Calcium di(octanoate)

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:

20 June 2018 to 27 September 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)

Version / remarks:

13 April 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Highest and lowest nominal concentrations, 100 mg/L and 19.8 mg/L, at the start and end of the test, alongside the control.
- Sampling method: Samples for analysis of test item concentrations were taken from the bulk solutions at the beginning of the test and from the pooled replicates at the end.
- Sample storage conditions before analysis: Not reported

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The highest nominal concentration of 100 mg/L was prepared by diluting 200.3 mg of the test item in 2000 ml of dilution medium (Elendt M4) and ultrasonicating for twenty minutes followed by 10 minutes on a magnetic stirrer. The remaining treatment solutions were prepared by serial dilution with a geometric factor of 1.5 (i.e. 1334 ml made up to 2000 ml or 667 ml made up to 1000 ml).
- Eluate: Elendt M4 medium
- Differential loading: Yes
- Controls: Untreated test medium
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): None reported

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Strain/clone: Not reported
- Source: Originally obtained from Sciento Ltd, UK and cultured at AgroChemex Environmental.
- Age of parental stock (mean and range, SD): Not reported
- Feeding during test None

ACCLIMATION
- Acclimation period: All daphnids used in this test were less than 24 hours old and not of first brood progeny.
- Acclimation conditions (same as test or not): Yes
- Type and amount of food: Stock cultures were fed with an aqueous suspension of the unicellular green alga Chlorella vulgaris
- Feeding frequency: Once daily
- Health during acclimation (any mortality observed): None reported

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

249 mg/L as CaCO3

Test temperature:

Test media: 18.7 – 20.1°C
Test area: 18.4 – 18.9°C (mean: 18.6°C)

pH:

Test start: 7.52 - 7.55
Test end: 7.75 - 7.81

Dissolved oxygen:

Test start: 8.79 - 9.15 mg/L
Test end: 8.62 - 9.19 mg/L

Salinity:

Not applicable

Nominal and measured concentrations:

Nominal: 0, 19.8, 29.6, 44.4, 66.7 and 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Tall form beakers
- Type (delete if not applicable): Glass
- Material, size, headspace, fill volume: 250 mL capacity, containing 100 mL of test solution or dilution medium
- Aeration: None
- 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: Elendt M4
- Culture medium different from test medium: No
- Intervals of water quality measurement: The pH and dissolved oxygen were determined in the bulk solutions at the start of the test and from pooled treatment vessels at the end.

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Photoperiod: 16 hours light / 8 hours dark
- Light intensity: 530 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Observation of immobilised daphnia assessments were made after 24 and 48 hours (± 1 hour).

RANGE-FINDING STUDY
- Test concentrations: 0.1 to 100 mg/L
- Results used to determine the conditions for the definitive study: Range finding results indicated no true immobilisation up to the maximum concentration of 100 mg/L. The EC50 (immobilisation – 48h) was therefore expected to be >100 mg/L. However, as unusual effects relating to trapping at the water surface were seen at lower concentrations after 24 hours (largely unseen at 48 hours), a full dose-response test was indicated. Therefore, the exposure concentrations were selected to test up to the maximum concentration required by the testing guideline, with a low spacing factor of 1.5.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Details on results:

- Behavioural abnormalities: A number of daphnids were observed to be trapped in the surface tension of the medium but there was no clear dose-related pattern to these observations and the trapped daphnids were readily reinstated to the water column by the gentle agitation of the test vessels used to assess for immobility. At 48 hours, the highest concentration exhibited the same proportion of surface trapping as the controls (10%).
- Other biological observations: None
- Mortality of control: 0
- 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 reported
- Effect concentrations exceeding solubility of substance in test medium: No

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: EC50 = 0.6 - 2.1 mg/L
- Limit test: No
- Dose-response test: Yes
- ECx: EC50 = 1.422 mg/L (conducted during August 2017)

Reported statistics and error estimates:

Although the immobilisation seen at 19.8 mg/L was statistically significant, the four higher treatment groups did not exhibit statistically significant immobilisation. Therefore, the immobilisation at 19.8 mg/L was not considered to be treatment-related and the higher NOEC/LOEC pairing was selected.

Table 1: Immobilisation results

Treatment Group	Nominal Concentration (mg/L)	24 hours		48 hours
		Mean no. Immobilised	% Immobilised	Mean no. Immobilised	% Immobilised
1 (control)	0	0	0	0	0
2	19.8	0.75	15	1.75	35^a
3	29.6	0	0	0.25	5
4	44.4	0	0	0	0
5	66.7	0	0	0	0
6	100	0	0	0	0

^a Immobilisation statistically significant but not considered treatment-related, as the four higher concentrations exhibited no statistically significant immobilisation

Table 2: NOEC and LOEC data

	mg Test Item/L
	NOEC	LOEC
24 hours	100	> 100
48 hours	100	> 100

Table 3: Acceptance Criteria

Parameter	Acceptable Range
pH	6 to 9
	+/- units from initial measurement
Dissolved Oxygen	≥ 3 mg/L
Hardness (as CaCO3)	140 - 250 mg/L

Table 4: pH results

Solution	0 hours	48 hours	Difference	Acceptable (Y/N)
Control	7.55	7.75	+0.20	Y
Solution 2 (19.8 mg/L)	7.53	7.80	+0.27	Y
Solution 3 (29.6 mg/L)	7.52	7.78	+0.26	Y
Solution 4 (44.4 mg/L)	7.54	7.77	+0.23	Y
Solution 5 (66.7 mg/L)	7.54	7.79	+0.25	Y
Solution 6 (100 mg/L)	7.54	7.81	+0.27	Y

Table 5: Dissolved Oxygen results (mg/L)

Solution	0 hours	48 hiurs	Acceptable (Y/N)
Control	8.79	9.13	Y
Solution 2 (19.8 mg/L)	9.08	8.62	Y
Solution 3 (29.6 mg/L)	9.15	8.80	Y
Solution 4 (44.4 mg/L)	9.14	8.83	Y
Solution 5 (66.7 mg/L)	9.14	8.92	Y
Solution 6 (100 mg/L)	9.15	9.19	Y

Table 6: Incubation Conditions

Parameter	Acceptable Range	Recorded
Temperature of test area	18 - 22 °C	18.4 - 18.9 °C
	+/- 1°C variation throughout test	Mean: 18.6 °C
Temperature of test media	18 - 22 °C	18.7 - 20.1 °C
Photoperiod	16 hours light / 8 hours dark, cycle	On: 06:00 Off: 22:00
Light Intensity	400 - 800 lux	530 lux

Table 7: Test item analysis

Time point	Solution ID	Nominal concentration (mg test item/L)	Measured Concentration (mg test item/L)	Recovery (%)
0 hours	Sol 1 (control)	0	ND	N/A
	Sol 2	19.8	18.5	94
	Sol 6	100	102.9	103
48 hours	Sol 1 (control)	0	ND	N/A
	Sol 2	19.8	18.5	94
	Sol 6	100	97.6	98

ND = not detected

N/A = not applicable

Validity criteria fulfilled:

yes

Remarks:

Immobilisation in controls: 0%, trapped in meniscus: 10% (must be <10%); dissolved oxygen concentrations 8.62 – 9.19 mg/L (must be ≥3 mg/L)

Conclusions:

Based on nominal concentrations, the 48-hour EC50 for the test item to Daphnia magna was > 100 mg test item/L. The 48-hour NOEC and LOEC were 100 mg test item/L and > 100 mg test item/L, respectively.

Executive summary:

The acute toxicity of the test item to Daphnia magna was determined in a static test following OECD guideline 202. Daphnia were exposed to nominal concentrations of 19.8, 29.6, 44.4, 66.7 and 100 mg/L, alongside a control. Based on nominal concentrations, the 48-hour EC50 was >100 mg test item/L. The 48-hour NOEC and LOEC were 100 mg test item/L and >100 mg test item/L, respectively.

 

The study is a GLP compliant, guideline experimental study and is available as an unpublished study report. There are no restrictions and the study is fully adequate for assessment.

Description of key information

Based on nominal concentrations, the 48-hour EC50 for the test item to Daphnia magna was > 100 mg test item /L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

The acute toxicity of the test item to Daphnia magna was determined in a static test following OECD guideline 202 (Cockroft 2018). Daphnia were exposed to nominal concentrations of 19.8, 29.6, 44.4, 66.7 and 100 mg/L, alongside a control. Based on nominal concentrations, the 48-hour EC50 was >100 mg test item/L. The 48-hour NOEC and LOEC were 100 mg test item/L and >100 mg test item/L, respectively.

 

The study is a GLP compliant, guideline experimental study and is available as an unpublished study report. There are no restrictions and the study is fully adequate for assessment.