Benzenesulfonic acid, 4-C10-13-sec-alkyl derivs., ammonium salts

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 June 2017 to 11 December 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted in acordance with international guideines and in accordance with GLP. All guideline validity criteria were met.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

April 2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Principles of method if other than guideline:

- Principle of test: Water accomadated fraction (WAF) was used in accordance with OECD series on testing and assessment no. 23.
- Short description of test conditions: Each concentration is prepared separately, whereby the test item is added to the medium at a specific loading rate. The material is left until fully dissolved (or as disolved as possible), then particulate matter is left to settle out of the water column. The remaining substance in the water column of the sample is transferred to the test vessels. This is done at each concentration. The concentration-response relationship is based on the initial loading of the vessel.
- Parameters analysed / observed: Tyndall's effect is monitored to ensure minimal particulate matter is transferred to the test system. Further, chemical analysis is conducted to confirm that the correct loading has occured, presence of the test item and to confirm there is a series of varying concentrations.

GLP compliance:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: confidential
- Expiration date of the lot/batch: 01 Jan 2019
- Purity test date: N/D

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: N/A
- Specific activity: N/A
- Locations of the label: N/A
- Expiration date of radiochemical substance: N/A

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- Stability under test conditions: sufficient for the test purpose
- Solubility and stability of the test substance in the solvent/vehicle: N/A
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: N/A

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: N/A
- Preliminary purification step (if any): N/A
- Final dilution of a dissolved solid, stock liquid or gel: N/A
- Final preparation of a solid: N/A

FORM AS APPLIED IN THE TEST (if different from that of starting material)

OTHER SPECIFICS: N/A

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All tested concentrations were analysed at 0, 24 hour fresh and 48 hours, the concentrations were 0, 1.21, 2.43, 4.85, 9.70, 19.4 mg/L.
- Sampling method: taken from water column
- Sample storage conditions before analysis: All samples were stored deep frozen until they were transferred to the analytical laboratory.

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: water accomodated fractions were used as per the OECD guidance on testing and assessment no. 23. Each concentration was seperately prepared. The prerequisite amount of test item was added to the test vessel containing the test medium and stirred for 48 h in the dark at room temperature. After the settling the necessary volumes for the test were withdrawn via a Teflon tube from the medium level of the stock solution. Approximately 50 mL of the prepared solutions were transferred to each test vessel. The preparation of the test solutions is shown below.
- Eluate: N/A
- Differential loading: N/A
- Controls: Elendt M4
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N/A
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): N/A
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): After a 5.5 hour settling phase the solutions were clear and transparent and no Tyndall's effect was observed.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Clone V
- Source (laboratory, culture collection): Laboratory culture; Federal Environment Agency in Berlin/Germany
- Age of inoculum (at test initiation): < 24 hour old
- Method of cultivation: Daphnia magna was bred as single culture (1 daphnid per 100 mL) in Elendt M4 medium.
Culture conditions were as follows:
Temperature: 18 - 22 °C
pH: 6 to 9
Dissolved oxygen: > 60 %
Hardness: 140-250 mg/L (CaCO3)
Fed: Desmodesmus subspicatus at least 3 times weeks

ACCLIMATION
- Acclimation period: N/A - bred in the same climate and medium as the test.
- Culturing media and conditions (same as test or not): same as test
- Any deformed or abnormal cells observed: no

Test type:

semi-static

Water media type:

freshwater

Remarks:

Elendt M4

Limit test:

no

Total exposure duration:

48 h

Remarks on exposure duration:

Guideline perscribed

Post exposure observation period:

none

Hardness:

140 - 250 mg/L (as CaCO3) °C

Test temperature:

20.6 ± 0.1

pH:

7.92 ± 0.10

Dissolved oxygen:

8.4 ± 0.2

Salinity:

N/A

Conductivity:

N/A

Nominal and measured concentrations:

Nominal concentrations: 0, 0.625, 1.25, 2.50, 5.00, 10.0 mg/L
Measured concentrations: 0.842, 1.59, 3.17, 5.43 and 9.43 mg/L

As the test was a WAF of a UVCB the entire test item can not be analysed, therefore measured concnetrations simply confirm the presence of the test item and a series of concentrations is present in the test.

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass vessel
- Type (delete if not applicable): covered with a glass plate
- Material, size, headspace, fill volume: 50 mL
- Aeration: None
- Type of flow-through (e.g. peristaltic or proportional diluter): None
- Renewal rate of test solution (frequency/flow rate): every 24 hours
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): No vehicle used
- Biomass loading rate: N/A

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: media prepared with
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides: n/a
- Chlorine: n/a
- Alkalinity: pH 7.8
- Ca/Mg ratio: 232 mg/L
- Conductivity: ND
- Salinity: ND
- Culture medium different from test medium: No
- Intervals of water quality measurement: 0, 24 hour old, 24 hour fresh, 48 hour old

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hour light: 8 hours dark
- Light intensity: ND

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : NOEL and EL50

VEHICLE CONTROL PERFORMED: No vehicle was used

RANGE-FINDING STUDY
- Test concentrations: 0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: 100 % mortality was seen at 10 and 100 mg/L.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

3.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOELR

Effect conc.:

0.625 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EL50

Effect conc.:

6.16 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

NOELR

Effect conc.:

2.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: none observed
- Observations on body length and weight: none observed
- Other biological observations: immobility
- Mortality of control: none
- Other adverse effects control: none
- Abnormal responses: none
- 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
- Relevant effect levels: effective concentration which causes 50 % immobility
- Limit test: no
- Dose-response test: yes
- ECx: EC50 between 1 and 2 mg/L. The effect level is in accordance with the requirements of the OECD guideline 202 and falls within the historical data generated with the reference item at the testing facility, therefore the daphnids were suitable for the determination of the toxicological effects of the test item.
- Other: N/A

Reported statistics and error estimates:

The effects were based on nominal loading rates as the test item is a UVCB. The values for EL50 values were determined by using Trimmed Spearman-Karber procedure. The evaluation of data was performed by ToxRat Professional 3.2.1. The data was presented with 95 % confidence intervals.

Table 1 Immobility at 24 hours

	Nominal test item loading rate [mg/L]
	Control	0.625	1.25	2.50	5.00	10.0
	Immobilised daphnids after 24 h
Group 1	0	0	0	0	3	5
Group 2	0	0	0	0	0	5
Group 3	0	0	0	0	1	5
Group 4	0	0	0	0	1	4
Σ	0	0	0	0	5	19
%	0	0	0	0	25	95

Table 2 Immobility at 48 hours

	Nominal test item loading rate [mg/L]
	Control	0.625	1.25	2.50	5.00	10.0
	Immobilised daphnids after 48 h
Group 1	0	0	1	1	4	5
Group 2	0	0	3	0	5	5
Group 3	0	0	0	0	4	5
Group 4	0	0	0	0	4	5
Σ	0	0	4	1	17	20
%	0	0	20	5	85	100

Table 3 Temperature of the test solutions

	Nominal test item loading rate [mg/L]
	Control	0.625	1.25	2.50	5.00	10.0
Time [h]	Temperature [°C]
0 fresh	20.0	21.0	21.7	21.7	21.8	21.7
24 aged	19.8	19.8	19.9	19.9	19.9	19.9
24 fresh	19.8	22.0	21.9	22.0	21.9	21.8
48 aged	19.8	19.8	19.8	19.8	19.9	19.9
Mean	19.9	20.7	20.8	20.9	20.9	20.8
Std. dev.	0.1	1.1	1.1	1.2	1.1	1.1
Mean	20.6
Std. dev.	1.0

Table 4 pH of the test solutions

	Nominal test item loading rate [mg/L]
	Control	0.625	1.25	2.50	5.00	10.0
Time [h]	pH
0 fresh	7.80	7.90	7.93	7.95	7.96	7.95
24 aged	7.90	7.92	7.92	7.94	7.95	7.97
24 fresh	7.58	7.75	7.85	7.89	7.91	7.93
48 aged	7.92	7.96	7.99	8.01	8.05	8.07
Mean	7.80	7.88	7.92	7.95	7.97	7.98
Std. dev.	0.16	0.09	0.06	0.05	0.06	0.06
Mean	7.92
Std. dev.	0.10

Table 5 Oxygen concentration of the test solutions

	Nominal test item loading rate [mg/L]
	Control	0.625	1.25	2.50	5.00	10.0
Time [h]	Oxygen [mg/L]
0 fresh	8.9	8.5	8.3	8.3	8.3	8.3
24 aged	8.7	8.4	8.3	8.4	8.4	8.4
24 fresh	8.7	7.9	8.2	8.1	8.1	8.1
48 aged	8.7	8.3	8.4	8.3	8.4	8.3
Mean	8.8	8.3	8.3	8.3	8.3	8.3
Std. dev.	0.1	0.3	0.1	0.1	0.1	0.1
Mean	8.4
Std. dev.	0.2

Validity criteria fulfilled:

yes

Conclusions:

A test was conducted to determine the effects of the Benzenesulfonic acid, 4-C10-13-sec-alkyl derivs., ammonium salts on the mobility of the water flea Daphnia magna. The study was conducted in accordance with the OECD guideline 202 the “Freshwater Alga and Cyanobacteria, Growth Inhibition Test”, as the test item is a UVCB of varying solubilities a WAF approach was used in accordance with OECD series on testing and assessment no. 23. There were no deviations from the guidance or study plan, and all validity criteria were met. The results are based on nominal loading rates as the test item is a UVCB.

According to the results of the test, the EL50 (48 h) was determined to be 3.30 mg/L (95 % CI: 2.75-3.96 mg/L) (nominal). The corresponding NOELR (48 h) was 0.625 mg/L (nominal).

Executive summary:

The objective of this study was to determine the effects of the test item on mobility of the water flea Daphnia magna, to determine the effect loading rate that casues 50 % immobility of the organism (EL₅₀). The study was conducted in accordance with the OECD guideline 202 the “Daphnia sp., Acute immobilisation test”, as the test item is a multi-constituent of varying solubilities a WAF approach was used in accordance with OECD series on testing and assessment no. 23.  A range finding study was used to determine definitive test concentrations and optimal stirring and settling time for the creation of the WAFs at each concntration. The definitive test concentrations were, 0, 0.625, 1.25, 2.50, 5.00 and 10 mg/L (all concentrations are based on test item added minus its water content as water is deemed to not contribute to the toxic effects observed, if any). WAFs were prepared at each concentration by adding a prerequisite amount of test item to the test vessel, stirring for 48 hours in the dark at room tempreature and then leaving any particulate to settle for 5.5 hours. The test solutions were clear and no particulate could be seen, this was further confirmed as no Tyndall's effect could be seen in any of the test solutions. There were no major deviations, however temperature during the study exceeded a change of 1 degree celcius (actual 19.8 to 21.8 degrees celcius), this was not assumed to impact the study and is within a normal range for the organism (18 to 20 degrees celcius as seen in OECD 202). All water quality parameters and study validity criteria were met as per OECD guideline 202. A positive control (potassium dichromate) is routinely run at the laboratory proving sensitivity of the test organism. The positive control EC50 was within the standard range. The test results were anaylsed using the Trimmed Spearmen-Karber Method. The results were based on loading of the test item as the test item is a UVCB.

According to the results the EL50 (48 hour) was estimated to be 3.30 mg/L (95 % CI: 2.75 -3.96 mg/L) (nominal). The NOELR (48 hour) was 0.625 mg/L (nominal)

Description of key information

48h EL50 = 3.30 mg/L, OECD 202 (Daphnia magna), Kummich, F. (2017)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

3.3 mg/L

Additional information

OECD 202 (2007) - The study was conducted in accordance with the OECD guideline 202 the “Daphnia sp., Acute immobilisation test”, as the test item is a multi-constituent of varying solubilities a WAF approach was used in accordance with OECD series on testing and assessment no. 23.  A range finding study was used to determine definitive test concentrations and optimal stirring and settling time for the creation of the WAFs at each concntration.  The definitive test concentrations were, 0, 0.625, 1.25, 2.50, 5.00 and 10 mg/L (all concentrations are based on test item added minus its water content as water is deemed to not contribute to the toxic effects observed, if any). WAFs were prepared at each concentration by adding a prerequisite amount of test item to the test vessel, stirring for 48 hours in the dark at room tempreature and then leaving any particulate to settle for 5.5 hours. The test solutions were clear and no particulate could be seen, this was further confirmed as no Tyndall's effect could be seen in any of the test solutions. There were no major deviations, however temperature during the study exceeded a change of 1 degree celcius (actual 19.8 to 21.8 degrees celcius), this was not assumed to impact the study and is within a normal range for the organism (18 to 20 degrees celcius as seen in OECD 202). All water quality parameters and study validity criteria were met as per OECD guideline 202. A positive control (potassium dichromate) is routinely run at the laboratory proving sensitivity of the test organism. The positive control EC50 was within the standard range. The test results were anaylsed using the Trimmed Spearmen-Karber Method. The results were based on loading of the test item as the test item is a UVCB.