Octadecanoic acid, reaction products with triethylenetetramine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-03-25 to 2018-03-29

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

The concentrations of the test item were analytically verified via LC-MS/MSat the start (0 hours) and at the end (72 hours) of the exposure in all loading levels and in the control.

Vehicle:

no

Details on test solutions:

One water accommodated fraction (WAF) were prepared with nominal loading of the test item of 100 mg/L. Dilutions of this WAF was set up in a geometric series with a factor of 2: 6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L.
For the highest loading level an appropriate amount of the test item was weighed. The test item was applied onto a glass slide.
The glass slide with the test item was inserted in a brown glass flask with an appropriate amount of dilution water.
A slow stirring procedure was applied for 24 hours at room temperature.
After a separation phase of 2 hours at room temperature, the aqueous phase or WAF was removed by siphoning (from the approximate centre of the glass flask).
The resulting water accommodated fraction (WAF) was checked via laser beam (Tyndall effect) for undissolved test item.
The Tyndall effect was positive and partieies were observed.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: HINDÁK, SAG 61.81
- Source (laboratory, culture collection): Laboratory culture, origin: Sammlung von Algenkulturen (SAG), Pflanzenphysiologisches Institut der Universität Göttingen, Nikolausberger Weg 18, D-37073 Göttingen
- Age of inoculum (at test initiation): A four days old preculture, prepared in dilution water, was used as inoculum.
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounts to 2567 – 5130 lux for 24 hours per day.

ACCLIMATION
- Acclimation period: A three days old preculture, prepared in dilution water, was used as inoculum.
- Culturing media and conditions (same as test or not): Nutrient medium Z according to LÜTTGE et al. (1994)
Microscopic evaluation of the cells at the start and the end of exposure revealed no morphological abnormalities.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

22.8 °C

pH:

7.88 - 9.26

Nominal and measured concentrations:

One water accommodated fraction (WAF) were prepared with nominal loading of the test item of 100 mg/L. Dilutions of this WAF was set up in a geometric series with a factor of 2: 6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L.
The measured test item concentrations in fresh media at the start of the exposure (0 hours) were in the range of 0.211 to 0.624% of the nominal concentrations and 0.0988 to 0.738% at the end (72 hours) of the exposure.

Details on test conditions:

TEST SYSTEM
- Test vessel: Sterile Erlenmeyer flasks, volume: 250 mL, sealed with cotton wool plugs.
- Aeration: Test containers were placed on a rotary shaker and oscillated at approximately 70 rpm.
- Initial cells density: 5523 cells/mL
- Control end cells density:
- No. of vessels per concentration (replicates): 3 replicates per loading level
- No. of vessels per control (replicates): 6


GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: 24 hours/day light
- Light intensity and quality: Approximately 4440 to 8880 lux, corresponding to 60 to 120 µE*m-2*s-1



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: fluorimeter
- Chlorophyll measurement: The cell density was measured daily via Chlorophyll a-fluorescence, excitation at 436 nm, emission at 685 nm.


TEST CONCENTRATIONS
- Range finding study: A preliminary range finding test under static conditions over aperiod of 72 hours was conducted at the test facility with a water accommodated fraction (WAF) of the test item with a
nominal loading of 100 mg/L. The WAF was also filtrated and tested as a water soluble fraction (WSF) with two dilution levels out of the WSF.

- Test concentrations: One water accommodated fraction (WAF) were prepared with nominal loading of the test item of 100 mg/L.
Dilutions of this WAF was set up in a geometric series with a factor of 2: 6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L.

Reference substance (positive control):

yes

Remarks:

Results of the most recent performed reference tests (potassium dichromate)

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

45.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): Microscopic evaluation of the cells at the start and end of exposure revealed no morphological abnormalities.

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: The Tyndall effect was positive and particles were observed in the WAF of 100 mg/L.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- EC50 48 hr growth: 0.535 mg/L;
- EC50 48 hr yield: 0.320 mg/L

Reported statistics and error estimates:

ELx-Calculation - Growth Rate (Nominal Test Item Concentrations)
Equation: Sigmoidal dose-response (variable slope)
Y=Bottom + (Top-Bottom)/{1 +10^({LogEC50-X)*HiliSlope))

Statistical Significance (NOEL/LOEL) _ Growth Rate
Shapiro-Wilk's Test on Normal Distribution
Levenes Test on Variance Homogeneity (with Residuals)
Multiple Sequentially-rejective Welsh-t-test After Bonferroni-Holm

Validity criteria fulfilled:

yes

Conclusions:

The EL50-value for inhibition of growth rate after 72 hours were 45.9 (43.5 - 48.3) mg/L for Octadecanoic Acid, reaction products with triethylenetetramine.

Executive summary:

The toxicity of Octadecanoic Acid, reaction products with triethylenetetramine to the unicellular freshwater green alga Pseudokirchnerie/la subcapitata was determined according to the principles of OECD 201 and Council Regulation (EC) No. 266/2016/Method C.3. The aim of the study was the determination of NOEL, LOEL, EL10- , EL20- and EL50-values of the growth rate and the yield over aperiod of 72 hours.

 

The study was conducted under static conditions with an initial cell density of 5523 celis/mL. One water accommodated fraction (WAF) was prepared with nominal loading of 100mg/L. Dilutions of this WAF were set up in a geometrical series with a factor of 2: 6.25 - 12.5 - 25.0 - 50.0 – 100 mg/L. The test solutions were clear with particles (Tyndall was positive) throughout the test period.

Three replicates were tested for each test item loading and six replicates for the control. The environmental conditions were within the acceptable limits.

 

The concentrations of the test item Octadecanoic Acid, reaction products with triethylenetetramine were analytically verified via LC-MS/MS at the start (0 hours) and at the end (72 hours) of the exposure in all loading levels and in the control. The measured test item concentrations in fresh media at the start of the exposure (0 hours) were in the range of 0.211 to 0.624% of the nominal concentrations and 0.0988 to 0.738% at the end (72 hours) of the exposure (highest concentration following lead substance C16/18or C18/18, whatever is higher).

 

All effect values are based on the nominal test item loadings of Octadecanoic Acid, reaction products with triethylenetetramine.  

Growth Rate Inhibition:

 

NOEC 72 hr.: 25 mg/L

LOEL 72 hr: 50 mg/L

ErC10 72 hr.: 23.8 mg/L

ErC20 72 hr.: 31.0 mg/L

ErC50 72 hr.: 45.9 mg/L

 

The study meets the validity criteria of the guideline.

Description of key information

The EL50-value for inhibition of growth rate after 72 hours were 45.9 (43.5 - 48.3) mg/L for Octadecanoic Acid, reaction products with triethylenetetramine.

Key value for chemical safety assessment

EC50 for freshwater algae:

45.9 mg/L

Additional information

The toxicity of Octadecanoic Acid, reaction products with triethylenetetramine to the unicellular freshwater green alga Pseudokirchnerie/la subcapitata was determined according to the principles of OECD 201 and Council Regulation (EC) No. 266/2016/Method C.3. The aim of the study was the determination of NOEL, LOEL, EL10- , EL20- and EL50-values of the growth rate and the yield over aperiod of 72 hours.

 

The study was conducted under static conditions with an initial cell density of 5523 celis/mL. One water accommodated fraction (WAF) was prepared with nominal loading of 100mg/L.Dilutions of this WAF were set up in a geometrical series with a factor of 2: 6.25 - 12.5 - 25.0 - 50.0 – 100 mg/L.The test solutions were clear with particles (Tyndall was positive) throughout the test period.

Three replicates were tested for each test item loading and six replicates for the control. The environmental conditions were within the acceptable limits.

 

The concentrations of the test item Octadecanoic Acid, reaction products with triethylenetetramine were analytically verified via LC-MS/MS at the start (0 hours) and at the end (72 hours) of the exposure in all loading levels and in the control.The measured test item concentrations in fresh media at the start of the exposure (0 hours) were in the range of 0.211 to 0.624% of the nominal concentrations and 0.0988 to 0.738% at the end (72 hours) of the exposure (highest concentration following lead substance C16/18or C18/18, whatever is higher).

 

All effect values are based on the nominal test item loadings of Octadecanoic Acid, reaction products with triethylenetetramine. 

Growth Rate Inhibition:

 

NOEC 72 hr.: 25 mg/L

LOEL 72 hr: 50 mg/L

ErC10 72 hr.: 23.8 mg/L

ErC20 72 hr.: 31.0 mg/L

ErC50 72 hr.: 45.9 mg/L

 

The study meets the validity criteria of the guideline.