Bos taurus, milk, pasteurized, homogenized, skimmed, fermented, spray-dried

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:

13 - 16 March 2018

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:

13 April 2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

30 May 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The following test vessels were set up:
- Test solution (Tn); containing Daphnia medium with test item (four replicates with five Daphnia each)
- Blank control (Bn); containing pure Daphnia medium (four replicates with five Daphnia each)
- Since the test item is not well soluble, the test solutions were prepared by stirring a 100 mg/L suspension of test item in Daphnia medium for about 1 h followed by about 45 min of ultra-sonication. The suspension was then filtered (MILLIPORE AP15 glass fiber filter). The resulting solution was used as the test solution in the toxicity test. A sample for the determination of the test concentration was taken from the remaining solution. Pure Daphnia medium served as blank controls.
- O2 concentration and pH were measured in the test medium for each concentration including the blank control. If the O2 concentration was too low (>5.0 mg/L required) the media was aerated by stirring. Afterwards, the media were filled into the test vessels (50 mL per vessel).
- Daphnia, aged less than 24 h and already acclimatized to the Daphnia medium, were then introduced into the test media, and the vessels were covered with a glass plate. The Daphnia were not fed during the test and the test vessels were not aerated.
- Semi-static exposure conditions

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISMS
- Test organism: Daphnia magna (Straus, 1820), derived from a healthy stock and not first brood progeny
- Breeding: Parental and young Daphnia held in 75 mL glass vessels (50 mL medium) at 18-22°C
- Illumination: 16 h per day
- Medium: Continuously aerated Elendt M4 medium prepared with ultra-pure water (conductivity <1.5 µS/cm)
- Feed: Suspension of Desmodesmus subspicatus in Elendt M4 medium with an optical density OD680 of about 15 units

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

not reported

Test temperature:

18–22°C, controlled at ±1 °C

pH:

6 to 9

Dissolved oxygen:

The minimum dissolved oxygen concentration in the controls and the test vessels at the end of each 24 h-period was >=3.52 mg O2/L (required >=3 mg O2/L).

Salinity:

not reported

Conductivity:

722 to 729 µS cm-1

Nominal and measured concentrations:

The concentration of Yoghurt powder in the test medium was determined by DOC analysis at the beginning and after 24 h (in the old and the fresh medium) and after 48 h of exposure. These analyses confirmed that the test item was partly dissolved (78 and 79% of the nominal loading rate in the freshly prepared test medium at 0 and 24 h). In the old test medium 63 and 51% of the loading rate was determined. Conclusions about the stability of the test item during the exposure phase cannot be drawn, since this quantification method is not substance-specific. The effective concentration EC50 was assessed based on the loading rate of the test item as well as on the arithmetic mean of the two 24-h periods’ geometric means of the measured concentrations (after 48 h).

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL beakers, all-glass, with 50 mL of test medium, covered with a glass plate to avoid evaporation and contamination of the test solutions with dust
- Test medium: Elendt M4 medium; prepared with ultra-pure water (conductivity <1.5 µS/cm)
- Number of Daphnia: 20 individuals per test concentration, 5 per vessel
- Age: Less than 24 h
- Light: 16 h photoperiod a day, supplied by overhead white fluorescent tubes
- Temperature: 18–22 °C, controlled at ±1 °C
- pH: 6 to 9. The pH should normally not vary by more than 1.5 units in one test.
- Feed: The Daphnia are not fed during the test
- Aeration: The test vessels were not aerated during the test.
- Test type: Semi-static exposure conditions
- Test duration: 48 h

RANGE-FINDING STUDY
Prior to the definitive test a non-GLP range finding test with a nominal concentration of 100 mg/L of Yoghurt powder was performed.
The DOC determinations indicated that only about 80% of the test item was dissolved. Even though this kind of analytics is not substance-specific and usually cannot give any indications about the stability of the test item, the present data shows that a significant loss of dissolved organic carbon occurred, which may be explained by fermentation or even feeding by the Daphnia.
In order to maintain the test item concentration as constant as possible semi-static conditions were applied in the definitive test.

EFFECT PARAMETERS MEASURED
Observed immobility (inability to swim) of the daphnids: Observations of immobile Daphnia were made after 24 and 48 h of exposure. Any abnormal behaviour or appearance was reported.

Reference substance (positive control):

yes

Remarks:

Acute reference test with potassium dichromate conducted twice a year

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

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

24 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

No significant effects (<=10% immobilization) were observed at 100 mg/L nominal concentration or in the blank controls, neither after 24 h of exposure nor after 48 h. Therefore, the median effect concentrations (EC50) of Yoghurt powder (EC no. 917-734-0) on Daphnia magna after 24 h as well as after 48 h of exposure were estimated to be >100 mg/L nominal concentration.
The NOEC values after 24 h and after 48 h of exposure were both 100 mg/L nominal concentration.

Results with reference substance (positive control):

Acute ref. test with K2Cr2O7 conducted twice a year. The EC50 value for the control of sensitivity for 24 h of exposure with K2Cr2O7 was estimated to be 1.4 mg/L (19 October 2017), which lies within the recomm. range of 0.6–2.1 mg/L acc. to OECD Guideline 202.

Reported statistics and error estimates:

The effective concentrations ECx were assessed based on the nominal concentrations.
No statistical analysis was performed.

Validity criteria fulfilled:

yes

Remarks:

All validity criteria i.e. immobilization in the controls <=10% and O2 concentrations at the end of the test >=3 mg/L were fulfilled.

Conclusions:

The acute toxicity of Yoghurt powder (EC no. 917-734-0) to Daphnia magna was determined in a 48 hour static test according to OECD guideline 202. The median effect concentrations (EC50) of Yoghurt powder on Daphnia magna were estimated to be >100 mg/L and >100 mg/L nominal concentration after 24 and 48 h of exposure, respectively. The NOEC values were determined to be 100 mg/L nominal concentration after 24 and 48 h of exposure, respectively. The results of the test can be considered reliable without restriction.

Executive summary:

The acute toxicity of Yoghurt powder (EC no. 917-734-0) to Daphnia magna was investigated according to test guideline OECD 202, under semi-static exposure conditions over a period of 48 h.

The test item Yoghurt powder is 100% pure and is not well soluble in Daphnia medium. The test media were prepared by stirring a 100 mg/L suspension of the test item in Daphnia medium for about 1 h followed by about 45 min of ultrasonication and subsequently filtered to separate the undissolved test item from the solution.

The single investigated loading rate was 100 mg/L.

Since the test concentration was not stable, the definitive test was performed under semi-static conditions, with a change of test medium at 24 hours after the start of the test.

20 individuals, divided in 4 groups of 5 individuals, were used for the test concentration and for the blank.

The concentration of Yoghurt powder in the test medium was determined by DOC analysis at the beginning and after 24 h (in the old and the fresh medium) and after 48 h of exposure. These analyses confirmed that the test item was partly dissolved (78 and 79% of the nominal loading rate in the freshly prepared test medium at 0 and 24 h). In the old test medium 63 and 51% of the loading rate was determined. Conclusions about the stability of the test item during the exposure phase cannot be drawn, since this quantification method is not substance-specific. The effective concentration EC50 was assessed based on the loading rate of the test item as well as on the arithmetic mean of the two 24-h periods’geometric means of the measured concentrations (after 48 h).

No significant effects (<=10% immobilization) were observed at 100 mg/L loading rate or in the blank controls after 48 h of exposure.

Therefore, the median effect concentration (EC50) of Yoghurt powder (EC no. 917 -734 -0) on Daphnia magna after 48 h of exposure was estimated to be >100 mg/L loading rate corresponding to 66.6 mg/L mean measured (based on DOC determination).

The NOEC value after 48 h of exposure was 100 mg/L nominal loading rate corresponding to 66.6 mg/L mean measured (based on DOC determination).

All validity criteria were fulfilled.

Description of key information

The acute toxicity of Yoghurt powder (EC no. 917-734-0) to Daphnia magna was determined in a 48 hour static test according to OECD guideline 202. The median effect concentrations (EC50) of Yoghurt powder on Daphnia magna were estimated to be >100 mg/L and >100 mg/L nominal concentration after 24 and 48 h of exposure, respectively. The NOEC values were determined to be 100 mg/L nominal concentration after 24 and 48 h of exposure, respectively. The results of the test can be considered reliable without restriction.