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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-04-22 to 2010-04-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes
Remarks:
no certificate
Specific details on test material used for the study:
- Name of test material (as cited in study report): Decanal, Ald C-10
- Substance type: Colourless to pale yellow liquid
- Physical state: Liquid
- Impurities (identity and concentrations): No data
- Composition of test material, percentage of components: No data
- Isomers composition: No data
- Purity test date: 2009-12-07
- Lot No: 0002825582
- Expiration date of the lot: 2010-12
- Stability under test conditions: Stable under normal storage conditions
- Storage condition of test material: Room temperature
Analytical monitoring:
yes
Vehicle:
no
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Source: In-house culture
- Age at study initiation: < 24 hr
- Food type: During the holding period, the daphnids were fed a suspension of the algal species, Pseudokirchneriella subcapitata (formerly Selenastrum capricornutum) at least once a day supplemented by an artificial diet that was prepared according to ABC standard operating procedure.
- No feeding during test

ACCLIMATION
- Acclimation period: None required.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
140 to 142 mg CaCO₃/L
Test temperature:
20.4 to 21.4 °C (table 4 in attached supporting materials)
pH:
8.1 to 8.5 (table 5 in attached supporting materials)
Dissolved oxygen:
5.5 to 8.9 mg/L (65 to 105 % saturation) (table 6 in attached supporting materials)
Salinity:
Conductivity: 331 to 340 μS
Nominal and measured concentrations:
Nominal: 0 (control), 0 (stabiliser control), 1.0, 1.5, 2.2, 3.3, and 5.0 mg/L
Mean measured: < MQL (control), < MQL (stabiliser control), 0.903, 1.30, 1.88, 2.85, and 4.24 mg/L
Details on test conditions:
DILUTION WATER
The dilution water was an aged laboratory freshwater prepared by blending naturally hard well water with well water that was demineralised by reverse osmosis. These waters were blended to yield a total hardness of approximately 130 to 160 mg CaCO₃/L and biologically aged (held in a tank containing aquatic organisms). The water was then passed through a sediment filter, UV irradiated, and aerated prior to use.

EXPOSURE SYSTEM
The range-finding test was conducted in 150 mL glass jars containing approximately 150mL of control or test solution and covered with plastic lids. The definitive test was conducted in 250 mL glass jars containing approximately 250 mL of control or test solution and covered with glass plates for minimal headspace conditions. The jars were maintained at 20 ± 1 °C in a temperature-controlled water bath. Fluorescent lighting was maintained on a 16-hour daylight photoperiod with 30-minute simulated dawn and dusk periods. The measured light intensity at the start of the definitive test was 611 lux as measured with a LI-COR Model LI-189 light meter equipped with a photometric sensor.

RANGE-FINDING TEST
A static-renewal range-finding test was conducted at nominal concentrations of 0 (control), 1.3, 2.5, 5.0, 10, and 20 mg/L. Dilutions were made from a 0.020 mg/mL primary stock solution which was prepared at test initiation and 24 hr by suspending approximately 0.0100 g decanal into a 500-mL volume of dilution water. After 24 hr, surviving daphnids were transferred to freshly prepared solutions. Daphnids were observed after 24 and 48 hours.

A static-renewal range-finding test with the stabiliser, BHA, was conducted at nominal concentrations of 0 (control) and 0.020 mg BHA/L. A 0.000020 mg/mL primary stock solution was prepared at test initiation and 24 hr by suspending approximately 0.00004 g BHA into a 2,000 mL volume of dilution water. The primary standard was used as the test treatment solution. After 24 hr, surviving daphnids were transferred to freshly prepared solutions. The control and test substance solution were clear and colourless with no visible signs of undissolved test substance, precipitate, or surface film throughout the study.

DEFINITIVE TEST
Based upon the results of the range-finding tests, the definitive test was performed at nominal concentrations of 0 (control), 0 (stabiliser control), 1.0, 1.5, 2.2, 3.3, and 5.0 mg a.i./L. A 0.020 mg a.i./mL primary stock solution was prepared at test initiation and 24 hr by weighing 0.4036 g decanal into a 21-L glass jar, correcting for purity, and bringing to a volume of 20,000 mL volume with dilution water. Aliquots of the primary stock solution were used to prepare each test concentration. The control consisted of dilution water only. Because the decanal contains a stabiliser, BHA, a stabiliser control was tested at a concentration approximating the concentration of stabiliser in the highest test substance treatment (i.e., 5.0 mg/L). The stabiliser control was prepared by weighing 0.000045 g of BHA into a 10-L glass jar and bringing to a volume of 9,000 mL with dilution water. Each treatment or control was replicated four times. All jars were labelled with the study number, treatment, and replicate. The jars were positioned by treatment in a water bath. No aeration was provided to any jar during the test.

5 neonates (< 24 hr old) were added to each jar. Daphnids were impartially added to a set of labelled containers with each container representing one treatment replicate. Each container was then randomly assigned to a treatment replicate by random number generator. The individuals within each container were then transferred into the corresponding test chamber below the surface of the solution using a pipette.

Renewal test chambers for the control and all test treatments were 250 mL glass jars (the same as those used at initiation) with 250 mL of the appropriate solutions prepared as above. Organisms were transferred after 24 hr from “old” to “new” solutions using a glass pipette. Transfers began with the control and then proceeded from the lowest concentration to the highest concentration. Observations for immobility and sublethal responses were made every 24 hr (± 1 hr) for the duration of the test.

Total hardness, total alkalinity, and conductivity were measured in the dilution water at test initiation. Total hardness and alkalinity were measured using titrimetric methods. Conductivity was measured with a WTW Cond 330i conductivity/salinity meter. Temperature, dissolved oxygen concentration, and pH were measured daily in all replicates. Dissolved oxygen concentrations were measured with a WTW Oxi 330i dissolved oxygen meter. The test solution temperature and pH were measured with a WTW pH 330i meter. A thermistor probe was positioned in the water bath to continuously record temperature.

EFFECT PARAMETERS MEASURED
The criterion for effect was immobilisation which is defined as those animals that are not able to swim or show movement within 115 seconds, after gentle agitation of the test chamber or gentle disturbance of the daphnid itself.

STATISTICS
All statistical analyses were performed with SAS software (version 9.1). Estimates of EC₅₀ values and their 95 % confidence limits were calculated using the probit method and Trimmed Spearman-Karber method. When the P value for Goodness of Fit was > 0.05 and there was no other evidence of questionable convergence, the probit method was selected for reporting. When this criterion was not achieved, the Trimmed Spearman-Karber method was selected for reporting. The no-observed-effect concentration (NOEC) was determined by using Fisher’s one-tailed exact test
Reference substance (positive control):
no
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
2.95 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % CL: 2.70 and 3.23 mg/L; Mean of measured fresh solution concentrations
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
1.94 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % CL: 1.76 and 2.15 mg/L; Mean of measured fresh solution concentrations
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
1.3 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Mean of measured fresh solution concentrations
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
2.36 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
act. ingr.
Basis for effect:
mobility
Remarks on result:
other: 95 % CL: 2.04 and 2.72 mg a.i./L, based on geometric mean of measured old and new solutions.
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
1.17 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
act. ingr.
Basis for effect:
mobility
Remarks on result:
other: 95 %CL: 0.999 and 1.40 mg a.i./L, based on geometric mean of measured old and new solutions.
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
0.588 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
act. ingr.
Basis for effect:
mobility
Remarks on result:
other: Based on geometric mean of measured old and new solutions.
Details on results:
RANGE-FINDING TEST
- The control and all test substance solutions were clear and colourless with no visible signs of undissolved test substance, precipitate, or surface film throughout the study.
- After 48 hrs of exposure, the immobility was 0, 0, 20, 100, 100, and 100 % in the 0 (control), 1.3, 2.5, 5.0, 10, and 20 mg/L treatment levels, respectively.

ANALYTICAL RESULTS
Analytical confirmation of the test substance, decanal, in test solutions was performed at 0, 24, and 48 hrs. Measured concentrations of decanal in the test solutions at test initiation were 0.879, 1.25, 1.82, 2.68, and 4.24 mg/L, which represented recoveries of 81 to 88 % of the nominal treatment concentrations. The measured concentrations in the 24 hr old test solutions were < MQL, < MQL, 1.48, 2.42, and 3.84 mg/L, which represented recoveries of 0 to 77 % of the nominal treatment concentrations. The measured concentrations in the 24 hr new test solutions were 0.926, 1.34, 1.94, and 3.02 mg/L, which represented recoveries of 88 to 93% of the nominal treatment concentrations. The measured concentrations in the 48 hr test solutions were < MQL, < MQL, < MQL, and 1.48 mg/L, which represented recoveries of 0 to 45% of the nominal treatment concentrations. The mean measured concentrations of the fresh solutions (0 and 24 hours) were 0.903, 1.30, 1.88, 2.85, and 4.24 mg/L which represented recoveries of 85 to 90% of the nominal treatment concentrations. The geometric mean measured concentrations of all test solutions during the 48 hr study were 0.491, 0.588, 1.09, 2.32, and 4.04 mg/L, which represented recoveries of 39 to 81 % of the nominal treatment concentrations. No residues of the test substance were detected in the dilution water or stabilizer controls above the minimum quantifiable limit (MQL) of 0.533 mg/L. Recoveries from quality control fortifications ranged from 86 to 110 % of the nominal concentrations. The biological response results are based upon the mean of the measured fresh solution concentrations at 0 and 24 hours.

BIOLOGICAL RESULTS
After 48 hrs of exposure, immobility was 0, 0, 0, 0, 50, 95, and 100% in the 0 (control), 0 (stabilizer control), 0.903, 1.30, 1.88, 2.85, and 4.24 mg/L treatments. The 24 hr EC₅₀ was estimated to be 2.95 mg/L with 95% confidence limits of 2.70 and 3.23 mg/L (based on the mean of measured fresh solution concentrations) and 2.36 mg/L with 95% confidence limits of 2.04 and 2.72 mg/L (based on the geometric mean of measured new and old solution concentrations). The 48 hr EC₅₀ was estimated to be 1.94 mg/L with 95% confidence limits of 1.76 and 2.15 mg/L (based on the mean of measured fresh solution concentrations) and 1.17 mg/L with 95% confidence limits of 0.999 and 1.40 mg/L (based on the geometric mean of new and old solution concentrations). The slope of the 48 hr concentration-response line was 11.4 (based on the mean of measured fresh solution concentrations) and 6.68 (based on the geometric mean of new and old solution concentrations). This indicates the endpoints calculated with fresh solution mean measured values are more indicative of the actual organism response to the test substance. There were no sublethal effects noted in the control or treatments during the definitive test. The 48 hr NOEC was 1.30 mg/L (mean of measured fresh solution concentrations) and 0.588 mg/L (geometric mean of new and old solution concentrations) based upon a lack of statistically significant immobility at this and all lower test substance concentrations.

Table 1: Measured concentrations of decanal

Nominal concentration

(mg a.i./L)

Measured concentration as mg a.i./L (percent nominal)

0 ha

24 h (old)a

24 h (new)a

48 h (old)a

Meanc

Geometric meand

Control

<MQLb

<MQLb

<MQLb

<MQLb

<MQLb

<MQLb

Stabiliser control

<MQLb

<MQLb

<MQLb

<MQLb

<MQLb

<MQLb

1.0

0.879 (88)

<MQLb

0.926 (93)

<MQLb

0.903 (90)

0.491 (49)

1.5

1.25 (83)

<MQLb

1.34 (89)

<MQLb

1.30 (87)

0.588 (39)

2.2

1.82 (83)

1.48 (67)

1.94 (88)

<MQLb

1.88 (85)

1.09 (50)

3.3

2.68 (81)

2.42 (73)

3.02 (92)

1.48 (45)

2.85 (86)

2.32 (70)

5.0

4.24 (85)

3.84 (77)

NA

NA

4.24 (85)

4.04 (81)

NA= not available due to the fact that all daphnids were immobilized prior to this time point.

aSamples collected and analyzed on the corresponding day.

bMQL = 0.533 mg a.i./L

cMean is for new solutions only (0-hour and 24-hour)

dGeometric Mean is for new and old solutions. When measured concentration is less than MQL, a value of ½ MQL (0.267 mg/L) was used to calculate geometric mean.

 

 

Table 2: Immobility of Daphnia magna exposed to decanal

Nominal concentration

(mg a.i./L)

Mean concentration of fresh solutions (geometric mean of old and new solutions as mg a.i./L)

Cumulative percent immobile (% sublethal observationsa)

Replicate

24 h

48 h

Treatment

mean (48 h)

0-Control

0-Control

A

0 (0)

0 (0)

0

B

0 (0)

0 (0)

C

0 (0)

0 (0)

D

0 (0)

0 (0)

0-Stabiliser Control

0-Stabiliser Control

A

0 (0)

0 (0)

0

B

0 (0)

0 (0)

C

0 (0)

0 (0)

D

0 (0)

0 (0)

1.0

0.903 (0.491)

A

0 (0)

0 (0)

0

B

0 (0)

0 (0)

C

0 (0)

0 (0)

D

0 (0)

0 (0)

1.5

1.30 (0.588)

A

0 (0)

0 (0)

0

B

0 (0)

0 (0)

C

0 (0)

0 (0)

D

0 (0)

0 (0)

2.2

1.88 (1.09)

A

0 (0)

40 (0)

50*

B

0 (0)

40 (0)

C

0 (0)

60 (0)

D

0 (0)

60 (0)

3.3

2.85 (2.32)

A

40 (0)

100 (--)

95*

B

40 (0)

100 (--)

C

60 (0)

100 (--)b

D

20 (0)

80 (0)

5.0

4.24 (4.04)

A

100 (--)

100 (--)

100*

B

100 (--)

100 (--)

C

100 (--)

100 (--)

D

100 (--)

100 (--)

aNo sublethal observations were noted.

bFour mysids immobile; one not found and considered immobile.

* Statistically significant reduction in survival as compared to the control (Fisher’s One-Tailed Exact Test, p < 0.05).

Note: Five daphnids were placed in each replicate at test initiation, totalling 20 daphnids per treatment.

Validity criteria fulfilled:
yes
Remarks:
The study met all the criteria for test acceptability (i.e. < 10 % of control animals were affected, dissolved O₂ remained > 3 mg/L over all concs and at >1 conc. exhibited < 50% response and 1 conc. exhibited >50 response).
Conclusions:
The 48 h EC50 for the test substance was estimated to be 1.17 mg a.i./L based on the geometric mean measured concentrations of old and new solutions. The 48 h NOEC, also based on geometric mean measured concentrations of old and new solutions, was 0.588 mg a.i./L.

Description of key information

Daphnia  48 hr EC₅₀ 1.17 mg a.i./L based on geometric mean measured concentrations of old and new solutions (OECD 202).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
1.17 mg/L

Additional information

A reliable study was performed in accordance with GLP and OECD Guideline No. 202 to assess the effect of Decanal to the freshwater invertebrate, Daphnia magna in a 48 hour semi-static toxicity test. The nominal concentrations used were 0, 1.0, 1.5, 2.2, 3.3 and 5.0 mg/L and the mean measured concentrations of the fresh solutions were 0, 0.903, 1.30, 1.88, 2.85 and 4.24 mg/L. No sublethal effects were noted in the control or treatment groups during the definitive test. The 48-hour EC50 was estimated to be

1.94 mg/L (based on the mean of measured fresh solution concentrations) and 1.17 mg/L (based on the geometric mean of new and old solution concentrations). The 48 h NOEC was 1.30 mg/L (mean of measured fresh solution concentrations) and 0.588 mg/L (geometric mean of new and old solution concentrations) based upon a lack of statistically significant immobility at this and all lower test substance concentrations.

The study met all the criteria for test acceptability (i.e.,<10% of control animals were affected, dissolved oxygen remained above 3 mg/L over all concentrations and at least one concentration exhibited < 50% response and one concentration exhibited >50% response).