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Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
The definitive test was performed with concentrations of 0 (control), 6.3, 13, 25, 50, and 100 % of an aged tin(IV) chloride pentahydrate solution prepared at the target nominal loading rate of 1.0 mg Sn/L. The prepared tin(IV) chloride solution were aged for three days, filtered using a 0.45 µm filter, and aged for an additional four days prior to use.

Details on properties of test surrogate or analogue material:
Tin (IV) chloride pentahydrate 98.9 % purity
Impurities:
Fe: <6ppm
K: <40ppm
Na: <40ppm
Pb: < 5ppm
SO4: <200ppm
Analytical monitoring:
yes
Details on sampling:
The concentration of dissolved and total tin was measured in the control and all test substance treatments at initiation (0 hour) and termination (96 hours) of the definitive test. MSSN-10PPM fortified samples (i.e., QC samples) were also prepared for analysis at each sample period. The analyses were completed on 27 September 2010.
Appropriate sample volumes (9.8 mL for unfiltered samples and 14 mL for filtered samples) were collected from the control and each test substance solution. To each 9.8 mL sample, 0.2 mL of concentrated nitric acid was added and the solution was mixed. Each 14 mL sample was filtered through a 0.45 µm Acrodisc 25 mm syringe filter. 9.8 mL of each filtered sample was then diluted with 0.2 mL of concentrated nitric acid and mixed. QC samples were prepared in a similar manner by fortifying 10-mL volumes of 2 % nitric acid in dilution medium with concentrations of 0.612, 1.02, and 86.7 ng Sn/mL. The samples were then mixed prior to analysis by ICP MS.
Vehicle:
no
Details on test solutions:
Not applicable
Test organisms (species):
Pimephales promelas
Details on test organisms:
Fathead minnows (Pimephales promelas) were obtained from an in-house culture. The fish while in culture were fed brine shrimp and/or a commercially available fish food twice each day. Periodic lots of the fish food are screened for contaminants. Records of fish food suppliers and the results of contaminant analyses are on file at ABC Laboratories, Inc. A daily record of fish observations during the culture period, along with any prophylactic or therapeutic disease treatments, was maintained. Fathead minnows were maintained in the dilution water for at least seven days and acclimated to the test temperature for at least 48 hours before the definitive test initiation. There was no mortality two weeks prior to initiation of the study. Food was withheld for 24 hours immediately before study initiation and no mortalities were observed. A representative group of acclimated fish was measured at initiation. The fish ranged from 17 to 22 mm in total length (mean and standard deviation = 19 ± 1.8 mm), and from 0.0456 to 0.1112 g in blotted wet weight (mean and standard deviation = 0.0769 ± 0.0253 g). The instantaneous biomass loading rate did not exceed 0.269 g of fish tissue per liter of test solution in the control.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
Not applicable
Hardness:
The total alkalinity and total hardness of the dilution water at test initiation were 158 and 146 mg CaCO3/L, respectively. Total organic carbon (TOC) of the dilution water was less than 2.0 mg/L as measured in the monthly facility records.
Test temperature:
Test solution temperatures during the 96-hour exposure ranged from 21.6 to 22.0 °C as measured in the individual test replicates
pH:
Test solution pH ranged from 6.9 to 8.6 throughout the test
Dissolved oxygen:
Dissolved oxygen varied between a minimum of 6.3 mg/L and a maximum of 8.2 mg/L throughout the test
Salinity:
The conductivity of the dilution water was 337 µS/cm.
Nominal and measured concentrations:
Nominal concentrations were 0 (control), 6.3, 13, 25, 50, and 100 % of an aged tin(IV) chloride pentahydrate solution prepared from the target nominal loading rate of 1.0 mg Sn/L. The prepared tin(IV) chloride solution were aged for three days, filtered using a 0.45 µm filter, and aged for an additional four days prior to use.
Details on test conditions:
The definitive test was conducted in 3.8-L glass jars containing approximately 2 L of control or test solution. All test chambers were covered with plastic Petri dishes. The fish were impartially added one at a time proceeding from the control to the high test substance treatment, and repeating these steps until seven fish had been distributed to each test substance treatment.
A 1.0 mg Sn/L (3.0 mg tin(IV) chloride pentahydrate/L) primary solution was prepared by transferring 12.1318 mg of tin(IV) chloride pentahydrate to a 4-L volume of blended freshwater in a 5-L glass carboy. The resulting solution was allowed to sit, undisturbed under ambient laboratory conditions for three days. After three days, the entire solution was filtered into a second 5-L carboy using a 0.45-μm cellulose filter. The filtered solution was allowed to age under laboratory conditions for an additional four days. After a total of 7 days aging, the primary solution was used as the highest test treatment (100 %) and aliquots of the primary solution were diluted to a volume of 2-L to prepare the four lower-level test treatments of 6.5, 13, 25, and 50%. The control solution consisted of dilution water only.
Observations for mortality and sublethal responses (e.g., discoloration, loss of equilibrium, animals lying on the bottom of the test chamber, irregular respiration, etc.) were made once every 24 hours (± 1 hour) for the remainder of the test.
Reference substance (positive control):
not required
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 12.4 µg/L
Nominal / measured:
meas. (initial)
Conc. based on:
element
Basis for effect:
mortality
Remarks on result:
other: Unable to calculate 95 % CI as no effects observed
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
12.4 µg/L
Nominal / measured:
meas. (initial)
Conc. based on:
element
Basis for effect:
mortality
Details on results:
There were no sublethal effects noted in the control or any treatments during the test. The control and test subtance solutions were clear and colourless with no visible signs undissolved test substance, precipiate or surface film throughout the study.
Results with reference substance (positive control):
Not applicable

Table 3.  Mortality of Fathead Minnow Pimephales promelas during a 96-Hour static exposure to aged solutions of Tin(IV) Chloride Pentahydrate

Inital Measured

Total Tin

(µg Sn/L)

Cumulative Number Dead (Percent Mortality)

24 Hours

48 Hours

72 Hours

96 Hours

Treatment Mean

(96 Hours)

0 (control)

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

0.483

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

1.21

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

2.49

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

5.63

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

12.4

0 (0)

0 (0)

0 (0)

0 (0)

0 (0)

Notes: Seven fish were placed into each test treatment at initiation. No sublethal effects were observed

Validity criteria fulfilled:
yes
Conclusions:
No toxicity or sublethal effects observed on tin (IV) solutions aged for 7 days at commencement of test.
Executive summary:

The conditions used in this test have been developed following difficulties encountered during earlier toxicity tests with precipitation of tin hydroxides. The objective of this and the other key aquatic toxicity tests is to assess any adverse effects from soluble tin released from tin metal. Attempts to investigate the potential toxicity from tin ions using salts such as tin (IV) chloride have been confounded by lack of stability of tin ions in solution. This instability produces dissolved tin concentrations of only 10 to 20 µg/L against a nominal total tin concentration of 1 mg/L. The high concentrations of precipitated tin hydroxides and the associated pH effects at higher doses do not reflect the true behaviour of tin ions released from the transformation and dissolution of tin metal. Therefore, to overcome these difficulties, aged solutions containing tin concentrations approaching a steady state condition have been prepared just prior to the testing. The maximum measured concentration of tin this test achieved after ageing is lower than the ABC 2010 growth inhibition test with Pseudokirchneriella subcapitata with an identical ageing protocol. This may be due to the effects of the specific water chemistry of the the different test media on the speed and degree of precipitation of tin (IV) from solution.

Description of key information

No mortality or sub-lethal effects were observed on Pimephales promelas at all strengths of aged solutions of tin (IV) chloride including at 100 %. This test is interpreted as demonstrating that acute effects will not be observed at concentrations of soluble tin that could arise from the dissolution of tin metal in the environment.

Key value for chemical safety assessment

Additional information

The objective of the Parametrix 2009 study was to determine the toxicity of tin (IV) ions to the fathead minnow Pimephales promelas under a variety of test conditions. Tests were run at two pH, each with five nominal loading rates. In addition, after preparation of the stock solution the test media was split in half, and one half used unfiltered, whilst the other was filtered before the addition of the test fish. Tin (IV) chloride was used to supply tin ions for the test and as a surrogate for tin ions released on the dissolution of tin metal. However, as in other preliminary toxicity tests using tin (IV) chloride the test results are difficult to interpret due to hydrolysis and precipitation effects which proceed over a matter of hours and days. This results in tests being undertaken in unstable conditions and makes it difficult to distinguish whether effects are due to soluble or particulate tin species.

For example, at the highest concentration treatment in the pH 8.5 filtered test, the average dissolved tin was 17.5 µg/L whereas the total measured tin was 828 µg/L. This was only concentration at which a survival effect was noted in the pH 8.5 filtered category. This study also has the added complication of uncertain analytical data with contamination effects observed possibly due to insufficient flushing of the analytical instrument between assays.

Due to these issues, the Parametrix 2009 study was used to inform the design of the key fat head minnow acute toxicity test (ABC 2010) but is not relevant for use in the aquatic toxicity assessment of tin metal.

The definitive short term toxicity fish test (ABC 2010) was undertaken in October 2010. The conditions used in this test have been developed following the difficulties encountered during earlier toxicity tests with precipitation of tin hydroxides. Earlier attempts to investigate the potential toxicity from tin ions using salts such as tin (IV) chloride have been confounded by lack of stability of tin ions in solution and consequential precipitation. This instability produces dissolved tin concentrations of 10 to 20 µg/L against a nominal tin concentration of 1 mg/L. The high concentrations of precipitated tin hydroxides and the associated low pH effects at higher doses do not reflect the true behaviour of tin ions released from the transformation and dissolution of tin metal.

The objective of this and the other key aquatic toxicity tests is to assess any adverse effects from soluble tin released from tin metal and not tin (IV) chloride. Therefore, to overcome these difficulties, aged solutions containing more stable tin concentrations have been prepared just prior to the testing. However, even after a total of seven days of ageing the tin (IV) ions are not yet at equilibrium conditions as slow precipitation and mineralisation reactions continue to take place.However, the aged solutions are considered to represent the maximum dissolved tin that can be achieved in the test media, and therefore a worst case exposure to dissolved tin.

No mortality or sub-lethal effects were observed on Pimephales promelas at all strengths of aged solutions of tin (IV) chloride including at 100 %. This test is interpreted as demonstrating that acute effects will not be observed at concentrations of soluble tin that could arise from the dissolution of tin metal in the environment.