuring the absorption of the solutions at 440 nm every 24 hours with a spectral photometer. The cell density of the cultures was calculated based on the correlation curve between the absorption and the cell density of the cultures determined by with a cell counter. Growth rate and the yield were determined from the cell number at the respective observation times.

Significant inhibition of algal growth was observed at the following concentrations: 100 / 32/ 10/3.2 mg/L. The following results for the test item ARCOT 31 35 were determined:

The 72h ErC50 is 17 mg/L (95% confidence interval 14 - 20 mg/L)

The 72 EyC50 is 5.1 mg/L (95% confidence interval 4.4 - 5.8 mg/L)