Glycerol trioctanoate

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:

02 May - 06 May 1994

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP Guideline study with acceptable restrictions: O2 concentration is measured to be < 60% in some test vessels.

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

yes

Remarks:

O2 concentration < 60% in some test vessels

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Samples were taken directly from the test vessels.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: 1 g test substance/L was dissolved in drinking water (Gelsenwasser AG), stirred for 18 h and filtered.
- Controls: yes, drinking water without addition of test substance

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebra fish
- Source: West Aquarium, Bad Lauterberg, Germany
- Length at study initiation: 3 cm ± 0.5 cm
- Feeding during test: none

ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions: same as test
- Type and amount of food: TetraMin®
- Feeding frequency: daily (approx. 1% of body weight)
- Health during acclimation (any mortality observed): 7 days before study initiation the mortality was ≤ 5%
- Other: Test organisms were treated with malachite green three times a week before the beginning of the 14-days acclimation period.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

approx. 12.3 °dH

Test temperature:

20 - 22 °C

pH:

7.2 - 8.4

Dissolved oxygen:

50 - 100% (50% after 48 h and 59% after 72 h in the exposure vessel with test substance)

Nominal and measured concentrations:

Nominal: 1 g/L
Measured: 23 mg/L (0 h), 60 mg/L (24 h), 100 mg/L (48 h), 74 mg/L (72 h)

Details on test conditions:

TEST SYSTEM
- Test vessel: aquaria
- Material: glass; Size: 30 L; Headspace: 20 L; Fill volume: 10 L
- Aeration: yes
- Renewal rate of test solution: daily
- No. of organisms per vessel: 10
- No. of vessels per concentration: 1
- No. of vessels per control: 1
- Biomass loading rate: approx. 0.29 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: drinking water from Gelsenwasser AG
- Culture medium different from test medium: yes, culture water was dechlorinated

OTHER TEST CONDITIONS
- Photoperiod: 16 h light / 8 h dark

EFFECT PARAMETERS MEASURED: Mortality was observed daily.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: limit test

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 64 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: filtered test solution

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 95 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtered test solution

Basis for effect:

mortality (fish)

Details on results:

- No mortality was observed in control or treatment group
- Effect concentrations exceeding solubility of substance in test medium: Used test substance concentration exceeded the water solubility. A saturated solution was used in the test.

Sublethal observations / clinical signs:

The O2-concentration was < 60% after 48 and 72 h in the test vessels with the test substance. Misleading results can be excluded since no mortality was observed.

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31 Jan- 4 Feb 1994

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions.

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: The concentration of the Water Accommodated Fraction was analysed

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution with 1 g/L test substance in drinking water was prepared. It was stirred for approx. 18h and afterwards filtrated. The filtrated solution (Water Accommodated Fraction - WAF) was used as test solution and prepared daily

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Source: West Aquarium, Bad Leuterberg
- Length at study initiation: 3 ± 0.5 cm
- Feeding during test: none


ACCLIMATION
- Acclimation period: The fish were treated with Malachitgreen three times a week and were in quarantine for 14 days before being used in tests.
- Type and amount of food: TetraMin (1% of bodyweight)
- Feeding frequency: daily
- Health during acclimation (any mortality observed): The fish were free of obvious illnesses. The mortality was 7 days before the start of the test < 5%.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

approx. 13 °dH

Test temperature:

20 ± 1 °C

pH:

7.3 - 8.3

Dissolved oxygen:

69 to 107% O2 saturation

Nominal and measured concentrations:

Preparation of Water Accommodated Fractions (WAF): Mean of measured concentrations is 53 mg/L (DOC measured)

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 30 L aquaria filled with 9 L
- Aeration: continuous
- Renewal rate of test solution: daily
- 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: drinking water from "Gelsenwasser AG"


OTHER TEST CONDITIONS
- Photoperiod: 16 h

EFFECT PARAMETERS MEASURED: mortality at 24, 48, 72 and 96 hours


TEST CONCENTRATIONS
- Justification for using less concentrations than requested by guideline: As substance is poorly water soluble, the test substance was only tested at water soluble concentration (WAF)

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 53 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: Measured concentrations ranged from 17 to 104 mg/L in freshly prepared test solutions (based on measured DOC).

Details on results:

- Mortality of control: 1 out of 10
- Effect concentrations exceeding solubility of substance in test medium: No, as Water Accommodated Fractions were prepared

Sublethal observations / clinical signs:

No toxicity of the test substance up to the limit of the water solubility under test conditions was observed.

WAF were prepared, but measured concentrations ranged from 17 to 104 mg/L in freshly prepared test solutions (based on measured DOC).

Table 2: Number of surviving and dead individuals during the test (absolute numbers and % values)

Concentration (mg/L)	Test period
	24			48			72			96
	Number		%	Number		%	Number		%	Number		%
	alive	dead	alive	alive	dead	alive	alive	dead	alive	alive	dead	alive
Control	10	0	0	10	0	0	10	0	0	9	1	10
WAF	10	0	0	10	0	0	10	0	0	10	0	0

 

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 Oct - 21 Dec 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Qualifier:

according to guideline

Guideline:

other: OECD Series on Testing and Assessment No. 126 "Threshold approach for acute fish toxicity" (May 31st, 2010)

Qualifier:

according to guideline

Guideline:

other: OECD Guidance Document No. 23 on Aquatic Toxicity of Difficult Substances and Mixtures, OECD 2000

GLP compliance:

yes (incl. QA statement)

Remarks:

LUBW Landesanstalt für Umwelt, Messungen und Naturschutz Baden-Württemberg, Karlsruhe, Germany

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the start and at the end of the experimental phase samples (4 mL) were taken for DOC measurement and filtered with a 0.45 µm cellulose acetate (CA) membrane filter (Whatman FP 30/0.45 µm). First the filter was washed with 100 mL bi-distilled water and with ca. 10 mL sample. Additional samples were not filtered for TOC measurement.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Due to the low water solubility of the test item, the study was performed with WAF ("water accommodated fraction") prepared with synthetic fish water according to OECD 203. The test item is a liquid and was weighted on pieces of polyethylene foil (rinsed with ethanol and H2O bidest. before) which was transferred together with the test item into a defined volume of synthetic fish water into a 5000 mL beaker. Two beakers were prepared like this and stirred for a period of 48 hours at 21 - 22 °C in diffuse light. For stirring an overhead high-grade stainless steel radial impeller was used (about 500 rpm). The impeller was placed about 4 cm (limit test) resp. 10 cm (LC50 test) above the bottom of the beakers. The depth of the vortex should was about 10% of the total height of the liquid. After stirring was stopped, un-dissolved material was allowed to sediment or float for a period of about 1 hour. After this sedimentation period the WAF with the highest loading rate was slightly turbid and oily drops were swimming on the surface. Also the second highest loading rate was slightly turbid. The three lower loading rates were clear. The test solutions were withdrawn from the middle of the suspension in the beakers using a glass tube and transferred into the test vessels (aquarium). The filtration of the WAF to obtain the water soluble fraction (WSF) was not taken into consideration to avoid unwanted impacts of the filtration process and to have a worst case scenario.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebra fish
- Source: Aquarium Dietzenbach Fischzucht, Dietzenbach, Germany; held in the Hydrotox laboratory since October 9th, 2012 (limit test) resp. November 20th, 2012 (LC50 test).
- Length at study initiation: 2 ± 1 cm
- Housing: After arrival in the lab, the fish were held at 21 - 25 °C in synthetic fish water according to OECD 203 with a light/dark cycle of 14:10 hours. The aquarium was cleaned at least once in two weeks and the water was changed at least once a week. Five days a week the fish were fed twice with commercially available fish feed (TetraMin from TetraWerke, Melle, Germany). The water temperature and the appearance of the fish were checked on a daily basis.
- Feeding during test: none

ACCLIMATION
- Acclimation period: 10 days before test start
- Acclimation conditions (same as test or not): same

Ten days before the test started the fish were held in synthetic fish water under test conditions with 1 litre water per fish. The water was renewed twice during this period. The temperature was in a range of 21.8 to 22.8 °C (LC50 test) resp. 22.6 to 23.1 °C (limit test). The target range of 21 to 25 °C ± 1 °C kept. The fish were kept at a light/dark cycle of 14:10 and were fed twice every working day. 24 hours before the start of the test, a last feeding was carried out. During this pre-test keeping period the fish behaved normally. One fish died during the limit test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

22.3 - 22.8 °C (limit test)
22.4 - 22.8 °C (LC50 study)

pH:

7.7 - 8.0 (limit test)
7.9 - 8.2 (LC50 study)

Dissolved oxygen:

4.5 - 8.2 mg O2/L (limit test)
6.8 - 8.5 mg O2/L (LC50 study)

Nominal and measured concentrations:

Nominal: control, 100 mg/L (WAF, nominal, limit test)
Nominal: control, 6.25, 12.5, 25, 50, and 100 mg/L (WAF, nominal, LC50 study)

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 8 L aquarium
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
Stock Solutions (synthetic fish water):
DF1: CaCI2 x 2 H2O: 294 g/L (2.0 mol/L)
DF2: MgSO4 x 7 H2O: 123.3 g/L (0.5 mol/L)
DF3: KCI: 2.3 g/L (0.03 mol/L)
DF4: NaHCO3: 25.9 g/L (0.3 mol/L)

The synthetic fish water is prepared as described in the following: To 10 mL each of solutions DF 1 and 2 and 25.5 mL each of solutions DF3 and DF4 deionised water was added up to 10 litres (concentration in the test: DF1: 2.0 mmol/L, DF2: 0.5 mmol/L, DF3: 0.07 mmol/L and DF4: 0.76 mmol/L).

- Intervals of water quality measurement: The pH of the test and the control aquarium was measured and recorded at the beginning and at the end of the test. The oxygen concentration (mg/L), the min-max temperatures and the behaviour of the fish were recorded on a daily basis.

OTHER TEST CONDITIONS
- Photoperiod: light/dark cycle of 14:10

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): mortality daily

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Results used to determine the conditions for the definitive study: A limit test with a nominal loading rate of 100 mg/L was performed. Because 6 of 7 fish died within the first 48 hours of the test a LC50 study comprising five loading rates between 100 mg/L and 6.25 mg/L differing by a spacing factor of 2 was performed.

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

75 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

mortality (fish)

Details on results:

- 100% mortality occurred at the highest loading rate of 100 mg/L, whereas no mortality was observed in any other treatment group.

Sublethal observations / clinical signs:

Results of the Fish, Acute Toxicity Test:

With 100 mg/L 85.7% (limit test) resp. 100% (LC50 study) of the fish introduced died during the first 48 hours of the test course. With the next lower concentration of 50 mg/L no fish died. The LL50 was calculated by interpolation between 100% effect and 0% effect. The LL50 is therefore 75 mg/L (lethal loading rate, nominal, WAF).

With the highest loading rate of 100 mg/L during the first hour of the test fish swam slowly at the bottom of the aquarium whereas the control fish were inconspicuous. After 24 hours five fish were strongly affected and laid on the ground of the aquarium. Another fish swam on its back and one fish was already almost dead on the ground. But all seven fish were still alive. After 43 hours six fish were dead, one fish swam at the surface and gasped. This fish died after 48 hours. It is striking that the oxygen concentration in the test solution decreased during 24 and 48 hours to 6.8 mg/L. This value corresponds to 79% of saturation and therefore oxygen lack is not the cause for fish's dead. A similar effect was observed in the limit test where oxygen concentration decreased to 4.5 mg/L.

With the second highest loading rate of 50 mg/L during the first hour of the test fish also swam slowly at the bottom of the aquarium as it could be observed with 100 mg/L. After 24 hours gill movement was increased and after 48 hours fish swam slower than those in the control. But no fish died with 50 mg/L during the 96 hours test course. With all three lower loading rates of 25 mg/L and 12.5 mg/L and 6.25 mg/L no conspicuous effects were observed. The fish were not affected.

Accompanying Analysis:

The data provided from the sponsor indicate that the test item is dispersible in water. Samples were analysed from the highest (100 mg/L), the middle (25 mg/L) and the lowest loading rate (6.25 mg/L) at the start (Oh) and at the end (96h) of test.

The results obtained are difficult to interpret. The TOC concentrations decreased during the 96 hours test course by a factor up to 3.2 in the highest loading rate. This could be attributed to the fact that on the glass surface of the aquarium a slippery coating was developed. Maybe the test item polymerised or changed otherwise its structure during the test course. On the other hand the DOC concentrations increased during the test course by a factor up to 3.4 in the highest loading rate. At the start of the test filtration for DOC determination was no problem, but at the end of the test (96 hours) the filter clogged already after filtration of about 3 - 5 mL test solution with the highest (100 mg/L) and the middle (25 mg/L) loading rate. A very high pressure had to be carried out to obtain enough sample material for DOC determination. Maybe polymerised material dissolved in the filter cake again by the high pressure during filtration which could explain the higher DOC values at the end of the experiment. This effect was not seen in the limit test, but the TOC/DOC concentrations in the test solutions at the start of the test were comparable.

Based on the TOC content of the test item of 64.3% a DOC/TOC concentration of 64.3 mg/L was expected in the highest loading rate (100 mg/L). But DOC analysed in the test solution at the start (Oh) was 14.1 mg/L corresponding to a recovery rate of 21.9% respectively to an test item concentration of 21.9 mg/L. For the middle concentration (25 mg/L) a DOC/TOC of 16.1 mg/L was expected and a DOC of 7.4 mg/L was analysed. This corresponds to a recovery rate of 46% respectively to an test item concentration of 11.5 mg/L. Due to the higher concentrations analysed at the end of the experiment and the deviating TOC concentrations the interpretation of the analytical data must be made with caution and it has to be kept in mind that a difficult substance was tested.

Reference 4

Endpoint:

short-term toxicity to fish

Type of information:

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

Remarks:

Summary of available data used for the endpoint assessment of the target substance

Adequacy of study:

key study

Justification for type of information:

Please refer to the analogue justification provided in IUCLID section 13

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 64 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: filtered test solution

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 620-67-7, Sasol, 1994, Danio rerio

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 95 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtered test solution

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 620-67-7, Sasol, 1994, Danio rerio

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

75 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test material / WAF

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 85536-07-8, Cremer, 2013, Danio rerio

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test material / WAF

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 85536-07-8, Cremer, 2013, Danio rerio

Duration:

96 h

Dose descriptor:

LOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

other: test material / WAF

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 85536-07-8, Cremer, 2013, Danio rerio

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

>= 53 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: Source: CAS 73398-61-5, Sasol, 1994, Danio rerio

Additional information

Since no studies investigating the short-term toxicity of Glycerol trioctanoate (CAS 538-23-8) to fish are available, in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5, a read-across to the structurally related source substances Glycerol triheptanoate (CAS 620-67-7), Glycerides, C8-10 mono and di (CAS 85536-07-8) and Triglycerides, mixed decanoyl and octanoyl (CAS 73398-61-5) was conducted. The source substances are representative to evaluate the short-term toxicity of the target substance to fish. This read-across is justified in detail within the analogue justification in IUCLID Section 13.

The target substance Glycerol trioctanoate (CAS 538-23-8) is a monoconstituent characterized by the alcohol component glycerol which is fully esterified with octanoic acid to give glycerol triesters of octanoic acid.

The source substance Glycerol triheptanoate (CAS 620-67-7) is a monoconstituent characterized by the alcohol component glycerol which is esterified to give triesters of C7 fatty acids.

The source substance Glycerides, C8-10 mono and di (CAS 85536-07-8) is a UVCB substance characterized by the alcohol component glycerol and C8-10 (even numbered) fatty acids. Esterification of glycerol gives mainly mono- and diesters of C8-10 (even numbered) fatty acids.

The source substance Triglycerides, mixed decanoyl and octanoyl (CAS 73398-61-5) is a UVCB substance characterized by the alcohol component glycerol and C8-10 (even numbered fatty acids). Esterification of glycerol gives mainly triesters of C8 and C10 fatty acids.

The study with read-across substance Glycerol triheptanoate (CAS 620-67-7) was performed in a semi-static design and under GLP (with acceptable restrictions) according to EU Method C.1 (Acute toxicity to fish). The freshwater zebrafish Danio reriowas used as test organism. The fish were exposed to a single nominal concentration of 1g/L (before filtration) over a period of 96 hours. Analytical measurements of the daily renewed test medium revealed actual concentrations of 23 mg/L (0 h), 60 mg/L (24 h), 100 mg/L (48 h) and 74 mg/L (72 h). No mortality was observed up to the limit of saturation (nominal: LC0≥95 mg/L, measured: LC0≥64 mg/L).

The study with read-across substance Glycerides, C8-10 mono and di (CAS 85536-07-8) was performed in a static design under GLP according to OECD Guideline 203 (Fish, Acute Toxicity Test). The freshwater zebrafish Danio reriowas used as test organism. The fish were exposed to nominal concentrations (WAF) of 6.25, 12.5, 25, 50, and 100 mg/L over a period of 96 hours. The highest test concentration resulting in 0% mortality was 50 mg/L. The lowest test concentration resulting in 100% mortality was 100 mg/L. The LL50 (96h) was determined as 75 mg/L.

The study with read-across substance Triglycerides, mixed decanoyl and octanoyl (CAS 73398-61-5) was performed as a limit test in a semi-static design and under GLP according to EU Method C.1 (Acute toxicity to fish). The freshwater zebrafish Danio reriowas used as test organism. The fish were exposed to a single measured concentration of 53 mg/L (WAF) over a period of 96 hours. No mortality was observed up to the limit of saturation (measured: LC0≥53 mg/L).

Based on the available results from structurally related read-across substances (in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5) which are characterized by a similar ecotoxicological profile and comparable structure, it can be concluded that Glycerol trioctanoate (CAS 538-23-8) shows no acute toxicity to fish up to the limit of saturation (lowest measured LC0≥53 mg/L).