α,2,2,6-tetramethylcyclohexene-1-butyraldehyde

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:

January 10- March 28, 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed under GLP according to OECD guideline, validity criteria met.

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Principles of method if other than guideline:

Not relevant

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

α,2,2,6-tetramethylcyclohexene-1-butyraldehyde

EC Number:

265-747-9

EC Name:

α,2,2,6-tetramethylcyclohexene-1-butyraldehyde

Cas Number:

65405-84-7

Molecular formula:

C14H24O

IUPAC Name:

2-methyl-4-(2,6,6-trimethylcyclohex-2-en-1-yl)butanal

Details on test material:

- Name of test material (as cited in study report): Dihydro-C14-aldehyde
- Physical state: liquid
- Storage condition of test material: At room temperature (at about 20 °C), in a tightly closed container

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Not relevant

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Duplicate samples were taken from each test medium (without daphnids) including the control (without daphnids) just before the start of the test and after 48 hours. For the 48-hour stability samples, the contents of the respective replicates were combinedprior to sampling.
- Sample storage conditions before analysis: Samples were deep-frozen (at about -20 °C) immediately after sampling. Based on preexperiments for investigation of the storage stability (without GLP) the test item is sufficiently stable in the test water under the storage conditions.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: a filtrate of a supersaturated stock dispersion of the test item with a loading rate of 100 mg/L was prepared. The test item was mixed into
the test water as homogeneously as possible using ultrasonic treatment for 15 minutes and intense stirring for 3 hours at room temperature in the
dark to dissolve a maximum concentration of the test item in the dispersion. Then, the supersaturated stock dispersion of the test item was filtered
through a membrane filter (Schleicher & Schuell, Type NC45, pore size 0.45 pm) after the 3-hour stirring period just before the preparation of the
test media. The test concentrations/dilutions were: the undiluted filtrate, and dilutions 1:2, 1:4, 1:8 and 1:16. Additionally, a control (test water
without test item) was tested in parallel. Adequate volumes of the filtrate were diluted with test water for the preparation of test media with lower test
item concentrations. The test media were prepared just before the start of the test (addition of daphnids). The preparation of the test medium was
performed as far as possible in the dark to avoid photolytic degradation of the test item.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Strain: clone 5
- Source: originally supplied by the University of Sheffield/UK in 1992
- Age at study initiation (mean and range, SD): 6-24 hours
ACCLIMATION
- Acclimation conditions (same as test or not): the clone is bred in reconstituted water of the quality identical to the water quality used in the tests (in respect of pH, main ions, and total hardness) and under temperature and light conditions identical to those of the tests.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

Not applicable

Test conditions

Hardness:

Nominal Water Hardness: 2.5 mmol/L (= 250 mg/L as CaCO3)

Test temperature:

20 °C during the test period. The test was performed in a temperature-controlled room(room temperature continuously monitored).

pH:

between 7.8 and 8.0

Dissolved oxygen:

The test water was aerated prior to the start of the study until oxygen saturation was reached. During the test period the test water was not aerated.
At the beginning and at the end of the test period, the dissolved oxygen concentrations in the test media and the control were at least 8.0 mg/L

Salinity:

no data

Nominal and measured concentrations:

At the start of the test, the measured test item concentrations in the analyzed test media samples (dilutions 1:16, 1:8, 1:4, 1:2 and the undiluted filtrate) amounted to 0.27, 0.61, 1.32, 1.61 and 2.39mg/L. In these test media, the measured concentrations of Dihydro-C14-Aldehyde were 0.18, 0.40, 0.78, 0.81 and 1.0 mg/L after 48 hours.

Details on test conditions:

TEST SYSTEM
- Test vessel:100 mL glass beakers filled with 50 mL of test medium
- Aeration: The test water was aerated prior to the start of the study until oxygen saturation was reached. During the test period the test water was not aerated.
- No. of organisms per vessel:5
- No. of vessels per concentration (replicates):4
- No. of vessels per control (replicates):2
- No. of vessels per vehicle control (replicates):2

TEST MEDIUM / WATER PARAMETERS
- Alkalinity:0.8 mmol/L
- Ca/Mg ratio:4 : 1 (based on molarity)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobilisation

OTHER TEST CONDITIONS
- Photoperiod: The test was performed ain 16-light to 8-hour dark photoperiod with a 30 minute transition period (light intensity during the light period was between approx. 570 and 740 Lux)

CaCl2 x 2H20 : 2.0 mmol/L (=294 mg/L)
MgSO4 x 7H20: 0.5 mmol/L (=123 mg/L)
NaHCO3: 0.75 mmol/L (= 65mg/L)
KCI: 0.075 mmol/L (=5.8 mg/L)
Na: K = 10 : 1 (based on molarity)

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

1.41 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Duration:

24 h

Dose descriptor:

EC0

Effect conc.:

0.49 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 3

Duration:

24 h

Dose descriptor:

EC100

Effect conc.:

1.55 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 4

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.92 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 5

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

0.22 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 6

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

1.14 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: no abnormalities
- Mortality of control: no mortality of control
- Other adverse effects control: no effects
- Abnormal responses: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no, clear test media
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

Results with reference substance valid?
Yes, Results for the reference substance are determined once a year, results were within the historical range.

Reported statistics and error estimates:

The reported biological results are based on the mean measured test item concentrations. The tabulated values represent rounded results obtained
by calculation of the geometric mean using the exact raw data.

Any other information on results incl. tables

No remarks

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The 48 hour EC50 is 0.92 mg/l and the 48 h NOEC (observed) is 0.22 mg/l.

Executive summary:

The acute toxicity of the test item Dihydro-C14-Aldehyde to Daphnia magna was determined

in a 48-hour static test according to the EU Commission Directive 92/69/EEC, Part C.2

(1992) and the OECD Guideline for Testing of Chemicals, No. 202 (2004).

Due to the low water solubility of the test item, a supersaturated dispersion of the test item

with a loading rate of 100 mg/L was continuously stirred at room temperature in the dark

over 3 hours. After the stirring period an oily film was present on the surface. Therefore, only

a part of the dispersion of the test item was taken out of the middle of the water column and

filtered through a membrane filter just before the preparation of the test media. This procedure was necessary to avoid a

penetration of the oily film through the filter into the test medium. The dispersion was filtered

and the undiluted filtrate with the maximum concentration of dissolved test item was used as

the highest test medium concentration and as stock solution for dilutions in a geometric

series differing by a constant factor of 2. Additionally, a control was tested in parallel.

The measured test item concentration in the analyzed test media samples (dilutions 1:16,

1.8, 1.4, 1:2 and the undiluted filtrate) amounted to 0.27, 0.61, 1.32, 1.61 and 2.39 mg/L at

the start of the test. In these test media, incubated under the conditions of the test during the

test period (but without daphnia), the measured concentrations of Dihydro-C14-Aldehyde

decreased to values of 0.18, 0.40, 0.78, 0.81 and 1.0 mg/L after 48 hours. This decrease of

the test item concentrations was considered to be caused by a loss by adsorption onto the

glass surfaces or by a degradation process.

The biological results were based on mean measured test item concentrations (calculated as

the geometric means of the concentrations measured at the start and the end of the test)

which were 0.22 (dilution 1:16), 0.49 (dilution 1:8), 1.02 (dilution 1:4), 1.14 (dilution 1:2) and

1.55 mg/L (undiluted filtrate).

The biological test results were as follows:

-24 hour EC0:                0.49 mg/l -24 hour EC100:            1.55 mg/l -48 hour EC50:               0.92 mg/l 95% confidence limits:   could not be determined    -48 hour EC0 and 48 hour NOEC (observed):       0.22 mg/l -48 hour EC100:                1.14mg/l