Carnosine

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

01 May - 21 Jun 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss Federal Office of Public Health, Bern, Switzerland

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control and 100 mg/L
- Sampling method: Samples were taken at test start and after 24 and 48 h (test end).
- Sample storage conditions before analysis: All samples were stored frozen (at -20 ± 5 °C) immediately after sampling until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: 50.04 mg of the test item were dissolved in 500 mL of test water using ultrasonic treatment for 15 min and stirring for 15 min at room temperature. After the dissolving procedure the single test concentration of 100 mg/L was a clear solution. The test medium was prepared just before the introduction of the daphnids.
- Controls: Yes
- Eluate: No
- Differential loading: Yes
- Evidence of undissolved material: No, test solutions were clear

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Feeding during test: none
- Food type: The green algae Desmodesmus subspicatus SAG or a mixture of this algal suspension and a commercial fish diet (Tetra Min® Hauptfutter; TETRA-Werke, Melle, Germany) were used.
- Others: At the start of the test, the organisms used in the test were 6-24 hours old and were not first brood progeny.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

2.5 mmol/L corresponds to 250 as CaCO3

Test temperature:

21 - 22 °C

pH:

7.8 - 8.0

Dissolved oxygen:

8.3 - 8.5 mg/L

Nominal and measured concentrations:

Control and 100 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
Test vessel:
- Type: open (covered with glass plates)
- Material, size, headspace, fill volume: 100 mL glass beakers; with 50 mL of test medium and 50 mL headspace
- Aeration: Oxygen saturation of the test water was adjusted before test start
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Test water according to OECD Guideline No. 202
- Alkalinity: 0.8 mmol/L
- Ca/mg ratio: 4:1
- Na:K ratio: 10:1
- Intervals of water quality measurement: start and end of the test medium renewal periods, the pH values, dissolved oxygen concentrations and water temperature were determined in each treatment. At the same observation dates, the appearance of the test media was visually controlled and documented at the same time.

OTHER TEST CONDITIONS
- Photoperiod: 16 h light to 8 h dark cycle with an 30 min transition period
- Light intensity: 19 and 20 μmol m-2 s-1

EFFECT PARAMETERS MEASURED: Immobilization after 24 and 48 h

RANGE-FINDING STUDY
- Test concentrations: 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: Since no statistically significant effect was determined up to 100 mg/L the main test was performed as limit test.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Other biological observations: No other adverse effects (symptoms) were observed.
- Mortality of control: 0%
- Any observations that might cause a difference between measured and nominal values: No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the test.
- Other: Biological and analytical results are summarized in the tables 1-3 within the section "Any other information on results incl. tables"

Results with reference substance (positive control):

The result of the latest positive control test in March 2017 24 h EC50: 1.2 mg/L, the test facility showed that the sensitivity of the test organisms was within the range given by the guideline (24 h EC50: 0.60-2.1 mg/L).

Analytical Results:

The measured concentrations of the test item in the test medium of the single test concentration of 100 mg/L were 97 and 98% of nominal value at the start and the end of the test, respectively.

Table1: Analytical Results

Sampling Day/ Sample Age	Nominal Concentration of Test Item cnom	Measured Concentration of Test Item x	Sample Preparation Factor F	Determined Concentration of Test Item c	% of Nominal Concentration
[d/h]	[mg/L]	[mg/L]		[mg/L]	[%]
0/0	Control	n.d.	1.0	<LOQ	n.a.
(fresh)	100	96.5	1.0	96.5	97
2/48	Control	n.d.	1.0	<LOQ	n.a.
(aged)	100	97.9	1.0	97.9	98

n.d. = no test item detected

n.a. = not applicable

LOQ: 24.2 mg/L

Biological Results:

Table 2: Effect of test item on the mobility of Daphnia magna

Nominal Test Item Concentration	No. of Daphnids Tested	Immobilized Daphnids after 24 Hours		Immobilized Daphnids after 48 Hours
[mg/L]		No.	[%]	No.	[%]
Control	20	0	0	0	0
100	20	0	0	0	0

 

Table 3: Validity criteria for OECD 202

Criterion from the guideline	Outcome	Validity criterion fulfilled
In the control, including the control containing the solubilising agent, not more than 10% of the daphnids should have been immobilized.	No immobile Daphnids were observed	Yes
The dissolved oxygen concentration at the end of the test should be ≥ 3 mg/L in control and test vessels.	8.3 - 8.5 mg/L	Yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Conclusions:

In conclusion, the test item had no acute toxic effects on Daphnia magna in a 48 h static test up to the nominal concentration of 100 mg/L.

Description of key information

EC50 (48 h) >100 mg/L (Daphnia magna, nominal, OECD 202)

Additional information

One experimental study is available investigating the short-term effects of the substance to aquatic invertebrates. The study was performed according to OECD 202 (GLP) with the water flea Daphnia magna. Since no effects were observed in the range-finding test, the main test was performed as limit test with a concentration of 100 mg/L. The initial and the aged measured test item concentrations were in the range of 80 to 120% of the nominal values. Therefore, the endpoints were evaluated using the nominal concentration. No immobilisation of Daphnia magna was observed at the limit concentration of 100 mg/L. Thus, the EC50 (48 h) was determined to be >100 mg/L.