Fatty acids, C16 and C18-22-unsatd., C16-18 and C18-unsatd. alkyl esters

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Remarks:

Summary of available data used for the endpoint assessment of the target substance.

Adequacy of study:

key study

Justification for type of information:

Refer to the analogue approach justification document provided in section 13.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Remarks on result:

other:

Remarks:

Source: CAS 36078-10-1, Dako, 2013, D. magna

Duration:

21 d

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Remarks on result:

other:

Remarks:

Source: CAS 36078-10-1, Dako, 2013, D. magna

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Remarks on result:

other:

Remarks:

Source: CAS 3687-45-4, Ecogreen, 2013, D. magna

Duration:

21 d

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

mortality

Remarks on result:

other:

Remarks:

Source: CAS 3687-45-4, Ecogreen, 2013, D. magna

Conclusions:

No long-term effects to aquatic invertebrates were observed up to the limit of water solubility (OECD 211, D. magna, RA to CAS 36078-10-1 + CAS 3687-45-4)

Reference 2

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 Aug - 11 Dec 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

LUBW Landesanstalt für Umwelt, Messungen und Naturschutz, Baden-Württemberg, Germany

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control, 6.75, 25 and 100 mg/L
- Sampling method: At the start (0 h), at the end (21 d) and at least once a week, 3.8 mL sample were taken from the test vessels A, C and E (100, 25 and 6.75 mg/L) and the control vessels at the beginning and at the end of a medium-change interval or chemical analysis.
- Sample storage conditions before analysis: Sample storage in 4 mL brown glass vials (filled up to the brim to avoid oxidative processes), maximum storage period of 8 d at 5 °C ± 3 °C before analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Due to the low water solubility of the test item, the study was performed with the water accommodated fraction (WAF) prepared with M4 medium according to OECD 202. The suspension was stirred for 24 hours at 20 – 22 °C in diffuse light. After stirring was stopped, the suspension was allowed to float and sediment for a period of 1 hour. After sedimentation and floatation period the WAF was clear and no particles could be observed. The test solution was taken from the middle of the suspension on the beaker using a glass tube and transferred into the test vessels. About 75 mL test solution was transferred into each test vessel (100 mL glass beakers). WAFs were prepared 3 x week.
- Controls: yes, M4 medium

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain/clone: Straus
- Source: German Federal Environmental Agency, department V 3.2, Berlin, Germany
- Age of parental stock: 0.5 – 21.5 h
- Feeding during test
- Food type: Suspension of the algae Desmodemus subspicatus
- Amount: 0 to 6 days: 0.1 mg C/Daphnia/day, 7 – 21 days: 0.2 mg C/Daphnia/day
- Frequency: daily

ACCLIMATION
- Acclimation conditions (same as test or not): The Daphnia are held at 20 ± 2 °C with a light : dark cycle of 16 : 8 h.
- Type and amount of food: Suspension of the algae Desmodesmus subspicatus, 0.1 mg C/Daphnia/days
- Feeding frequency: Fed during working days.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Test temperature:

20.6 - 21.8 °C

pH:

7.6 - 8.2

Dissolved oxygen:

7.7 - 8.5 mg O2/L

Nominal and measured concentrations:

Nominal: control, 6.75, 12.5, 25, 50, 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL beakers
- Type (delete if not applicable): covered with a transparent plastic foil
- Material, size, headspace, fill volume: glass, 100 mL, headspace: 25 mL, fill volume: 75 mL
- Renewal rate of test solution (frequency/flow rate): three times per week
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt-M4-medium, according to OECD 202. Before the addition of the M4 stock solutions the water was aerated for 2 hours to allow the dissolved oxygen concentration to reach its saturation.
- Intervals of water quality measurement: With every medium renewal the pH and oxygen content were measured.

OTHER TEST CONDITIONS
- Adjustment of pH:
- Photoperiod: 16 h light : 8 h dark
- Light intensity: 6.73 µmol/m2

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Immobility and reproduction (cumulative offspring was recorded daily.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Details on results:

- Mortality of parent animals: Only in one vessel (B: 50 mg/L) one dead Daphnia was found among the parent animals. This animal died because of an accident during the medium change and not because of influence of the test item.

- Body length and weight of parent animals: Control: mean 4.282 ± 0.152 mm; 6.8 mg/L: mean 4.285 ± 0.120 mm; 12.5 mg/L: mean 4.211 ± 0.146 mm; 25 mg/L: mean 4.284 ± 0.139 mm; 50 mg/L: mean 4.373 ± 0.122 mm; 100 mg/L: mean 4.278 ± 0.114 mm

- Time to first brood release or time to hatch: Control: mean 8.50 ± 0.00 day; 6.8 mg/L: mean 9.20 ± 1.16 day; 12.5 mg/L: mean 9.20 ± 0.82 day; 25 mg/L: mean 9.70 ± 1.14 day; 50 mg/L: mean 9.60 ± 2.62 day; 100 mg/L: mean 9.20 ± 1.64 day

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50 (24 h): 1.92 mg/L; EC50 (48 h): 1.50 mg/L

Reported statistics and error estimates:

Williams Multiple t-test procedure using ToxRat v2.09 (ToxRat solutions, Alsdorf, Germany)

Table 1: Effective Concentrations for Mean Cumulative Offspring per parent animal

Concentration [mg/L]	Mean offspring	Standard deviation	n	% Reduction
Control	99.0	6.80	10	0.0
6.8	97.0	8.80	10	2.3
12.5	102.0	6.10	10	-3.1
25.0	103.0	10.60	10	-4.6
50.0	97.0	37.60	9	1.9
100.0	108.0	10.40	10	-9.5

 

The analytical determination of the test substance concentration indicated that in almost all vessels no test substance concentration could be measured up to the limit of quantification (LLOQ) of 5 µg/L reflecting the very low water solubility of the test substance. Only in three test samples the test substance could be detected with a recovery rate of ≤ 4.4%.

Reference 3

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

05 Jan - 22 May 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Qualifier:

according to guideline

Guideline:

EU Method C.20 (Daphnia magna Reproduction Test)

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss Federal Office of Public Health, Bern, Switzerland

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control, 100 mg/L
- Sampling method: Duplicate samples were taken from the freshly prepared test medium of the single test concentration and from the control at one treatment period of the first, second and last week of the test (day 0, 2, 7 and 16, respectively). To determine the maintenance of the test substance concentration in test medium, stability samples were taken at the end of two test medium renewal periods of 48 h (day 2 and 9) and at the end of two renewal periods of 72 h (days 5 and 19).
- Sample storage conditions before analysis: All samples were stored deep-frozen (at about -20 °C) and protected from light immediately after sampling.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A dispersion with the nominal concentration of 100 mg/L was prepared prior to the start of the test and prior to each medium renewal by dispersing 400 and 200 mg of the test item (dosed in the range of 400 to 401 mg on days 0, 7 and 16 and in the range of 200 to 201 mg on days 2, 5, 9, 12, 14 and 19) in 4000 and 2000 mL of test water under intense stirring for 96 h at room temperature in the dark to avoid photolytic degradation. After the 96 h stirring period, the test item was filtered through a membrane filter (Schleicher & Schuell, Type NC45, pore size 0.45 µm). The test medium was prepared just before the start of the test (= addition of daphnids) and before each test medium renewal.
- Eluate: no
- Differential loading: no
- Controls: yes, test medium control

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Source: Originally supplied by the University of Sheffield/UK in 1992. Since this date, the clone is successfully bred at Harlan Laboratories.
- Age of parental stock (mean and range, SD): < 24 h
- Feeding during test
- Food type: food mixture containing a suspension of green algae of the species Scenedesmus subspicatus (freshly grown at the laboratory) and a fish food suspension. The fish food suspension was prepared by dispersing 10 g of powdered commercial fish diet (TETRA MIN Hauptfutter, obtained from TETRA-Werke, Melle, Germany) in 500 mL of test water. The suspension was allowed to stand for 4 hours. Then, 400 mL of the supernatant were taken, diluted 1:1 with test water and boiled. The suspension was stored deep frozen in small quantities until use.
- Amount: 0.20 mg TOC per Daphnia and day
- Frequency: daily
- Method of breeding: Each stock animal was maintained separately in a 100 mL glass beaker filled with about 80 mL culture medium and was transferred twice a week to fresh medium. The condition of the stock animals was frequently checked. No signs of stress were observed and the brood stock was healthy.

ACCLIMATION
- Acclimation period: The daphnids used for the test originated form parental daphnids that were at least 14 d old but not older than four weeks and were not first brood progeny.
- Acclimation conditions (same as test or not): same as test
- Type and amount of food: same as test
- Feeding frequency: three times a week

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

21 d

Test temperature:

20 - 21 °C

pH:

7.7 - 8.1

Dissolved oxygen:

8.2 - 9.2 mg O2/L

Nominal and measured concentrations:

nominal: control, 100 mg/L
measured: < LOQ in the control and treatment

Details on test conditions:

TEST SYSTEM
- Test vessel: beakers
- Type (delete if not applicable): covered with glass plates to reduce loss of water by evaporation and to avoid the entry of dust into the solutions.
- Material, size, headspace, fill volume: glass, 100 mL, headspace: 20 mL, fill volume: 80 mL
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): renewed on days 2, 5, 7, 9, 12, 14, 16 and 19.
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 20
- No. of vessels per control (replicates): 20

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water (Elendt M7 medium)
- Alkalinity: 0.9 mmol/L
- Water hardness: 250 mg/L as CaCO3
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light / 8 h dark (30 min transition period)
- Light intensity: 400 - 540 lux

EFFECT PARAMETERS MEASURED: Mortality and reproduction on days 0-2 and thereafter at least three times per week before renewal of test media. On the same dates, the test repliacates were observed for the number of living and dead offspring and for the presence of aborted eggs.

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

mortality

Details on results:

- Mortality of parent animals: 0% in control; 5% in treatment
- Type and number of behavioural abnormalities: No visible abnormalities were observed during the test.
- No. of offspring produced after 21 d per female: 142 (mean, control); 137 (mean, treatment)
- Time to first brood release or time to hatch: day 9 in control and treatment

Reported statistics and error estimates:

Student t-test (one-sided smaller, α = 0.05) with the mean values of living offspring.

Description of key information

No effects up to the limit of water solubility (OECD 211, D. magna); read-across

Additional information

There is no study available investigating the long-term toxicity of Fatty acids, C16 and C18-22 - unsatd., C16-18 and C18 unsatd. alkyl ester (CAS 90990-29-7) to aquatic invertebrates. Therefore, a read-across to the two structurally related source substances dodecyl oleate (CAS 36078-10-1) and (Z)-octadec-9-enyl oleate (CAS 3687-45-4) was conducted in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5.

Both source substances are characterized by fatty acids and fatty alcohols of similar chain lengths as the target substance. Therefore, they are considered suitable representatives for the assessment of the long-term toxicity of the target substance to aquatic invertebrates. A detailed analogue approach justification is provided in IUCLID section 13 of the technical dossier.

 

Both studies were conducted according to the OECD guideline 211 and GLP.

In the key study testing with the source substance dodecyl oleate (CAS 36078-10-1), Daphnia magna was exposed to water accommodated fractions (WAFs) with nominal loading rates of 6.75, 12.5, 25, 50, and 100 mg/L test item in a semi-static test for 21 d. No effects on the reproduction of D. magna were observed after 21 d. One dead animal was recorded at 50 mg/L but it was reported that it was no substance specific effect (accident during medium change). Therefore, a NOELR (21 d) of ≥ 100 mg/L (nominal) was derived. The chemical analysis resulted in concentrations below the analytical detection limit of 0.005 mg/L, reflecting the very low water solubility of the test item. Consequently, it can be concluded that the source substance dodecyl oleate will not exhibit chronic effects on aquatic invertebrates up to the limit of water solubility.

 

The second study investigating (Z)-octadec-9-enyl oleate (CAS 3687-45-4) was performed with the water flea D. magna under semi-static conditions. Due to the very low water solubility (< 0.05 mg/L) a limit test with a WAF of a nominal loading concentration of 100 mg/L was performed. No effects on reproduction and mortality were recorded after 21 d resulting in a NOELR (21 d) of ≥ 100 mg/L. 0% mortality was recorded in the control whereas 5% mortality was observed in the treatment. This mortality is regarded as natural and is tolerated by the test guideline. The chemical analysis resulted in concentrations below the analytical detection limit of 0.0423 mg/L, reflecting the very low water solubility of the test item. Thus, it can be concluded that the source substance (Z)-octadec-9-enyl oleate will not exhibit chronic effects on aquatic invertebrates up to the limit of water solubility.

 

Based on the structural and chemical similarity of the target and source substances, the target substance is expected to exhibit a similar ecotoxicological profile. Therefore, it can be concluded that the target substance Fatty acids, C16 and C18-22 - unsatd., C16-18 and C18 unsatd. alkyl ester (CAS 90990-29-7) will not exhibit long-term effects to aquatic invertebrates up to the limit of water solubility (< 0.05 mg/L at 20 °C, EU Method A.6).