Alkenes, C15-18-branched and linear, maleated, hydrolyzed

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:

09 February 2009 to 29 May 2009

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Principles of method if other than guideline:

In view of the difficulties associated with the evaluation of aquatic toxicity of poorly water soluble test materials, a modification of the standard method for the preparation of aqueous media was performed. An approach endorsed by several important regulatory authorities in the EU and elsewhere (ECETOC 1996, OECD 2000 and Singer et al 2000), is to expose organisms to a Water Accommodated Fraction (WAF) of the test material in cases where the test material is a complex mixture and is poorly soluble in water and in the permitted auxiliary solvents and surfactants. Using this approach, aqueous media are prepared by mixing the test material with water for a prolonged period. Pre-study work showed that a preparation period of 24 hours was sufficient to ensure equilibration between the test material and water phase. At the completion of mixing, the test material phase is separated by siphon and the test organisms exposed to the aqueous phase or WAF (which may contain dissolved test material and/or leachates from the test material). Exposures are expressed in terms of the original concentration of test material in water at the start of the mixing period (loading rate) irrespective of the actual concentration of test material in the WAF.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 10, 18, 32, 56 and 100 mg/L
- Sampling method: Samples were taken at 0 and 48 hours
- Sample storage conditions before analysis: Sample volumes required for chemical analysis precluded the storage of duplicate samples at 72 hours.

Vehicle:

no

Details on test solutions:

Due to the low aqueous solubility and complex nature of the test material for the purposes of the test, the test material was prepared as a Water Accomodated Fraction (WAF). Amounts of test material (25, 45, 80, 140 and 250 mg) were each separately added to the surface of 2.5 L of reconstituted water to give the nominal concentrations of 10, 18, 32, 56 and 100 mg/L loading rates respectively. After the addition of the test material, the reconstituted water was stirred by a magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. Visual inspection of the WAFs indicated that a significant amount of dispersed test material was present in the water column and hence it was justifiable to remove the WAFs by filtering through a glass wool plug. A wide bore glass tube, covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed through the Nescofilm seal. A glass wool plug was inserted into the opposite end of the tubing and the WAF was removed by mid-depth siphoning to give the 10, 18, 32, 56 and 100 mg/L loading rate WAFs.

Test organisms (species):

Daphnia magna

Details on test organisms:

Adult Daphnia were maintained in polypropylene vessels containing approximately 2 litres of reconstituted water in a temperature controlled room at approximately 20 ºC.
The reconstituted water used for both the range-finding and definitive tests was the same as that used to maintain the stock animals.
The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a suspension of algae (Chlorella sp.). Culture conditions ensured that reproduction was by parthenogenesis.
Gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing.
The diet and diluent water are considered not to contain any contaminant that would affect the integrity or outcome of the study.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

No post-exposure observation was carried out

Hardness:

The reconstituted water had an approximate theoretical total hardness of 250 mg/L as CaCO3.

Test temperature:

Temperature was maintained at 21 ° C throughout the test.

pH:

The pH was recorded at the start and termination of the test, the pH ranged from pH 7.6 - 8.0.

Dissolved oxygen:

Dissolved oxygen concentrations were recorded at the start and termination of the test, the oxygen concentration ranged from 8.4 mg O2/L to 8.5 mg O2/L. The air saturation value ranged from 94 % to 96 %.

Salinity:

Not measured.

Nominal and measured concentrations:

Chemical analysis of the test loading rates of 10, 18, 32, 56 and 100 mg/L at 0 hours showed measured concentrations of 0.183, 0.442, 0.392, 0.396 and 0.347 mg/L respectively. Analysis of the test loading rates of 10, 18, 32, 56 and 100 mg/l at 48 hours showed measured concentrations of 0.0593, 0.179, 0.202, 0.173 and 0.249 mg/l respectively.

Details on test conditions:

In the definitive test 250 mL glass jars containing approximately 250 mL of test preparation were used. At the start of the test 10 daphnids were placed in each test and control vessel at random, in the test preparations. Two replicate test and control vessels were prepared. The test vessels were then covered to reduce evaporation and maintained in a temperature controlled room at approximately 20 ° C with a photoperiod of 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods. The daphnids were not individually identified, received no food during exposure and the test vessels were not aerated.

The control group was maintained under identical conditions but not exposed to the test material.

The test preparations were not renewed during the exposure period.

Mobility of the test species was monitored at 24 and 48 hours after the start of the exposure.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

95 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL = 80-131 mg/l

Duration:

48 h

Dose descriptor:

NOELR

Effect conc.:

32 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

There were no treatment related differences for oxygen concentration or for pH.

Results with reference substance (positive control):

The results from the positive control with potassium dichromate were within the normal range for this reference material. The mean 48-Hour EC50 value calculated from all positive controls was 0.78 mg/L (sd=0.21). The NOEC after 24 hours and 48 hours was 0.32 mg/L. The NOEC was based upon zero immobilisations at this concentration.

Reported statistics and error estimates:

The EC50 values and associated confidence limits at 24 and 48 hours and the slope of the response curves and standard errors were calculated by the maximum-likelihood probit method (Finney, 1971) using the ToxCalc computer software package (Toxcalc 1999).

 Cumulative Immobilisation Data in the Definitive Test

Nominal Loading Rate (mg/l)	Cumulative Immobilised Daphnia (Initial Population: 10 Per Replicate)
	24 Hours				48 Hours
	R1	R2	Total	%	R1	R2	Total	%
Control	0	0	0	0	0	0	0	0
10	0	0	0	0	0	0	0	0
18	0	0	0	0	0	0	0	0
32	0	0	0	0	0	0	0	0
56	0	1	1	5	1	1	2	10
100	3	3	6	30	5	6	11	55

R₁¿ R₂= Replicates 1 and 2

Cumulative Immobilisation Data in the Positive Control

Nominal Concentration (mg/l)	Cumulative Immobilised Daphnia (Initial Population: 10 Per Replicate)
	3 Hours				24 Hours				48 Hours
	R1	R2	Total	%	R1	R2	Total	%	R1	R2	Total	%
Control	0	0	0	0	0	0	0	0	0	0	0	0
0.32	0	0	0	0	0	0	0	0	0	0	0	0
0.56	0	0	0	0	1	1	2	10	2	2	4	20
1.0	0	0	0	0	7	8	15	75	9	9	18	90
1.8	0	0	0	0	10	10	20	100	10	10	20	100
3.2	0	0	0	0	10	10	20	100	10	10	20	100

R₁¿ R₂= Replicates 1 and 2

Validity criteria fulfilled:

yes

Conclusions:

The 48-Hour EL50 for Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes to Daphnia magna based on nominal loading rates was 95 mg/L loading rate WAF with 95% confidence limits of 80 - 131 mg/L loading rate WAF. The No Observed Effect Loading rate was 32 mg/L loading rate WAF.

It is assumed that the Effective Loading rate (EL) is equivalent to the Effective Concentration (EC).
According to Regulation (EC) No. 1272/2008 and based on the results of this study, Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes is placed in Acute Category 3 for acute aquatic hazards.

Executive summary:

An acute toxicity test was carried out to determine the toxicity of Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes to Dapnia magna. The Daphnia magna were exposed to the nominal loading rates of 10, 18, 32, 56 and 100 mg/L. The 48-Hour EL₅₀ for Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes to Daphnia magna under static test conditions based on nominal loading rates was 95 mg/L loading rate WAF with 95% confidence limits of 80 - 131 mg/L loading rate WAF. The No Observed Effect Loading rate was 32 mg/L loading rate WAF.

It is assumed that the Effective Loading rate (EL) is equivalent to the Effective Concentration (EC).

According to Regulation (EC) No. 1272/2008 and based on the results of this study, Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes is placed in Chronic Category 3 for acute aquatic hazards.

Description of key information

A short-term toxicity study on Daphnia magna was carried out according to OECD Guideline 202 and EU Method C.2. Daphnia magna were exposed to Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes over a period of 48 hours.  The EL50 (48 h) was 95 mg/L. The NOELR (48 h) was 32 mg/L. 
 - Water Solubility: insoluble 
 - Pow: log10 Pow range: 6.78 to 9.23.
 - Biodegradation in water: Not readily biodegradable: 22% degradation after 28 days (OECD 301B).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

95 mg/L

Additional information

An acute toxicity test was carried out to determine the toxicity of Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes toDapnia magna. TheDaphnia magna were exposed to the nominal loading rates of 10, 18, 32, 56 and 100 mg/L in a freshwater static system. The 48-Hour EL₅₀for Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes toDaphnia magnaunder static test conditions based on nominal loading rates was 95 mg/L loading rate WAF with 95% confidence limits of 80 - 131 mg/L loading rate WAF. The No Observed Effect Loading rate was 32 mg/L loading rate WAF.

It is assumed that the Effective Loading rate (EL) is equivalent to the Effective Concentration (EC).

According to Regulation (EC) No. 1272/2008 and based on the results of this study, Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes is placed in Acute Category 3 for acute aquatic hazards.

According to Directive 67/548/EEC, Hydrolysed reaction products of furan-2,5-dione and C15-18-(linear and branched)-alkenes is classified as R52/53: Harmful to aquatic organisms and May cause long-term adverse effects in the aquatic environment.