Chlorotrioctylstannane

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:

01 June 2004 to 16 June 2004

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:

other: ISO Standard 7346-2: "Water quality - Determination of the acute lethal toxicity of substances to a freshwater fish [Brachydanio rerio Hamilton-Buchanan (Teleostei, Cyprinidae)]" - Part 2: Semi-static method

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Physical appearance: Slightly yellow liquid
- Storage conditions of test material: In the freezer in the dark
- Stability under storage conditions: Stable
- Density: 1.04 g/cm³
- Expiry date: 31 December 2004

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Duplicate samples for analysis were taken from the control and the WAF prepared at a loading rate of 100 mg/L. Additionally, singular reserve samples of 100 mL were taken from the control and the WAF prepared at 100 mg/L for possible analysis.
- Sampling method: At t = 0 h and t = 72 h samples were taken from freshly prepared solutions and at t = 24 h and t = 96 h samples were taken from 24-hour old solutions. 100 mL was taken from the approximate centre of the test vessels.
- Sample storage conditions before analysis: Samples were stored at room temperature for a maximum of 10 days until transportation to the testing facility for analysis. If not already used, the reserve samples were stored at room temperature for possible analysis for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test material was not soluble in test medium at the concentrations tested. Preparation of test solutions started with two dispersions of the stock solution in test medium at a nominal loading of 100 mg/L. The dispersions were magnetically stirred for approximately 24 hours in two volumetric flasks (volumes of 2 and 5 litres), after which they were left to stabilise for another 24 hours. The water fractions were then separated from the undissolved fraction of test material and both transferred to one 10-litre separation funnel by means of siphoning. The Water Accommodated Fraction (WAF) collected from the separatory funnel was used for testing. The lower test concentrations were prepared by subsequent dilution of the WAF in test medium. All test vessels were pre-conditioned for 30 minutes with approximately 1 litre of the corresponding test concentrations prior to the start of the test. Test solutions were renewed daily.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Species: Brachydanio rerio, Teleostei, Cyprinidae
- Common name: Zebra-fish
- Mean length: 2.8 ± 0.2 cm. Ten fish of the batch used for the test were measured prior to the start of the test.
- Mean weight: 0.42 ± 0.14 g. Ten fish of the batch used for the test were weighed prior to the start of the test.
- Feeding during test: No

ACCLIMATION
- Acclimation period: At least 12 days after delivery.
- Acclimation conditions (same as test or not): Yes, fish were held in ISO-medium (ISO International standard 734).
- Type of food: Trouvit
- Feeding frequency: Daily. Fish were not fed from 48 hours prior to the test.
- Health during acclimation (any mortality observed): In the batch of fish used for the test, mortality during the seven days prior to the start of the test was less than 5 %.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

250 mg CaCO3 per litre

Test temperature:

21.9 to 20.9 °C

pH:

7.2 to 7.9

Dissolved oxygen:

7.0 to 9.9 mg/L

Nominal and measured concentrations:

- Nominal test concentrations: A WAF prepared at a loading rate of 100 mg/L and dilutions containing 0.1, 1.0 and 10 % of the WAF were prepared.
- Measured (100 % WAF): Average of 84 µg/L over t = 0 to 96 h

Details on test conditions:

TEST SYSTEM
- Test vessel: 6.52 litres, all-glass
- Fill volume: 4 litres
- Aeration: The test media were not aerated during the test.
- Renewal rate of test solution (frequency/flow rate): Semi-static with daily renewal of test solutions
- No. of organisms per vessel: Control and WAF: 7 fish per concentration; WAF dilutions: 3 fish per concentration
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: Control and WAF: 0.74 g fish/litre, i.e. 7 fish per 4 litres of test medium; WAF dilutions: 0.32 g fish/litre, i.e. 3 fish per 4 litres of test medium.
- Euthanasia: At the end of the test the surviving fish were rapidly killed by exposing them to ca. 1.2% ethylene glycol monophenylether in water.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO-medium (ISO International Standard 7346), aerated until the dissolved oxygen concentration had reached saturation and the pH had stabilised. After aeration the hardness was and the pH was 7.9. The medium contains 293.8 mg/L CaCl2.2H2O, 123.3 mg/L MgSO4.7H2O, 64.8 mg/L NaHCO3 and 5.8 mg/L KCl dissolved in tap water purified by reverse osmosis. Fish were introduced within one hour after preparation of the test media.
- Culture medium different from test medium: No
- Intervals of water quality measurement: Dissolved oxygen content, pH and temperature were measured daily in all vessels, beginning at the start of the test (day 0).

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hour photoperiod daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
Mortality and other effects were evaluated at 3, 24, 48, 72 and 96 hours following the start of exposure.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Range finding study: No, the test was conducted as a combined limit/range finder.

Reference substance (positive control):

yes

Remarks:

Pentachlorophenol (PCP) 0.10, 0.22 and 0.46 mg/L in ISO-medium.

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 0.084 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

No mortality or visible clinical effects were observed in any of the solutions during the 96-hour test period. All test solutions were clear and colourless during the entire 96-hour test period when visually assessed.

DETERMINATION OF AVERAGE EXPOSURE CONCENTRATIONS
The average exposure concentrations for the periods between refreshment were calculated as (Ct = 0*Ct= 24)^1/2, being the geometric means of the mean concentrations of the test material measured in the duplicate samples taken at the start (Ct = 0) and the end of the 24-hour refreshment periods (Ct = 24). This was applied for the periods 0 to 24 hours and 72 to 96 hours of exposure.
The (rounded) arithmetic mean of the geometric mean exposure concentrations determined during these two refreshment periods was taken as the average exposure concentration for the total 96-hour test period.

MEASURED CONCENTRATIONS
The measured concentrations decreased by 25 or 50 % (first and last renewal period, respectively) relative to the initial measured concentrations during the 24-hour renewal periods. Analyses showed good repeatability of test solution preparation. The observed decrease in test concentrations was not unexpected considering the extremely low solubility of the test material in the test medium in combination with the difficulties encountered with the separation of the undissolved from the dissolved fraction, which is a common feature when testing WAFs. In addition, adsorption to organic matter or surface areas (i.e. glass walls) might have contributed to the observed decrease.
In some of the control samples Sn was measured in largely varying concentrations between duplicate samples. It is however most unlikely that the Sn measured originates from the test material. This is indicated by the fact that the measured concentrations in the 24-hour old solutions were generally higher than in the freshly prepared solution in combination with the large differences between duplicate measurements. Hence, measured concentrations at these low levels may not be firmly reliable. The origin of the measured Sn is unclear. Even if Sn originated from the test material, this is not considered to invalidate the test result, since no effects were observed in the control and/or in the much higher limit concentration tested.
An overview of the individual measurements for the control and the mean concentrations measured in the WAF (including the average exposure concentrations) is presented in Table 1.

Results with reference substance (positive control):

Under the conditions of the present test with zebra-fish exposed to PCP, the 96 hour LC50 was estimated to correspond to 0.11 mg/L based on nominal concentrations, with 20 % mortality at 0.10 mg/L and 100 % mortality at 0.22 mg/L. This effect was already reached within 48 hours of exposure.
The range of the 96 hour LC50 for zebra-fish is generally between 0.10 and 0.46 mg/L based on historical data of reference tests performed. Hence, the sensitivity of zebra-fish originating from the present batch for PCP is relatively high in comparison to the sensitivity generally observed previously.

Sublethal observations / clinical signs:

Table 1: Mean measured test material concentrations and average exposure concentrations

% of WAF at 100 mg/L	Test Material Concentration (µg/L)
	t = 0 h	t = 24 h	Average (t = 0 - 24 h)	t = 72 h	t = 96 h	Average (t = 72 - 96 h)	Average (t = 0 - 96 h)
0	0.10	32	N/A	0.10	17	N/A	N/A
	<0.05	0.52		0.08	<0.05
100	114	85	98	99	50	70	84

  

Table 2: Incidence of mortality and total mortality

Test Group	Initial number of Fish	Cumulative Mortality					Total Mortality (%)
		3 hours	24 hours	48 hours	72 hours	96 hours
Blank (Control)	7	0	0	0	0	0	0
0.1*	3	0	0	0	0	0	0
1.0*	3	0	0	0	0	0	0
10*	3	0	0	0	0	0	0
100*	7	0	0	0	0	0	0

* Test Groups represent percentages of a WAF prepared at 100 mg/L

 

Table 3: Effect parameters

Parameter	Loading rate (% of a WAF at a loading rate of 100 mg/L)	Concentration of Test Material (mg/L)
NOEC	100	0.084
96 h LC0	100	0.084
96 h LC50	>100	>0.084
96 h LC100	>100	>0.084

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the study, the 96 hour LC50 was determined to be >0.084mg/L.

Executive summary:

The acute toxicity of the test material to zebra-fish (Brachydanio rerio) was investigated in a study which was conducted in accordance with the standardised guidelines OECD 203, EU Method C.1 and ISO 7346-2 Part 2 under GLP conditions.

A limit test was combined with a range-finding test in a semi-static system with daily renewal. Seven zebra-fish per test group were exposed to a blank-control and a Water Accommodated Fraction (WAF) prepared at a test material loading rate of 100 mg/L in the limit test, while three zebra-fish per test group were exposed to dilutions containing 0.1, 1.0 and 10 % of the WAF in the additional range-finding test.

Samples for analytical confirmation of actual exposure concentrations were taken from freshly prepared solutions of the blank-control and the undiluted WAF at t = 0 h and t = 72 h and from the 24-hour old solutions at t = 24 h and t = 96 h.

The test material induced no visible effects in zebra-fish at or below a WAF prepared at a loading rate of 100 mg/L.

Analysis showed that the measured concentrations decreased by 25 or 50 % (first and last renewal period, respectively) relative to the initial measured concentrations during the 24-hour renewal periods. Analyses showed good repeatability of test solution preparation. The study met the acceptability criteria prescribed by the guidelines and was considered valid.

Based on average measured concentrations, the NOEC corresponded to 0.084 mg of test material per litre. Consequently, the 96 hour LC50 for zebra-fish exposed to the test material exceeded an average exposure concentration of 0.084 mg/L.

Description of key information

96 hour LC50 >0.084 mg/L (zebra fish), OECD 203, EU Method C.1 and ISO 7346-2 Part 2, Migchielsen & de Haan (2004)

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

0.084 mg/L

Additional information

The acute toxicity of the test material to zebra-fish (Brachydanio rerio) was investigated in a study which was conducted in accordance with the standardised guidelines OECD 203, EU Method C.1 and ISO 7346-2 Part 2 under GLP conditions. The study was awarded a reliability score of 1 in accordance with the principles for assessing data quality set forth by Klimisch et al. (1997).

A limit test was combined with a range-finding test in a semi-static system with daily renewal. Seven zebra-fish per test group were exposed to a blank-control and a Water Accommodated Fraction (WAF) prepared at a test material loading rate of 100 mg/L in the limit test, while three zebra-fish per test group were exposed to dilutions containing 0.1, 1.0 and 10 % of the WAF in the additional range-finding test.

Samples for analytical confirmation of actual exposure concentrations were taken from freshly prepared solutions of the blank-control and the undiluted WAF at t = 0 h and t = 72 h and from the 24-hour old solutions at t = 24 h and t = 96 h.

The test material induced no visible effects in zebra-fish at or below a WAF prepared at a loading rate of 100 mg/L.

Analysis showed that the measured concentrations decreased by 25 or 50 % (first and last renewal period, respectively) relative to the initial measured concentrations during the 24-hour renewal periods. Analyses showed good repeatability of test solution preparation. The study met the acceptability criteria prescribed by the guidelines and was considered valid.

Based on average measured concentrations, the NOEC corresponded to 0.084 mg of test material per litre. Consequently, the 96 hour LC50 for zebra-fish exposed to the test material exceeded an average exposure concentration of 0.084 mg/L.