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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:

24 - 26 May 2016

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

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD series on testing and assessment No. 23

Version / remarks:

2000

GLP compliance:

yes (incl. QA statement)

Remarks:

Staatliches Gewerbeaufsichtsamt Hildesheim, Germany

Analytical monitoring:

yes

Details on sampling:

- Concentrations: control, 2.5, 5.0, 10.0, 20.0, 40.0 mg/L
- Sampling method: Sampling was performed after preparation of the test concentrations after 0 and 24 h and at the end of the exposure intervals at 24 and 48 h. Samples were directly taken from the test vessels. All sampling vials were completely filled having no headspace and were tightly closed.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until the start of the analysis, if necessary. Prepared samples were stored in the autosampler at room temperature until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A stock solution (40 mg/L) was prepared with dilution water one day prior to the test start (at 0 and 24 h). The stock solution was stirred without headspace at room temperature for 24 h with approx. 1100 rpm. Five test concentrations were prepared by dilution of the stock solution.
- Eluate: no
- Differential loading: no
- Controls: yes, test medium control
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no, all test solutions appeared to be clear and colorless. No Tyndall effect was recorded in the stock solution of 40 mg/L.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain/clone: Clone 5
- Feeding during test: none
- Culture: Daphnids were cultured in Elendt M4 medium in glass vessels (2-3 L capacity) with approx 1.8 L of medium at 20 ± 2 °C and 16 h light/8 h darkness (max. 1340 lux). The culture was fed at least 5 times per week ad libitum with a mix of unicellular algae (e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus) with an algae cell density of > 10E+06 cells/mL.
- Age at study initiation: < 24 h (no first brood progeny was used)

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

174 - 178 mg CaCO3/L (dilution water)

Test temperature:

20.5 - 20.6 °C

pH:

8.01 - 8.98

Dissolved oxygen:

7.10 - 9.32 mg O2/L

Conductivity:

489 - 491 µS/cm (dilution water)

Nominal and measured concentrations:

nominal: control, 2.5, 5.0, 10.0, 20.0, 40.0 mg/L
measured: < LOQ, 1.93, 4.25, 7.22, 13.9, 30.1 mg/L (geometric mean)

Details on test conditions:

TEST SYSTEM
- Test vessel: Sealed glass glasks with screw caps (polypropylene) filled woth approx. 130 mL of test medium
- Type: closed
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): after 24 h
- 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: Elendt M4 medium
- Culture medium different from test medium: no, same as test
- Intervals of water quality measurement: Water quality parameters were measured prior to the start (0 h) and after medium renewal (24 h).

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light / 8 h darkness
- Light intensity: diffuse light, light intensity of max 20 µE*m-2*s-1 (max. 1340 lux)

EFFECT PARAMETERS MEASURED
- Immobilisation: Immobilisation was measured after 24 and 48 h of exposure

RANGE-FINDING STUDY
- Test concentrations: control, 10, 40 mg/L
- Results used to determine the conditions for the definitive study: 0% immobilisation in control, 10% immobilisation at 10 mg/L, 100% immobilisation at 40 mg/L after 48 h

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

12.9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CI

Remarks:

7.28 - 24.4 mg/L

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

13.7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CI

Remarks:

7.60 - 24.9 mg/L

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

30.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Mortality of control: 0%
- 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: EC50: 2.08 mg/L (95% CI: 1.00 - 4.00 mg/L)
- Dose-response test: 1.00, 2.00, 4.00 mg/L

Reported statistics and error estimates:

The EC10- and EC50-values (after 24 and 48 h of exposure) were calculated by sigmoidal dose-response regression with the software GraphPad Prism 5. The respective 95% confidence limits were calculated from the standard error and the t-distribution. All calculations were carried out from the best-fit values with the software GraphPad Prism 5. The EC50-value for the reference item was calculated accordingly.

Table 1: Immobilisation after 24 and 48 h.

Geomatric mean measured concentration [mg/L]	Immobilisation [%]
	24 h					48 h
	Replicates					Replicates
	1	2	3	4	Mean	1	2	3	4	Mean
30.1	100	80	60	80	80	100	100	100	100	100
13.9	0	0	20	20	10	60	40	60	80	60
7.22	0	0	0	0	0	0	0	0	0	0
4.28	0	0	0	0	0	0	0	0	0	0
1.93	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

Table 2: Measured concentrations and percent of nominal concentrations of test item.

Nominal concentration [mg/L]	0 h of exposure		24 h end of exposure		24 h start of exposure		48 h end of exposure		Geometric mean [mg/L]
	Meas. Concentration [mg/L]	%	Meas. Concentration [mg/L]	%	Meas. Concentration [mg/L]	%	Meas. Concentration [mg/L]	%
40	30.2	76	23.0	57	36.7	92	32.3	81	30.1
20	14.0	70	11.2	56	18.4	92	12.8	64	13.9
10	7.52	75	5.80	58	9.73	97	6.40	64	7.22
5	4.17	83	3.70	74	5.32	106	4.08	82	4.28
2.5	1.94	78	1.47	59	2.67	107	1.84	73	1.93
control	< LOQ		< LOQ		< LOQ		< LOQ

LOQ = 0.1 mg/L

Validity criteria fulfilled:

yes

Conclusions:

This study resulted in an EC50 (48 h) of 13.7 mg/L (geom. mean measured concentration) based on mobility of D. magna (OECD 202, GLP).

Executive summary:

The study was performed according to OECD 202 (GLP) with the water flea Daphnia magna. Nominal concentrations of 2.5, 5.0, 10, 20 and 40 mg/L were tested under semi-static conditions. Test concentrations were verified by GC/MS analysis at the beginning and after each 24 h of the exposure period. The measured concentrations were between 70 - 83% after 0 h, 57-74% after 24 h (old), 92 - 107% after 24 h (fresh) and 64 - 82% of nominal after 48 h of exposure. Since the measured concentrations were not within 80 - 120% of nominal the geometric mean concentrations were calculated resulting in 1.93, 4.28, 7.22, 13.9, 30.1 mg/L. Immobilisation of D. magna was observed after 48 h resulting in an EC50 (48 h) of 13.7 mg/L based on the geometric mean of measured concentrations.

Description of key information

EC50 (48 h): 13.7 mg/L based on immobilisation of D. magna (geometric mean measured concentration, OECD 202)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

13.7 mg/L

Additional information

One experimental study is available investigating the short-term effects of the substance to aquatic invertebrates. The study was performed according to OECD 202 (GLP) with the water flea Daphnia magna. Nominal concentrations of 2.5, 5.0, 10, 20 and 40 mg/L were tested under semi-static conditions. Test concentrations were verified by GC/MS analysis at the beginning and after each 24 h of the exposure period. The measured concentrations were between 70 - 83% after 0 h, 57-74% after 24 h (old), 92 - 107% after 24 h (fresh) and 64 - 82% of nominal after 48 h of exposure. Since the measured concentrations were not within 80 - 120% of nominal the geometric mean concentrations were calculated resulting in 1.93, 4.28, 7.22, 13.9, 30.1 mg/L. Immobilisation of D. magna was observed after 48 h resulting in an EC50 (48 h) of 13.7 mg/L based on the geometric mean of measured concentrations.