Fatty acids, C16-18 and C18-unsatd., compds. with triethanolamine

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:

2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Guideline compliant study with good documentation, minor deviation on the validity critera

Qualifier:

according to guideline

Guideline:

ISO 10253 (Water quality - Marine Algal Growth Inhibition Test with Skeletonema costatum and Phaeodactylum tricornutum)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Samples of the water accommodated fractions with loading 100 and 1000 mg/L and the control medium were taken at the start of the test and analysed for content of total organic carbon (TOC).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:
The test substance is a multi-component substance of low water solubility. Therefore the test was performed on water accommodated fractions (WAF) as recommended by OECD. Separate WAFs were prepared for 5 loading rates from 100 to 1000 mg/L. The test substance was transferred to 0.5 L of the test medium in 1 L sealed glass bottles. The flasks were placed on a reciprocating shaker. The WAF extraction was performed in the darkness in a room controlled at 21±1 °C. After 23 hours, the agitation was terminated and approximately one hour later, 200 mL of each WAF was drawn with a pipette, avoiding the settled material at the bottom of the flasks.

Test organisms (species):

Skeletonema costatum

Details on test organisms:

TEST ORGANISM
- Common name: Skeletonema costatum
- Strain: NIVA strain BAC 1
- Source (laboratory, culture collection): NIVA culture collection
- Age of inoculum (at test initiation): Inoculum culture was set up 1 day before test initiation in the same medium as used in the test. Incubation conditions were the same as during the test. The cell density in the inoculum culture increased 4.7 times during 1 days.
- Method of cultivation: Cultured in natural seawater with 10 % Z8 medium (3) on reciprocating shaker and continuous light at approximately 20°C.

ACCLIMATION
- Acclimation period: 1 day
- Culturing media and conditions (same as test or not): No, therefore adapted 1 day before tesing
- Any deformed or abnormal cells observed: Not reported

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

72 h

Salinity:

34 S

Nominal and measured concentrations:

Nominal concentrations: Control, 100, 180, 320, 560 and 1000 mg/L, measured concentrations in the control and 100 and 1000 mg/L test solutions were 4.9, 10.9 and 29.4 mg TOC/L.

The results indicate that less than 10 % of the test substance partitioned to the water phase as dissolved organic matter. (The estimate is based on the assumption that the carbon content of the test substance is 70% by weight).

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL flat bottom flasks, covered with inverted, perforated plastic beakers. The culture volume was 50 mL.
- Initial cells density: 5*10^6 cells/L
- Control end cells density: 1696 *10^6 cells/l
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: Yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test medium is ISO 10253 medium, based on natural seawater taken at 60 m depth at Solbergstrand Research Station in the outer Oslo Fjord and filtered through a membrane filter with 0.45 μm porosity. The salinity was 34.4 S. Nutrients were added from concentrated stock solutions as described in ISO 10253.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No
- Photoperiod: Continuous
- Light intensity and quality: Daylight-type fluorescent tubes 75μM m-2 s-1 direct irradiation (PAR), and 49 μM m-2 s-1 reflected from the white support
- Salinity (for marine algae): 34.4 S.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Electronic particle counter
- Chlorophyll measurement: No


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Justification for using less concentrations than requested by guideline: NA
- Range finding study: Not reported

Reference substance (positive control):

no

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): The curves show that growth in the control cultures was close to exponential during the first two days and declining the last day.
- Observation of abnormalities (for algal test): Not reported
- Any stimulation of growth found in any treatment: The WAFs with loading up to 1000 mg/L caused a slight stimulation of the growth.
- Effect concentrations exceeding solubility of substance in test medium: Yes (WAF)

The cell density was measured after 24.5, 48 and 72 hours The growth curves show that growth in the control cultures was close to exponential during the first two days and declining the last day. The WAFs with loading up to 1000 mg/L caused a slight stimulation of the growth. No inhibition of the growth rate was observed at any WAF loading.

In a separate document it was checked if the control growth validity criteria as outlined in the updated OECD 201 (2006) were met.

·        The increase of the algal cell concentration was 320 and thus within the validity criterion (at least 16 fold increase during the exposure period).

·        The CV Average specific growth rate was with 1.24 % within the acceptable range (i.e., less than 7%).

·        The CV- section-by section growth rate was with 42.1 % outside the acceptable range (i.e., higher than 35%).

The higher variation of the section-by-section growth rate was mentioned in the report even if the validity parameter was not calculated since the updated OECD 201 was not available at the time when the test was conducted. However, the effect of the reduced growth was visible in the graphical presentation of the data. However, this deviation is not considered as critical for the general outcome of the study. The reduced growth from 48 to 72 hours is most likely caused by the high growth of the algae and potential light or nutrient limitation during the last 24 hours. Based on the cell concentration until hour 48 and the graphical presentation of the data in the report there is no adverse effect of the test item visible at 48 hours. Therefore, this deviation to the validity criteria of the updated OECD 201 is considered to have no impact on the relevance of general interpretation of the study.

Validity criteria fulfilled:

no

Remarks:

but not considered as relevant (see text)

Executive summary:

In the Klimisch 2 study from Källqvist (2004) the growth inhibition of the test material (Oleic salt of triethanolamine) to the marine algae Skeletonema costatum was determined in a 72 h static GLP algae growth inhibition test. The test design was based on ISO 10253: "Marine algal growth inhibition test with Skeletonema costatum and Phaeodactylum tricornutum". Since the test substance is a UVCB substance of low water solubility the test was performed on water accommodated fractions (WAF) as recommended by OECD. The nominal loading rates were 0 (control), 100, 180, 320, 560 and 1000 mg test item/L. The WAF extraction was performed in the darkness in a room controlled at 21±1 °C. After 23 hours, the agitation was terminated and approximately one hour later, 200 mL of each WAF was drawn with a pipette, avoiding the settled material at the bottom of the flasks. Six control replicates and 3 replicates from each test solution were set up. Dose verification analysis was performed by TOC analysis of the control, 100 and 1000mg/L test solutions at the start of the test.

 

In order to determine the growth of the cultures, aliquots were counted by an electronic particle counter. The cell density was determined at 0, 24, 48 and 72 hours. The growth of the control cultures almost fulfilled the validity criteria from OECD 201 (2006). The deviation was observed for the section-by section growth rate. This was considered to be due to the high growth rate of the algae in all cultures and is considered to have no effects on the general outcome of the study.

 

Since no inhibition was observed at the highest loading tested, the loading causing 50% reduction of the average growth rate (E_rL₅₀) may be expressed as >1000 mg/L. No WAF loading caused a significant reduction of the growth rate, thus the no observable effect loading (NOEL) is >1000 mg/L.

 

This study is considered to be acceptable for the risk assessment despite minor shortcomings in the validity criteria.

Description of key information

72h-NOELR: > 1000 mg/L

72h-ErL50: > 1000 mg/L

Key value for chemical safety assessment

EC50 for marine water algae:

1 000 mg/L

EC10 or NOEC for marine water algae:

1 000 mg/L

Additional information

In the Klimisch 2 study from Källqvist (2004) the growth inhibition of the test material (Oleic salt of triethanolamine) to the marine algae Skeletonema costatum was determined in a 72 h static GLP algae growth inhibition test. The test design was based on ISO 10253: "Marine algal growth inhibition test with Skeletonema costatum and Phaeodactylum tricornutum". Since the test substance is a UVCB substance of low water solubility the test was performed on water accommodated fractions (WAF) as recommended by OECD. The nominal loading rates were 0 (control), 100, 180, 320, 560 and 1000 mg test item/L. The WAF extraction was performed in the darkness in a room controlled at 21±1 °C. After 23 hours, the agitation was terminated and approximately one hour later, 200 mL of each WAF was drawn with a pipette, avoiding the settled material at the bottom of the flasks. Six control replicates and 3 replicates from each test solution were set up. Dose verification analysis was performed by TOC analysis of the control, 100 and 1000 mg/L test solutions at the start of the test.

 

In order to determine the growth of the cultures, aliquots were counted by an electronic particle counter. The cell density was determined at 0, 24, 48 and 72 hours. The growth of the control cultures almost fulfilled the validity criteria from OECD 201 (2006). The deviation was observed for the section-by section growth rate. This was considered to be due to the high growth rate of the algae in all cultures and is considered to have no effects on the general outcome of the study.

 

Since no inhibition was observed at the highest loading tested, the loading causing 50% reduction of the average growth rate (E_rL₅₀) may be expressed as >1000 mg/L. No WAF loading caused a significant reduction of the growth rate, thus the no observable effect loading (NOEL) is >1000 mg/L.

 

This study is considered to be acceptable for the risk assessment despite minor shortcomings in the validity criteria.