Dialkyl C18 and C18-unsaturated phosphonates

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:

From2017-06-09 to 2017-10-05

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

2015-11-03

Test material

Test material information

Constituent 1

Reference substance name:

Dialkyl C18 and C18-unsaturated phosphonates

EC Number:

701-298-1

Cas Number:

64051-29-2

Molecular formula:

Not applicable for a UVCB Substance

IUPAC Name:

Dialkyl C18 and C18-unsaturated phosphonates

Test material form:

liquid

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

During the final test samples for possible analysis were taken from both test groups according to the schedule below.
Frequency at t=0 h, t=24 h and t=96 h
Volume 1.0 mL
Storage Samples were stored in a freezer (≤-15°C) until analysis.

Test solutions

Vehicle:

yes

Details on test solutions:

Preparation of Test Solutions
The batch of Alkenyl phosphonate tested was a colourless liquid and a UVCB substance. No correction was made for the purity/composition of the test item.
For the range-finding test, preparation of test solutions started with loading rates of 1.0, 10 and 100 mg/L. A 3- day period of magnetic stirring was applied to ensure maximum dissolution of the test item in medium. The obtained mixtures were allowed to settle for a period of 1 hour. Thereafter, the aqueous Water Accommodated Fractions (WAFs) were collected by means of siphoning through glass wool and used as test concentrations. The WAF prepared at a loading rate of 1.0 mg/L was clear and colorless, the WAF prepared at a loading rate of 10 mg/L was slightly hazy and the WAF prepared at a loading rate of 100 mg/L was hazy.
For the final test, preparation of test solutions started with a stock solution of 100 mg/L in acetone. A 15- minute period of magnetic stirring was applied to ensure maximum dissolution of the test item in acetone. Thereafter, 800 µL of the stock was spiked into 8 litres of medium to reach a test item concentration of 10 µg/L.

Test organisms

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM
- Common name: Cyprinus carpio, Teleostei, Cyprinidae
- Source: Zodiac, proefacc, "De Haar Vissen", Wageningen University and Research Centre, The Netherlands.
- Mean length
Range-finding test: 3.8 ± 0.2 cm
Final test: 3.1 ± 0.1 cm
Mean weight1
Range-finding test: 0.78 ± 0.18 g
Final test: 0.45 ± 0.03 g
Characteristics F1 from a single parent-pair bred in UV-treated water.
Reason for selection This system has been selected as an internationally accepted species.
Total fish used 23

- 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%.

ACCLIMATION
- Acclimation period: At least 12 days after delivery
- Feeding frequency: Daily with pelleted fish food (Essence (300-500 um), Coppens International bv, Helmond, The Netherlands)

FEEDING DURING TEST
- No feeding from 24 hours prior to the test and during the total test period

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Test conditions

Hardness:

180 mg/L CaCO3

Test temperature:

20.6°C to 21.2 °C

pH:

7.6 to 8.0

Dissolved oxygen:

6.1 to 8.9 mg/L

Salinity:

Not applicable, fresh water

Details on test conditions:

TEST SYSTEM
- Test vessel: glass vessels
- Type : open
- Material, size, headspace, fill volume: 10 litres, containing 8 litres of test solution
- Aeration: No
- 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

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
- Adjusted ISO medium

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16h illumination per day

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Mortality and other effects At 3, 24, 48, 72 and 96 hours following the start of exposure. In addition, every afternoon to observe for any dead or severely distressed fish. Dead fish were removed when observed.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Range finding study: Yes
- Test concentrations: 1, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study:
The initial concentration was 8.7 mg/L. The measured concentration decreased to 3.3% of initial at the end of the test
The measured concentrations strongly exceeded solubility of the test item in water. This was confirmed by the observation that the two highest concentrations were slightly hazy to hazy and a floating layer was observed at these concentrations from the 24 hour of exposure. Therefore, it was decided to perform the final test as a limit with concentration exceeding water solubility of the test item by factor not higher than 100, i.e. 10 µg/L.

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 8.7 µg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Mortality of control: No
- Abnormal responses: No
- Effect concentrations exceeding solubility of substance in test medium: Yes

Results with reference substance (positive control):

- Results with reference substance valid ?: Yes
- Mortality: Yes
- LC50: 0.32 mg/L

Any other information on results incl. tables

Sublethal observations / clinical signs:

Incidence of Mortality and Total Mortality During the Range-Finding Test

Alkenyl phosphonate Loading rate (mg/L)	Initial number of fish	Cumulative mortality					Total Mortality (%)
		2½h	24h	48h	72h	96h
1.0	3	0	0	0	0	0	0
10	3	0	0	0	0	0	0
100	3	0	0	0	0	0	0

 

 

Incidence of Mortality and Total Mortality During the Limit Test

Alkenyl phosphonate Average exposure concentration (µg/L)	Initial Number of fish	Cumulativemortality					Total Mortality (%)
		3h	24h	48h	72h	96h
Solvent-control	7	0	0	0	0	0	0
8.6	7	0	0	0	0	0	0

 

pH-Values and Dissolved Oxygen Concentrations (mg/L) During the Limit Test

Alkenyl phosphonate Average exposure concentration (µg/L)	Day 0		Day 1		Day 2		Day 3		Day 4
	pH^1	O2	pH	O2	pH	O2	pH	O2	pH	O2
Solvent-control	8.0	8.9	7.7	7.5	7.8	7.3	7.7	6.4 (6.5)	7.7	6.2
8.6	8.0	8.9	7.6	7.5	7.7	7.2	7.6	6.3 (6.4)	7.6	6.1

¹  pH of culture medium was: 7.8 , () – additional measurement performed at the end of the day to check whether there was no further decrease of the concentration

Temperatures (°C) Measured During the Limit Test

Alkenyl phosphonate Average exposure concentration (µg/L)	Day 0^1	Day 1	Day 2	Day 3	Day 4
Solvent-control	21.2	20.6	20.7	20.6	20.7
8.6	20.9	20.7	20.8	20.8	20.7

¹  Temperature of culture medium was: 21.6 °C.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

In conclusion, the 96h-LC50 was beyond the range tested, i.e. exceeded the average exposure concentration of 8.6 µg/L (this concentration was consider to exceed solubility of the test item in both medium and water).

Executive summary:

The objective of the study was to evaluate Alkenyl phosphonate for its ability to generate acute toxic effects in Cyprinus carpio during an exposure period of 96 hours and, if possible, to determine the LC50 at all observation times.

The study procedure described in this report was based on the OECD guideline No. 203, 1992. In addition, procedures were based on the test methods described in the OECD series on testing and assessment number 23, 2000.

The batch of Alkenyl phosphonate tested was a colourless liquid and a UVCB substance.

Stock solution was prepared in acetone at a factor 10,000 higher than the final test concentration. Thereafter, 800 µL of the stock was spiked into 8 litres of medium to reach a test item concentration of 10 µg/L.

A final test was performed as a limit test, based on the results of a preceding range-finding test and information regarding solubility of the test item in water.

Seven fish per group were exposed to an untreated control, a solvent control and to a limit concentration of 10 µg/L. The total exposure period was 96 hours and samples for analytical confirmation of exposure concentrations were taken at the start of the test, after 24 hours and at the end of the test.

Analysis of the samples taken at the start of the final test showed a measured concentration of 15.4 µg/L at the nominal concentration of 10 µg/L. The measured concentration remained stable during the first 24 hours of exposure (89% of initial). Thereafter, it decreased to 3.3 µg/L (21 % of initial) at the end of the test. The calculated average exposure concentration was 8.6 µg/L.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

In conclusion, the 96h-LC50 was beyond the range tested, i.e. exceeded the average exposure concentration of 8.6 µg/L (this concentration was consider to exceed solubility of the test item in both medium and water).