Sulfonic acids, C6-8-alkane, sodium salts

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:

29 April - 02 May, 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Name of test material (as cited in study report): Bio-Terge PAS 7S-DRY
- Analytical purity: 100 % (UVCB)
- Lot/batch No.: PG12122011 (dried from industrial lot 7476775)
- Expiration date of the lot/batch: 12 December 2013
- Storage condition of test material: Controlled Room Temperature (15-25°C, below 70 RH%)

Analytical monitoring:

yes

Details on sampling:

Test concentrations were analytically determined at the start and at the end of the test.

Details on test solutions:

An amount of 100 mg test item was dissolved in one litre vehicle (OECD medium) by mechanical dispersion in order to give the 100 mg/L test concentration. The test solutions were prepared by the appropriate diluting of this stock solution and distributed into test vessels prior to introduction of algae.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: 61.81 SAG (identical strains: CCAP 278/4; UTEX 1648; ATCC 22662)
- Source (laboratory, culture collection): SAG: Collection of Algal Cultures, Inst. Plant Physiology, University of Göttingen, GERMANY. Cultured under standardised conditions (see OECD 201) in the Ecotoxicological Laboratory of CiToxLAB Hungary Ltd.
ACCLIMATION
The pre-culture was intended to give an amount of alga suspension suitable for the inoculation of test cultures. The pre-culture was incubated under the conditions of the study in an aerated Algal Growth Medium and used when still exponentially growing (after an incubation period of 3 days). The cell count of above culture was determined by microscopic method and this cell suspension was diluted with Algal Growth Medium to 1.E07 cells/mL

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

not applicable

Hardness:

no data

Test temperature:

The temperature was in the range of 22.3 – 22.6 °C measured in the flask and between 22.0 and 22.9 °C measured within the climate chamber.

pH:

The range of the pH was 7.41 – 9.25 during the experiment.

Dissolved oxygen:

no data

Salinity:

not applicable

Nominal and measured concentrations:

nominal concentrations of test item used in the main experiment were: 6.25; 12.5; 25; 50 and 100 mg/L.
measured geometric mean test item concentrations were: 5.9; 11.8; 24.7; 48.8 and 93.8 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flask
- Type (delete if not applicable): air-permeable stopper
- Material, size, headspace, fill volume: 100 mL algal suspension
- Initial cells density: 1.E04 algal cells/mL test medium
- Control end cells density: 1.E04 algal cells/mL test medium
- 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: Reconstituted algal growth medium (OECD medium, according to OECD 201) was used as dilution water for both the range finding and definitive tests.
- Culture medium different from test medium: No
OTHER TEST CONDITIONS
- Sterile test conditions: no data
- Adjustment of pH: No
- Photoperiod: Continuous light
- Light intensity and quality: The light intensity at the position occupied by algal culture flasks during the test was about 8116 lux (equivalent to 110 E/m2/s), which was ensured with fluorescent lamps (with a spectral range of 400-700 nm). The differences in light intensity between the test vessels did not exceed  15 % and therefore provided equal conditions for each test vessel.
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :growth rate, yield, biomass (24, 48 & 72 h)
- Determination of cell concentrations: [counting chamber]
- Chlorophyll measurement: No
TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2x
- Range finding study
- Test concentrations: 0, 0.1, 1.0, 10, 100 mg/L
- Results used to determine the conditions for the definitive study:
Results of the Preliminary Range-Finding Test
Nominal concentrations
{mg/L] Untreated
control 0.1 1.0 10.0 100.0
Average of cell number
at 72 hours (x 104cell/mL) 63.33 66.50 47.00 43.50 45.00

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

6.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

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: no
- 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):

- Results with reference substance valid? yes
The date of the last study (Study Code: 13/019-022AL) with the reference item Potassium dichromate is (Batch Number: 0769128): 05 - 08 February 2013.
The 72h ErC 50 : 0.87 mg/L, (95 % confidence limits: 0.79 – 0.96 mg/L)
The 72h EbC 50 : 0.48 mg/L, (95 % confidence limits: 0.44 – 0.53 mg/L)
The 72h EyC 50 : 0.45 mg/L, (95 % confidence limits: 0.41 – 0.50 mg/L)

Reported statistics and error estimates:

The section-by-section specific growth rates in the control cultures were assessed (calculated as the specific growth rates for each day during the course of the test (days 0-1, 1-2 and 2-3) and to demonstrate exponential growth for the entire study period.

The inhibition of alga growth was determined from the biomass (area under the growth curves, A), the average specific growth rate (r) and from the yield (y). Mean values and standard deviations were calculated for each concentration at the start, and at the end of the test using Excel for Windows software (Microsoft Co./One Microsoft Way/Redmond, WA 98052-6399).

Statistical comparisons of biomass, average specific growth rates and yield in control and in treated groups were carried out using analysis of variance (ANOVA) and Bonferroni t-Test ( = 0.05) by TOXSTAT software.
For the determination of the LOEC and NOEC, the calculated mean biomass, growth rates and yield at the test concentrations were tested on significant differences to the control values by Bonferroni t-Test.

As the analytically measured concentration deviated not more than 20 per cent from the nominal in all cases, the biological results are based on the nominal concentrations.

Validity

 The cell density in the control cultures increased by a factor of 68.00 within three days.

The mean coefficient of variation for section-by-section specific growth rates (days 0-1; 1-2; 2-3) in the control cultures was 14.21 %.

The coefficient of variation of average specific growth rates during the whole test period (day 0-3) in the control cultures was0.80 %.

All validity criteria were met, therefore the study can be considered as valid.

Average Specific Growth Rates

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h average specific growth rate was statistically significantly different from the untreated control value at the concentration range of 12.5 – 100 mg/L (nominal), correspondingly the No Observed Effect Concentration (NOEC)was determined as 6.25 mg/L (nominal). The 72 h E_rC₅₀value was determined as higher than 100 mg/L(nominal).

Growth Rates (m) and Percentage Inhibition ofmduring the Test Period

NominalConcentration [mg/L]	Growth rate (m) and % inhibition ofm
	0–24 h		0–48 h		0–72 h
	m	%	m	%	m	%
Control	0.0655^+	0.0	0.0599	0.0	0.0586	0.0
6.25	0.0671	-2.4	0.0597	0.3	0.0587	-0.1
12.5	0.0609	7.1	0.0606	-1.1	0.0561*	4.3
25	0.0578*	11.8	0.0606^+	-1.1	0.0556*	5.1
50	0.0640	2.4	0.0571^+	4.8	0.0553*^+	5.6
100	0.0578*	11.8	0.0577	3.7	0.0554*	5.5

*: statistically significantly different compared to the control values (Bonferroni t-Test;a= 0.05)

⁺          :          at this value the rounding of the EXCEL and TOXSTAT software was different. The table contains the values calculated with EXCEL.

AREAS UNDER THE GROWTH CURVES

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h areas were statistically significantly different from the untreated control value at the examined concentration range of50 – 100 mg/L (nominal), correspondingly the No Observed Effect Concentration (NOEC) was determined as 25 mg/L (nominal).The 72 h E_bC₅₀value was determined as higher than100 mg/L(nominal).

 Area under the Growth Curves (A) and Percentage Inhibition of A during the Test Period

NominalConcentration [mg/L]	Area under the Growth Curves (A) and Percentage Inhibition of A
	0–24 h		0–48 h		0–72 h
	A	%	A	%	A	%
Control	46.0	0.0	294.0	0.0	1300.0	0.0
6.25	48.0	-4.3	296.0	-0.7	1304.0	-0.3
12.5	40.0	13.0	288.0	2.0	1164.0	10.5
25	36.0*	21.7	280.0	4.8	1136.0	12.6
50	44.0	4.3	264.0	10.2	1076.0*	17.2
100	36.0*	21.7	252.0	14.3	1068.0*	17.8

*: statistically significantly different compared to the control values (Bonferroni t-Test;a= 0.05)

YIELD

  

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h yield was statistically significantly different from the untreated control value at the tested concentration range of 12.5 –100 mg/L (nominal), correspondingly the
No Observed Effect Concentration (NOEC) was determined as 6.25 mg/L (nominal).The 72 h E_yC₅₀value was determined as higher than100 mg/L(nominal).

 

Yield (Y) and Percentage Inhibition of Y during the Test Period

NominalConcentration [mg/L]	Yield (Y) and % inhibition of Y
	0–72 h
	Y	%
Control	67.0	0.0
6.25	67.3	-0.5
12.5	55.7*	16.9
25	54.0*	19.4
50	53.0*	20.9
100	53.0*	20.9

*: statistically significantly different compared to the control values (Bonferroni t-Test;a= 0.05)

Validity criteria fulfilled:

yes

Executive summary:

The toxicity of the test item to aquatic algae was assessed in a study performed according to OECD TG 201 over a period of 72 hours in a static test system using Pseudokirchneriella subcapitata.

Test concentrations used during the main study were nominal concentrations of test item used in the main experiment were 6.25, 12.5, 25, 50 and 100 mg/L (measured geometric mean test item concentrations 5.9, 11.8, 24.7, 48.8 and 93.8 mg/L, respectively). As the measured concentrations deviated not more than 20 per cent from the nominal in all cases, biological results are based on the nominal test item concentration.

Three replicates were used per test concentration and six replicates for the controls.

All validity criteria were met therefore the study can be considered as valid.

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h average specific growth rate was statistically significantly different from the untreated control value at the concentration range of 12.5 – 100 mg/L (nominal), correspondingly the NOEC was determined as 6.25 mg/L (nominal). The 72 h ErC50 value was determined as higher than100 mg/L(nominal).

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h areas were statistically significantly different from the untreated control value at the examined concentration range of 50 – 100 mg/L (nominal), correspondingly NOEC was determined as 25 mg/L (nominal). The 72 h EbC50 value was determined as higher than 100 mg/L(nominal).

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h yield was statistically significantly different from the untreated control value at the tested concentration range of 12.5 –100 mg/L (nominal), correspondingly the NOEC was determined as 6.25 mg/L (nominal).The 72 h EyC50 value was determined as higher than 100 mg/L(nominal).

Description of key information

Toxicity to aquatic algae: 72-h ErC50 > 100 mg/L, NOErC 6.25 mg/L (Pseudokirchneriella subcapitata; OECD TG 201)

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

6.25 mg/L

Additional information

The toxicity of the test item to aquatic algae was assessed in a study performed according to OECD TG 201 over a period of 72 hours in a static test system using Pseudokirchneriella subcapitata.

Test concentrations used during the main study were nominal concentrations of test item used in the main experiment were 6.25, 12.5, 25, 50 and 100 mg/L (measured geometric mean test item concentrations 5.9, 11.8, 24.7, 48.8 and 93.8 mg/L, respectively). As the measured concentrations deviated not more than 20 per cent from the nominal in all cases, biological results are based on the nominal test item concentration.

Three replicates were used per test concentration and six replicates for the controls.

All validity criteria were met therefore the study can be considered as valid.

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h average specific growth rate was statistically significantly different from the untreated control value at the concentration range of 12.5 – 100 mg/L (nominal), correspondingly the NOEC was determined as 6.25 mg/L (nominal). The 72 h ErC50 value was determined as higher than100 mg/L(nominal).

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h areas were statistically significantly different from the untreated control value at the examined concentration range of 50 – 100 mg/L (nominal), correspondingly NOEC was determined as 25 mg/L (nominal). The 72 h EbC50 value was determined as higher than 100 mg/L(nominal).

The results of the statistical evaluation (based on Bonferroni t-Test; a=0.05) show that the 0-72 h yield was statistically significantly different from the untreated control value at the tested concentration range of 12.5 –100 mg/L (nominal), correspondingly the NOEC was determined as 6.25 mg/L (nominal).The 72 h EyC50 value was determined as higher than 100 mg/L(nominal).