Reaction mass of 2-O-α-L-Rhamnopyranosyl-α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid and α-L-Rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid

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:

1 - 3 March 2016

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

GLP compliance:

no

Remarks:

, but the study was conducted in accordance with ISO/IEC 17025.

Test material

Test material information

Constituent 1

Reference substance name:

Reaction mass of 2-O-α-L-Rhamnopyranosyl-α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid and α-L-Rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid

EC Number:

604-610-3

Cas Number:

147858-26-2

IUPAC Name:

Reaction mass of 2-O-α-L-Rhamnopyranosyl-α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid and α-L-Rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoic acid

Test material form:

liquid

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION OF TEST SOLUTION
Since the test item is soluble, the test solution was prepared by dilution of a stock solution with Daphnia medium.
100 ml of the respective dilution were filled in one of the two replicate test vessels. O2 concentration and pH were measured. The media was aerated by stirring for about 0.5 h if the O2 concentration is too low (target >5.0 mg/l) and the pH was adjusted to 7.0±0.5, if necessary. Afterwards, half of the media (50 ml) was filled in the second replicate test vessel.
The following test flasks were set up:
- Test solution (Tn); containing test medium and test item ; two replicate samples per concentration, with 20 Daphnia each;
- Blank control (Bn); containing test medium ; three replicate samples with 20 Daphnia each.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

- Test organism: Daphnia magna (Straus, 1820)
- Breeding: Parental and young Daphnia held in 4 l glass aquaria (3 l medium) at 20 ± 2°C.
- Illumination: 16 h per day
- Medium: Continuously aerated Elendt M4 medium; prepared with deionised water (conductivity <1.5 μS/cm)
- Feed: Suspension of Desmodesmus subspicatus (green algae) in Elendt M4 medium with an optical density OD680 of about 15 units

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

none

Test conditions

Nominal and measured concentrations:

100 mg/l (nominal concentration)

Details on test conditions:

TEST SYSTEM
- Test vessel 100 ml flasks, all-glass, with 50 ml of test medium
- Test medium Elendt M4 medium (Table 1 and Table 2); prepared with ultra-pure water (conductivity <1.5 μS/cm)
- Number of Daphnia: 40 individuals per test concentration, 20 per vessel
- Age: Less than 24 h
- Temperature (target): 18–22 °C, controlled at ±1 °C
- Test type: Static exposure conditions
- Test duration: 48 h.

TEST MEDIUM / WATER PARAMETERS
- Medium: Continuously aerated Elendt M4 medium, prepared with deionised water (conductivity <1.5 μS/cm).
- Analyses of the Elendt M4 Daphnia medium (mean values of three independent medium batches): total hardness (as sum of Ca and Mg): 2.52 mmol/l; carbonate hardness: 0.32 mmol/l; alkalinity (p-value/m-value):–0.08/0.70 mmol/l; Ca/Mg ratio: 4.11; Na/K ratio: 9.13.

OTHER TEST CONDITIONS
- Adjustment of pH: the pH was adjusted to 7.0±0.5, if necessary
- Photoperiod: 16 h photoperiod a days upplied by overhead white fluorescent tubes
- Light intensity: unknown

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

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

Effect concentrations 2

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

Details on results:

After 24 and 48 h of incubation no significant effects (≤10% of a total of 40 individuals) were observed at the nominal concentration of 100 mg/l and in the blank control.

Results with reference substance (positive control):

The EC50 for the control of sensitivity for 24 h of exposure with potassium dichromate was estimated to be 1.1 mg/l. The resulting EC50 value of the reference item was within the recommended range of 0.6–2.1 mg/L (according to OECD Guideline 202).

Reported statistics and error estimates:

No statistical analysis was performed.

Any other information on results incl. tables

EVALUATION OF THE TEST RESULTS

1) Definition and determination of the effect concentrations (ECx)

The effective concentration ECx is the concentration leading to immobilization of x% of the Daphnia after 24 or 48 h of exposure. Those individuals not able to swim within 15 s after gentle agitation of the test vessel are considered to be immobile.

The median effect concentrations (EC50) for 24 and 48 h was estimated based on the concentrations tested and immobilization observed.

2) Determination of the no observed effect concentration (NOEC)

The no-observed effect concentration (NOEC) is the highest test concentration for which the test item is observed to have no biologically significant effects when compared to the blank controls. Immobilization rates of <=10% are regarded as not biologically significant.

3) Validity criteria

The test is considered valid if not more than 10% of the Daphnia in the control are immobilized or trapped at the surface of the water. The dissolved oxygen concentration at the end of the test should be ≥3 mg O2/l.

4) Tables

Table 1: Immobilization of Daphnia magna after 24 and 48 h of exposure to the test item

Nominal concentration (mg/l)	Replicate	Number of individuals per test vessel	Immobile Daphnia (No./vessel) 24 h	Immobile Daphnia (No./vessel) 48 h	% immobile Daphnia after 24 h of exposure	% immobile Daphnia after 48 h of exposure
0	A	20	0	0	0	0
	B	20	1	1	5	5
	C	20	0	0	0	0
100	A	20	0	0	0	0
	B	20	0	0	0	0

Table 2: pH-values, oxygen concentrations and conductivity values

Nominal concentration (mg/l)	Replicate	pH-values 0 h	pH-values 48 h	Oxygen concentration (mg/l) 0 h	Oxygen concentration (mg/l) 48 h	Conductivity (µS.cm^-1) 0 h
0	A B C	6.6	7.3	7.6	7.8	685
100	A B	6.6	7.3	8.0	7.1	687

Values were determined in the pooled samples only.

The pH in the control and the test vessels did not vary by more than 0.7 units (OECD Guideline 202: pH should

normally not vary by more than 1.5 units).

The dissolved oxygen concentration at the end of the test in the control and the test vessels was ≥7.8 mg O2/l

(required ≥3 mg O2/l).

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

- (48h) EC50 > 100 mg/L (nominal);
- NOEC: 100 mg/L (nominal).
The criteria for classification of a substance in category Acute 1 are defined on the basis of acute aquatic toxicity data only (EC50 or LC50); classification is required for values ≤ 1 mg/L. Therefore, the substance does not need to be classified for acute (short-term) aquatic hazard.
Further, classification as aquatic chronic category 3 or worse is required if the 96 hr LC 50 (for fish), and/or 48 hr EC 50 (for crustacea) and/or 72 or 96 hr ErC 50 (for algae or other aquatic plants) values are > 100 mg/l (and the substance is not rapidly degradable and/or the experimentally determined BCF ≥ 500 (or, if absent, the log Kow ≥ 4). The determined 48h EC50 is > 100 mg/l, so noc classification as aquatic chronic Cat. 3 or worse is required.

Executive summary:

The acute toxicity of the test item 'Decanoic acid, 3-[[6-deoxy-2-O-(6-deoxy-α-L-mannopyranosyl)-α-L-mannopyranosyl]oxy]-, 1-(carboxymethyl)octyl ester, mixt. with 1-(carboxymethyl)octyl 3-[(6-deoxy-α-L-mannopyranosyl)oxy]decanoate' to Daphnia magna was investigated under static exposure conditions in a screening test over a period of 48 h.

Since the test item is soluble, the test solution was prepared by dilution of a stock solution with algal medium. The only investigated nominal concentration was 100 mg/l (limit test).

40 individual Daphnia divided in 2 test vessels were exposed to the test item, and 60 Daphnia divvied into 3 vessels were used in the blank control.

No analytical verification of the test item was conducted. The evaluation of the test was based on the nominal concentration of the test item, assuming the test item to be stable in water over 48 h.

After 24 and 48 h of incubation no significant effects (≤10% of a total of 40 individuals) were observed at the nominal concentration of 100 mg/l and in the blank control.

Based on these immobilization data, the median effect concentration (EC50) of the test material to Daphnia magna for 24 and 48 h was estimated to be >100 mg/l nominal concentration.

The experimentally determined no-effect concentration (NOEC) is 100 mg/l nominal concentration after an exposure of 24 and 48 h.