2-ethylhexyl mercaptoacetate

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:

From 15th May to 23rd June 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

yes

Remarks:

Test system used was closed, no growth medium was described.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Sampling method: 20 mL are sampled using syringe and pipette
- Sample storage conditions before analysis: no data

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: a solution is prepared by adding 100 mg of substance in 1 L of water. This solution was agitated vigorously for 24 h. Then solution is filtered for eliminating substance in excess and obtain the saturated solution. Test solutions are then prepared using water added with NaHCO3 at 0.3 g/L and 6mM HEPES buffer.
- Differential loading: test solutions are then added with algae pre-culture (10E4 cells/mL at t0)
- Controls: Control consisted of algae culture in medium without treatment
- Evidence of undissolved material (e.g. precipitate, surface film, etc): solutions were observed to be clear at both the beginning and end of the assay. Only slight cloudy aspects were observed at the end of the assays in the two highest tested concentrations.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green algae Pseudokirchnerella subcapitata
- Strain: CCAP 278/4
- Source (laboratory, culture collection): Culture Center of Algae and Protozoa

ACCLIMATION
- Culturing media and conditions (same as test or not): 2 recipients of 100 mL each of algae stock culture are maintained at 23 +/- 1°C in a light cycle of 16 hours light/ 8 hours dark under filtered air. Each week, 2 new cultures are issued from the preceeding cultures. 4 days prior testing, 2 pre-cultures are prepared by dilution of each stock culture and incubated in the same condition as stock culture. Only one of the two cultures is uesd for assays.
- Any deformed or abnormal cells observed: no abnormal cells were reported to have been observed.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

no

Hardness:

not reported

Test temperature:

23 +/- 1°C

pH:

Around 9 (8.81 - 9.31)

Dissolved oxygen:

Around 9 mg/L (8.0 - 9.2) except the measure in the non diluted saturated solution (100% volume) at the begining of the assay: pH 5.9

Salinity:

not reported

Conductivity:

not reported

Nominal and measured concentrations:

Nominal concentrations: 0, 1.94, 4.2, 9.4, 20.6, 45.4 and 100% v/v in the absence of algae
Measured concentrations: see table 1
For statistical analysis and expression of the results, geometric mean concentrations were used. Measured concentrations falling between LOD and LOQ were replaced by LOQ/2 while measured concentrations falling below LOD were replaced by LOD, in accordance with OECD guidance n°23 on aquatic toxicity testing of difficult substances and mixtures (2000).
Geometric mean concentrations: 0 ; 0.052 ; 0.067 ; 0.5 ; 0.8 ; 7.8 ; 10.8 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type: closed
- Material, size, headspace, fill volume: 120 ml glass bottles completely filled with test solution and stoppered with PTFE bungs and sealed with aluminum caps
- Aeration: no
- Initial cells density: 10E4 cells/mL
- Control end cells density: 2.1 x 10E5 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: no
- Detailed composition if non-standard medium was used:
Test solutions are prepared using water added with NaHCO3 at 0.3 g/L and 6mM HEPES buffer.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Ultrapure water sterilised by autoclave prior testing.

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light/ 8 hours

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Optic Microscopy with haemacytometer

TEST CONCENTRATIONS
- Spacing factor for test concentrations: about 2.2
- Range finding study
- Test concentrations: from 0.1 to 100% vol
- Results were claimed to have been used to determine the conditions for the definitive study but they were not reported.

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.62 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.42 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Any stimulation of growth found in any treatment: the 2 lowest tested concentrations were found to have a slight stimulation effect on the growth.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

No test conducted

Reported statistics and error estimates:

Results were analysed and ECx were determined with Toxrat 3.2.1, using the provided template for the OECD 201 study. The statistical report is attached in the adequate section of the RSS.

Table 2: Algal concentration, biomass, and growth rate

Nominal Concentration [% vol]	Mean density of Algal Cells (x 10000/mL)				Area under the curve (biomass indicator)	Growth rate
	0 h	24 h	48 h	72 h
Control	1.0	1.9	6.9	20.9	4.1 E+6	0.0422
1.94	1.0	2.5	9.0	41.0	7.0 E+6	0.0514
4.2	1.0	2.1	7.3	27.7	5.0 E+6	0.0461
9.4	1.0	1.6	3.5	12.9	2.2 E+6	0.0355
20.6	1.0	1.4	2.3	2.1	0.5 E+6	0.0107
45.4	1.0	1.6	8.7	0.9	0.1 E+6	- 0.0015
100	1.0	7.3	1.3	1.2	0.024 E+6	0.0025

Validity criteria fulfilled:

no

Remarks:

In the control group: Biomass was multiplied by a factor > 16 at the end of the test. The CV for growth rate (0-72h) was < 7%. The CV of the section-by-section growth rate was 37%, which is sligthly higher than 35%. The study was still considered valid.

Conclusions:

In this study, the EC10 and EC50 at 72 h were found to be 0.42 and 0.62 mg/L for growth rate inhibition.

Executive summary:

The determination of the growth inhibition of the freshwater algae Pseudokirchneriella subcapitata exposed to 2 -ethylhexyl thioglycolate for 72 hours was performed following the method C3 described in Directive 92/69/EEC of the European Commission and in the guideline 201 of the OECD. The study was performed using 120 ml glass bottles stoppered with PTFE bungs and sealed with aluminium caps, containing 50 ml of test solution inoculated with an algal suspension so that the initial cell concentration was equal to 10 000 cells/mL. The validity criteria were all fulfilled (control cell multiplication > 16 and control CV for growth rate < 7%), except the CV for section-by-section which slightly exceeded 35% in the control group (found value was 37%). This was not considered affecting the outcome of the study which was therefore considered valid. The concentrations of test substance causing 10% and 50 % growth rate inhibition (ErC10 and ErC50) were 0.42 and 0.62 mg/L, respectively.

Description of key information

The determination of the growth inhibition (chronic toxicity) of the freshwater algae Pseudokirchneriella subcapitata exposed to 2-ETHYLHEXYL THIOGLYCOLATE for a duration of 72 hours was performed following the method C3 described in Directive 92/69/EEC of the European Commission and in the guideline 201 of the OECD.

The study was performed using 120 ml glass bottles stoppered with PTFE bungs and sealed with aluminium caps, containing 50 ml of test solution inoculated with an algal suspension so that the initial cell concentration was equal to 1x10⁴cells/ml.

The concentration of test substance causing a 50 % reduction in biomass (EbC50) and in growth rate (ErC50) were determined to be 0.41 and 0.91 mg/L, respectively. Respective NOEC values were both 0.5 mg/L.

The validity criteria related to the increase of cell density, in the control solution between the end and the beginning of the the test, was respected with a factor of 21 which is greater than 16.

The final concentrations of 2-ETHYLHEXYL THIOGLYCOLATE were not maintained within the designated limit of 80 % of the initial concentrations in inoculated and non inoculated flasks. Thus the validity criteria related to the stability of the test substance was not respected.

Key value for chemical safety assessment

EC50 for freshwater algae:

0.62 mg/L

EC10 or NOEC for freshwater algae:

0.42 mg/L