2-hexyldecanoic acid

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:

2019-10-09 to 2019-10-17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP compliant

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted 13. April 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

GLP Certificate: Statement of GLP Compliance from Staatliches Gewerbeaufsichtsamt Hildesheim, Germany, dated 03.01.2017

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all concentration levels and the control were analytically verified
- Sampling method: samples were taken in the fresh media at the start of the exposure and at the renewal of the test solutions (0 and 24 hours) after preparation of the test item concentrations as well as in 24-hours old media at the renewal and at the end of the exposure (24 and 48 hours), samples of the 24-hours old media were taken directly from the test vessels
- Sample storage conditions before analysis: at room temperature until the start of the analysis, if necessary

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Preparation of saturated solution with a nominal concentration of 1.00 mg/L 4 days prior to start of exposure as well as 4 days prior to the renewal of the test solutions in a glass flask filled with the appropraite amount of dilution water (ISO Test Water acc. to guideline). The test item (density of 0.8779 g/cm3 taken into account) placed by pipette onto the water surface, bottle closed with a screw cap, then slow stirring procedure was applied (gentle stirring to avoid formation of emulsion carried out for 72 ± 1 hour with a magnetic stirrer at 30 ± 1 °C). After completion of stirring, dispersion was allowed to stand for at least 24 hours at 20 ± 1 °C for separation of undissolved test item. Thereafter, clear water phase removed by siphoning from the bottom of the glass flask and centrifuged at 17000 g for 20 minutes at 20 ± 1 °C. Mixing regime selected based on observations during preliminary testing. This saturated solution was used as highest test concentration level and as stock solution for preparation of further dilutions.
- Evidence of undissolved material: No, the saturated test item solution checked via laser beam (Tyndall effect) for undissolved test item (formation of an emulsion). The Tyndall effect was negative.
- Other relevant information: none

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Strain/clone: Daphnia magna STRAUS, obtained from continuous laboratory cultures
- Age at study initiation: less than 24 hours old
- Stage and instar at study initiation: to achieve this juvenile daphnids were removed from culture vessels at the latest 24 hours before start of exposure and discarded, juveniles born within the following period were used for the test, no first brood progeny was used
- Method of breeding: in glass vessels (2-3 l capacity) with approximately 1.8L culture medium (Elendt M4 acc. to guideline), feeding at least 5 times per week ad libitum with a mix of unicellular green algae (e.g. Pseudokirchneriella and Desmodesmus subspicatus)
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), Berlin, Germany
- Feeding during test : none

ACCLIMATION
- Acclimation period: at least 2 hours in dilution water
- Acclimation conditions: same as test

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

273 mg CaCO3/L

Test temperature:

19.0 - 20°C

pH:

7.15 - 7.85

Dissolved oxygen:

7.16 - 8.57 mg/L

Conductivity:

649 - 668 µS/cm

Nominal and measured concentrations:

Nominal: 6.25, 12.5, 25.0, 50.0, 100% saturated solution (test concentrations based on results of non-GLP preliminary range finding test)
Measured (geometric mean): 20.0, 52.4, 112, 203, 448 µg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: beaker, loosely covered with watch glasses
- Material: glass, (4 (id) x 7 (h) cm), 50 mL capacity
- Volume of solution: 20 mL
- Aeration: no
- Renewal rate of test solution (frequency): 1 renewal after 24 hours
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Test Water (according to guideline OECD 202, Annex 3)
- Culture medium different from test medium: yes, Elendt M4
- Intervals of water quality measurement: prior to start of exposure (0 hours) and renewal of test solutions (24 hours) ), the water quality parameters (i.e. pH value, dissolved oxygen concentration, temperature, conductivity and total hardness) of the dilution water were measured.
At the start of the exposure and at the renewal (0 and 24 hours), the water quality parameters of the fresh media (i.e. pH value, dissolved oxygen concentration) were measured in one additional replicate per concentration level and control. At the renewal and at the end of the exposure (24 and 48 hours), the water quality parameters of the old media were measured in one appropriate replicate (containing daphnids) per concentration level and control. The replicate with the highest immobilization rate per concentration level and control was measured.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16/8 hours light/dark cycle
- Light intensity: diffuse light, light intensity of max. 1500 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): immobilization determined in all groups after 24 and 48 hours (an animal considered immobile, if not able to swim in the water phase within 15 s after gentle agitation of the test vesel), determination of other adverse, sub-lethal effects (e.g. discolouration, abnormal behaviour, etc.)

RANGE-FINDING STUDY
- Test concentrations: control, 10 and 100% saturated solution
- Results used to determine the conditions for the definitive study: 90% immobile daphnids at 100% saturated solution (geometric mean measured concentration: 735 µg/l), 0% immoblie daphnids at 10% saturated solution (geometric mean measured concentration: 69 µg/l), 0% immobile daphnids at control

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 448 µg/L

Nominal / measured:

meas. (geom. mean)

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

302 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Results with reference substance (positive control):

EC50-value (with Confidence Limits) of the Reference Item Potassium dichromate based on nominal concentrations mg/L, (0 - 24 hours)
EC50 = 2.03 mg/L (valid range: 0.6-2.4 mg/L)
confidence limits: 1-4 mg/L

Reported statistics and error estimates:

After 48 hours only 0 and 100% mortality occurred. Therefore the EC50-values were calculated by straight line regression. The concentrations enclosed to the EC50 - values were chosen as lower and upper confidence limits. All calculations were carried out from the best-fit values with the software GraphPad Prism.

All data were computer-processed and rounded for presentation. Consequently, minor variations may occur from the original figures if manual calculations based on the original figures are made subsequently. Calculations were made using the following software:
- Excel, MICROSOFT CORPORATION
- GraphPad Prism, Version 5.03, GraphPad Software, Inc.

Table 2: Measured Concentrations of the Test Item Isocarb 16 during the Definitive Test

Sampling date	Fresh media 0 hours	Old media, 24 hours		Fresh media, 24 hours	Old media, 48 hours		Geometric mean measured test item concentration/µg/L
Percentage of the saturated solution/%	Isocarb 16
	Meas. conc./µg/L	Meas. conc./µg/L	%	Meas. conc./µg/L	Meas. conc./µg/L	%
100	547	568	104	333	391	117	448
50.0	224	258	115	164	178	109	203
25.0	137	141	103	92.8	87.5	94	112
12.5	65.1	55.8	86	50 .8	40.9	81	52.4
6.25	< LOQ	< LOQ	< LOQ	< LOQ	< LOQ	< LOQ	20
Control	< LOQ	< LOQ	< LOQ	< LOQ	< LOQ	< LOQ

Table 3:       EC10-, EC50- and EC100-Values

Effect concentrations	Test duration/hours	2 -hexyldecanoic acid Geometric mean measured concentration of the test item/µg/L
EC10 (with confidence limits	24	> 448
	48	203 < EC10 < 448
EC50 (with confidence limits	24	> 448
	48	302 (203 -448)*
EC100	24	> 448
	48	448

*= determined via straight line regression due to no partial response

Table 4: Immobilization Rates after 24 and 48 hours of Exposure in the Definitive Test

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

Geometric mean           measured test item concentration/µg/L	Immobilization/%
	24 hours					48 hours
	Replicates					Replicates
	1	2	3	4	MV	1	2	3	4	MV
448	0	0	20	0	5	100	100	100	100	100
203	0	0	0	0	0	0	0	0	0	0
112	0	0	0	0	0	0	0	0	0	0
52.4	0	0	0	0	0	0	0	0	0	0
20.0	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

Table 5: Absolute Numbers of immobile Daphnids after 24 and 48 hours of Exposure in the Definitive Test

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

Geometric mean           measured test item concentration/µg/L	Number of immobile daphnids/total number of daphnids
	24 hours					48 hours
	Replicates					Replicates
	1	2	3	4	Sum	1	2	3	4	Sum
448	0/5	0/5	1/5	0/5	1/20	5/5	5/5	5/5	5/5	20/20
203	0/5	0/5	0/5	0/5	0/20	0/5	0/5	0/5	0/5	0/20
112	0/5	0/5	0/5	0/5	0/20	0/5	0/5	0/5	0/5	0/20
52.4	0/5	0/5	0/5	0/5	0/20	0/5	0/5	0/5	0/5	0/20
20.0	0/5	0/5	0/5	0/5	0/20	0/5	0/5	0/5	0/5	0/20
Control	0/5	0/5	0/5	0/5	0/20	0/5	0/5	0/5	0/5	0/20

Validity criteria fulfilled:

yes

Remarks:

control group: no daphnids immobilized or showed any signs of disease or stress, dissolved O2 concentration in the 24-hours old media was ≥ 7.16 mg/L) in all concentration levels and the control.

Conclusions:

Based on the geometric mean measured concentrations of the test item 2-hexyldecanoic acid, the 48 hours EC50 for Daphnia magna was 302 µg/L (confidence limits: 203 – 448 µg/L).

Executive summary:

In the acute immobilization test with Daphnia magna (STRAUS), the effects of the test item 2-hexyldeccanoic acid were determined according to OECD 202 (2004).

The study was conducted under semi-static conditions over a period of 48 hours with a saturated solution of the test item and a further four dilution levels in a geometric series with a separation factor of 2 (nominal concentrations 6.25 to 100% of the saturated solution, corresponding to the geometric mean measured concentrations 20.0 – 52.4 – 112 – 203 – 448 µg/L).

The test item is a colorless liquid with a water solubility of 71 µg/L (ECHA factsheet, QSAR calculation). A saturated solution of the test item was prepared before the start of the exposure (day 0) and before the renewal of the test solutions (day 1). The saturated solution was used as the highest concentration level and as the stock solution for the preparation of all further tested concentration levels.

The tested concentration levels were colorless and visually clear throughout the exposure period. A negative Tyndall effect was observed in the saturated solution. The test solutions were colourless and clear. Twenty daphnids (divided into 4 replicates with 5 daphnids each) were exposed to each concentration level and the untreated control.

The test item concentrations were analytically verified via LC-MS/MS in the fresh media at the start of the exposure and at the renewal of the test solutions (0 and 24 hours) as well as 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 measured concentrations of the test item in the old media (24 and 48 hours)were in the range of < LOQ to 117% of the initially measured concentrations.

Based on the geometric mean measured concentrations of the test item 2-hexyldecanoic acid, the 48 hours EC50 for Daphnia magna was 302 µg/L (confidence limits: 203 – 448 µg/L).

Description of key information

In the acute immobilization test with Daphnia magna (STRAUS), the effects of the test item 2-hexyldeccanoic acid were determined according to OECD 202 (2004). The study was conducted under semi-static conditions over a period of 48 hours with a saturated solution of the test item and a further four dilution levels in a geometric series with a separation factor of 2 (nominal concentrations 6.25 to 100% of the saturated solution, corresponding to the geometric mean measured concentrations 20.0 – 52.4 – 112 – 203 – 448 µg/L). Based on the geometric mean measured concentrations of the test item 2-hexyldecanoic acid, the 48 hours EC50 for Daphnia magna was 302 µg/L (confidence limits: 203 – 448 µg/L).

 

In addition, a reliable calculated value from ECOSAR 2.0 is available. The calculated daphnid 48 hour LC50 was 0.29 mg/L for the substance.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

0.32 mg/L

Additional information

In the acute immobilization test with Daphnia magna (STRAUS), the effects of the test item 2-hexyldeccanoic acid were determined according to OECD 202 (2004).

The study was conducted under semi-static conditions over a period of 48 hours with a saturated solution of the test item and a further four dilution levels in a geometric series with a separation factor of 2 (nominal concentrations 6.25 to 100% of the saturated solution, corresponding to the geometric mean measured concentrations 20.0 – 52.4 – 112 – 203 – 448 µg/L).

The test item is a colorless liquid with a water solubility of 71 µg/L (ECHA factsheet, QSAR calculation). A saturated solution of the test item was prepared before the start of the exposure (day 0) and before the renewal of the test solutions (day 1). The saturated solution was used as the highest concentration level and as the stock solution for the preparation of all further tested concentration levels.

In addition, the ECOSAR 2.0,model calculated a daphnid 48 hour LC50 was 0.29 mg/L for the substance.

In toxicity studies with linear C16 acid, no toxicity to daphnids was observed at the level of water solubility.