[No public or meaningful name is available]

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 - 13 Mar 2020

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Measured concentrations were below the limit of quantitation.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1075 (Freshwater and Saltwater Fish Acute Toxicity Test)

Version / remarks:

1996

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

2009

GLP compliance:

yes (incl. QA statement)

Remarks:

National Institute of Pharmacy and Nutrition, Budapest, Hungary (date of issue: 03 August 2018)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control and 100 mg/L nominal loading rate WAF was analytically determined at the start and at the end of the first and at the last renewal periods.
- Sampling method: Four samples were taken (2 x 2 x ~15 mL) from the control and at the applied test concentration level
- Sample storage conditions before analysis: Samples were frozen and kept approximately at -20°C at the Test Facility. One set of the samples was sent to the Test Site for analysis and one set was retained as a back-up at the Test Facility, if required for any confirmatory analyses.

Vehicle:

no

Details on test solutions:

- Method: The test item is poorly soluble in water, so test solutions were prepared using a saturated solution method (nominal loading rate of 100 mg/L, water accommodated fraction, WAF) according to the Guidance Document on Aqueous-Phase Aquatic Toxicity Testing of Difficult Test Chemicals, OECD No. 23. The saturated test item solution was prepared by dispersing/dissolving the amount of test item into the test medium (ISO-medium) two days before the start of the study. This solution was shaken for about 24 hours at approximately 30°C and then equilibrated for about 24 hours at approximately 20°C. The non-dissolved test material was removed by filtration through a fine (0.22 μm) filter. Prior to treatment of each renewal period, test item solutions were prepared by the method described above.
- Controls: yes, dilution water (circulated and filtered)

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebrafish
- Source: István Szent University, 2100 Gödöllő, Páter Károly u. 1, Hungary
- Length at study initiation (length definition, mean, range and SD): 2.4 – 2.7 cm
- Weight at study initiation (mean and range, SD): calculated mean weight of 1 fish = 0.25 - 0.27 g
- Method of breeding: Fish were bred in a well-known fish farm, under disease- and parasite-controlled conditions. Fish were observed during the acclimatisation and test periods, the fish were healthy. There was no mortality of the population for seven days before the test.

ACCLIMATION
- Acclimation period: more than 12 days

FEEDING DURING TEST
- Food type: No

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

186 mg/L (as CaCO3)

Test temperature:

20.6 - 22.6°C

pH:

7.59 – 8.32

Dissolved oxygen:

66 – 100% of the air saturation value

Nominal and measured concentrations:

- Nominal concentration: 100 mg/L nominal loading rate WAF
- Measured concentration 0 h (new): < LOQ
- Measured concentration 72 h (new): < LOQ
- Measured concentration 24 h (old): < LOQ
- Measured concentration 96 h (old): < LOQ

Details on test conditions:

TEST SYSTEM
- Test vessel: aquarium
- Fill volume: 2.5 L
- Renewal rate of test solution (frequency/flow rate): 24 h
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: Control = 0.77 g fish/L, 100 mg/L test concentration = 0.69 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: circulated and filtered water
- Intervals of water quality measurement: The water temperature, oxygen concentration and pH were measured at the beginning and at the end of the renewal periods during the experiment in each test aquarium.

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light and 8 hours darkness

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality or any sub-lethal effects were observed at 3, 6, 24, 48, 72 and 96 hours.

TEST CONCENTRATIONS
- Range finding study: Yes
- Test concentrations: 0.1, 1, 10, 100 mg/L nominal loading rate WAF
- Results used to determine the conditions for the definitive study: No effects in range finding study, NOEC = 100 mg/L nominal loading rate WAF

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other:

Remarks:

based on nominal loading rate WAF

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other:

Remarks:

based on nominal loading rate WAF

Details on results:

- Behavioural abnormalities: No, there were no clinical signs observed in the fish at any of the concentrations during the experiment.
- Observations on body length and weight: There was no considerable difference observed concerning body weights between the groups.

Reported statistics and error estimates:

No statistical analysis was performed because of the lack of toxic effects.

Sublethal observations / clinical signs:

Table 1: Calculation of exposure concentrations

Nominal Loading rate WAFs (mg/L)	Measured concentrations 1st renewal period (mg/L)		Measured concentrations 2nd renewal period (mg/L)
	start	end	start	end
Control	<LOQ*	<LOQ*	<LOQ*	<LOQ*
100.0	<LOQ*	<LOQ*	<LOQ*	<LOQ*

*0.038 mg/L sample concentration (counting with 50x enrichment and 1.05x dilution during the sample preparation)

 

Table 2: Cummulative mortality data of danio rerio

Nominal Loading rate WAF mg/L	Total number of animals	Cummulative mortality of fish
		3 h	6 h	24 h	48 h	72 h	96 h
Control	7	0	0	0	0	0	0
100	7	0	0	0	0	0	0

 

Table 3: Measured and calculated data of bodyweight

Test Group	Measured weight of 7 fish (g)	Calculated mean weight of 1 fish (g)	Loading of testing aquarium (g fish/L testing liquid)
Control	1.92	0.27	0.77
100.0 mg/L nominal loading rate	1.72	0.25	0.69

 

Table 4: Body length of test fish at mortality or at the end of the test

Loading rate WAF (mg/L)	Body length [cm]
Nominal
Control	2.6	2.4	2.7	2.6	2.6	2.5	2.6
100.0	2.4	2.5	2.5	2.6	2.7	2.5	2.6

 

Table 5: Validity criteria

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	66 – 100% air saturation throughout the test	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 per cent of the nominal concentration throughout the test. If the deviation from the nominal concentration is greater than 20 per cent, results should be based on the measured concentration.	The results were based on nominal concentrations, measured concentrations were below the limit of quantitation in this study	No

 

Validity criteria fulfilled:

no

Remarks:

See Table 5 at "any other information on results incl. tables".

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Refer to analogue justification document provided in IUCLID section 13.

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

2 300 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test mat. WAF

Basis for effect:

mortality (fish)

Remarks on result:

other: source, RA-A, CAS 91050-80-5, Emery, 2008, Cyprinodon variegatus, 96h

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

> 2 300 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test mat. WAF

Basis for effect:

mortality (fish)

Remarks on result:

other: source, RA-A, CAS 91050-80-5, Emery, 2008, Cyprinodon variegatus, 96h

Conclusions:

No effects are expected up to the limit of water solubility.

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 Nov - 10 Nov 2007

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Basic data given (comparable to guidelines/standards)

Qualifier:

according to guideline

Guideline:

other: OSPARCOM (2005-11) protocol for a fish acute toxicity test

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Principles of method if other than guideline:

Test methods were consistent with OECD 203 guideline as adapted by OSPARCOM (2005-11) for marine testing of offshore chemicals.

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test materials were characterised following Opus Plus SOP 402, where a 1000 mg/L stock was prepared and stirred for approximately 3 hours. This preparation was then left undisturbed for approximately one hour before behaviour in seawater classification was assigned and subsequently prepared for testing according to Opus Plus SOP 403. Poorly soluble materials were prepared as water accommodated fractions (WAFs). Solutions of 4 litres were prepared, stirred for 20 hours and then siphoned into the test vessels.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): floating or settled particles at 1000 mg/L in seawater

Test organisms (species):

Cyprinodon variegatus

Details on test organisms:

TEST ORGANISM
- Common name: Sheepshead minnow, killifish
- Source: reared and maintained at Opus Plus Ltd
- Age at study initiation (mean and range, SD):
- Length at study initiation: 1.5-2.5 cm

ACCLIMATION
- Acclimation conditions: Sheepshead minnow were held under flow-through conditions in 50 litre capacity glass tanks, containing 1 µm filtered ultra-violet treated seawater and supplied with gentle aeration.
- Type and amount of food: food in flake form ad libitum
- Feeding frequency: at least twice daily

Test type:

semi-static

Water media type:

saltwater

Limit test:

yes

Total exposure duration:

96 h

Test temperature:

19.7 - 20.4 °C

pH:

7.80 - 8.19 (control)
7.89 - 8.11

Dissolved oxygen:

94 - 97%

Salinity:

34 - 36‰

Nominal and measured concentrations:

2300 mg/L (nominal, WAF)

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 15 litre capacity glass tanks containing 3 litres of test media
- Aeration: yes
- Renewal rate of test solution (frequency/flow rate): replacement of media after 48h
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Natural local seawater is pumped to holding tanks from Scapa Flow, Orkney. The treatment process was: sand filtered to remove particulate, filtered to 1 µm, heated to 16 ºC, UV treated, degassed and held at a quantity of 3000 to 5000 litres. The water is constantly pumped through the system and trickle replenished as the water is used.


OTHER TEST CONDITIONS
- Adjustment of pH: yes


TEST CONCENTRATIONS
- Range finding study: Range-finding tests were conducted under static conditions over 96h with five animals exposed per concentration (one replicate); no data on concentrations provided
- Results used to determine the conditions for the definitive study: The test concentration is based on the LC/EC50 of the test material between most sensitive species Skeletonema costatum and Acartia tonsa in other tests. The algal species was noted to be the more sensitive from allied studies.

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

2 300 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test mat. WAF

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

> 2 300 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test mat. WAF

Basis for effect:

mortality (fish)

Reported statistics and error estimates:

Mortalities were determined in each vessel every 24h period. The number of dead organisms were expressed as a proportion of the total number exposed. Where sufficient response was observed for Range-finding and definitive tests, the 24h, 48h, 72h and 96h LC50values, 96h LC90and 96h NOEC values were calculated from the proportional response data using an appropriate method (ToxCalc V5, Tidepool Scientific Software).

Description of key information

LL50 (96 h) > 100 mg/L (nominal, Danio rerio)

LL50 (96 h) > 2300 mg/L (nominal, Cyprinodon variegatus); read-across

Additional information

There is one study available in which the short-term toxicity of Fatty acids, C16-18 (even numbered), esters with glycerol oligomers to fish was assessed.

The GLP study was performed according to OECD 203 using Danio rerio as test organism. A limit test with a nominal test concentration of 100 mg/L was conducted in a semi-static exposure regime. This test concentration was prepared as Water Accommodated Fraction (WAF). The test concentration was analytically determined at the start and at the end of the first and at the last renewal periods. The test item concentration could not be analytically quantified during the whole experiment as all measured concentrations were below the Limit of Quantification (LOQ = 0.038 mg/L). Therefore, all results are expressed as the nominal loading rate WAF. No effects were observed after 96 h for the tested concentration. Therefore, a LL50 (96 h) of > 100 mg/L (nominal) was derived.

This result is supported with a study for the structurally and chemically related source substance Fatty acids, C16-18, tetraesters with 3,3'-oxybis[1,2-propanediol] (CAS 91050-80-5). This read-across was performed in accordance with Regulation (EC) No 1907/2006 Annex XI, section 1.5 in order to fulfill the data requirements for this substance. The source substance is characterized by comparable chemical structure and similar physico-chemical properties and is therefore considered as suitable representative for the assessment of the short-term toxicity of the target substance to fish. A read-across justification in provided in IUCLID section 13.

The GLP study conducted with the source substance Fatty acids, C16-18, tetraesters with 3,3'-oxybis[1,2-propanediol] (CAS 91050-80-5) was performed according to “OSPARCOM (2005-11) protocol for a fish acute toxicity test”, equivalent to OECD 203, using the marine fish Cyprinodon variegatus. A limit test with a nominal test concentration of 2300 mg/L was conducted in a semi-static exposure regime. This test concentration was prepared as Water Accommodated Fraction (WAF). No mortality was observed after 96 h for the tested concentration. Therefore, a LL50 (96 h) of > 2300 mg/L (nominal) was derived.

Based on the available results from the target substance and a structurally related read-across substance (in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5) which is characterized by a similar ecotoxicological profile and comparable structure, it can be concluded that the target substance Fatty acids, C16-18 (even numbered), esters with glycerol oligomers is not expected to show toxicity to fish up to the limit of its water solubility (91.9 µg/L).