Glycerides, C8-18 and C18-unsatd., reaction products with ethylene oxide and glycerol

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:

From February 06, 2012 to March 23, 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 7346/1-3 (Determination of the Acute Lethal Toxicity of Substances to a Freshwater Fish [Brachydanio rerio Hamilton-Buchanan (Teleostei, Cyprinidae)])

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all concentrations.
- Sampling method: On the day of analysis, the samples were defrosted at room temperature. The test samples were diluted in a 1:3 (v:v) ratio with acetonitrile and analysed. If necessary, the samples were further diluted with 75/25 (v/v) acetonitrile/ISO-medium to obtain concentrations within the calibration range.
- Sample storage conditions before analysis: Samples were stored in a freezer (≤ -15°C) until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The test substance was not completely soluble in test medium at the initial loading rates prepared. Preparation of test solutions started with individually prepared loading rates between 1.0 and 100 mg/L that were magnetically stirred for a period of 20-22 hours. The resulting aqueous mixtures were then left to settle for 2 hours. The resulting Water Accommodated Fractions (WAFs) were then collected by means of siphoning the middle fractions. The final test solutions were in general slightly hazy for the WAFs of 10 mg/L and higher and appeared clear and colourless at 1.0 and 4.6 mg/L WAFs.

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM
- Common name: carp.
- Source: Zodiac, proefacc, "De Haar Vissen", Wageningen University and Research Centre, The Netherlands.
- Length at study initiation (mean):
Range-finding test: 2.5 ± 0.2 cm
Final test: 2.1 ± 0.2 cm
- Weight at study initiation (mean):
Range-finding test: 0.37 ± 0.07 g
Final test: 0.71 ± 0.12 g
- Feeding during test: No feeding from 24 hours prior to the test and during the total test period.

ACCLIMATION
- Acclimation period: at least 12 days after delivery.
- Type of food: pelleted fish food (Cyprico Crumble Excellent (300-500 um), Coppens International bv, Helmond, The Netherlands)
- Feeding frequency: daily.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

180 mgCaCO3/L

Test temperature:

21.0 - 22.6 ºC

pH:

7.2 - 8.0

Dissolved oxygen:

2.7 - 9.6 mg O2/L

Nominal and measured concentrations:

WAFs prepared at loading rates of 4.6, 10, 22, 46 and 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 10L
- Material, size, headspace, fill volume: all-glass, containing 9.8 L of test solution.
- Aeration:The test media were not aerated during the test.
- Renewal rate of test solution (frequency/flow rate): daily.
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.51 g fish/L, i.e. 7 fish per 9.8 L of test medium.

TEST MEDIUM / WATER PARAMETERS
- Intervals of water quality measurement:
Dissolved oxygen content, pH and temperature: Daily in all vessels with surviving fish, beginning at the start of the test (day 0).

OTHER TEST CONDITIONS
- Photoperiod: 16 hours photoperiod daily.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Mortality and other effects: At 2, 24, 48, 72 and 96 hours following the start of exposure. In addition, every afternoon from day 0 and every morning from day 1 to observe for any dead or severely distressed fish.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: The expected LC50 was between concentrations present in WAFs prepared at 10 and 100 mg/L

Reference substance (positive control):

yes

Remarks:

pentachlorophenol

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

>= 46 - <= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

22 mg/L

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: (95% confidence interval 17 - 58 mg/L). The average exposure concentrations were calculated as mathematical means of the concentrations of test item measured during the first and the last refreshment period.

Details on results:

- Mortality: No mortality was observed in the control and WAFs prepared at loading rates of 4.6, 10, 22 and 46 mg/L. In the WAF prepared at 46 mg/L one fish died after 72 h of exposure, and 2 fish died after 96 h of exposure. In the WAF prepared at 100 mg/L 7 fish died after 24, 48, 72 and 96 h of exposure.

Other clinical effects: There were no clinical effects observed in the control and WAFs prepared at 100 mg/L during the test period.

Results with reference substance (positive control):

- Results with reference substance valid? Yes.
- LC50: The 96h-LC50 was 0.25 mg/L based on nominal concentrations, with a 95% confidence interval between 0.20 and 0.41 mg/L. This effect was already reached within 24 hours of exposure.The range of the 96h-LC50 for carp is generally between 0.10 and 0.46 mg/L

Sublethal observations / clinical signs:

Table 1. Incidence of mortality and total mortality during the final test.

Loading rate WAF* (mg/l)	Average meas. conc. (mg/l)	Initial number of fish	Cumulativemortality					Total Mortality (%)
			2h	24h	48h	72h	96h
Control	Control	7	0	0	0	0	0	0
4.6	0.50	7	0	0	0	0	0	0
10	2.9	7	0	0	0	0	0	0
22	10	7	0	0	0	0	0	0
46	18	7	0	0	0	1	2	29
100	68	7	0	7	7	7	7	100

* WAF prepared at the given Glycerides, coco mono- and di-, ethoxylated loading rate.

Table 2. Effect parameters.

Parameter	Loading rate WAF* (mg/l)	Average measured conc. (mg/l)	95%-confidence interval (mg/l)
NOEC	22	10	-
24 & 48h-LC50	>46, <100	35	18-68
72h-LC50	>46, <100	26	19-57
96h-LC50	>46, <100	22	17-58

  * WAF prepared at the given Glycerides, coco mono- and di-, ethoxylated loading rate.

Table 3. Clinical effects observed during the final test.

Loading rate WAF* (mg/l)	Time of recording (hours)	Specification of effects	Relative number
0, 4.6, 10, 22   46           100	0-96   24-48 72   96     2	No abnormalities   Swimming at the surface Swimming at the surface. Fins are red coloured (possibly internal haemorrhage) Swimming at the surface. Fins are red coloured (possibly internal haemorrhage)   Immobile Loss of equilibrium Swimming at the surface	7/7   7/7   6/6   5/5   4/7 2/7 1/7

* WAF prepared at the given Glycerides, coco mono- and di-, ethoxylated loading rate.

Validity criteria fulfilled:

yes

Remarks:

(No mortality in the control. Test conditions maintained constant during the test. Dissolved oxygen > 5mg/L. Measured conditions derivated 20% from nominal, results based on average exposure concentrations.)

Conclusions:

The 96h-LC50was between WAFs prepared at loading rates of 46 and 100 mg/L. The 96h-LC50 based on average exposure concentrations corresponded to 22 mg/L (95% confidence interval between 17 and 58 mg/L).

Executive summary:

 Assessment of acute toxic effects of Glycerides, coco mono- and di-, ethoxylated in Cyprinus carpio (Semi-static Method) was determinated according to the OECD Guideline 203, with GLP.

A final test was performed with daily renewal of test solutions based on the result of a preceding static range-finding test. Seven carp per test group were exposed to a control and WAFs prepared at loading rates of 4.6, 10, 22, 46 and 100 mg/L. Samples for analytical confirmation of actual exposure concentrations were taken during the first and last renewal interval. Samples were taken from freshly prepared solutions at the start and after 72 hours of exposure and from 24-hour old solutions at 24 and 96 hours of exposure.

 

Analysis of the samples taken from the WAFs during the two renewal intervals showed that measured concentrations decreased significantly over the 24-hour test periods with a significantly higher decrease during the last renewal interval. Based on these results, the average exposure concentrations were calculated. The range tested based on average exposure concentrations was 0.50, 2.9, 10, 18 and 68 mg/l for the WAFs prepared at 4.6, 10, 22, 46 and 100 mg/L, respectively.

 

The study generally met the acceptability criteria prescribed by the protocol and was considered valid.

The test substance induced no visible or lethal effects in carp at or below a WAF prepared at a loading rate of 22 mg/L. This corresponded to an average measured exposure concentration of 10 mg/L (NOEC).

 

The 96h-LC₅₀was between WAFs prepared at loading rates of 46 and 100 mg/L. The 96h-LC₅₀based on average exposure concentrations corresponded to 22 mg/L (95% confidence interval between 17 and 58 mg/L).

Description of key information

Key study: Experimental results: OECD Guideline 203 and EU method C.1. GLP study. 
The 96h-LC50was between WAFs prepared at loading rates of 46 and 100 mg/L. The 96h-LC50 based on average exposure concentrations corresponded to 22 mg/L (95% confidence interval between 17 and 58 mg/L).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

22 mg/L

Additional information

Key study: Experimental results: OECD Guideline 203 and EU method C.1. GLP study.

The 96h-LC50was between WAFs prepared at loading rates of 46 and 100 mg/L. The 96h-LC50 based on average exposure concentrations corresponded to 22 mg/L (95% confidence interval between 17 and 58 mg/L).