Fatty acids, C10-12, esters with polylactic acid, sodium salts

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 Sep. - 04 Apr. 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

2019

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD 126, Short Guidance on the Threshold Approach for Acute Fish Toxcitiy

GLP compliance:

yes (incl. QA statement)

Remarks:

Staatliches Gewerbeaufsichtsamt Hildesheim, Niedersachsen, Germany

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all concentrations
- Sampling method: fresh media after 0 and 72 h, aged media after 24 and 96 h (first and second definitve test)
- Sample storage conditions before analysis: All original samples were stored at room temperature before preparation.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: An appropriate amount of the test item was weighed and test medium transferred into a test vessel filled with an appropriate amount of the dilution water. The test media were mixed with an laboratory blender (1 min, 17000 rpm). This procedure was repeated for every
water renewal.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Source: Umweltbundesamt, Berlin, Germany
- Age at study initiation (mean and range, SD): juveniles

ACCLIMATION
- Acclimation period: at least 9 days of acclimatization
- Acclimation conditions (same as test or not): same
- Type and amount of food during acclimation: Sera Vipan; SERA GmbH, Germany, ad libitum
- Feeding frequency during acclimation: 3 times a week
- Health during acclimation (any mortality observed): no mortality observed

FEEDING DURING TEST
no feeding

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

55 mg/L (first and second definitve test)

Test temperature:

20.8 - 22.4 °C (first definitve test)
20.9 - 22.7 °C (second definitve test)

pH:

7.15 - 7.69 (first definitve test)
7.21 - 7.98 (second definitve test)

Dissolved oxygen:

62 - 100% (second definitve test)
85 - 100% (second definitve test)

Conductivity:

149 μS/cm (recent measurement 2020-01-21)

Nominal and measured concentrations:

Nominal test concentration: 34.0 mg/L (first definitve test)
Nominal test concentration: 2.13, 4.25, 8.50, 17.0 mg/L (second definitve test)

Geometric mean measured concentrations: 25.1 mg/L (first definitve test)
Geometric mean measured concentrations: 1.54, 3.76, 8.63, 14.9 mg/L (second definitve test)

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass aquaria
- Type (delete if not applicable): covered with a glass plates.
- Material, size, headspace, fill volume: glass, 3 L (dimensions: 11.5/13/20 cm, depth of water: approx. 17 cm)
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): daily renewal
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.0812 g/L (first definitve test); 0.101 g/L (second definitve test)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap water of local origin. The water was filtered on activated charcoal and aerated for at least 24 h to remove chlorine.
- Total organic carbon: < 2 mg/L (first and second definitve test)
- Alkalinity: 0.6 mmol/L (recent measurement 2020-01-21)
- Culture medium different from test medium: no
- Intervals of water quality measurement: measured at 0, 24, 48, 72 and 96 hours

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16:8 h light/dark cycle
- Light intensity: 772 to 871 Lux (first definitve test), 676 to 972 Lux (second definitve test)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Mortality and symptoms after 2 ± 0.5 h, 5 ± 1 h and 24 ± 2 h. On days 2-4 of the test, all vessels with living fish were inspected twice per day.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study : not given

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

19.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Mortality of control: no
- Other adverse effects control: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no

Reported statistics and error estimates:

The LC50-value was calculated with straight line regression of the concentration causing 0 and 86 or 100 % mortality using the software GraphPad Prism. The concentrations enclosed to the LC50-value were chosen as lower and upper confidence limits.
Excel, MICROSOFT CORPORATION
GraphPad Prism, GRAPHPAD SOFTWARE, INC.

Sublethal observations / clinical signs:

Table 1: Cumulative Mortality [%] in the First Definitive Test

(number of fish: n = 7)

Geometric mean measured test item concentration [mg/L]	Cumulative mortality [%] at observation time [hours]
	Day 0		Day 1	Day 2		Day 3		Day 4
	2	5	24	48	52	72	76	93	97
25.1	0	0	86	100	--	--	--	--	--
Control	0	0	0	0	0	0	0	0	0

Table 2: Observations in the First Definitive Test

(number of fish: n = 7)

Geometric mean measured test item concentration [mg/L]	Effect*	Cumulative mortality [%] at observation time [hours]
		Day 0		Day 1	Day 2		Day 3		Day 4
		2	5	24	48	52	72	76	93	97
25.1	(E)	0	0	6	7	--	--	--	--	--
	(1)	7	7	1	--	--	--	--	--	--
Control	(1)	7	7	7	7	7	7	7	7	7

*) The numbers in brackets correspond to the following observations:

(1) = Normal behavior

E = Exitus letalis

Table 3: Cumulative Mortality [%] in the Second Definitive Test

(number of fish: n = 7)

Geometric mean measured test item concentration [mg/L]	Cumulative mortality [%] at observation time [hours]
	Day 0		Day 1	Day 2		Day 3		Day 4
	2	5	24	48	52	72	76	93	97
1.54	0	0	0	0	0	0	0	0	0
3.76	0	0	0	0	0	0	0	0	0
8.63	0	0	0	0	0	0	0	0	0
14.9	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0

Table 4: Observations in the Second Definitive Test

(number of fish: n = 7)

Geometric mean measured test item concentration [mg/L]	Effect*	Cumulative mortality [%] at observation time [hours]
		Day 0		Day 1	Day 2		Day 3		Day 4
		2	5	24	48	52	72	76	93	97
14.9	(1)	7	7	7	7	7	7	7	7	7
8.63	(1)	7	7	7	7	7	7	7	7	7
3.76	(1)	7	7	7	7	7	7	7	7	7
1.54	(1)	7	7	7	7	7	7	7	7	7
Control	(1)	7	7	7	7	7	7	7	7	7

*) The numbers in brackets correspond to the following observations:

(1) = Normal behavior

E = Exitus letalis

Table 5: Measured Concentrations of the Test Item during the first

Definitive Test

Sampling date:	0 hours, Fresh medium		24 hours, Old medium			72 hours, Fresh medium		96 hours, Old medium
Nominal concentration of the test item [mg/L]	Test item									Geometric mean measured concentration of the test item [mg/L]
	Meas. conc. [mg /L]	%	Meas. conc. [mg /L]		%	Meas. conc. [mg /L]	%	Meas. conc. [mg /L]	%
Control	<LOQ		<LOQ			<LOQ		<LOQ		-
34.0	33.2	98	19.0	56		n.d.		n.d.		25.1

Table 6: Measured Concentrations of the Test Item during the second

Definitive Test

Sampling date:	0 hours, Fresh medium		24 hours, Old medium		72 hours, Fresh medium		96 hours, Old medium
Nominal concentration of the test item [mg/L]	Test item								Geometric mean measured concentration of the test item [mg/L]
	Meas. conc. [mg /L]	%	Meas. conc. [mg /L]	%	Meas. conc. [mg /L]	%	Meas. conc. [mg /L]	%
Control	<LOQ		<LOQ		<LOQ		<LOQ		-
2.13	2.41	113	< LOQ		2.35	111	< LOQ		1.54
4.25	5.00	118	3.42	80	5.09	120	2.29	71	3.76
8.50	9.54	112	8.10	95	9.86	116	7.27	86	8.63
17.0	19.5	115	12.5	74	17.0	100	12.1	71	14.9

Table 7: Validity criteria for OECD 203.

Criterion from the guideline	Outcome	Validity criterion fulfilled
The mortality in the control(s) should not exceed 10% (or one fish if less than ten are used) at the end of the test	0%	yes
The dissolved oxygen concentration must have been at least 60 per cent of the air saturation value throughout the test	first definitive test: ≥ 62% and second definitive test ≥ 85%	yes
There must be evidence that the concentration of the substance being tested has been satisfactorily maintained, and preferably it should be at least 80% of the nominal concentration throughout the test. If the deviation from the nominal concentration is greater than 20%, results should be based on the measured concentration.	80 to 120% of nominal	yes

 

 

Validity criteria fulfilled:

yes

Remarks:

Please refer to "Any other information on results incl. tables"

Description of key information

LC50 (96 h): 19.3 mg/L (meas. geom. mean, OECD 203)

Additional information

The acute toxicity to fish (Danio rerio) was investigated in a study following OECD 203 and GLP. Two definitive tests were carried out. The first test was set up as a threshold approach with a nominal test concentration of 34 mg/L. A semi-static test system with a daily renewal rate was set up in this first test. The test concentrations in the fresh and aged media were analytically determined by LC-MS. The geometric mean measured concentration was 25.1 mg/L. A mortality of 100% was observed after 48 h of exposure. Therefore, a second definitive test was performed with nominal test concentrations of 2.13, 4.25, 8.50 and 17.0 mg/L. The test medium was renewed daily and the test concentrations were analytically verified by LC-MS (fresh and aged media). The geometric mean measured concentrations in the second definitive test were 1.54, 3.76, 8.63, 14.9 mg/L. Mortality or symptoms of intoxication were not observed in the second test at the tested concentrations. A LC50 (96 h) of 19.3 mg/L (meas. geom. mean) was reported based on the combined results of the two tests.