(Z)-N-octadec-9-enylhexadecan-1-amide

Bioaccumulation: aquatic / sediment

Link to relevant study record(s)

Reference

Endpoint:

bioaccumulation in aquatic species: fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 May 2020 - 17 Feb 2023

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 305 (Bioaccumulation in Fish: Aqueous and Dietary Exposure) -III: Dietary Exposure Bioaccumulation Fish Test

Version / remarks:

2 October 2012

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Details on sampling:

- Sampling intervals/frequency for test organisms: day 0, 14 (uptake phase), 15, 18, 21, 28, 35, 42 (depuration phase)
- Sampling intervals/frequency for test medium samples: day 0, 14 (uptake phase)
- Sample storage conditions before analysis: chilled conditions
- Details on sampling and analysis of test organisms and test media samples: Food samples: extraction with tetrahydrofuran, vortex mixing for 1 min, sonication for 15 min, centrifugation at 3000 rpm for 15 min. Fish samples: homogenization by mincing with scissors, extraction with tetrahydrofuran by mixing with a blender for 5 min, sonication for 5 min. centrifugation at 2500 rpm for 15 min.

Vehicle:

yes

Remarks:

corn oil

Details on preparation of test solutions, spiked fish food or sediment:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: evaporation to dryness and solution in corn oil
- Controls: corn oil without test substance
- Concentration of vehicle in test medium: Test substance: 2.49 mL corn oil in 120.06 g food; Control: 2.4 mL corn oil in 120.16 g food
- Evidence of undissolved material: none reported

PREPARATION OF SPIKED FISH FOOD
- Details on fish food: Inicio 917 1.1 mm trout pellets (Biomar, Denmark) (50% protein and 16% fat content)
- Details of spiking: test substance dissolved in corn oil
- Quantity of corn oil vehicle per unit mass of fish food: Test substance: 2.49 mL corn oil in 120.06 g food; Control: 2.4 mL corn oil in 120.16 g food
- Method of mixing: manual shaking followed by rotational agitation for 5 min

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rain bow trout
- Source: Commcercial supplier, UK
- Length at study initiation: mean 8.13 cm
- Weight at study initiation: mean 5.37 g
- Weight at termination (mean and range, SD): 5.28 - 9.66 g
- Lipid content at test initiation: 4.26% (tissue weight)
- Health status: no abnormalities reported
- Description of housing/holding area: same as test, acclimatization for at least 2 weeks
- Feeding during test
- Food type: Inicio 917 1.1 mm trout pellets (Biomar, Denmark)
- Amount: 2% of body weight (on day 27 2.7% were fed in error)
- Frequency: daily

ACCLIMATION
- Acclimation period: at least 2 weeks
- Acclimation conditions: same as test
- Type and amount of food: Inicio 917 1.1 mm trout pellets (Biomar, Denmark)
- Feeding frequency: daily
- Health during acclimation: no mortality reported

Route of exposure:

feed

Justification for method:

dietary exposure method used because stable, measurable water concentrations cannot be maintained

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

14 d

Total depuration duration:

28 d

Hardness:

174 - 176 mg CaCO3

Test temperature:

13.9 - 14.3°C

pH:

7.6 - 8.1

Dissolved oxygen:

72 - 95% air saturation

TOC:

<1.0 - 1.3 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: aquaria
- Material, fill volume: glass, 100 L
- Aeration: continuous aeration
- Renewal rate of test solution: nominal 800 mL/min equal to approximately 1152 L per d
- No. of organisms per vessel: 50
- No. of vessels per concentration: 1
- No. of vessels per control / vehicle control: 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water, dechlorinated by activated carbon, softened to the desired water hardness
- Metals: none
- Pesticides: none
- Chlorine: none
- Holding medium different from test medium: no
- Intervals of water quality measurement: pH, oxygen, temperature daily

OTHER TEST CONDITIONS
- Photoperiod: 16 h light/ 8 h dark, with 20 min dawn and dusk transition periods
- Overall daily feeding rate used in the study: 2 % body weight, adjusted after every sampling interval (on day 27 erroneously 2.7% was fed)
- Number of feeds per day: 1

RANGE-FINDING / PRELIMINARY STUDY
The solubility in the vehicle corn oil was determined with nominal test substance concentrations of 10, 100, 1000 µg/g. The 1000 µg/g concentration contained undissolved material, while the lower concentrations were observed to be clear.

The homogeneity of the test substance on food was tested with the concentrations 10, 100 µg/g. The test substance dissolved in corn oil was administered to fish food same as in the test five times. The test substance concentrations were in the range of ±15%.

The palability of treated food was determined for 7 days. The feeding was considered to be normal. With a test substance concentration of 100 µg/g, radioactivity in fish was in the range of 1.05 - 1.91 µg equivalents/g after 7 d. This was determined to be sufficient for measuring a 95% reduction in body burden. Therefore, the concentration of 100 µg/g food was chosen for the test.

Nominal and measured concentrations:

Nominal 0 (control), 100 µg test substance/g food
Measured at d0: 0 (control), 112 µg/g
Measured at d14: 0 (control), 104 µg/g

Reference substance (positive control):

no

Lipid content:

5.33 %

Time point:

other: end of depuration

Lipid content:

4.65 %

Time point:

end of exposure

Remarks on result:

other: end of uptake/start of depuration

Lipid content:

4.26 %

Time point:

start of exposure

Conc. / dose:

100 µg/g food

Type:

BMF

Value:

0.386 dimensionless

Basis:

whole body w.w.

Calculation basis:

kinetic

Conc. / dose:

100 µg/g food

Type:

BMF

Value:

0.582 dimensionless

Basis:

whole body w.w.

Calculation basis:

other: Lipid normalized

Conc. / dose:

100 µg/g food

Type:

BMF

Value:

0.15 dimensionless

Basis:

whole body w.w.

Calculation basis:

kinetic, corrected for growth

Elimination:

yes

Parameter:

DT50

Depuration time (DT):

7 d

Rate constant:

growth-corrected depuration rate constant (d-1)

Value:

0.012

Rate constant:

growth-corrected half-life (d)

Value:

59.2

Details on kinetic parameters:

- Uptake rate constant k(f): 0.00176 kg food / kg fish / day
- Depuration rate constant k(e): 0.0455
- Indication of bi- or multiphasic kinetics: none

Metabolites:

2 metabolites could be identified:
M1: Palmitic acid, C16H32O2
M2: C32H63O3N

Details on results:

- Mortality of test organisms: None
- Behavioural abnormalities: None
- Observations on feeding behavior: Normal feeding bevaior
- Observations on body length and weight: No abnormalities
- Mortality and/or behavioural abnormalities of control: None
- Loss of test substance during test period: None

Reported statistics:

Data was analysed using GraphPad Prism (ver. 9.0.1) and R software with package OECD-TG305 bcmfR (v0.4-18). Three optimization models were tested against the data. Linear regression, a Box-Cox transformation and a Natural Log transformation. Box-Cox showed optimum results and was ultimately used for evaluation.

Table 1: Validity criteria for OECD 305-III

Criterion from the guideline	Outcome	Validity criterion fulfilled
The temperature variation is less than ± 2 °C	13.9 – 14.3°C	yes
The concentration of dissolved oxygen does not fall below 60% saturation	≥72%	yes
The concentration of the test substance in fish food before and at the end of the uptake phase is within a range of ± 20% (based on at least three samples at both time points)	104 – 112 % recovery	yes
The homogeneity of test item in feed is confirmed to be less than ±15 % from the mean (by analysis at least triplicate portions)	109%	yes
Concentrations of test substance are not detected, or are present only at typical trace levels, in un-spiked food or control fish tissues relative to treated samples	No test substance in untreated food or control fish reported	yes
The mortality or other adverse effects/disease in both control and treated fish is less than 10% at the end of the test; where the test is extended over several weeks or months, death or other adverse effects in both sets of fish should be less than 5% per month and not exceed 30% in all.	No mortality or adverse effects reported	yes

 

Validity criteria fulfilled:

yes

Remarks:

see Table 1 in "Any other information on results incl. tables" for further details

Description of key information

BMF: 0.582 kg food / kg fish (lipid normalised, OECD 305, dietary exposure)

Additional information

Experimental data on bioaccumulation of (Z)-N-Octadec-9-enylhexadecan-1-amide (CAS No. 16260-09-6) is available from an OECD 305 GLP-study by dietary route. 

Radio-labeled substance was administered to rainbow trout (Oncorhynchus mykiss) via food over a period of 14 days, followed by a 24 days depuration phase. The concentration of the test substance in food during the uptake phase was 100 µg/g.

The mean concentration of total radioactivity in fish tissue was 2.54 µg equivalents/g after 14 days of exposure, but declined to 0.514 µg equivalents/g. C hromatographic analysis of the extracts showed that the test substance had been metabolized, accounting for 42.0% sample radioactivity (1.07 µg/g) after 14 days of exposure. After 28 days depuration, crodamide 203 accounted for 5.8% sample radioactivity (0.03 µg/g).
The test substance was metabolized to palmitic acid (≤22.6% sample radioactivity), Met 5 (≤14.3%), Met 7 (≤56.8%), polar material (≤3.3%) and up to eight low level, unidentified metabolites (each comprising ≤4.5% sample radioactivity).

The calculation of effect levels and kinetic parameters showed a BMF of 0.582 kg food/kg fish (lipid normalized) and a growth corrected half-life of 59.2 d.