[1,2,4]Triazolo[1,5-c]pyrimidine, 2,2'-dithiobis[5-ethoxy-7-fluoro-

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29 November 1993 to 3 December 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted to GLP in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. No analytical determination of test substance concentration.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: EPA OTS 797.1440 (Fish Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

5-ethoxy-2-({5-ethoxy-7-fluoro-[1,2,4]triazolo[1,5-c]pyrimidin-2-yl}disulfanyl)-7-fluoro-[1,2,4]triazolo[1,5-c]pyrimidine

EC Number:

695-187-4

Cas Number:

166524-75-0

Molecular formula:

C14H12F2N8O2S2

IUPAC Name:

5-ethoxy-2-({5-ethoxy-7-fluoro-[1,2,4]triazolo[1,5-c]pyrimidin-2-yl}disulfanyl)-7-fluoro-[1,2,4]triazolo[1,5-c]pyrimidine

Test material form:

other: solid (unspecified)

Details on test material:

- Appearance: light tan/yellow solid

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The 15 L of water in treated tanks were dosed by the addition of 3.483, 3.574, 2.297, 2.236, 1.120, 1.138, 0.528, 0.562, 0.279 and 0.282 grams of test material dissolved in 7 mL of acetone. No correction for the test material purity was made, resulting in nominal test concentrations of 235, 151, 75.5, 36 and 19 mg/L

Test organisms

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow trout
- Rearing and acclimation: eggs were fertilised then hatched, acclimated, and reared in a temperature-controlled environment having a 16 hour light / 8 hour dark photoperiod with approximately 15 minutes dusk and dawn transitions. The fish were observed for at least 14 days before testing. During the holding period, the fish received a standard laboratory diet at least once daily. The test organisms used for the definitive test were acclimated to 12 ± 2 °C and held without food for at least 24 hours before testing. Mortality of the test lot was less than 0.1 % per day in the 14 days prior to testing.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test conditions

Test temperature:

12.6 - 13.3 °C

pH:

7.4 - 7.8

Dissolved oxygen:

7.7 - 8.8 mg/L (greater than or equal to 74 % of saturation)

Nominal and measured concentrations:

0 (acetone and water controls), 19, 36, 75.5, 151 and 235 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 18 L borosilicate glass aquaria (approximately 20 cm wide x 39.5 cm long x 24.5 cm deep)
- Type: closed (with glass or plexiglass covers to retard evaporation and prevent fish escaping)
- Fill volume: 15 L
- Aeration: none
- No. of organisms per vessel: 10 (except for 151 mg/L treatment level which was inadvertently set with 9 fish in the first replicate)
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2
- Biomass loading rate: 0.33 g fish/ L test solution

DILUTION WATER: softened Lake Huron water
- Supplier: the City of Midland Water Treatment Plant
- Source: the upper Saginaw Bay of Lake Huron off Whitestone Point
- Pre-treatment of water: the water was limed and flocculated with ferric chloride prior to arrival at the test facility. The water was subsequently sand-filtered, pH adjusted with CO2, carbon-filtered, passed through 5 µm filters and UV irradiated in the laboratory before use.
- Water analysis: the water was monitored weekly for alkalinity, total chlorine, conductivity, hardness and pH. Analyses were conducted three times per year for total organic carbon, total suspended solids and selected inorganic and organic chemicals.
> Analysis of dilution water:
- Hardness (as CaCO3): 70 mg/L
- Alkalinity (as CaCO3): 49 mg/L
- pH: 7.8
- Conductivity: 250 µmhos/cm
- Chlorine: 4.0 ppb

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light / 8 hours dark with 15 minute dawn and dusk transitions
- Light intensity: approximately 1025 lux

EFFECT PARAMETERS MEASURED
The fish were observed daily for mortality (no response to gentle prodding and no opercular movement) and sublethal effects e.g. swimming at the surface, complete or partial loss of body equilibrium, lethargy, erratic movement and exophthalmia. Dead fish, if any, were removed when observed.
All surviving test organisms were euthanised at test termination using tricaine methanesulfonate. Weight and standard length measurements were made on the laboratory water control fish at test termination.

WATER PARAMETERS MEASURED
Measurements of dissolved oxygen, pH and temperature were made at 2, 48 and 96 hours after initiation of testing in all test and control vessels.

TEST CONCENTRATIONS
- Justification for test concentrations: test concentrations were based on preliminary estimates of test material toxicity which were obtained using the USEPA Structure Activity Relationship (SAR) computer program (Version 1.01, 1993).

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 235 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

No fish died at any concentration level at any time during the test. Only one fish exhibited adverse effects. This fish was in the 151 mg/L treatment. Since the adverse effects observed for that one fish were not seen in the 151 mg/L replicate or 235 mg/L treatment replicates it is unlikely that the effect was treatment related.

Reported statistics and error estimates:

The LC50 was calculated using probit analysis and moving average methods. Furthermore, a point estimate of the LC50 was obtained using non-linear interpolation, i.e. log transformation of the concentration and angle transformation of the number dead.

Any other information on results incl. tables

Sublethal observations / clinical signs:

Length and weight of control fish at test termination:

- Mean standard length (range): 33.6 mm (30 - 38 mm)

- Mean weight: 0.500 g

Observations of test solutions:

The test material had not completely dissolved at 24 hours in all dose levels.

Applicant's summary and conclusion

Validity criteria fulfilled:

not specified

Conclusions:

Under the conditions of this study, the LC50 was determined to be in excess of 235 mg/L.

Executive summary:

The acute toxicity of the test material to fish was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guideline EPA OTS 797.1440.

During the 96 hour static study, groups of 20 rainbow trout were exposed to 19, 36, 75.5 and 235 mg/L of test material; a group of 19 fish was exposed to 151 mg/L test material. Further groups of 20 fish were exposed to acetone and dilution water controls. Mortality and sublethal effects were observed daily.

None of the fish died during the course of the study. Under the conditions of this study, the LC50 was determined to be in excess of 235 mg/L, based on nominal test concentrations.