Magnesium glycerophosphate

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:

2018-03-09 to 2018-07-10

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD, EU Guideline Study: GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Magnesium glycerophosphate

EC Number:

213-149-3

EC Name:

Magnesium glycerophosphate

Cas Number:

927-20-8

Molecular formula:

C3H9O6P.Mg

IUPAC Name:

magnesium glycerophosphate

Test material form:

solid: particulate/powder

Details on test material:

Batch: INVG003917

Specific details on test material used for the study:

Name: Magnesium glycerophosphate
Batch No.: INVG003917
Active Ingredient Purity: Magnesium glycerophosphate: 96.5 % (w/w) (calculated)
α-Glycerol phosphate magnesium salt: 96.5 % (w/w) (13C-NMR)
β- Glycerol phosphate magnesium salt : 0.0 % (w/w) (13C-NMR)
according to certificate of analysis and analytical report no. 17-011608
Certificate of Analysis Date: September 01, 2017
Aggregate State at Room Temperature: Solid (fine powder)
Colour: White
Expiry Date: June 10, 2021
Storage Conditions at Test Facility: at 20 +/- 5 °C, in the dark

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Duplicate samples from the freshly prepared test media of the only test concentration and the control were taken at the start of the test and at day 3.
For the determination of the stability of the test item under the test conditions, respectively the maintenance of the test item concentration during the test period, samples were taken in duplicate of all treatment groups at day1 and at the end of the test (96 hours) from the approximate centre of the aquaria.
All samples remained undiluted until analysis.

Test solutions

Vehicle:

no

Details on test solutions:

The test medium of the only test item concentration of nominal 100 mg/L was prepared by dissolving
500 mg test item into 5000 mL test water (day 0)
501.1 mg test item into 5011 mL test water (day 1)
503.3 mg test item into 5033 mL test water (day 2)
500.1 mg test item into 5001 mL test water (day 3)
by intense stirring for 15 minutes.
The test media was prepared just before introduction of the test fish (= start of the test and test medium renewal on day 1, 2 and 3).

Test organisms

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Species: Zebrafish (Danio rerio)
Size and Weight: Juveniles;
The mean body length of the fish* in the test was 1.82 ± 0.12 cm (Mean ± SD), the mean body wet weight 0.05 ± 0.01 g (Mean ± SD).
* 10 fish from the test fish batch were measured before the start of the test
Sex: Male and female
Origin: The test fish were obtained from ibacon's inhouse laboratory culture
Holding Conditions: All fish were obtained and held in the laboratory for at least 12 days before the start of the test. They were held in water of the quality to be used in the test for at least seven days immediately before testing under the following conditions:
Light: 16 hours photoperiod daily
Temperature: 21- 25 °C
Oxygen concentration: at least 80 % of the air saturation value
Feeding: three times per week or daily until 24 hours before the test was started
During the last 7 days prior to the start of the test no fish died in the test fish batch. Therefore the mortalities in the fish batch were below 5 % and the fish batch was accepted.

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Post exposure observation period:

no

Test conditions

Hardness:

2.5 mmol/L (= 250.0 mg/L) as CaCO3

Test temperature:

13.8 - 14.8 °C

pH:

7.2 to 8

Dissolved oxygen:

98 to 102 %

Nominal and measured concentrations:

100 mg test item/L and a control

Details on test conditions:

Test Environment: Controlled environmental room
Measurement of pH-Values, Dissolved Oxygen and Water Temperature:
The water temperature, pH-values and the dissolved oxygen concentrations were determined daily in the test media of each treatment group.
Water Temperature: 13.8 to 14.8 °C
pH value of the Test Water: 7.5 to 7.8
Dissolved Oxygen Concentrations: 87 to 99 % of the air saturation value
Light Regime: 16 h light : 8 h dark;
30 min dawn/dusk period was provided
Light Intensity: 450 to 550 lux Aeration of the Test Water: The test media were slightly aerated during the test.

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Effect concentrations 2

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

In the control and the only test concentration of 100 mg test item/L, all fish survived until the end of the experiment and showed no sublethal effects during the exposure time.

Reported statistics and error estimates:

No statistical analysis was performed.
The LC50, NOEC and the LOEC were determined directly from the raw data.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

In the control no fish died during the test. The dissolved oxygen concentration in the test media did not fall below 98 % of air saturation value during the test.

Conclusions:

The toxic effect of the test item Magnesium glycerophosphate to Zebrafish (Danio rerio) was assessed in a semi-static limit test. Based on the test results the 96-hour LC50 was determined to be >100 mg test item/L based on nominal concentrations. The NOEC was determined to be ≥ 100 mg test item/L also based on nominal concentrations. The initial concentrations and the maintenance of the exposure concentrations during the test were verified in the analytical part. All reported results refer to nominal values since the concentrations of the test item were within ± 20 % of the nominal concentrations during the test.

Executive summary:

Title: Magnesium glycerophosphate: Acute Toxicity to Zebrafish (Danio rerio) in a 96-hour Semi-Static Limit Test

Purpose:

The purpose of this study was to evaluate the acute toxicity of the test item Magnesium glycerophosphate to fish. For this purpose, juvenile Zebrafish were exposed in a semi static test to the only concentration under defined conditions for 96 hours. The recorded effects were the mortality and sublethal effects on the fish. A threshold approach was pursued.

The test method of application and the test system are recommended by the test guidelines and Zebrafish is one of the recommended test species.

This threshold approach was performed in order to demonstrate that the test item has no toxic effects on the test fish up to the threshold concentration or at 100 mg (active ingredient)/L. The lowest EC50 value of existing relevant algae or acute invertebrate (e.g. daphnia) tests was set as threshold concentration.

In the interest of animal welfare and efficient use of resources, it is important to avoid the unnecessary use of animals whenever possible. In the field of aquatic toxicology, this especially applies to the acute toxicity testing of fish according to OECD 203. The threshold approach described hereafter addresses fish toxicity by using a single-concentration test (limit test) requiring less fish compared to the full acute fish toxicity study. It is based on the derivation of a threshold concentration from relevant algae and acute invertebrate (e.g. daphnia) tests at which fish toxicity is tested. If no mortality occurred in the limit test using the threshold concentration, the threshold concentration might be used as a surrogate LC50 value in the further hazard or risk assessment.

The purpose of the analytical part of this study was to verify the concentrations of the test item in the test medium.

Guidelines:

 Commission Regulation (EC) No 440/2008, Annex, Part C, C.1.: "Acute Toxicity for Fish", Official Journal of the European Union, May 30, 2008

OECD Guideline for Testing of Chemicals, Section 2, No. 203, "Fish, Acute Toxicity Test", adopted July 17, 1992

OECD Guidance Document, Series on Testing and Assessment No. 126, “Short Guidance in the Threshold Approach for Acute Fish Toxicity”, May 31, 2010

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

Materials and Methods

Test Item: Magnesium glycerophosphate; batch no.: INVG003917;

purity:

- Magnesium glycerophosphate: 96.5 % (w/w) (calculated)

-α-Glycerol phosphate magnesium salt: 96.5 % (w/w) (13C-NMR)

-β- Glycerol phosphate magnesium salt : 0.0 % (w/w) (13C-NMR)

according to certificate of analysis and analytical report no. 17-011608

Test Species: Juvenile Zebrafish (Danio rerio) mean length: 1.82 cm ± 0.12 cm; source: inhouse laboratory culture

Test Design:

This study encompassed two treatment groups (one test item concentration at nominal 100 mg/L and one control) each containing 7 individuals. The acute toxicity to unfed juvenile zebrafish was determined in an aerated, semi-static, 96-hour test. The test fish were observed at test start and after approximately 2, 24, 48, 72 and 96 hours test duration for sublethal effects and mortality.

Endpoints: NOEC after 96 h, LOEC after 96 h; LC50: lethal concentration producing 50 % mortality after 96 h of exposure.

Test Concentration: 100 mg test item/L and a control

Test Conditions: Water temperature: 23 to 25 °C; pH value: 7.2 to 8.0; dissolved oxygen concentration: 98 to 102 % of the air saturation value; photoperiod: 16 h light - 8 h dark; light intensity: 400 to 470 lux and thus were within the ranges requested by guideline OECD 203.

Results:

Biological test results:

In the control and the only test concentration of 100 mg test item/L, all fish survived until the end of the experiment and showed no sublethal effects during the exposure time.

Analytical test results:

The quantification of the test item Magnesium glycerophosphate in the test samples was performed using liquid chromatography with MS/MS detection. In the freshly prepared test media at the start of the test and at the renewal of the test media 92 % of the nominal test concentrations were found (average of all samples of the only test concentration). In the aged test media after 24 and 96 hours test duration, 93 % of the nominal value was determined (average of all samples of the only test concentration). During the test the test organism were exposed to a mean of 92 % of nominal.

Conclusion:

The toxic effect of the test item Magnesium glycerophosphate to Zebrafish (Danio rerio) was assessed in a semi-static limit test. Based on the test results the 96-hour LC50 was determined to be >100 mg test item/L based on nominal concentrations. The NOEC was determined to be ≥ 100 mg test item/L also based on nominal concentrations. The initial concentrations and the maintenance of the exposure concentrations during the test were verified in the analytical part. All reported results refer to nominal values since the concentrations of the test item were within ± 20 % of the nominal concentrations during the test.

This study is classified acceptable and satisfies the guideline requirements for fish acute test studies.