Dibutyl [(dipropoxyphosphinothioyl)thio]succinate

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:

17 November 2017 to 16 December 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes

Specific details on test material used for the study:

-Description: Yellow, clear liquid
- Storage: room temperature, in the dark

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control and 10 mg/L loading rate WAF.
- Sampling method: Individual samples were collected by pipet at 0, 24, 48, and 72 Hour (new) and 24, 48, 72, and 96 Hour (aged) from the approximate midpoint from the surface, bottom, and sides of the vessel
- Sample storage conditions before analysis: Analyzed upon collection. Subsamples also were stored frozen, but later determined to not be sufficiently stable under the storage conditions.
Quality Control (QC) Samples: Three samples per sampling interval were prepared in dilution water at nominal concentrations approximating the test concentration range.
- Sample processing: extraction in hexane with acetone rinse; extracts reconstituted in 50/50 acetonitrile/purified reagent water (v/v), then diluted into the calibration standard range with 50/50 acetonitrile/purified reagent water (v/v).

Vehicle:

no

Details on test solutions:

The treated solution was prepared as a water accommodated fraction (WAF) by adding the aliquot of weighed test substance, without correction for purity, directly to the total volume of dilution water in a glass and silicone aquarium to achieve a nominal loading rate of 10 mg/L. The solution was mixed slowly overnight for approximately 19.5 hours with a magnetic stir plate and Teflon covered stir bar. After mixing, the solution was allowed to settle for approximately 30 minutes. The solution was observed to be clear and colorless with undissolved oily globules of the test substance visible on the surface. The solution was then siphoned into the test vessel, avoiding the solution’s surface. The resulting solution was observed to be clear and colorless with no visible undissolved test substance. This solution was used as the treatment level for the limit test exposure. A control aquarium was also established containing dilution water and no test substance and was maintained under the same conditions as the treatment level solution.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Source: Troutlodge, Inc. (Sumner, Washington, USA)
- Length at study initiation: 42 mm (40 to 45 mm)
- Weight at study initiation: 0.96 g (0.77 to 1.2 g)

ACCLIMATION
- Acclimation period: at least 12 days
- Acclimation conditions: same as test conditions
- Type and amount of food during acclimation: Fish were fed Trout Chow at least once daily until 24 hours prior to exposure initiation
- Health during acclimation (any mortality observed): 0.54% mortality observed in the stock batch of fish in the seven days prior to test initiation.

FEEDING DURING TEST
The fish were not fed during the 24 hours before the start of the definitive test and were not fed throughout the test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

82 mg/L as CaCO3

Test temperature:

14 +/- 1˚C

pH:

6.9 – 7.1

Dissolved oxygen:

7.4 – 11 mg/L (>60% saturation)

Salinity:

not applicable

Conductivity:

680 µS/cm

Nominal and measured concentrations:

Nominal: 0 (control) and 10 mg/L loading rate WAF
Geometric mean measured:

Details on test conditions:

TEST SYSTEM
- Test vessel: 19.5-L glass aquaria (39x20x25 cm)
- Volume of test media: 15 L
- Aeration: none
- 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): not applicable
- Biomass loading rate: 0.45 g/L per day

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Laboratory well water from a 100-meter bedrock well supplemented with dechlorinated Town of Wareham well water
- Total organic carbon (TOC): 0.92 mg/L (March 2018)
- Culture medium different from test medium: same
- Intervals of water quality measurement: 0, 24 (new and aged solutions), 48 (new and aged solutions), 72 (new and aged solutions), and 96 hours; total organic carbon (TOC) is analyzed monthly.

OTHER TEST CONDITIONS
- Adjustment of pH: none
- Photoperiod: 16 hour light: 8 hours dark lighting cycle with a 30 minute transition period
- Light intensity: 65 to 82 footcandles (700 to 880 lux) measured with a Fisher Scientific traceable light meter

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): The biological observations, number of mortalities, observation of morbidity, and observations of the physical characteristics of the exposure and control solutions were recorded at 0, 24, 48, 72, and 96 hours of exposure, if applicable. The concentrations tested and the corresponding mortality data derived from the definitive toxicity exposure were used to estimate the 24-, 48-, 72-, and 96-hour median lethal loading rate or concentration (LL50 or LC50) and No-Observed-Effect Loading rate or Concentration (NOEL or NOEC).

TEST CONCENTRATIONS
Due to the very low solubility of the test substance and based on the results of a preliminary exposure in which no effects were observed, the definitive exposure was conducted at a limit/threshold concentration of 10 mg/L loading rate as a water-accommodated fraction (WAF) solution. Testing at a higher concentration was not considered necessary since additional testing with other freshwater organisms yielded greater sensitivity. As a conservative measure, the test design was adjusted from a 48-hour solution renewal scheme in the preliminary testing to a daily solution renewal test design to maintain the test concentration in the definitive exposure.

Key result

Duration:

96 h

Dose descriptor:

EL50

Effect conc.:

> 10 other: mg/L loading rate WAF

Nominal / measured:

nominal

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

0.014 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

act. ingr. (dissolved fraction)

Basis for effect:

mortality (fish)

Details on results:

- Other biological observations: none
- Mortality of control: none
- Other adverse effects control: none
- Abnormal responses: none
- Any observations that might cause a difference between measured and nominal values: Analysis of the freshly prepared and aged exposure solutions resulted in test substance concentrations ranging from 0.0093 to 0.033 mg a.i./L. Solution concentrations decreased by approximately 10 to 67% over each 24-hour period during the exposure. The most frequent renewal scheme (daily) was used for this exposure to maintain test concentrations as much as possible. Analysis of the QC samples resulted in measured concentrations which were consistent with the predetermined recovery range. The biological endpoint results are based on nominal loading rate WAF concentrations and geometric mean measured concentrations.
- Effect concentrations exceeding solubility of substance in test medium: yes

Reported statistics and error estimates:

Statistical analysis was not performed given that no mortality and no sublethal effects were observed during the test. The LL50 and LC50 values, no observed effect loading rate, NOEL (or concentration, NOEC) were estimated empirically by observation of the data.

Sublethal observations / clinical signs:

Mortality of fish recorded during the definitive test

Nominal Concentration	Geometric Mean Measured Concentration	Cumulative number of mortalities recorded*
(mg/L loading rate WAF)	(mg a.i./L)	24 hours	48 hours	72 hours	96 hours
Control	0	0	0	0	0
10	0.014	0	0	0	0

* Presented as percent of mortality (actual number of mortalities).

Validity criteria fulfilled:

yes

Conclusions:

The test showed no toxicity to rainbow trout (Oncorhynchus mykiss) at the limit of solubility of the test substance in the test media (0.014 mg a.i./L based on mean measured concentration). The 96-hour LL50 was determined to be greater than 10 mg/L loading rate WAF.

Executive summary:

The study was conducted in accordance with the requirements of OECD Chemicals Testing Guideline No. 203 to assess the toxicity the 96-hour acute toxicity of the substance to rainbow trout (Oncorhynchus mykiss). Test solutions were prepared as Water Accommodated Fractions (WAFs) based on the recommendations of OECD Series on Testing and Assessment, No. 23. 

The definitive test was conducted as a ‘Limit Test’ under semi-static conditions (daily renewal) at a single nominal loading rate of 10 mg/L with a concurrent control group, each consisting of a single 19.5 L constructed glass aquaria containing 15 L of media. Following acclimation to the test conditions for at least 12 days, seven fish were added to each test vessel. TheDaphnia magnain each test vessel were observed for mortality and sub-lethal effects at 24-hour intervals (24, 48, 72 and 96 hours) throughout the duration of the test. The fish were not fed during the test.

Test solutions were analysed at 0, 24, 48 and 72 hours (fresh media) and at 24, 48, 72 and 96 hours (old media) using liquid chromatography with tandem mass spectrometry detection (LC-MS/MS). The geometric mean measured concentration in the WAF test solution was 0.014 mg a.i./L.

The test showed no toxicity to rainbow trout (Oncorhynchus mykiss) at the limit of solubility of the test substance in the test media. The 96-hour LL50 was determined to be greater than 10 mg/L loading rate WAF.

Description of key information

In an OECD Guideline No. 203 study, conducted according to GLP, to assess the toxicity of the substance to the freshwater fish species, Oncorhynchus mykiss, the 96-hour LL50 was determined under daily static-renewal test conditions to be greater 10 mg/L loading rate WAF, corresponding to an EC50 of greater than 0.014 mg a.i./Lbased on geometric mean measured concentration(Smithers Viscient, 2019).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

10 mg/L