p-menth-1-en-8-ol

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:

03 December 2018 – 11 December 2018

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)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 (control), 10, 100 and 180 mg/L.
- Sampling method: 3 test concentrations along with the negative control were analyzed for the test item concentration at the beginning and at the end of the test. One test tube (1*10 mL) for each concentration was used for chemical analysis.An aliquot was sampled at the middle of the flask.
- Sample storage conditions before analysis: All the solutions were kept in refrigerator.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test item was soluble in the test medium (M4 Elendt) with a mixing protocol carried out before the experimentation, according to the OECD nº23 guidance document on aquatic toxicity testing of difficult substances and mixtures.
The method consisted in stirring each test item nominal concentrations in test medium during 24 hours, in the dark and 20±2ºC prepared by adding directly the test item in flasks containing test medium. After this step, a settling period of 30 minutes was performed before sampling.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus, clone number 5, from at least the third generation (i.e. the third brood) obtained with a non-cyclic parthenogenesis reproduction.
- Source: no data
- Feeding during test: No

ACCLIMATION
- Acclimation period: not specified.
- Acclimation conditions (same as test or not): The daphnids breeding was carried out in closed bottles placed in a climatic chamber free from any toxic vapour and in medium M4. Daphnids used in this experimentation were previously filtered in order to obtain animals from a size between 560 μm and 800 μm (corresponding to daphnids less than 24 hours old).

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

250 mg/L as CaCO3

Test temperature:

19.8 - 20.0ºC

pH:

8.5 (T0) and 7.8 to 8 (T48)

Dissolved oxygen:

9.1 to 9.3 mg O2/L

Nominal and measured concentrations:

Nominal concentrations: 0 (control), 10, 18, 32.4, 58.3, 100 and 180 mg/L.
Measured concentrations for 0, 10, 100 and 180 mg/L (initial time, T0h): <1 (control), 6.94, 82.3 and 128 mg/L.
Measured concentrations for 0, 10, 100 and 180 mg/L (final time, T48h): <1 (control), 7.64, 76.5 and 135 mg/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: closed tube
- Material, size, headspace, fill volume: glass,10 mL fill volume
- Aeration: not specified.
- Renewal rate of test solution (frequency/flow rate): Not applicable (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): Not applicable
- Biomass loading rate: 2 mL of test media per daphnia.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Medium M4 ELENDT was prepared from concentrated stock solutions in ultrapure water as described in the OECD 202 guideline.
- Intervals of water quality measurement: pH and dissolved oxygen were measured at the start and at the end of the test (including Controls).

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: complete darkness

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Observations of the daphnid’s immobility and behaviour were done after 24 and 48 hours in control and exposed groups. Any sign of stress observed on the daphnids after 48 hours of exposure to the test item was reported.

VEHICLE CONTROL PERFORMED: Not applicable

RANGE-FINDING STUDY
- Test concentrations: 1, 10 and 100 mg test item/L along with negative control.
- Results used to determine the conditions for the definitive study: In the range finding test the per cent immobilisation of daphnia at 48h was 0, 0, 0, and 90% at the tested concentrations of 0, 1, 10 and 100 mg/L, respectively. Based on these results, the main study was conducted on six concentrations levels: 10.0 – 18.0 – 32.4 – 58.3 – 100.0 and 180.0 mg/L.

Reference substance (positive control):

yes

Remarks:

(potassium dichromate)

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

123.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: Confidence interval = [106.2– 146.0]

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

83.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: Confidence interval = [73.2 – 94.0]

Details on results:

- Behavioural abnormalities:
- Mortality of control: No mortality was observed over a period of 48 h in the control group.
- Abnormal responses: no signs of stress or other abnormal behaviour was reported.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not reported.

Results with reference substance (positive control):

- Results with reference substance valid? Yes, the last toxic response obtained on the reference item K2Cr2O7 (November 22, 2018) indicates that the daphnids sensitivity is correct (within the accepted interval of 0.6–2.1 mg/L fixed by the International Standard ISO 6341 December 2012 as mentioned in the OECD 202 Guideline) and in accordance with historical data obtained by the laboratory.
- EC50-24 h: 1.40 mg/L (1.27 – 1.55).

Table 1: pH and dissolved oxygen values during the test

Nominal test item concentration (mg/L)	pH
	T0	T48h
Control	8.5	8.0
10.0	8.5	7.9
18.0	8.5	7.8
32.4	8.5	7.8
58.3	8.5	7.8
100.0	8.5	7.8
180.0	8.5	7.8

Nominal test item concentration (mg/L)	O2 (mg/L)
	T0 (Air: 9.5 mg/L)	T48h (Air: 9.5 mg/L)
Control	9.3	9.2
10.0	9.2	9.2
18.0	9.3	9.1
32.4	9.2	9.2
58.3	9.2	9.2
100.0	9.2	9.1
180.0	9.3	9.3

Table 2: Results on daphnia immobilization

Nominal test item concentration ( mg/L)	Number of daphnids exposed at the start	Immobilised daphnids (%)
		at 24h	at 48h
Control	20	0 (0%)	0 (0%)
10.0	20	0 (0%)	0 (0%)
18.0	20	0 (0%)	0 (0%)
32.4	20	0 (0%)	0 (0%)
58.3	20	0 (0%)	1 (5%)
100.0	20	6 (30%)	16 (80%)
180.0	20	17 (85%)	20 (100%)

Table 3: Conclusion on the chemical analysis

	Concentration (mg test item / L)		Deviation (%)
Nominal test item concentration (mg/L)	Initial time (T0h)	Final time (T48h)
control	<1	<1	NA
10	6.94	7.64	+10.1
100	82.3	76.5	-7.0
180	128	135	+5.5

According to the results, the measured concentrations of test item in samples were maintained within ± 20 per cent of the measured initial concentration. Consequently, the test item “P-MENTH-1-EN-8-OL was considered as stable in the testing conditions after 48 hours of exposure. Therefore, and in agreement with the guideline, concentrations were expressed in nominal test item concentration.

Validity criteria fulfilled:

yes

Remarks:

(mortality in control < 10% (no immobilisation observed); dissolved oxygen concentration > 3 mg O2/L)

Conclusions:

In a short-term toxicity test to daphnia magna, the 48h-EC50 of the test substance was found to be 83.3 mg/L.

Executive summary:

An acute aquatic toxicity study with Daphnia magna was conducted on test item according to OECD guideline 202, following GLP. In a preliminary range finding study the per cent immobilisation of daphnia was 0, 0, 0 and 90% at the tested concentrations of 0, 1, 10 and 100 mg/L, respectively. Based on these results, the definitive test was conducted with nominal test item concentrations of 0 (control), 10, 18, 32.4, 58.3, 100 and 180 mg/L for a period of 48 h in static conditions in 4 replicates of 5 daphnids per replicate. Elendt M4 medium was used as test medium. No vehicle was used since the test item was found soluble in test medium at the highest concentration tested. Negative control consisted of test medium alone and a solution of potassium dichromate was used as reference substance. A validated analytical method based on GC/FID was used to monitor the concentration and stability of the active ingredient in the test solution. The test item was considered as stable in the testing conditions after 48 hours of exposure as the measured concentrations in samples were maintained within ± 20 per cent of the measured initial concentration. There was no immobilization of daphnia in the negative control and dissolved oxygen was higher than 3 mg/L in control and test vessels at the end of the test. The immobilization of daphnia was 0, 0, 0, 0, 5, 80 and 100 % at 48 hours exposure at 0 (control), 10, 18, 32.4, 58.3, 100 and 180 mg/L. Based on these results, the 48h-EC50 value was 83.3 mg/L (nominal) with 73.2 to 94.0 mg/L as confidence interval.

Description of key information

Key study. Test method according to OECD 202, GLP study. The 48h-EC50 of the test substance to Daphnia magna was found to be 83.3 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

83.3 mg/L

Additional information

Key study: An acute aquatic toxicity study with Daphnia magna was conducted on test item according to OECD guideline 202, following GLP. In a preliminary range finding study the per cent immobilisation of daphnia was 0, 0, 0 and 90% at the tested concentrations of 0, 1, 10 and 100 mg/L, respectively. Based on these results, the definitive test was conducted with nominal test item concentrations of 0 (control), 10, 18, 32.4, 58.3, 100 and 180 mg/L for a period of 48 h in static conditions in 4 replicates of 5 daphnids per replicate. Elendt M4 medium was used as test medium. No vehicle was used since the test item was found soluble in test medium at the highest concentration tested. Negative control consisted of test medium alone and a solution of potassium dichromate was used as reference substance. A validated analytical method based on GC/FID was used to monitor the concentration and stability of the active ingredient in the test solution. The test item was considered as stable in the testing conditions after 48 hours of exposure as the measured concentrations in samples were maintained within ± 20 per cent of the measured initial concentration. There was no immobilization of daphnia in the negative control and dissolved oxygen was higher than 3 mg/L in control and test vessels at the end of the test. The immobilization of daphnia was 0, 0, 0, 0, 5, 80 and 100 % at 48 hours exposure at 0 (control), 10, 18, 32.4, 58.3, 100 and 180 mg/L. Based on these results, the 48h-EC50 value was 83.3 mg/L (nominal) with 73.2 to 94.0 mg/L as confidence interval.