Alcohols, C6-8, ethoxylated

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:

2012-10-17 to 2012-10-23

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Alcohols, C6-C8-(even numbered, linear)-ethoxylated (<2,5 EO)

EC Number:

800-182-9

Cas Number:

1426148-68-6

Molecular formula:

R-O-R' , whereas R=C6-8-(even numbered, linear, saturated)-alkyl and R'=H or xEO wheras x=1-12, mean=2,3

IUPAC Name:

Alcohols, C6-C8-(even numbered, linear)-ethoxylated (<2,5 EO)

Details on test material:

- Name of test material (as cited in study report): Alcohols, C6-8 alkyl-(even, linear), ethoxylated (< 2.5 EO)
- Physical state: Liquid, colourless
- Analytical purity: 90 % (w/w)
- Purity test date: 2012-06-15
- Lot/batch No.: ESD0006420 (19.05.2011)
- Expiration date of the lot/batch: 2013-09-01
- Stability under test conditions: Not specified
- Storage condition of test material: Room temperature, protected from light, in original container.

Sampling and analysis

Analytical monitoring:

no

Details on sampling:

- Concentrations: 6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L
- Sampling method: All test concentrations and the control were analytically verified by LC-MS/MS in the fresh media (at 0 hours) and old media (after 48 hours) as specified below.
Sampling from freshly prepared media was carried out immediately after homogenisation. Samples of the old media were taken directly from the test vessels.
- Sample storage conditions before analysis: All samples were stored at room temperature before preparation and before analysis, if necessary.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The water accommodated fraction (WAF) was prepared with a nominal loading of the test item of 100 mg/L as specified below. 100 mg of the test item were weighed out and transferred with 1000 mL of the dilution water into a measuring flask. This dispersion was shaken for 24 h with 20 rpm at room temperature. After a separation phase of 1 h, the WAF* was taken from the homogeneous liquid phase (the approximate centre of the glass flask).

- Eluate: Dilution water
- Differential loading: Water accommodated fraction prepared with 100 mg/L test item/L
- Controls: Dilution water without test item tested under the same conditions as the test groups.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna (STRAUS)
- Strain: Clone 5
- Source: Institut für Wasser-, Boden- Lufthygiene (WaBoLu), further own breeding
- Age at study initiation (mean and range, SD): 2 to 24 h
- Method of breeding: In 2-3 L glass vessels with appr. 1.8 L culture medium, at 20 +/- 2 °C, in an incubator, 16 h illumination, illumination strength max. 20 µExm-2 x s-1
- Feeding during test: No feeding

ACCLIMATION
- Acclimation period: 2 h
- Acclimation conditions (same as test or not): Same as test
- Type and amount of food: No feeding
- Health during acclimation (any mortality observed): Healthy, no mortality observed

DILUTION WATER:
Dilution water ISO test water, according to OECD 202, Annex 3:
Component Concentration [mg/L]
KCl 5.76
NaHCO3 64.8
CaCl2  2 H2O 294
MgSO4  7 H2O 123
pH-value: 7.8  0.2

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

No

Test conditions

Hardness:

Dilution water at test start: 231 (mg CaCO3/L)

Test temperature:

20.7 °C

pH:

Water Quality Parameters at 0 h (Fresh Media)
(measured in one additional replicate per concentration level and control)
Nominal
Test Item Concentration
[mg/L] pH-Value


100 8.09
50.0 8.09
25.0 8.08
12.5 8.07
6.25 8.04
Control 7.99

Water Quality Parameters after 48 h (Old Media)
(measured in all replicates)
Nominal
Test Item Concentration
[mg/L] pH-Values
Replicates
1 2 3 4
100 7.93 7.93 7.92 7.94
50.0 7.98 7.98 7.95 7.95
25.0 7.97 7.98 7.97 7.96
12.5 7.96 7.96 7.97 7.96
6.25 7.95 7.95 7.96 7.95
Control 7.90 7.93 7.90 7.95

Dissolved oxygen:

Water Quality Parameters at 0 h (Fresh Media)
(measured in one additional replicate per concentration level and control)
Nominal
Test Item Concentration
[mg/L] Dissolved O2-Concentration

[mg/L]
100 8.60
50.0 8.57
25.0 8.53
12.5 8.45
6.25 8.14
Control 8.76

Water Quality Parameters after 48 h (Old Media)
(measured in all replicates)
Nominal
Test Item Concentration
[mg/L] Dissolved O2 -Concentration [mg/L]
Replicates
1 2 3 4
100 7.78 7.84 7.91 7.96
50.0 8.26 8.31 7.70 8.06
25.0 8.43 8.36 8.40 8.44
12.5 8.34 8.35 8.40 8.35
6.25 8.29 8.39 8.38 8.43
Control 8.30 8.21 8.37 8.34

Salinity:

Not measured, freshwater

Nominal and measured concentrations:

please refer to "Any other information on materials and methods"

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers, 50 mL
- Type (delete if not applicable): closed with parafilm
- Material, size, headspace, fill volume: Glass beakers (4 cm ID x 7 cm H), 50 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): 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): None


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: see above
- Conductivity: Dilution water at test start:632 µS/cm
- Culture medium different from test medium: Elendt M4 acc. to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
- Intervals of water quality measurement: At test start and test end


OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16:8 light:dark cycle
- Light intensity: Diffuse light, illumination strength max. 20 µE x m-2 x s-1


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Immobility was determined in all groups after 24 h and 48 h.

TEST CONCENTRATIONS
- Range finding study:
Immobilization Rates in the non GLP Preliminary Test
(n = 20, divided into 2 replicates with 10 daphnids each)
Nominal Test Item Concentration
[mg/L] IMMOBILIZATION [%]
24 h 48 h
Replicates Replicates
1 2 MV 1 2 MV
1000 100 100 100 100 100 100
100 70 70 70 90 90 90
10 0 0 0 0 0 0
1 0 0 0 0 0 0
Control 0 0 0 0 0 0

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

59.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CI (52.6-66.6)

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

37.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CI (26.2-44.7)

Details on results:

- Mortality of control: 0 %
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50/LC50: 1.92 (CI 1.74 - 2.10) mg/L

Reported statistics and error estimates:

EC100-values after 24 and 48 h, the NOEC and the LOEC after 48 h were deduced directly from the observation data, without mathematical calculation.
EC10- and the EC50-values were calculated by sigmoidal dose-response regression. The confidence intervals were calculated from the best-fit values, the standard error and the t-distribution with the software GraphPad Prism5.
The concentration effect relationships are shown graphically.

Any other information on results incl. tables

Immobilization Rates in the Definitive Test after 24 and 48 h of Exposure

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

Nominal Test Item Concentration [mg/L]	IMMOBILIZATION [%]
	24 h					48 h
	Replicates					Replicates
	1.	2.	3.	4.	MV	1.	2.	3.	4.	MV
100	100	80	80	80	85	100	100	100	80	95
50.0	0	20	40	40	25	0	20	40	60	30
25.0	0	0	0	0	0	20	0	0	0	5
12.5	0	0	0	0	0	0	0	0	0	0
6.25	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Based on the nominal concentrations of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (<2.5 EO) the 48-h EC50 for Daphnia magna was calculated to be 59.2 mg/L (95 % confidence limits: 52.6 – 66.6 mg/L). The NOEC after 48 h was 25.0 mg/L. The LOEC after 48 h was 50.0 mg/L.

Executive summary:

In the acute immobilization test with Daphnia magna (STRAUS), the effects of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO)(batch number: ESD0006420) were determined according to OECD 202 (2004) and Council Regulation (EC) No.440/2008 Method C.2 (2008) from 2012-10-17 to 2012-10-23, with the definitive exposure phase from 2012-10-17 to 2012-10-19, at Dr.U.Noack-Laboratorien, Käthe-Paulus-Str.1, 31157 Sarstedt, Germany.

The study was conducted under static conditions over a period of 48 h with 5 concentration levels of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO)in the range of 6.25 to 100 mg/L prepared with dilution water in a geometric series with a dilution factor of 2. Twenty daphnids were exposed to each concentration level and the control.

The concentrations of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO) wereanalytically verified by LC-MS/MS in the fresh media (0 h) and in the old media (48 h) at all concentration levels and the control.

The measured concentrations of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO) in the fresh media (0 h) were in the range of 99 to 106 % of the nominal values. The measured concentrations of the test item in the old media (48 h) were in the range of 99 to 102 % of the nominal values, indicating that the concentrations of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO) were successfully maintained for the duration of the test.

The endpoints were assessed based on the nominal concentrations of the test item Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO) (Table 1).

The water quality parameters (pH-value and dissolved oxygen concentration), measured at 0 and 48 h, were within the acceptable limits. The validity criteria of the test guideline were fulfilled.

Table 1:    EC₁₀-, EC₅₀- (with Confidence Interval), EC₁₀₀ - Values, NOEC and LOEC

(based on the nominal concentrations of the test item)

Alcohols, C6-8 alkyl-(even, linear), ethoxylated (2.5 EO)
Endpoints (with 95 % confidence limits)	Test Duration [h]	Nominal Test Item Concentration [mg/L]
EC10	24	40.2 (Cl: 34.9 – 45.8)
	48	37.5 (Cl: 26.2 – 44.7)
EC50	24	63.0 (Cl: 56.9 – 69.3)
	48	59.2 (Cl: 52.6 – 66.6)
EC100	24	> 100
	48	> 100
NOEC	48	25.0
LOEC	48	50.0