[3R-(3α,3aβ,6α,7β,8aα)]-octahydro-6-methoxy-3,6,8,8-tetramethyl-1H-3a,7-methanoazulene

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:

10 October - 20 November 2015

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)

Version / remarks:

(2008)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Test concentrations were verified by chemical analysis. Water samples (3 mL, in duplicate) were taken from the approximate centre of the test vessels of the control and each exposure level at the start and end of both 24-h (renewal) periods. Samples were stored in the freezer until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
Preparation of test solutions started with a loading rate of 100 mg/L applying a two-day period of magnetic stirring followed by an one-hour settlement period. The aqueous Water Soluble Fraction (WSF) was siphoned off and used as the highest test concentration or as a stock solution. The lower test concentrations were prepared by subsequent dilutions of the WSF in test medium. The final test solutions were all clear and colourless

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna (Straus 1820)
- Strain: not stated
- Source: in-house laboratory culture with a known history (at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions)
- Age at study initiation: young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old
- Method of breeding: start of each batch with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium in an all-glass culture vessel. Renewal of the cultures: after 7 days of cultivation half of the medium twice a week.
- Feeding during test: no

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

180 mg/L (as CaCO3)

Test temperature:

20°C

pH:

7.9 - 8.4

Dissolved oxygen:

8.8 - 9.4 mg O2/L

Nominal and measured concentrations:

- Nominal concentrations: 0 (control), 1.0, 2.2, 4.6, 10, 22, 46% 100% of the WSF prepared at 100 mg/L (based on range-finding test).
- Measured concentrations (geom. mean): n.d., 0.030, 0.061, 0.15, 0.33, 0.69 and 1.3 mg/L

Measured concentrations range from 73 - 92 % of inital at the end of the first renewal period (t=0-24 hours) and 70-116 % of initial at the end of the second renewal period (t=24-48 hours).

Details on test conditions:

TEST SYSTEM
- Tests were performed in closed airtight all-glass vessels (50 mL) completely filled up with test solutions.
- Six test concentrations (1.0, 2.2, 4.6, 10, 22 and 46% of WSF prepared at 100 mg/L) plus one control each in 4-fold with 5 animals/test vessel (i.e. 20 animals/concentration).
- Loading rate: 10 mL test solution per organism
- Test vessels were maintained at 20 °C under a photoperiod of 16 hours light : 8 hours dark and without supplementary aeration or feeding during the 48 hour exposure period.
- The temperature was measured continuously and the pH and dissolved oxygen levels were recorded at the start and at the end of the study.

TEST MEDIUM / WATER PARAMETERS
- Test medium: adjusted ISO
- Culture medium different from test medium: yes (M7)

OTHER TEST CONDITIONS
- Adjustment of pH: no

EFFECT PARAMETERS MEASURED:
Immobility at 0, 24 and 48h. Daphnia were considered to be immobilised if they were unable to swim for approximately 15 seconds after gentle agitation.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.48 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL 0.40 - 0.57 mg/L

Details on results:

No immobility was observed up to a concentration of 0.15 mg/l. At higher concentrations the effect increased with increased concentration ranging from 15% at 0.33 mg/l to complete inhibition at 1.3 mg/L.

Results with reference substance (positive control):

- 48-h EC50: 0.39 mg/L (September 2015)
- Results with reference substance valid? yes

Reported statistics and error estimates:

The 24-h and 48-h EC50 values were calculated from the weibits of the percentages of affected daphnids and the logarithms of the corresponding test substance concentrations (average exposure) using the maximum likelihood estimation method.

Table: Immobility (percentage of 20 animals introduced at t=0h)

Average measured exposure concentration (mg/L)	Percentage immobility
	t=0h	t=24h	t=48h
0 (control)	0	0	0
0.030	0	0	0
0.061	0	0	0
0.15	0	0	0
0.33	0	0	15
0.69	0	15	85
1.3	0	55	100

Table: Percentage of WSF prepared at 100 mg/L,initial and mean initial exposure concentrations  

Nominal Concentration (mg/L)*	Measured concentration (mg/L)
	t=0h (fresh)	T=24h (old)	t=24h (fresh)	T=48h (old)	Average
2.2	0.032	0.027	0.035	0.028	0.030
4.6	0.0802	0.064	0.047	0.054	0.061
10	0.172	0.127	0.152	0.146	0.15
22	0.347	0.321	0.335	0.305	0.33
46	0.777	0.568	0.769	0.661	0.69
100	1.48	1.21	1.54	1.07	1.3

* As % of the WSF prepared at 100 mg/L

Validity criteria fulfilled:

yes

Remarks:

See 'Any other information on materials and methods incl. tables'.

Conclusions:

The 48 h EC50 (immobilisation) value for Daphnia magna is 0.48 mg/L.

Executive summary:

A study was performed to assess the acute toxicity of the substance to Daphnia magna under semi-static conditions (renewal period: 24 hours). The study was conducted in accordance with OECD Guideline for Testing of Chemicals No. 202.

Groups of twenty daphnids (less than 24 hours old) were exposed for 48 hours to six concentrations of the substance dispersed in test water (1.0 - 46 % of a WSF prepared at a loading rate of 100 mg/L) and a control. The incidence of immobilisation was recorded for each test and control group at 24 hours and at 48 hours and the following endpoint was determined: 48-h EC50 = 0.48 mg/L. All results are expressed in terms of geometric mean measured concentrations. Measured concentrations ranged from 73 - 92 % of inital at the end of the first renewal period (t=0-24 hours) and 70-116 % of initial at the end of the second renewal period (t=24-48 hours).

Description of key information

A study was performed to assess the acute toxicity of the substance to Daphnia magna under semi-static conditions (renewal period: 24 hours). The study was conducted in accordance with OECD Guideline for Testing of Chemicals No. 202. Groups of twenty daphnids (less than 24 hours old) were exposed for 48 hours to six concentrations of the substance dispersed in test water (1.0 - 46 % of a WSF prepared at a loading rate of 100 mg/L) and a control. The incidence of immobilisation was recorded for each test and control group at 24 hours and at 48 hours and the following endpoint was determined: 48-h EC50 = 0.48 mg/L. All results are expressed in terms of geometric mean measured concentrations. Measured concentrations ranged from 73 - 92 % of inital at the end of the first renewal period (t=0-24 hours) and 70-116 % of initial at the end of the second renewal period (t=24-48 hours).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

0.48 mg/L