(5E)-3,5-dimethyloct-5-en-4-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:

29-05-2018 to 01-06-2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study performed under GLP. All relevant validity criteria were met.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

An Acute Immobilisation Test to Daphnia magna STRAUS was carried out to determine the EC50-values of the test item after 24 and 48 hours of exposure under semi-static conditions in a closed system and in darkness.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

inspected: June 2016 ; signature: January 2017

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All relevant concentration levels and the control were analytically verified via GC-MS at the start and at the renewal of test solutions (0 and 24 hours) as well as 24-hour old media at the renewal and at the end of the exposure intervals (24 and 48 hours).
- Sampling method: At the start of the exposure and at renewal intervals (0 and 24 hours), sampling was carried out after preparation of the test concentrations. At the renewal and at the end of the exposure intervals (24 and 48 hours), samples were taken directly from the test vessels. The method was validated prior to this study according to SANCO 3029/99 rev.4 (2000).
- Sample storage conditions before analysis: All original samples were stored at room temperature, until analysis if necessary. Prepared samples were stored in an autosampler at room temperature until analysis.

Vehicle:

no

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphinds
- Strain: Daphnia magna STRAUS (Clone 5)
- Justification for species other than prescribed by test guideline: Not applicable.
- Age at study initiation (mean and range, SD): < 24 hours old daphnids from a healthy stock were used for the study
- Weight at study initiation (mean and range, SD): Not applicable.
- Length at study initiation (length definition, mean, range and SD): Not applicable.
- Stage and instar at study initiation: Juvenile ; < 24 hours
- Valve height at study initiation, for shell deposition study (mean and range, SD): Not applicable.
- Peripheral shell growth removed prior to test initiation: Not applicable.
- Method of breeding: Not reported.
- Source: in-house laboratory cultures
- Age of parental stock (mean and range, SD): Not applicable.
- Feeding during test: No. The daphnids were not fed during the study. During culture: The culture daphnids are fed at least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus, with an algae cell density of > 10^6 cells/mL. The algae are cultured at the test facility
- Food type: Not applicable.
- Amount: Not applicable.
- Frequency: Not applicable.

ACCLIMATION
- Acclimation period: At least 2 hours in dilution water.
- Acclimation conditions (same as test or not): Yes.
- Type and amount of food: Not applicable.
- Feeding frequency: Not applicable.
- Health during acclimation (any mortality observed): No.

QUARANTINE (wild caught)
- Duration: Not applicable.
- Health/mortality: Not applicable.

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES, INCLUDING CULTURING CONDITIONS: Not applicable.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

Dilution water quality parameters: 0 hours: Total Hardness: 293 mg CaCO3/L ; 24 hours Total Hardness: 245 mg CaCO3/L

Test temperature:

Dilution water quality parameters: 0 hours: Temperature 19.4 °C ; 24 hours: Temperature 19.2 °C

pH:

0 hours: pH 8.40 ; 24 hours: pH 8.08 ; mean pH at 48 hours all replicates: did not differ by more than 1.5 units

Dissolved oxygen:

Dilution water quality parameters: 0 hours: O2 dissolved: 9.04 mg/L ; 24 hours: O2 dissolved: 8.84 mg/L; dissolved oxygen at 24 and 48 hours all replicates: did not decrease to less than 3 mg/L.

Conductivity:

Dilution water quality parameters: 0 hours: Conductivity : 608 µS/cm ; 24 hours: Conductivity : 611 µS/cm

Nominal and measured concentrations:

Range finding tests were performed (non-GLP) prior to the definitive test: 0 (control), 0.01, 1.0, 10.0, 100 mg/L nominal concentration.
Definitive test: dilution level of saturated solutions: 0 (control), 6.25, 12.5, 25.0, 50.0, 100 mg/L analytically confirmed nominal concentration.

Details on test conditions:

TEST SYSTEM
- Test vessel: 4.5 (ID) x 9.5 (H) cm, ca. 130 mL
- Type: A semi-static test design with a daily renewal of the test solutions in glass flasks sealed with screw caps.
- Material, size, headspace, fill volume: Sealed glass flasks with screw caps. Fill volume ca. 130 mL.
- Aeration: No aeration of the test solutions.
- No. of organisms per vessel: Control and test item: 20 per concentration, 5 per vessel (divided into 4 replicates).
- No. of vessels per concentration (replicates): 4 (four replicates, 5 daphnia per vessel).
- No. of vessels per control (replicates): 4 (four replicates, 5 daphnia per vessel).
- No. of vessels per vehicle control (replicates): Not applicable.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4, according to OECD 202, Annex 3
- Culture medium different from test medium: No. Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L

The test vessels were filled up with the test solutions. The daphnids were inserted with a small amount of dilution water (start of the exposure) or test solution (water renewal) by pipette. Thereafter, the test vessels were closed immediately with screw caps. The test solutions were renewed after 24 hours. For this purpose, a second set of test vessels were filled with the freshly prepared test solutions and the daphnids were transferred by pipette.

OTHER TEST CONDITIONS
- Adjustment of pH: No. The test vessels were not pH adjusted during or after test item exposure.
- Photoperiod: 16/8 h light/dark cycle during culture conditions. During testing: Light exclusion.
- Light intensity: Diffuse light, light intensity of max. 1500 lx during testing.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Immobility (including mortality), 24 hours and at 48 hours. An organism was considered to be immobile, if it was not able to swim in the water phase within 15 seconds after gentle agitation of the test vessel. Other observations were not made.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.0
- Justification for using less concentrations than requested by guideline: Not applicable.
- Range finding study
- Test concentrations: Range finding tests were performed (non-GLP) prior to the definitive test: 0 (control), 0.01, 1.0, 10.0, 100 mg/L nominal concentration.
Definitive test: dilution level of saturated solutions: 0 (control), 6.25, 12.5, 25.0, 50.0, 100 mg/L analytically confirmed nominal concentration. See table 1.
- Results used to determine the conditions for the definitive study: Yes.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

24 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL: 20.0 - 26.2 mg/L ; analytically confirmed nominal concentrations

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

18.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL: 16.3 - 20.5mg/L ; analytically confirmed nominal concentrations

Details on results:

- Behavioural abnormalities: None reported.
- Observations on body length and weight: Not applicable.
- Other biological observations: 100% mortality at 50 mg/L and 100 mg/L nominal after 24 hours.
- Mortality of control: No mortalities in control.
- Other adverse effects control: None reported.
- Abnormal responses: None reported.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: None. Prior to start of the exposure intervals, the test solutions were checked for undissolved test item via laser beam (Tyndall effect). Presence of undissolved test item during preparation and during the test was not observed.
- Effect concentrations exceeding solubility of substance in test medium: No.

Results with reference substance (positive control):

- Results with reference substance valid?: Yes.
- Mortality: None. Acute immobilisation observed only.
- EC50/LC50: 24h-EC50 was 2.03 mg/L (C.I. 1.00 – 4.00 mg/L).
- Other: The EC50-value of the reference item potassium dichromate after 24 hours is within the prescribed concentration range of 0.6 - 2.4 mg/L of quality criteria according to AQS P 9/2 (02/2000) listed in DIN 38412 - L 30 for daphnids clone 5 cultured in Elendt M4 medium. The EC50-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD Guideline TG 202

Reported statistics and error estimates:

The EC50-value after 24 and 48 hours of exposure were calculated by sigmoidal dose-response regression. The respective 95% confidence limits were calculated from the standard error and the t-distribution.

Table 1. Immobilisation Rates in the non GLP Preliminary Range Finding Test

(n = 20, divided into 2 replicates with 10 daphnids each)

Nominal test item concentration [mg/L]	Immobilisation [%]
	24 hours			48 hours
	Replicates			Replicates
	1	2	MV	1	2	MV
100	100	100	100	100% mortality
10	0	0	0	0	0	0
1	0	0	0	0	0	0
0.01	0	0	0	0	0	0
Control	0	0	0	0	0	0

 

Table 2. Immobilisation Rates after 24 and 48 h of Exposure in the Definitive Test

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

Nominal test item concentration [mg/L]		Immobilisation [%]
		24 hours										48 hours
		Replicates										Replicates
		1		2		3		4		MV		1		2		3		4		MV
100		100		100		100		100		100		100% mortality
50		100		100		100		100		100
25		40		60		40		80		55		100		100		80		80		90
12.5		0		0		0		20		5		0		0		0		20		5
6.25		0		0		0		0		0		0		0		0		0		0
Control	0		0		0		0		0		0		0		0		0		0

 

Table 3. Measured Concentrations and Percent of the Nominal and the Initially Measured Concentrations of the Test Item during the Definitive Test

Sampling	0 hours Start of the exposure interval		24 hours End of the exposure interval		24 hours Start of the exposure interval		48 hours End of the exposure interval
Nominal test item concentration [mg/L]	Test item concentration
	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%
100	96.7	97	101	101	ND	ND	ND	ND
50	51.2	102	47.8	96	ND	ND	ND	ND
25	26.5	106	23.6	94	22.1	88	22.4	89
12.5	13.2	105	12.6	101	12.0	96		92
6.25	7.00	112	6.84	109	7.47	119		111
Control	< LOQ		< LOQ		< LOQ		< LOQ

Meas. conc. = measured concentration of the test item, dilution factors taken into account

% = percent of the initial concentration of the test item

LOQ = limit of quantification of the analytical method (0.1 mg/L of the test item)

ND = not determined due to 100% mortality after 24 hours

Validity criteria fulfilled:

yes

Conclusions:

The test item 48h-EC50 was 18.6 mg/L (C.I: 16.3 – 20.5 mg/L) based on analytically confirmed nominal concentrations.

Executive summary:

The acute toxicity to Daphnia magna was carried out according to OECD TG 202 Daphnia sp., Acute Immobilisation Test and EU Method C.2 guidelines under GLP. The study was conducted in a closed system (sealed glass flasks) without headspace under semi-static conditions over a period of 48 hours with five concentration levels of the test item in the 6.25 to 100 mg/L nominal concentration range: prepared in a geometric series with a separation factor of 2. To reduce losses of the test item, the study was conducted under semi-static conditions in a closed system. A stock solution of 100 mg/L test item was prepared with Elendt M4 test water (according to OECD 202, Annex 3), prior to the start of the exposure (at 0 hours) and on the renewal of the test solutions (at 24 hours). The stock solution was mixed thoroughly by manual agitation until the solution was visually clear. Additionally, the stock solution was checked via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion). The Tyndall effect was negative. All test concentrations were visually clear throughout the exposure intervals. Twenty daphnids were exposed to each concentration level and the control. The test solutions were renewed after 24 hours. For this purpose, a second set of test vessels was filled with the freshly prepared test solutions and the daphnids were transferred by pipette. The test item was analytically verified via GC-MS in the fresh media at the start of the exposure and at the renewal of the test solutions (0 and 24 hours) and in the old media at the renewal and at the end of the test (24 and 48 hours) in all tested concentration levels and the control. The concentrations in the fresh media varied between 88 and 119% of the nominal concentrations. The measured concentrations in old media at renewal and at the end of the test (24 and 48 hours) were in the range of 89 to 111% of the nominal concentrations. The concentrations of the test item were analytically verified via GC-MS at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours) in all relevant concentration levels and the control. Since the measured test item concentrations remained within ± 20% of the nominal concentrations, the EC50-values are based upon the nominal concentrations that were analytically confirmed. The water quality parameters (i.e. pH-value and dissolved oxygen concentration), measured at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours), were within the acceptable limits. The validity criteria of the test guideline were fulfilled. The 48h-EC50 was 18.6 mg/L (C.I: 16.3 – 20.5 mg/L) based on analytically confirmed nominal concentrations of the test item.

Description of key information

EC50 (invertebrates) = 18.6 (C.I. 16.3 - 20.5) mg/L based on analytically confirmed nominal concentrations, 48-hour, freshwater, OECD TG 202, 2018

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

18.6 mg/L

Additional information

Key study : OECD TG 202, 2018 : The acute toxicity to Daphnia magna was carried out according to OECD TG 202 Daphnia sp., Acute Immobilisation Test and EU Method C.2 guidelines under GLP. The study was conducted in a closed system (sealed glass flasks) without headspace under semi-static conditions over a period of 48 hours with five concentration levels of the test item in the 6.25 to 100 mg/L nominal concentration range: prepared in a geometric series with a separation factor of 2. To reduce losses of the test item, the study was conducted under semi-static conditions in a closed system. A stock solution of 100 mg/L test item was prepared with Elendt M4 test water (according to OECD 202, Annex 3), prior to the start of the exposure (at 0 hours) and on the renewal of the test solutions (at 24 hours). The stock solution was mixed thoroughly by manual agitation until the solution was visually clear. Additionally, the stock solution was checked via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion). The Tyndall effect was negative. All test concentrations were visually clear throughout the exposure intervals. Twenty daphnids were exposed to each concentration level and the control. The test solutions were renewed after 24 hours. For this purpose, a second set of test vessels was filled with the freshly prepared test solutions and the daphnids were transferred by pipette. The test item was analytically verified via GC-MS in the fresh media at the start of the exposure and at the renewal of the test solutions (0 and 24 hours) and in the old media at the renewal and at the end of the test (24 and 48 hours) in all tested concentration levels and the control. The concentrations in the fresh media varied between 88 and 119% of the nominal concentrations. The measured concentrations in old media at renewal and at the end of the test (24 and 48 hours) were in the range of 89 to 111% of the nominal concentrations. The concentrations of the test item were analytically verified via GC-MS at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours) in all relevant concentration levels and the control. Since the measured test item concentrations remained within ± 20% of the nominal concentrations, the EC50-values are based upon the nominal concentrations that were analytically confirmed. The water quality parameters (i.e. pH-value and dissolved oxygen concentration), measured at the start (0 and 24 hours) and at the end of the exposure intervals (24 and 48 hours), were within the acceptable limits. The validity criteria of the test guideline were fulfilled. The 48h-EC50 was 18.6 mg/L (C.I: 16.3 – 20.5 mg/L) based on analytically confirmed nominal concentrations of the test item.