Reaction mass of sodium hydrogen N,N-dibutyl-10-(sulphonatooxy)octadecanamidate and sodium sulphooctadecanoate

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:

2012--09-19 to 2012-12-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted April 13, 2004

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

May 30, 2008

Qualifier:

according to guideline

Guideline:

other: OECD No 23; Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures

Version / remarks:

December 15, 2000

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

One sample of the freshly prepared stock solutions and duplicate samples from the freshly prepared test media of all test concentrations and the control were taken at the start of the test and at day 1.
For the determination of the stability of the test item under test conditions and of the maintenance of the test item concentrations during the test period, duplicate samples from the aged test media of all test concentrations and the control were collected at the end of the test and at day 1 (after 24 hours of exposure) by pouring together the contents of the test beakers of each treatment.
The concentrations of the test item were analysed in unicate samples of the stock solution and in duplicate samples of the test media from all sampling times (0, 24 hours and 48 hours). From the control samples only one of the duplicate samples was analysed from each sampling time.

Vehicle:

no

Details on test solutions:

A stock solution of 50 mg/L was prepared by dissolving 23.6 and 21.7 mg test item into 472 and 434 mL test water, respectively, by intense stirring for 10 minutes. Adequate volumes of this stock solution were diluted with test water to prepare the test media of the desired test concentrations.
The test media were prepared just before introduction of the daphnids (= start of the test) and at test medium renewal on Day 1. The appearance of the test item in the test media was observed at the start of the test, on Day 1 in the old and the new test media and at the end of the test in the test media of all test concentrations.

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: Daphnia magna (Straus), clone 5
Age at Test Start: From 2.25 to 17.75 hours old
Sex: Female
Origin: The daphnids introduced in the test were taken from an in house laboratory culture.
Breeding Conditions: The daphnids were bred in the laboratories of IBACON under similar temperature and light conditions as used in the test. The cultivation of the parental daphnids was performed in reconstituted water of a similar quality with regards to pH, the constituent salts and total hardness as the test water used in the test. The test organisms were not first brood progeny. The daphnids in the stock culture were fed at least on all working days with green algae (Desmodesmus subspicatus) freshly grown in the laboratories of IBACON.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

250 mg CaCO3/L

Test temperature:

20 °C in the freshly prepared test media and 19 to 20 °C in the aged test media

pH:

7.9 to 8.1 in the freshly prepared media;
7.7 to 7.8 in the aged test media; and thus the pH-value did not vary by more than 1.5 units

Dissolved oxygen:

8.4 to 9.8 mg/L in the freshly prepared media;
8.1 to 8.8 mg/L in the aged test media

Nominal and measured concentrations:

Nominal concentrations: 10, 4.5, 2.1, 0.94 and 0.43 mg test item/L and a control
Measured Concentration: below LOQ

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beakers
- Type: closed
- Material, size, fill volume: glass, 100 mL, 60 mL
- Volume of solution: 60 mL
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Preparation of dilution water: Reconstituted water (Elendt "M4")

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light: 8 h dark
- Light intensity: 860 - 980 lux

EFFECT PARAMETERS MEASURED: The mobility of the daphnids was determined by visual observation after 24 and 48 hours. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).

RANGE-FINDING STUDY
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

no

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

4.58 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

2.51 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

0.94 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

2.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

After 48 hours of exposure no immobilisation of the test animals was observed in the control and up to and including the test item concentration of nominal 0.94 mg test item/L. At the nominal concentration of 2.1 mg test item/L two animals were immobile and 7 animals were immobile at the nominal concentration of 4.5 mg test item/L after 48 hours of exposure. At the highest test concentration of nominal 10 mg test item/L all 20 animals were immobile after 48 hours.
After 24 hours of test duration no immobilisation of the test animals was observed in the control and in the concentrations up to and including nominal 0.94 mg test item/L. At the nominal concentration of 2.1 mg test item/L one Daphnia was immobile and two daphnids were immobile at the nominal concentration of 4.5 mg test item/L. At the highest test concentration of nominal 10 mg test item/L 15 daphnids were immobile after 24 hours test duration.

Reported statistics and error estimates:

The 24-hour and 48-hour EC50, EC20 and EC10 and the 95 % confidence limits were calculated by Probit analysis. The NOEC and LOEC after 24 and 48 hours were determined directly from the raw data. The software used to perform the statistical analysis was ToxRat Professional, Version 2.10.05, ToxRat® Solutions GmbH.

Table 1: Summary of Biological Results

Nominal concentration	% of immobilised daphnids after
[mg test item/L]	24 hours	48 hours
Control	0	0
0.43	0	0
0.94	0	0
2.1	5	10
4.5	10	35
10	75	100
EC50 [mg/L]:	7.38	4.58
95 % CI [mg/L]:	5.81 – 10.24	3.70 – 5.70
EC20 [mg/L]:	4.52	3.08
95 % CI [mg/L]:	3.00 – 5.74	2.19 – 3.80
EC10 [mg/L]:	3.50	2.51
95 % CI [mg/L]:	1.96 – 4.61	1.60 – 3.19
NOEC [mg/L]:	0.94	0.94
LOEC [mg/L]:	2.1	2.1

Values refer to nominal test concentrations.

CI: Confidence interval

n.d.: could not be determined

NOEC and LOEC were determined directly from the raw data.

Table 2: Analytical Results

Determination of the Test Item:	Based on the results of TOC measurements the concentration of the test item was determined using a calibration curve.
Calibration Range:	TC: 2.5 to 100 mf Carbon/L IC: 1 to 25 mg Carbon/L
Linearity of Response:	Correlation of peak area of different standard solutions with their corresponding concentrations, using a linear regression
Regression Coefficient (r^2):	TC: at least 0.9991 IC: at least 0.9935
Typical Calibration Curves:	TC: y = 4.2877 * x – 1.9182 IC: y = 4.4053 * x +1.5142
Limit of Detection:	TC: 7.7074 mg Carbon/L IC: 3.5314 mg Carbon/L
Limit of Quantification:	20 mg test item/L 96 % (n = 9, RSD 25 %)
Mean Recovery in the Fortified Samples:	96 % (n = 18, RSD 19 %)
Mean Recovery in the Stock Solutions:	At day 0: 111 % (n = 1) At day 1: 90 % (n = 1) All other values were below the Limit of Quantification.

Table 3: Summary of Analytical Results

sample description [mg/L]	% of nominal^1	RSD	n
Control	< LOQ	n.a.	4
0.43	< LOQ	n.a.	8
0.94	< LOQ	n.a.	8
2.1	< LOQ	n.a.	8
4.5	< LOQ	n.a.	8
10	< LOQ	n.a.	8
stock solution	101	15	2

¹mean value of all measured samples per treatment group

RSD: relative standard deviation per treatment group

n: number of analysed samples

n.a.: not applicable

Validity criteria fulfilled:

yes

Conclusions:

The toxic effect of the test item to Daphnia magna was assessed in a semi-static dose-response test. The 48-hour NOEC was determined to be 0.94 mg test item/L. The 48-hour LOEC was determined to be 2.1 mg test item/L and the 48-hour EC50 value was calculated to be 4.58 mg test item/L.

Executive summary:

The purpose of this study was to evaluate the acute toxicity of the test item on the mobility of Daphnia magna according to OECD 202 test guideline. Young daphnids (< 24 hours old) were exposed in a semi-static test to the test item for 48 hours, added to test water at a range of concentrations. The study comprised of 6 treatment groups (5 dose rates of the test item, control) each containing 20 individuals. Under otherwise identical test conditions, different concentrations of the test item result in different percentages of daphnids being no longer capable of swimming at the end of the test. The mobility of the daphnids was determined in a semi-static 48-hour test by visual observation after 24 and 48 hours. The samples collected at test start, after 24 hours and after 48 hours were analysed via TOC-method. The purpose of the analytical part of this study was to verify the concentrations of the test item in the test water. At day 0, 111 % of nominal was determined in the freshly prepared stock solution. At day one, 90 % were determined in the freshly prepared stock solution. All other values were below the Limit of Quantification. Since correct dosing of the stock solutions could be demonstrated and no further information is available, all reported results refer to nominal concentrations. After 48 hours of exposure no immobilisation of the test animals was observed in the control and up to and including the test item concentration of nominal 0.94 mg test item/L. At the nominal concentration of 2.1 mg test item/L two animals were immobile and 7 animals were immobile at the nominal concentration of 4.5 mg test item/L after 48 hours of exposure. At the highest test concentration of nominal 10 mg test item/L all 20 animals were immobile after 48 hours. The 48-hour NOEC was determined to be 0.94 mg test item/L. The 48-hour LOEC was determined to be 2.1 mg test item/L and the 48-hour EC₅₀ value was calculated to be 4.58 mg test item/L.

Description of key information

The toxic effect of the test item to Daphnia magna was assessed in a semi-static dose-response test. The 48-hour NOEC was determined to be 0.94 mg test item/L. The 48-hour LOEC was determined to be 2.1 mg test item/L and the 48-hour EC₅₀ value was calculated to be 4.58 mg test item/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

4.58 mg/L

Additional information

The purpose of this study was to evaluate the acute toxicity of the test item on the mobility of Daphnia magna according to OECD 202 test guideline. Young daphnids (< 24 hours old) were exposed in a semi-static test to the test item for 48 hours, added to test water at a range of concentrations. The study comprised of 6 treatment groups (5 dose rates of the test item, control) each containing 20 individuals. Under otherwise identical test conditions, different concentrations of the test item result in different percentages of daphnids being no longer capable of swimming at the end of the test. The mobility of the daphnids was determined in a semi-static 48-hour test by visual observation after 24 and 48 hours. The samples collected at test start, after 24 hours and after 48 hours were analysed via TOC-method. The purpose of the analytical part of this study was to verify the concentrations of the test item in the test water. At day 0, 111 % of nominal was determined in the freshly prepared stock solution. At day one, 90 % were determined in the freshly prepared stock solution. All other values were below the Limit of Quantification. Since correct dosing of the stock solutions could be demonstrated and no further information is available, all reported results refer to nominal concentrations. After 48 hours of exposure no immobilisation of the test animals was observed in the control and up to and including the test item concentration of nominal 0.94 mg test item/L. At the nominal concentration of 2.1 mg test item/L two animals were immobile and 7 animals were immobile at the nominal concentration of 4.5 mg test item/L after 48 hours of exposure. At the highest test concentration of nominal 10 mg test item/L all 20 animals were immobile after 48 hours. The 48-hour NOEC was determined to be 0.94 mg test item/L. The 48-hour LOEC was determined to be 2.1 mg test item/L and the 48-hour EC₅₀ value was calculated to be 4.58 mg test item/L.

The reported effect concentrations refer to the commercial formulation containing 46.2% (w/w) of the REACH registration substance which reflects the test item used in the study.