Cyclododecane

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-07-26 to 2010-07-30

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Cyclododecane

EC Number:

206-033-9

EC Name:

Cyclododecane

Cas Number:

294-62-2

Molecular formula:

C12H24

IUPAC Name:

cyclododecane

Details on test material:

- Name of test material (as cited in study report): Cyclododecan
- Molecular formula: C12H24
- Molecular weight: 168.32 kg/kmol
- Physical state: Solidified melt, colourless
- Analytical purity: 99.3 - 99.8 %
- Purity test date: 2010-05-20
- Lot/batch No.: 3347/27173
- Expiration date of the lot/batch: 2011-05-20
- Storage condition of test material: Room temperature, protected from moisture and light

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Samples of the saturated test item solution and the control from the fresh media (0 and 24 h) and old media (24 and 48 h) were analytically verified by GC-MS.
- Sampling method: For the fresh media, samples of the freshly prepared test solutions were taken and analysed. Samples of the old media were taken directly from the test vessels.
The effect levels are given based on the geometric mean measured concentrations.
- Sample storage conditions before analysis:All samples were stored at room temperature until start of analysis.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Following the recommendations of OECD guidance document no 23 (2000), a column elution system as generator system was applied. Two generator columns were prepared once for the test prior to application, one for the preparation of the saturated solution used at test start (0 h) and one for the preparation of the saturated solution used at the medium renewal after 24 h. For the preparation of a generator column 10 g glass beads (size 0.75 – 1.00 mm) were spiked with the test item. The test item (100 mg was weighed out) was dissolved in 5 mL acetone and mixed thoroughly with the glass beads in a glass beaker. After complete evaporation of the solvent, the dry glass beads were filled in the generator column.
For preparation of the saturated test solution the generator column was rinsed with 1 L of dilution water in a closed circuit within 24 h by a tubing pump with a flow rate of 2 mL/min.
- Eluate: Dilution water
- Differential loading: A limit test with the saturated solution (maximum solubility level of the test item in the test medium) was carried out.
- Controls: Four replicates of dilution water without test item tested under the same conditions as the test groups.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna (STRAUS)
- Strain: Clone 5
- Source: Institut für Wasser-, Boden- Lufthygiene (WaBoLu), further own breeding
- Age at study initiation (mean and range, SD): 2 to 24 h
- Method of breeding: In 2-3 L glass vessels with appr. 1.8 L culture medium, at 20 +/- 2 °C, in an incubator, 16 h illumination, illumination strength max. 20 µExm-2 x s-1
- Feeding during test: No feeding

ACCLIMATION
- Acclimation period: 2 h
- Acclimation conditions (same as test or not): Same as test
- Type and amount of food: No feeding
- Health during acclimation (any mortality observed): Healthy, no mortality observed

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

No

Test conditions

Hardness:

Dilution water: 231 anf 244 (mg CaCO3/L)

Test temperature:

19.6 and 19.4 °C

pH:

Water Quality Parameters in the Old Media (24 h)
(measured in all replicates)
pH-Values
Replicates
1 2 3 4
Saturated test item solution 7.56 7.57 7.56 7.56
Control 7.60 7.71 7.65 7.59


: Water Quality Parameters in the Old Media (48 h)
(measured in all replicates)
pH-Values
Replicates
1 2 3 4
Saturated test item solution 7.60 7.58 7.57 7.57
Control 7.42 7.75 7.70 7.64

Dissolved oxygen:

Water Quality Parameters in the Old Media (24 h)
(measured in all replicates)
Dissolved O2 -Concentration [mg/L]
Replicates
1 2 3 4
Saturated test item solution 8.00 7.94 8.08 8.00
Control 8.65 8.69 8.70 8.73

Water Quality Parameters in the Old Media (48 h)
(measured in all replicates)
Dissolved O2 -Concentration [mg/L]
Replicates
1 2 3 4
Saturated test item solution 8.35 8.25 8.26 8.14
Control 8.77 8.85 8.82 8.81

Salinity:

Freshwater

Nominal and measured concentrations:

Please refer to "Any other information on materials and methods"

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers, 50 mL
- Type (delete if not applicable): closed with parafilm
- Material, size, headspace, fill volume: Glass beakers (4 cm ID x 7 cm H), 50 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): One application at test start, static test
- 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


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Acc. to Directive 92/69/EEC Method C.2, Annex 1
- Conductivity: Dilution water at test start: 640 µS/cm
- Culture medium different from test medium: Elendt M4 acc. to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
- Intervals of water quality measurement: At test start and test end


OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16:8 light:dark cycle
- Light intensity: Diffuse light, illumination strength max. 20 µE x m-2 x s-1


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :


TEST CONCENTRATIONS
- Spacing factor for test concentrations: None, limit test
- Range finding study
Immobilization Rates of the non GLP Preliminary Range Finding Test
(n = 20, divided into 2 replicates with 10 daphnids each)
Dilution Level
of the Saturated
Test Item Solution IMMOBILIZATION [%]
24 h 48 h
Replicates Replicates
1 2 MV 1 2 MV
Saturated solution 0 0 0 0 0 0
1:10 0 0 0 0 0 0
Control 0 0 0 0 0 0

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

11.6 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 11.6 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: None
- Mortality of control: 0%
- Other adverse effects control: None
- Abnormal responses: None
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: None

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50/LC50: 1.82 (CI 1.70 - 1.95) mg/L

Reported statistics and error estimates:

The EC50-value was calculated for the reference item by sigmoidal dose-response regression. The confidence interval of the EC50 was calculated from the best-fit values, the standard error and the t-distribution with the software GraphPad prism.

Any other information on results incl. tables

Immobilization Rates in the Definitive Test after 24 and 48 h of Exposure

                     (n = 20, divided into 4 replicates with 5 daphnids each)

Cyclododecane	IMMOBILIZATION [%]
Geometric Mean Measured Concentration [µg/L]	24 h					48 h
	Replicates					Replicates
	1.	2.	3.	4.	MV	1.	2.	3.	4.	MV
11.6 (Saturated test item solution)	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

At the saturated solution of the test substance Cyclododecane, which corresponded to the geometric mean measured concentration of 11.6 µg/L, no effects were observed on Daphnia magna. The results indicate that at the saturated solution of the test substance no significant hazard to aquatic invertebrates is expected.

Executive summary:

In the acute immobilization test to Daphnia magna (STRAUS) the effects of the saturated solution* of the test item Cyclododecane (batch number: 3347/27173) as a limit concentration were determined according to OECD 202 (2004). The limit test was conducted under semi-static conditionsfrom 2010-07-26 to 2010-07-30, with the definitive exposure phase of 48 h from 2010-07-28 to 2010-07-30 at Dr.U.Noack-Laboratorien-
Twenty test organisms were each exposed to the saturated solution* and the control.
The concentrations of Cyclododecane were analysed in both the saturated test item solution* and the control by GC-MS in samples of the fresh media (0 and 24 hours) and the old media (at 24 and 48 hours).
The recoveries of Cyclododecane in the saturated test item solution were determined to be 66 and 72 % of the initially measured values at 24 and 48 hours, respectively. The overall geometric mean measured concentration of Cyclododecane was calculated to be 11.6 µg/L. No immobilization was observed at the maximum concentration of the test item in this system (Table 1).

The overall geometric mean measured concentration was used to assess the effect levels. The validity criteria of the test guideline were fulfilled.

EC₅₀-Values, NOEC and LOEC

                      (based on the geometric mean measured concentration of Cyclododecane)

	Test Duration [h]	Geometric Mean Measured Concentration of Cyclododecane [µg/L]	Confidence Interval p = 95% [µg/L]
EC50	24	> 11.6	not applicable
	48	> 11.6	not applicable
NOEC	48	11.6	---
LOEC	48	> 11.6	---

At the saturated solution* of the test item Cyclododecane, which corresponded to the geometric mean measured concentration of 11.6 µg/L,
no effects were observed on Daphnia magna.