Pentaerythritol tetrakis(3-mercaptopropionate)

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:

02 February 2009 - 20 March 2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Analytical monitoring:

yes

Details on sampling:

Samples were taken from solvent control and each surviving test group at 0 /fresh media), 24, 48, 72 (old and fresh media), and 96 hours (old media). Due to the instability of the test material the samples were prepared and analysed immediately after sampling. Two samples of the solvent control and each surviving test group were taken at each occasion. One sample was analysed untreated and one sample after centrifugation. Further samples were taken (in duplicate) and stored at approx. -20°C for further analysis, if necessary

Vehicle:

yes

Details on test solutions:

Pre-study media preparation trial:

Pre-study solubility work showed that the highest attainable test concentration (by visual inspection) that could be prepared was 1.0 mg/L with an aliquot (100 µl/L) of a solvent stock solution prepared in tetrahydrofuran. At higher concentrations precipitation was observed. The test substance thus falls in the category of a 'difficult substance' as defined by OECD guidance documents (OECD 23, 2000). A media trial was conducted to determine the solubility of the test material under test conditions:

(1) Saturated solution preparation:
550 mg test substance were dispersed in 11 L water with the aid of propeller stirring at approx. 1500 rpm at a temperature of approx. 21°C for either 24 or 48 hours. Samples were then analysed after the following pre-treatments:
- untreated
- centrifugation at 10000g for 30 minutes
- centrifugation at 40000g for 30 minutes
- Filtration through a 0.2 µm Sartorius Sartopore filter (initial approx. 500 mL discarded)
- Filtration through a 0.2 µm Sartorius Sartopore filter (initial approx. 1 L discarded)

(2) Solvent spike preparations:
100 mg test material was dissolved in tetrahydrofuran and the volume adjusted to 10 mL to give a 100 mg/10 mL solvent stock solution. An aliquot (1000 µL) of this solvent stock solution was dispersed in 10 L of reconstituted water with the aid of magnetic stirring for approx. 10 min. to give a 1.0 mg/L test concentration. Samples were then analysed after the following pre-treatments:
- untreated
- centrifugation at 10000g for 30 minutes
- centrifugation at 40000g for 30 minutes
- Filtration through a 0.2 µm Gelman AcroCap filter (initial approx. 100 mL discarded)
- Filtration through a 0.2 µm Gelman AcroCap filter (initial approx. 500 mL discarded)

The remainder of the 1.0 mg/L test concentration was returned to the magnetic stirrer and stirred for a further 48 hours with samples being taken for analysis after both 24 ad 48 hours stirring.

Range finding test:
100 mg test material was dissolved in tetrahydrofuran and the volume adjusted to 10 mL to give a 100 mg/10 mL solvent stock solution from which a dilutions was prepared in tetrahydrofuran to give a 10 mg/10 mL solvent stock solution. An aliquot (2.0 mL) of these solvent stock solutions were each seperately dispersed in a final volume of 20 L of dechlorinated tap water and stirred with the aid of a flat bladed mixer for approx. 1 min. to give the nominal test concentrations of 0.10 and 1.0 mg/L, respectively. Each of the stock solutions was inverted several times to ensure adequate mixing and homogeneity.

Definitive test:
200 mg test material was dissolved in tetrahydrofuran and the volume adjusted to 20 mL to give a 200 mg/20 mL solvent stock solution. Aliquots (1.0, 1.8, 3.2, 5.6, and 10 mL) of this solvent stock solution were each seperately diluted in tetrahydrofuran and the volume each adjusted to 10 mL to give further solvent stock solutions of 10, 18, 32, and 56 mg/10 mL. An aliquot (2.2 mL) of the 10, 18, 32 and 56 mg/10 mL and the 2000 mg/20 mL solvent stock solutions was each added seperately to a final volume of 22 L of dechlorinated tap water, and stirred with the aid of a flat bladed mixer for approx. 1 min. to give the 0.010, 0.18, 0.32, 0.56, and 1.0 mg/L test concentrations, respectively. Each of the stock solutions was inverted several times to ensure adequate mixing and homogeneity.


- Chemical name of vehicle (organic solvent, emulsifier or dispersant): tetrahydrofuran

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Source: Brow Well Fisheries Limited, Hebden, near Skipton, Yorkshire, UK; maintained in-house since 9 February 2009
- Age at study initiation (mean and range, SD): juvenile, ≤ 1 year
- Weight at end of study: 1.82 g ± 0.32 g
- Housing: 14°C water temp.; >= 10.1 mg O2/L dissolved oxygen; 16 h light : 8 hours dark, 20 min of dawn and dsuk transition periods; glass fibre tanks with single pass water renewal systemat least 1L volume per fish; 80% air saturation value; laboratory tap water dechlorinated; commercial trout pellets
- Feeding during test: no

ACCLIMATION
- Acclimation period: 12 days
- Acclimation conditions (same as test or not): yes
- Health during acclimation (any mortality observed): no mortaliyt observed

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

140 mg/L as CaCO3

Test temperature:

14°C

pH:

7.5 - 7.8

Dissolved oxygen:

10.3 - 13.1 mg O2/L

Nominal and measured concentrations:

0.010, 0.018, 0.032, 0.56 and 1.0 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 20 L glas vessels
- Aeration: yes, via narrow bore glass tubes
- 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
- No. of vessels per vehicle control (replicates): 1
- Biomass loading rate: 0.64 g bw/L


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: laboratory tap water, dechlorinated by passage through an activated carbon filter and partly softened
- Total organic carbon: 0.810 - 1.380 (average 1.041) mg/L
- Pesticides: 0.00 µg/L
- Chlorine: 0.080 - 0.610 (average 0.297) mg/L
- Conductivity: 251.000 - 542.000 (average 421.385) µS/cm at 20°C
- Culture medium different from test medium: no
- Intervals of water quality measurement: daily


OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light, 8 hours dark, 20 min transition periods of dusk and dawn




TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0.10 and 1.0 mg/L

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

0.42 mg/L

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL: 0.32-0.56 mg/L

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.32 mg/L

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

0.034 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

dissolved

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL: 0.017-0.40 mg/L

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.017 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

dissolved

Basis for effect:

mortality (fish)

Details on results:

Sublethal effects were observed at a test concentration of 0.066 mg/L. This response was a slightly increased rate of respiration.

Sublethal observations / clinical signs:

Analysis of the freshly prepared 0.32, 0.56 and 1.0 mg/L test concentrations at 0, 24, 48, and 72 hours showed measured test concentrations to range from less than the LOQ to 0.580 mg/L for the untreated samples and less than the LOQ to 0.507 mg/L for the centrifuged samples. A decline in measured concentration was observed in the old media at 24, 48, 72 and 96 hours with measured concentrations in the range of less than the LOQ to 0.465 mg/L for the untreated samples and less than the LOQ to 0.308 mg/L for the centrifuged samples.

Based on the decline in measured test concentrations it was considered justifiable to base the results on the time-weighted mean measured test concentrations of the centrifuged test media to give a "worst case" analysis of the data.

The time-weighted mean measured test concentrations (TWA) for the 0.32, 0.56 and 1.0 mg/L nominal test concentrations were calculated to be: 0.017, 0.066 and 0.40 mg/L. TWAs for the nominal concentrations of 0.10 and 0.18 mg/L were not claculated given that all analyses were less than the the LOQ. This was not considered to impact the study given that mortalities were not observed at these concentrations and that an LC50 value could be calculated from the three calculated TWA concentrations.

Table: Cumulative mortality data in the definitive test

Nominal concentration (mg/L)	Time-weighted mean measured concentration (mg/L)	Cumulative mortality (Initial population = 7)						% Mortality
		3 hours	6 hours	24 hours	48 hours	72 hours	96 hours	96 hours
Control	Control	0	0	0	0	0	0	0
Solvent control	Solvent control	0	0	0	0	0	0	0
0.10	-	0	0	0	0	0	0	0
0.18	-	0	0	0	0	0	0	0
0.32	0.017	0	0	0	0	0	0	0
0.56	0.066	0	0	0	0	4	7	100
1.0	0.40	0	0	7	7	7	7	100

Validity criteria fulfilled:

yes

Conclusions:

The acute toxicity of the test material to the freshwater trout (Oncorhynchus mykiss) has been investigated. The 96-hour LC50 based on nominal test concentrations was 0.42 mg/L.

Executive summary:

A study according to OECD TG 203 was performed to assess the acute toxicity of the test material to rainbow trout (Oncorhynchus mykiss).

Information supplied by the Sponsor indicated that the test material had a low solubility in water. Based on this information the test material fell into the category of a
‘difficult substance’ as defined by the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures (OECD 23, 2000). Therefore a media
preparation trial was conducted in order to determine the solubility of the test material under test conditions.
The pre-study media preparation trial indicated that the use of an auxiliary solvent to spike the test material into the test media followed by centrifugation to remove the
undissolved test material was the most appropriate method of preparation for the test material. Due to the volumes required, the test media was not centrifuged prior to the exposure of the test organisms. The test samples were analysed untreated and following centrifugation at 40000 g for 30 minutes in order to determine the dissolved,
and hence bioavailable, amount of test material.
Following a preliminary range-finding test, fish were exposed, in groups of seven, to an aqueous solution of the test material over a range of concentrations of 0.10, 0.18, 0.32, 0.56 and 1.0 mg/l for a period of 96 hours at a temperature of approximately 14°C under semi-static test conditions. The number of mortalities and any sub-lethal effects of exposure in each test and control vesse! were determined 3 and 6 hours after the start of exposure and then daily throughout the test until termination after 96 hours.
The 96-Hour LC50 based on nominal test concentrations was 0.42 mg/l with 95% confidence limits of 0.32 - 0.56 mg/l.

Analysis of the nominal 0.10 and 0.18 mg/l test concentrations showed measured concentrations in the untreated and centrifuged samples to be less than the limit of
quantitation (LOQ) of the analytical method throughout the test.
Analysis of the freshly prepared 0.32, 0.56 and 1.0 mg/l test concentrations at 0, 24, 48 and 72 hours showed measured test concentrations to range from less than the LOQ to 0.580 mg/l for the untreated samples and less than the LOQ to 0.507 mg/l for the centrifuged samples. A decline in measured concentration was observed in the old media at 24, 48, 72 and 96 hours with measured concentrations in the range of less than the LOQ to 0.465 mg/l for the untreated samples and less than the LOQ to 0.308 mg/l for the centrifuged samples. The decline in measured concentrations observed over the test period was in line with the preliminary stability analyses conducted.

However, effect levels based on mean measured concentrations don't represent the real environmental conditions because the test substance is rapidly oxidized by the oxygen content in the aqueous phase. Therefore, nominal values are used for hazard assessment.

Description of key information

LC50 (96h) = 0.42 mg/L (nominal)(Oncorhynchus mykiss)

 

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

0.42 mg/L

Additional information

A study according to OECD TG 203 was performed to assess the acute toxicity of the test material to rainbow trout (Oncorhynchus mykiss).

Information supplied by the Sponsor indicated that the test material had a low solubility in water. Based on this information the test material fell into the category of a
‘difficult substance’ as defined by the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures (OECD 23, 2000). Therefore a media
preparation trial was conducted in order to determine the solubility of the test material under test conditions.
The pre-study media preparation trial indicated that the use of an auxiliary solvent to spike the test material into the test media followed by centrifugation to remove the
undissolved test material was the most appropriate method of preparation for the test material. Due to the volumes required, the test media was not centrifuged prior to the exposure of the test organisms. The test samples were analysed untreated and following centrifugation at 40000 g for 30 minutes in order to determine the dissolved,
and hence bioavailable, amount of test material.
Following a preliminary range-finding test, fish were exposed, in groups of seven, to an aqueous solution of the test material over a range of concentrations of 0.10, 0.18, 0.32, 0.56 and 1.0 mg/l for a period of 96 hours at a temperature of approximately 14°C under semi-static test conditions. The number of mortalities and any sub-lethal effects of exposure in each test and control vesse! were determined 3 and 6 hours after the start of exposure and then daily throughout the test until termination after 96 hours.
The 96-Hour LC50 based on nominal test concentrations was 0.42 mg/l with 95% confidence limits of 0.32 - 0.56 mg/l.

Analysis of the nominal 0.10 and 0.18 mg/l test concentrations showed measured concentrations in the untreated and centrifuged samples to be less than the limit of
quantitation (LOQ) of the analytical method throughout the test.
Analysis of the freshly prepared 0.32, 0.56 and 1.0 mg/l test concentrations at 0, 24, 48 and 72 hours showed measured test concentrations to range from less than the LOQ to 0.580 mg/l for the untreated samples and less than the LOQ to 0.507 mg/l for the centrifuged samples. A decline in measured concentration was observed in the old media at 24, 48, 72 and 96 hours with measured concentrations in the range of less than the LOQ to 0.465 mg/l for the untreated samples and less than the LOQ to 0.308 mg/l for the centrifuged samples. The decline in measured concentrations observed over the test period was in line with the preliminary stability analyses conducted.

However, effect levels based on mean measured concentrations don't represent the real environmental conditions because the test substance is rapidly oxidized by the oxygen content in the aqueous phase. Therefore, nominal values are used for hazard assessment.