Methyl 4(or 1)-isopropyl-1(or 4)-methylbicyclo[2.2.2]oct-5-ene-2-carboxylate

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:

5th February 2003 - 6th February 2003

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:

1999 (Draft)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All test concentrations and control
- Sampling method: Samples taken at Day 0 and Day 28 for DOC analysis

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 50 mg/L stock solution stirred for 24 h on magnetic stirrer (600 rpm) after which undissolved particles of the test item were removed by centrifugation (20 minutes, 3000rpm) to ensure a saturated solution
- Differential loading: Test dilutions of the saturated solution were used at 1:1, 1:2, 1:4, 1:8 and 1:16
- Controls: Dilution water
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): None used
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): Undissolved particles were removed by centrifugation. The test item was clearly dissolved in all tested dilutions.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Giant water flea
- Strain: STRAUS (Clone 5)
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu)
- Age at study initiation: 2 to 24 h
- Feeding during test: None

ACCLIMATION
- Acclimation period: 2 h
- Acclimation conditions: same as test
- Type and amount of food: ad libitum Desmodesmus subspicatus and Chlorella vulgaris
- Feeding frequency: 5 x weekly

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

261 mg CaCO3/L for dilution water

Test temperature:

19 ± 1°C

pH:

7.66 - 7.81

Dissolved oxygen:

8.66 - 10.16 mg/L

Nominal and measured concentrations:

Nominal concentrations: 1:1, 1:2, 1:4, 1:8, 1:16 dilutions of the saturated solution
Measured concentrations; 11.6, 5.92, 3.12, 1.86, 1.31 mg/L (based on carbon content of 0.73 mg C/mg test item)

Details on test conditions:

TEST SYSTEM
- Test vessel: Beaker
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: Glass, 5 cm ID x 8 cm H, 50 mL
- 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): None included

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dilution water pH 7.8, hardness 261 mg CaCO3/L
- Conductivity: 646 µS/cm
- Culture medium different from test medium: Culture medium Elendt M4, dilution water not stated
- Intervals of water quality measurement: Dissolved oxygen and pH measured at the start of the test in one replicate and at the end in all replicates

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: Diffuse light, illumination range max. 20 µ mol.m-2 . s-1


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobilisation measured after 24 and 48 hours

VEHICLE CONTROL PERFORMED: Not required

RANGE-FINDING STUDY
- Test concentrations: Dilution levels 1:1 to 1:100, factor of 10
- Results used to determine the conditions for the definitive study: Yes, based on rangefinder results dilutions of 1:1 to 1:16 used in the main study

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

5.69 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

7.98 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL lower: 7.26 95% CL higher: 8.76

Results with reference substance (positive control):

24 h EC10: 0.83 mg/L
24 h EC50: 2.17 mg/L (CL 1.66-2.83)
24 h EC100: 5.80 mg/L

DOC analysis of new (0 h) and Old Media (48 h)

Dilution of the saturated solution*	DOC (mg/L)						Real Test Item Concentration (mg/L)
	Parallel Determination    (0 h) (New Media)		Parallel Determination    (48 h) (Old Media)		Overall Mean	Corrected vs. Control
1:1	14.23	14.82	6.35	6.86	10.57	8.49	11.6
1:2	8.82	9.03	3.92	3.89	6.42	4.34	5.92
1:4	6.05	6.24	2.37	2.80	4.37	2.29	3.12
1:8	4.82	4.89	2.15	1.91	3.44	1.36	1.86
1:16	4.22	4.31	2.14	1.50	3.04	0.96	1.31
Control	3.15	3.06	1.03	1.08	2.08	-	-

* Saturated solution: the maximum dissolved concentration of the test item that can be achieved under the test conditions in the test medium, according to OECD Series No. 23.

1) The real test concentrations were calculated based on the carbon content of 0.73 mg C/mg test item

Percentage of Daphnids Incapable of Swimming after 24 and 48 hours of Exposure (n = 20)

Dilution of the saturated solution*	Test item concentration	Immobilisation (%)
		24 h					48 h
	mg/L	Replicates					Replicates
		1	2	3	4	MV	1	2	3	4	MV
1:1	11.6	40	60	80	80	65	60	100	100	100	90
1:2	5.92	0	0	0	20	5	20	20	20	20	20
1:4	3.12	0	0	0	0	0	0	0	0	0	0
1:8	1.86	0	0	0	0	0	0	0	0	0	0
1:16	1.31	0	0	0	0	0	0	0	0	0	0
Control		0	0	0	0	0	0	0	0	0	0

MV = Mean value

* Saturated solution: The maximum dissolved concentration of the test item that can be achieved under the test conditions in the test medium, according to OECD Series No. 23

EC₁₀ -, EC₅₀ - (with confidence interval), EC₁₀₀ - Values based on mean measured concentrations

	Test duration (h)	Concentration (mg/L)	Confidence interval (p = 95 %)
EC10	24	6.75	--
	48	5.69	--
EC50	24	10.2	9.08 - 11.4
	48	7.98	7.26 - 8.76
EC100	24	>11.6	--
	48	> 11.6	--

Validity criteria fulfilled:

yes

Remarks:

No immobilisation in control group; reference item EC50 >1 <2.5 mg/L; dissolved oxygen >5.21 mg/L; pH did not vary by >1 unit

Conclusions:

The 48 h EC50 value for Daphnia Magna immobilisation is 7.98 mg/L.

Executive summary:

This study is conducted in accordance with OECD guideline 202 and EU Method C.2 and is GLP compliant. Dilutions of a saturated solution were tested and measured concentrations analysed based on DOC. A dilution water control was included and all validity criteria were met. Therefore, the study is considered to be reliable without restrictions (Klimisch 1). The 48 h EC50 value for Daphnia Magna immobilisation is 7.98 mg/L.

Description of key information

The 48 h EC50 value for Daphnia Magna immobilisation is 7.98 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

7.98 mg/L

Additional information

This study is conducted in accordance with OECD guideline 202 and EU Method C.2 and is GLP compliant (2003). Dilutions of a saturated solution were tested and measured concentrations analysed based on DOC. A dilution water control was included and all validity criteria were met. Therefore, the study is considered to be reliable without restrictions (Klimisch 1). The 48 h EC50 value for Daphnia Magna immobilisation is 7.98 mg/L.