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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017-06-02 to 2017-08-21
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD Guideline; EU Method; GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Qualifier:
according to guideline
Guideline:
other: OECD Series on Testing and Assessment, No. 23, "Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures"
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Reference substance name:
Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters
Molecular formula:
unspecified
IUPAC Name:
Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters
Test material form:
liquid
Specific details on test material used for the study:
Name: Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters
Batch No.: RP GW 915 MU
Aggregate State at Room Temperature: Liquid
Colour: Light amber

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Duplicate samples from the freshly prepared test medium of the only test concentration and the control were taken at the start of the test and at day 1.
For the determination of the stability of the test item under the test conditions and of the maintenance of the test item concentrations during the test period, duplicate samples from the aged test media of the only test concentration and the control were collected at day 1 (after 24 hours of exposure) and at the end of the test by pouring together the contents of the test beakers of each treatment.

Test solutions

Vehicle:
no
Details on test solutions:
Test Concentration: The only test item concentration was nominal 100.1 mg test item/L. Additionally; a control was tested in parallel (test water without addition of the test item).
Control: In the control, test water was used without addition of the test item.
Dosage of Test Item: The test medium of the only test item concentration of nominal 100.1 mg/L was prepared by dissolving 50.5 and 52.4 mg test item into 504.5 and 523.5 mL test water by intense stirring for 24 hours and 15 minutes.
The test media was prepared just before introduction of the daphnids (= start of the test) and test medium renewal on Day 1.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Species: Daphnia magna (Straus), clone 5
Age at Test Start: From 2.0 to 19.25 hours old
Sex: Female
Origin: The daphnids introduced in the test were taken from ibacon's in house laboratory culture.
Breeding Conditions: The daphnids were bred in the laboratories of ibacon under similar temperature and light conditions as used in the test. The cultivation of the parental daphnids was performed in Elendt M4 medium. The test organisms were not first brood progeny. The daphnids in the stock culture were fed at least on all working days with green algae (Desmodesmus subspicatus) freshly grown in the laboratories of ibacon.

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h

Test conditions

Hardness:
Water Hardness: 2.5 mmol/L (= 250 mg/L) as Calcium Carbonate
Test temperature:
20.0 to 21.4 °C in the freshly prepared media;
20.3 to 20.4°C in the aged test media
pH:
6.9 to 7.9 in the freshly prepared media;
7.5 to 7.9 in the aged test media; and thus the pH-value did not vary by more than 1.5 units
Dissolved oxygen:
8.4 to 8.8 mg/L in the freshly prepared media;
8.2 to 8.8 mg/L in the aged test media
Nominal and measured concentrations:
nominal 100.1 mg test item/L and a control
Details on test conditions:
Test Environment: Controlled environment room
Measurement of pH, Dissolved
Oxygen and Water Temperature:
The water temperature, pH-values and the dissolved oxygen concentrations were determined in all freshly prepared and aged test media of each treatment group. Therefore the residual test media, which were not used for the test, were filtrated through folded filters.
Water Temperature: 20.0 to 21.4 °C in the freshly prepared media;
20.3 to 20.4°C in the aged test media
pH-Values: 6.9 to 7.9 in the freshly prepared media;
7.5 to 7.9 in the aged test media; and thus the pH-value did not vary by more than 1.5 units
Dissolved Oxygen Concentration: 8.4 to 8.8 mg/L in the freshly prepared media;
8.2 to 8.8 mg/L in the aged test media
Light Regime: 16 h light : 8 h dark
Light Intensity: The light intensity was 630 to 900 lux (measured once during the test).

Reference substance (positive control):
yes
Remarks:
For the evaluation of the quality of the Daphnia clone and the consistency of the experimental conditions, the reference item potassium dichromate is tested at least twice a year to demonstrate satisfactory test conditions.

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
>= 100.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
> 100.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
After 48 hours of exposure no immobilisation of the test animals was observed in the control and in the only test concentration of nominal 100.1 mg test item/L.

The quantification of the test item Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters in the test samples was performed using liquid chromatography with MS/MS detection.
To determine the test item concentration in the test samples a single-point calibration through the standard 200 µg test item/L was used because the validity criteria according to SANCO/3029/99 rev.4 11/07/00 were not met. Therefore, the following values can only be considered as a rough indication of the test item concentration. In the freshly prepared test media at the start of the test and at the renewal of the test media 0.1% of the nominal test concentrations were found (of the nominal concentrations of 100.1 mg test item/L). In the aged test media after 24 and 48 hours test duration, 0.1% of the nominal value was determined of the nominal concentrations of 100.1 mg test item/L). These results confirm the poor solubility of the test item in test water. As also the undiluted part was applied to the test system and the test item is an UVCB substance the reported results refer to nominal test concentrations.
Results with reference substance (positive control):
In the most recent test with the reference item potassium dichromate performed in January 2017 the EC50 after 24 hours was determined to be 1.48 mg test item/L, indicating that the sensitivity of the daphnids was consistent with the level proposed by the OECD 202 guideline (EC50-24 h between 0.6 and 2.1 mg potassium dichromate/L).
Reported statistics and error estimates:
No statistical analysis was performed.
The NOEC and the LOEC were determined directly from the raw data.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
Control Immobilisation Rate: 0 % and furthermore no daphnid showed signs of disease or stress; Dissolved Oxygen Concentration: 8.2 mg O2/L in the control and test vessels at the end of the test
Conclusions:
The toxic effect of the test item Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters to Daphnia magna was assessed in a semi-static limit test. The 48-hour NOEC was determined to be ≥ 100.1 mg test item/L. The 48-hour LOEC was determined to be > 100.1 mg test item/L and the 48-hour EC50 value was determined to be > 100.1 mg test item/L.
Executive summary:

Title: Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters: Acute Toxicity to Daphnia magna in a Semi-Static 48-hour Immobilisation Limit Test

Purpose: The purpose of this study was to determine the influence of the test item Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters on the mobility of Daphnia magna. For this purpose, young daphnids (< 24 hours old) were exposed in a semi-static test for 48 hours to the only concentration of nominal 100.1 mg/L under defined conditions for 48 hours. The recorded effects were the mobility of the daphnids after 24 and 48 hours. The test method of application and the test system are recommended by the test guidelines and Daphnia magna is the recommended test species. The purpose of the analytical part of this study is to verify the concentrations of the test item in the test medium.

Guidelines:

Commission Regulation (EC) No 440/2008, Annex, Part C, C.2.: "Daphnia sp. Acute Immobilisation Test", Official Journal of the European Union (EN), dated May 30, 2008

OECD Guideline for Testing of Chemicals No. 202: "Daphnia sp., Acute Immobilisation Test" adopted April 13, 2004

OECD Series on Testing and Assessment, No. 23, "Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures", December 15, 2000

SANCO/3029/99 rev.4 11/07/00: Residues: Guidance for generating and reporting methods of analysis in support of pre-registration data requirements for Annex II (part A; Section 4) and Annex III (part A; Section 5) of directive 91/414

Material and Methods

Test Item: Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters; batch no.: RP GW 915 MU

Test Species: Female Daphnia magna, clone 5; 2.0 to 19.25 hours old

Source: The daphnids introduced in the test were taken from ibacon's in-house laboratory culture.

Test Design: This study encompassed two treatment groups (one test item concentration at nominal 100.1 mg/L, and one control) each containing 20 individuals. The mobility of the daphnids was determined in a semi-static 48-hour test by visual observation after 24 and 48 hours.

Endpoints: Number of immobile organisms after 24 and 48 hours

Test Concentrations: The only test item concentration was nominal 100.1 mg test item/L. Additionally; a control was tested in parallel (test water without addition of the test item).

Test Conditions: Water temperature: 20.0 to 21.4 °C; pH value: 6.9 to 7.9; dissolved oxygen concentration: 8.2 to 8.8 mg/L; photoperiod: 16 h light - 8 h dark; light intensity: 630 to 900 lux; and thus were within the ranges requested by guideline OECD 202

Results / Biological test results: After 48 hours of exposure no immobilisation of the test animals was observed in the control and in the only test concentration of nominal 100.1 mg test item/L.

Analytical test results: The quantification of the test item Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters in the test samples was performed using liquid chromatography with MS/MS detection. To determine the test item concentration in the test samples a single-point calibration through the standard 200 µg test item/L was used because the validity criteria according to SANCO/3029/99 rev.4 11/07/00 were not met. Therefore, the following values can only be considered as a rough indication of the test item concentration. In the freshly prepared test media at the start of the test and at the renewal of the test media 0.1% of the nominal test concentrations were found (of the nominal concentrations of 100.1 mg test item/L). In the aged test media after 24 and 48 hours test duration, 0.1% of the nominal value was determined of the nominal concentrations of 100.1 mg test item/L). These results confirm the poor solubility of the test item in test water. As also the undiluted part was applied to the test system and the test item is an UVCB substance the reported results refer to nominal test concentrations.

Conclusion: The toxic effect of the test item Phosphoric acid, mono C16-20 (branched, even numbered) alkyl esters to Daphnia magna was assessed in a semi-static limit test. The 48-hour NOEC was determined to be ≥ 100.1 mg test item/L. The 48-hour LOEC was determined to be > 100.1 mg test item/L and the 48-hour EC50 value was determined to be > 100.1 mg test item/L.

This study is classified as acceptable and satisfies the guideline requirements for an acute toxicity study with freshwater invertebrates.