2-Butene-1,4-diol, 1,4-diacetate, (2Z)-

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:

2015-07-21 to 2015-07-23

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:

2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control, 6.25, 12.5, 25.0, 50.0 and 100 mg/L
- Sampling method: At the start of the exposure intervals (0 and 24 hours), sampling was carried out after preparation of the test concentrations. At the end of the exposure intervals (24 and 48 hours), samples were taken from additional replicates, prepared with test media, but without daphnids, for the sampling of the aged media, because volumes of the test replicates were insufficient for the analytical determination of the test concentrations. These replicates were incubated under test conditions until sampling. The method was validated prior to this study according to SANCO 3029/99 rev.4 (2000).
- Sample storage conditions before analysis: All original samples were stored at room temperature before preparation. Prepared samples were stored in an autosampler at room temperature until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION:
A stock solution (100 mg/L of the test item were weighed out) was freshly prepared with the dilution water before the start of each exposure interval (at 0 and 24 hours). The stock solution was mixed thoroughly by hand until the test item was completely dissolved.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain/clone: Daphnia magna STRAUS (Clone 5)
- Age at study initiation: 2 to 24 hours old daphnids from a healthy stock were used for the study. Juvenile daphnids were removed from the culture vessels latest 22 hours before the start of the exposure and discarded. The juveniles
born within this period of max. 22 hours preceding the exposure were used for the test after an acclimatisation phase of at least 2 hours in the dilution water. No first brood progeny was used for the test.
- Method of breeding: The algae are cultured at the test facility. In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 + 2°C, in an incubator and 16 h illumination. The culture daphnids are fed at least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus, with an algae cell density of > 10^6 cells/mL.
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), 14195 Berlin, Germany
- Feeding during test : The daphnids were not fed during the study.

ACCLIMATION
- Acclimation period: At least 2 hours in dilution water

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

265 mg CaCO3/L (0 hours)
262 mg CaCO3/L (24 hours)

Test temperature:

20.5 °C (0 hours)
20.8 °C (24 hours)

pH:

7.52 - 8.2

Dissolved oxygen:

8.65 - 9.01 mg/L

Salinity:

not applicable

Conductivity:

671 µS/cm (0 hours)
669 µS/cm (24 hours)

Nominal and measured concentrations:

Nominal: Control, 6.25, 12.5, 25.0, 50.0 and 100 mg/L
Mean measured: The measured test item concentrations were all within ± 20 % of the nominal concentrations throughout the exposure period (see table 2).

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers
- Type: closed , loosely covered with watch glasses
- Material, size, fill volume: glass, 50 m L, 20 mL
- Volume of solution: 20 mL
- Renewal rate of test solution: Yes, daily renewal of the test solutions
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Test water, according to OECD 202.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: Diffuse light, light intensity of max. 20 µE/m²/s (1340 Ix)

EFFECT PARAMETERS MEASURED: Immobilisation was determined in all groups after 24 and 48 hours. An animal was considered to be immobile, if it was not able to swim in the water phase within 15 seconds after gentle agitation of the test vessel. Other observations were not made.

VEHICLE CONTROL PERFORMED: no t applicable

RANGE-FINDING STUDY
- Test concentrations: 1.00, 10.0 and 100 mg/L.
- Results used to determine the conditions for the definitive study: The results of the non GLP Preliminary Range Finding Test are shown in Table 1 in the section "Any other information on material and methods incl. tables"

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

35.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% confidence limit: 25.0 - 50.0

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

34.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% confidence limit: 25.0 - 50.0

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Immobilisation of control: no
- other: The tested concentration levels were all visually clear throughout the exposure in both exposure intervals.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: EC50: 1.88 mg/L (CI: 1.62 - 2.19 mg/L)

Reported statistics and error estimates:

The EC100-values (after 24 and 48 hours) as well as the NOEC and the LOEC (after 48 hours) were estimated directly from the observation data, without mathematical calculation.
The EC10-values (after 24 and 48 hours) were calculated by sigmoidal dose-response regression. The EC10-values (after 24 and 48 hours) were calculated as the geometric mean of the highest concentration level resulting in a mean immobilisation rate of 0 % and the lowest concentration level resulting in a mean immobilisation rate of 100 %, because only a partial effect was observed. Both concentration levels mentioned above were also used as 95 % confidence limits.
The EC50-value for the reference item was calculated by sigmoidal dose-response regression. The respective 95 % confidence limits were calculated from the standard error and the (-distribution with the best-fit values with the software GraphPad Prism5.
All data were computer processed and rounded for presentation. Consequently, minor variations may occur from the original figures if manual calculations based on the original figures are made subsequently. Calculations were made using the following software:
- GraphPad Prism5, GraphPad Software, Inc.
- Excel, Microsoft Corporation

Measured Exposure Concentrations during the Definitive Test

The concentrations of the test item were analytically verified via GC-MS/MS at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours) in all concentration levels and in the control. The measured concentrations of the test item at the start of the exposure intervals (0 and 24 hours) were in the range of 83 to 108 % of the nominal values. At the end of the exposure intervals (24 and 48 hours), the measured concentrations of the test item were in the range of 82 to 107 % of the nominal values. The analytical results are presented in Table 2. The measured test item concentrations were all within ± 20 % of the nominal concentrations throughout the exposure period. This indicates, that the test item concentrations were successfully maintained for the duration of the test.

Table 2: Measured Concentrations and Percent of the Nominal Concentrations of the Test Item during the Definitive Test

Sampling	2015-07-21 Start of the exposure interval		2015-07-22 24 hours End of the exposure interval		2015-07-22 Start of the exposure interval		2015-07-23 24 hours End of the exposure interval
Start of analysis	2015-07-21		2015-07-22		2015-07-22		2015-07-23
Nonminal test item concentration [mg/L]	Art. 278717
	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%
100	108*	108	107	107	103	103	82.7	83
50.0	44.2	88	47.8	96	51.5	103	51.9	104
25.0	25.6	102	23.7	95	26.5	106	24.0	6
12.5	10.4	83	12.9	103	13.2	106	11.6	93
6.2	6.56*	105	6.50	104	6.08	97	5.14	82
Control	<LOQ		<LOQ		<LOQ		<LOQ

Meas. conc.: measured concentration of the test item, single injection, enrichment and dilution factors taken into account

%: percent of the nominal concentration of the test item

LOQ: limit of quantification of the analytical method (0.50 mg/L of the test item)

* = reanalysed on 2015-07-22, mean value of two replicates, single injection, enrichment and dilution factors taken into account

 

Validity Criteria

The study was performed according to OECD Guideline 202 (2004), which is equivalent to the Council Regulation (EC) No.440/2008 Method C.2 (2008). The validity criteria were fulfilled:

• In the control group, no daphnids were immobilised or showed any signs of disease or stress, e.g. discoloration or unusual behaviour such as trapping on the surface of the water, during the 48-hour test period (required: not more than 10 percent of the daphnids in the control).

• The dissolved O2 concentration at the end of the exposure intervals was > 8.28 mg/L (required: > 3 mg/L at the test end of the exposure intervals) in the test vessels of all tested concentration levels and the control.

Validity criteria fulfilled:

yes

Conclusions:

Based on the analytically confirmed nominal concentrations of the test item the 48 hour-EC50 for Daphnia magna was 35.4 mg/L (95 % confidence limits: 25.0 - 50.0 mg/L). The NOEC after 48 hours was 25.0 mg/L. The LOEC after 48 hours was 50.0 mg/L.

Executive summary:

The effects of the test item to Daphnia magna (STRAUS) were determined in an acute immobilisation test according to OECD 202 (2004). The study was conducted under semi-static conditions over a period of 48 hours with nominal test item concentrations of 6.25, 12.5, 25.0, 50.0 and 100 mg/L. Twenty daphnids were exposed to each concentration level and the control. The concentrations of the test item were analytically verified via GC-MS/MS at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours) in all concentration levels and in the control. The measured test item concentrations were all within ± 20 % of the nominal concentrations throughout the exposure period. This indicates, that the test item concentrations were successfully maintained for the duration of the test. Therefore, the effect levels were based on the nominal concentrations of the test item. The validity criteria of the test guideline were fulfilled. Based on the analytically confirmed nominal concentrations of the test item the 48 hour-EC50 for Daphnia magna was 35.4 mg/L (95 % confidence limits: 25.0 - 50.0 mg/L). The NOEC after 48 hours was 25.0 mg/L. The LOEC after 48 hours was 50.0 mg/L.

Description of key information

Based on the analytically confirmed nominal concentrations of the test item the 48 hour-EC50 for Daphnia magna was 35.4 mg/L (95 % confidence limits: 25.0 - 50.0 mg/L). The NOEC after 48 hours was 25.0 mg/L. The LOEC after 48 hours was 50.0 mg/L (reference 6.1.3-1).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

35.4 mg/L

Additional information

The effects of the test item to Daphnia magna (STRAUS) were determined in an acute immobilisation test according to OECD 202 (2004). The study was conducted under semi-static conditions over a period of 48 hours with nominal test item concentrations of 6.25, 12.5, 25.0, 50.0 and 100 mg/L. Twenty daphnids were exposed to each concentration level and the control. The concentrations of the test item were analytically verified via GC-MS/MS at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours) in all concentration levels and in the control. The measured test item concentrations were all within ± 20 % of the nominal concentrations throughout the exposure period. This indicates, that the test item concentrations were successfully maintained for the duration of the test. Therefore, the effect levels were based on the nominal concentrations of the test item. The validity criteria of the test guideline were fulfilled. Based on the analytically confirmed nominal concentrations of the test item the 48 hour-EC50 for Daphnia magna was 35.4 mg/L (95 % confidence limits: 25.0 - 50.0 mg/L). The NOEC after 48 hours was 25.0 mg/L. The LOEC after 48 hours was 50.0 mg/L (reference 6.1.3-1).