Dibenzyl sulphoxide

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:

05.09. – 07.10.2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Preliminary test: The analytical determination of the test substance concentrations was performed at the beginning and at the end of the test. The samples for analysis were taken from the highest (100 mg.L-1) and the lowest (1 mg.L-1) test concentrations.
The samples for analysis (0 hours) were prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. The samples were analyzed on the day of delivery. The samples for analysis at the end of the test (48 hours) were delivered to analytical laboratory immediately after the end of testing. The samples were analysed on the day of delivery. All samples were stored at laboratory temperature.
The analytical results showed that the test substance was stable but poorly soluble in dilution water at the conditions of the test.


Definitive test: The analytical determination of the test substance concentrations was performed at the beginning and at the end of the test. The samples for analysis were taken from the highest (100 mg·L-1) and the lowest (21 mg·L-1) test concentrations. The samplesfor analysis (0 hours) were prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. The samples were analysed on the day of delivery. The samples for analysis at the end of the test (48 hours) were delivered to analytical laboratory immediately after the end of testing. The samples were analysed on the day of delivery. All samples were stored at laboratory temperature.

Details on test solutions:

The test substance is poorly soluble in dilution water at concentrations needed for the test. For this reason the preliminary experiment was performed for finding suitable period for achieving the highest solubility.
For the preliminary experiment the solutions were prepared as follows: 100 mg of the test substance in 1000 mL of dilution water was ultrasonicated for 30 minutes and stirred for period 3, 24, 48 and 72 hours on a shaft stirrer, subsequently filtered through 0.45 μm filter and the concentrations of the test substance in resulted solutions were determined.
On the basis of the measured concentrations of the test substance, the period of 72 hours stirring was chosen.
Final procedure for the preparation of the test solutions was then set as follows: The stock solutions of the test substance were prepared in the dilution water. 100 mg of the test substance was weighed into 1 000 mL of the dilution water for the preliminary and definitive test. The stock solutions were ultrasonicated for 30 minutes then stirred for period 72 hours and subsequently filtered through 0.45 μm filter.
The concentrations of test solutions used in the preliminary and definitive test were obtained by dilution of the stock solution with dilution water.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia Magna
- Strain: Straus
- Source: own laboratory breeding
- Age at study initiation: young daphnia born in 24 hours
- Method of breeding: aperiodic parthenogenesis
- Feeding during test: no

ACCLIMATION
- Acclimation period and conditions:
The grow-up gravid females are separated into dilution water. Young daphnia born in 24 hours are segregated to the handling container from which they entered the experiment. At handling a care should be taken of not to penetrate the air under animal’s shell.


- Type and amount of food: a mixture of algae
- Health during acclimation: no mortality observed
The sensitivity of the test species is periodically monitored by testing with the reference substance, potassium dichromate.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

The content of sum Ca + Mg ions should have the value of 2.50 +/- 0.10 mmol/L, values measured in this study were 2.47 mmol·L-1 in the preliminary test and the definitive test.

Test temperature:

20.0 ± 2.0 °C; with a variation of ± 1°C at each individual test

pH:

values measured in this study were 7.9 and 8.0 in preliminary test and 8.0 in the definitive test

Dissolved oxygen:

higher than 3 mg/L
7.9 mg·L-1 (definitive test)

Conductivity:

2.73 µS·cm-1(definitive test)

Nominal and measured concentrations:

100, 77, 59, 45, 35, 27, 21 mg·L-1
21 20.84 (0 h) 21.87 (48 h)mg·L-1
100 100.10 112.03

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beakers 50ml
- Type (delete if not applicable): open
- Aeration: no
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- Biomass loading rate: 50 mL per 10 individuals

For holding and culturing of the daphnia the M4 medium was used (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202).For test was used dilution water (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202).
Dilution water was prepared just before the test by admixing 2.5 mL of stored solutions A, B, C and D into 1 000 mL of deionized water.
A. 117.6 g CaCl2.2H20 in 1 L of deionized water
B. 49.3 g MgSO4.7H2O in 1 L of deionized water
C. 25.9 g NaHCO3 in 1 L of deionized water
D. 2.3 g KCl in 1 L of deionized water

OTHER TEST CONDITIONS
- Adjustment of pH: The dilution water should have of pH 7.8 +/- 0.2, values measured in this study were 8.0 in the definitive test.
- Photoperiod: daylight

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
immobilisation of daphnia after 24 and 48 hours
Daphnia is considered as immobile (incapable of swimming) when exhibit no motion during 15 seconds after a gentle agitation of the test container. The values of 24hour-EC50 and 48hour-EC50 are obtained by calculation. The values of EC0 and EC100 are identified by direct observation.

RANGE-FINDING STUDY
- Test concentrations: 100, 50, 10, 5 and 1 mg·L-1 + C(control)
- Results used to determine the conditions for the definitive study:
Immobilisation % (48 h) 100, 45, 0, 0, 0, 0 %

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

71.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC0

Effect conc.:

21 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

21 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Results with reference substance (positive control):

Reference test
The sensitivity of the test species and correctness of test performance is periodically verified in six-month period by testing with the reference substance, potassium dichromate.
The results of the verification test with K2Cr2O7, carried out in period from 13.04. to 15.04. 2015 are the following:
24 hour – EC50 = 1.17 mg.L-1(95% confidence limit: 1.03 – 1.37 mg.L-1)
48 hour – EC50 = 0.62 mg.L-1(95% confidence limit: 0.52 – 0.72 mg.L-1)

Interlaboratory test data (13 laboratories)
48 hour – EC50 = 0.46 – 1.08 mg·L-1

The immobilisation of the test organism by the reference substance (48hour – EC50) obtained from our last reference test meets the calculated range from the interlaboratory test.

Validity criteria fulfilled:

yes

Remarks:

1. Immobilisation in the controls < 10 % at the end of the test. 2. The dissolved oxygen concentrations (throughout the test) > 3 mg·L-1.

Conclusions:

The value of EC50 and the confidence intervals of the EC50 were calculated using the software ToxRat Professional Version 3.2.1.
The EC0 and EC100 values were identified by direct observation.
There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 % of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values.
Because concentrations of the test substance have been satisfactorily maintained within ± % of the measured initial concentration throughout the test, the nominal concentrations were used for all evaluations and results.

24 hour–EC50 = 71.8 mg·L-1
(95% confidence limit: 63.1 – 85.4 mg·L-1)
48 hour–EC50 = 50.0 mg·L-1
(95% confidence limit: 44.0 – 57.4 mg·L-1)

Executive summary:

The test substance, Dibenzylsulfoxide, was tested in acute immobilisation test on Daphnia magna.

The test was performed according to method C.2 – Daphnia sp. Acute Immobilisation Test, Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008.

The preliminary test was performed in a range of the test substance nominal concentrations

1 – 100 mg·L-1. Based on toxicity of the test substance found in the preliminary test, the definitive test was performed in appropriate concentration range.

The analytical results showed, that the test substance Dibenzylsulfoxide was sufficiently stable in dilution water in the conditions of the test, therefore the definitive test was performed as static one.

The definitive test was performed in a range of the test substance nominal concentrations

21 – 100 mg·L-1. Samples for analytical determination were taken at the beginning and at the end of the test.

There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 percent

of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values.

The nominal concentrations were used for all evaluations and results.

 

Test results:

24 hour–EC50 = 71.8 mg·L-1                         (nominal concentration)

(95% confidence limit: 63.1 – 85.4 mg·L-1)

48 hour–EC50 = 50.0 mg·L-1                          (nominal concentration)

(95% confidence limit: 44.0 – 57.4 mg·L-1)

 

24 hour–EC0 = 21 mg·L-1                                (nominal concentration)

48 hour–EC0 = 21 mg·L-1                                (nominal concentration)

 

24 hour–EC100 ˃ 100 mg·L-1                         (nominal concentration)

48 hour–EC100 ˃ 100 mg·L-1                         (nominal concentration)

Description of key information

The value of EC50 and the confidence intervals of the EC50 were calculated using the software ToxRat Professional Version 3.2.1.

The EC0 and EC100 values were identified by direct observation.

There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 % of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values.

Because concentrations of the test substance have been satisfactorily maintained within ± % of the measured initial concentration throughout the test, the nominal concentrations were used for all evaluations and results.

24 hour–EC50  = 71.8 mg·L-1

(95% confidence limit: 63.1 – 85.4 mg·L-1)

48 hour–EC50 = 50.0 mg·L-1

(95% confidence limit: 44.0 – 57.4 mg·L-1)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

50 mg/L