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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:

2020-04-16 - 2021-04-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Also conducted according to GLP.

Justification for type of information:

See attached justification.

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

October 20212

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Details on sampling:

- Sampling intervals/frequency for test organisms: fish were sampled five times during the uptake phase (7, 13, 21, 36 and 60d).
- Sampling intervals/frequency for test medium samples: at the start of the uptake phase, and at every fish sampling time for the test concentration. At the end of the uptake phase for the control.
- Details on sampling and analysis of test organisms and test media samples (e.g. sample preparation, analytical methods):
Four fish were taken at each sampling time and two were pooled for the analysis. The analysis was carried out at intervals of at least tow days with the final analysis conducted on day 60. The fish from the control were analyzed once, at the end of the uptake phase.

Vehicle:

yes

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

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Fresh stock solution was prepared every two weeks. The test item was dissolved in DMF to prepare the stock solution.
- Controls: Vehicle control prepared using DMF
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N,N-Dimethylformamide

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM
- Common name: Common carp
- Source: CERI Kurume
- Age at study initiation (mean and range, SD): yearling fish
- Length at study initiation (length definition, mean, range and SD): 7.2-7.6 cm
- Lipid content at test initiation (mean and range, SD): 5.22%
- Feeding during test
- Food type: feed for colored carp, Kyorin Food Industries
- Amount: 2% of total body weight
- Frequency: Daily, two feedings

ACCLIMATION
- Acclimation period: 33d
- Acclimation conditions (same as test or not): same
- Type and amount of food: feed for colored carp, Kyorin Food Industries, 2% of total body weight
- Feeding frequency: Daily, two feedings
- Health during acclimation (any mortality observed): < 5% mortality observed

Route of exposure:

aqueous

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

60 d

Total depuration duration:

7 d

Hardness:

18.4-18.6 mgCaCO3/L

Test temperature:

24.5 - 25.6 °C

pH:

7.8-8.0

Dissolved oxygen:

7.0 - 7.5 mg/L

TOC:

9.7-10.3 mgC/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 70-L glass vessel
- Type (delete if not applicable): open
- Aeration: yes
- Renewal rate of test solution (flow rate): 2880L/day
- No. of organisms per vessel: 34 (test concentration), 16 (control)
- No. of vessels per concentration (replicates): 1
- No. of vessels per control / vehicle control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Groundwater
- Particulate matter: < 1 mg/L
- Metals: Below detection limits
- Pesticides: Below detection limits
- Chlorine: < 0.02 mg/L
- Alkalinity: 110 mg/L
- Conductance: 280 µS/cm

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 14h light:10h dark

RANGE-FINDING / PRELIMINARY STUDY
- Test concentrations: 0, 0.428, 0.940, 2.078, 4.55, 10.0 mg/L (acute toxicity fish)
- Results used to determine the conditions for the definitive study: 96h LC50 3.07 mg/L

Nominal and measured concentrations:

Nominal: 0.025 mg/L
Measured: 0.0214 - 0.0244 mg/L

Lipid content:

6.75 %

Time point:

end of exposure

Remarks on result:

other: average lipid content after the uptake phase

Conc. / dose:

0.025 mg/L

Type:

BCF

Value:

210 - 750 L/kg

Basis:

whole body w.w.

Remarks on result:

other: Range based on all four constituents of the reaction mass. (Not normalised)

Conc. / dose:

0.025 mg/L

Value:

260 L/kg

Basis:

normalised lipid fraction

Calculation basis:

steady state

Remarks on result:

other: BCF SSL for one of the 4 isomers. Could not be calculated for the others due to variability amongst fish.

Details on kinetic parameters:

- Uptake rate constant k(s):
- Depuration rate constant k(e):
- Indication of bi- or multiphasic kinetics:
- Computation / data analysis:

Metabolites:

The metabolites could not be identified. Therefore, a depuration test was conducted after the uptake phase and the bioconcentration potential of the metabolites estimated from their depuration rate.
The metabolites were no longer detected in the fish after 7 days. the depuration constant (k2) was calculated to be 0.38/day, which results in a calculated BCF of 810 L/kg.

Details on results:

- Mortality of test organisms: No
- Behavioural abnormalities: No
- Mortality and/or behavioural abnormalities of control: No
- Loss of test substance during test period: No

The BCF was calculated according to

BCF = Cf/ av. Cw

BCF: bioconcentration factor, L/kg

Cf: concentration of test item in test fish (µg/g) substracting the average concentration of control fish

Av. Cw: average concentration of test item in test water (mg/L)

 

The steady-state was defined as having been reached when four successive measured concentrations of the test item in fish made at intervals of at least two days were within +- 20% of each other and there was no significant increase of concentration of the test item in test fish in between the first and last successive measurements.

The steady-state lipid-normalized BCF was calculatd according to the following:

BCFss = Av. Cfs / av. Cw 

BCFss: bioconcentration factor at steady-state, L/kg

Av. Cf: average concentration of test item in test fish at steady-state (µg/g) substracting the average concentration of control fish

Av. Cw: average concentration of test item in test water (mg/L)

 

The 5% lipid normalized BCFss (BCFssL) was calculated as follows

BCFssL = BCFss X 5 / Lave

Lave: Average fish lipid conctent in test fish after the uptake phase (%)

 

The growth rate constant was calculated based on weight measurements at the beginning and end of the uptake phase.

Test fish: 0.0127/day

Control: 0.0126/day

 

Table. Measured concentrations of test item in test water

	Measured concentration (mg/L)
Peak	Day 3	Day 7	Day 13	Day 21	Day 36	Day 60	Average (SD)	% of nominal
1	0.0201	0.0201	0.0216	0.0223	0.0217	0.0214	0.0214 (0.00080)	85.6
2	0.0221	0.022	0.0227	0.0232	0.0233	0.0234	0.0229 (0.00060)	91.6
3	0.0229	0.0234	0.0239	0.0241	0.0243	0.0242	0.0240 (0.00033)	95.9
4	0.0233	0.024	0.0246	0.0244	0.0246	0.0245	0.0244 (0.00028)	97.8

 

Table. Measured concentration of test item in test fish (µg/g)

Peak	Day 7	Day 13	Day 21	Day 36	Day 60
1	10.2 9.56	16.0 10.4	7.36 9.47	11.0 11.8	6.59 8.39
2	6.70 5.89	9.12 6.80	4.91 6.63	6.92 7.70	4.70 5.67
3	8.58 8.50	11.7 9.22	7.03 9.16	8.82 9.68	5.40 7.25
4	8.85 8.81	11.8 8.88	6.74 9.12	8.74 9.48	6.15 8.44

 

Table. BCF (L/kg; () average)

Peak	Day 7	Day 13	Day 21	Day 36	Day 60
1	480 450 (460)	750 490 (620)	340 440 (390)	510 550 (530)	310 390 (350)
2	290 260 (270)	400 300 (350)	210 290 (250)	300 340 (320)	210 250 (230)
3	360 350 (360)	490 380 (440)	290 380 (340)	370 400 (390)	230 300 (260)
4	360 360 (360)	480 360 (420)	280 370 (320)	360 390 (370)	250 350 (300)

 

Table. BCFss and BCFssL of peak 4

	BCFss (L/kg)	BCFssL (L/kg)
Peak 4	350	260

The BCFss for peaks 1 to 3 could not be obtained because the test item concentration of those peaks in test fish varied by more than 20 %. The tes item concentration was within +-20% of the nominal value and there wer no significant differences in fish lipid content or growth rate. therefore, the variation is considered to be individual difference of organisms.

 

Validity criteria:

- The temperature was maintained within +-2 °c of the set water temperature of 25 °C.

- The concentration of dissolved oxygen did not fall below 60%.

- The concentration of the test item in the water was maintained within +-20% of the mean measured concentration during the test period.

- No mortality or other adverse effect was observed in the control and test fish.

- The concentration of the test item was below the limit of solubility.

Validity criteria fulfilled:

yes

Conclusions:

The BCF values obtained are all less than 1000 L/kg. Therefore it is concluded that the test item is not bioaccumulative.

Executive summary:

The bioaccumulation potential of the test item was investigated in an OECD TG 305-I study conducted according to GLP. The test was conducted under flow-through conditions at 1 single concentration of 0.025 mg/L. The test concentration was maintained within +- 20% of the nominal concentration throughout the duration of the test. A stock solution of the test item was prepared using dimethyl formamide (DMF) and dose via a computer-controlled system. The uptake phase lasted 60 days while the depuration phase lasted 7 days. The BCF, the BCFss and the BCFssL were calculated for each peak of the test item individually.

 

The BCF of all four constituents as well as the those estimated for the metabolits were all below 1000 L/kg. The BCFss and ssL calculated for the only peak for which it was possible were below 500 L/kg.

	BCF (L/kg)
Peak 1	310 - 750
Peak 2	210 - 400
Peak 3	230 - 490
Peak 4	250 - 480

 

	BCFss (L/kg)	BCFssL (L/kg)
Peak 4	350	260

 

Therefore, the test item is not considered bioaccumulative.

Description of key information

The BCF values obtained are all less than 1000 L/kg. Therefore it is concluded that the test item is not bioaccumulative.

 

Key value for chemical safety assessment

BCF (aquatic species):

750 L/kg ww

Additional information

The bioaccumulation potential of the test item was investigated in an OECD TG 305-I study conducted according to GLP. The test was conducted under flow-through conditions at 1 single concentration of 0.025 mg/L. The test concentration was maintained within +- 20% of the nominal concentration throughout the duration of the test. A stock solution of the test item was prepared using dimethyl formamide (DMF) and dose via a computer-controlled system. The uptake phase lasted 60 days while the depuration phase lasted 7 days. The BCF, the BCFss and the BCFssL were calculated for each peak of the test item individually.

 

The BCF of all four constituents as well as the those estimated for the metabolits were all below 1000 L/kg. The BCFss and ssL calculated for the only peak for which it was possible were below 500 L/kg.

	BCF (L/kg)
Peak 1	310 - 750
Peak 2	210 - 400
Peak 3	230 - 490
Peak 4	250 - 480

 

	BCFss (L/kg)	BCFssL (L/kg)
Peak 4	350	260

 

Therefore, the test item is not considered bioaccumulative.