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Environmental fate & pathways

Bioaccumulation: aquatic / sediment

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Reference
Endpoint:
bioaccumulation in aquatic species: fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2004-11-19 to 2004-12-17
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
This study was performed according to OECD Guideline 305. The use of a reference substance and the validity of the test were not reported in the study report.
Qualifier:
according to
Guideline:
OECD Guideline 305 (Bioconcentration: Flow-through Fish Test)
Deviations:
yes
Remarks:
Reference substance and validity of the test not reported
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Remarks:
GLP statement in date of December 27, 2004.
Details on sampling:
Analysis of test water and fish: Analysis of the test item in test water and test fish was performed with gas chromatography-mass spectrometry (GC-MS) analysis.
Frequency of analysis:
- Test water analysis: The test water of Level 1 and Level 2 were analyzed once before first analysis of test fish and at the same time as analysis of test fish thereafter. One sample was analyzed at each sampling time.
- Test fish analysis: The test fish of Level 1 and 2 was analyzed five times in duration of exposure. Four fish were taken out at each sampling time and divided into two groups (two fish per group), then both were analyzed individually.
The control fish was analyzed before the experimental starting and after the experimental completion. Four fish were taken out at each sampling time and divided into two groups (two fish per group), and then each group was analyzed individually. In addition, two fish were taken out and three groups (two fish per group) were used for measurement of lipid contents.
Vehicle:
yes
Details on preparation of test solutions, spiked fish food or sediment:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Level 1: Test sample and HCO-40 (10 times amount of test sample) were dissolved in N,N-dimethylformamide to prepare 1000 mg/L stock solution, which was diluted with N,N-dimethylformamide to prepare 200 mg/L stock solution of the test item.
- Level 2: Test sample and HCO-40 (10 times amount of test sample) were dissolved in N,N-dimethylformamide to prepare 1000 mg/L stock solution, which was diluted with N,N-dimethylformamide to prepare 20 mg/L stock solution of the test item.
- Controls: HCO-40 was dissolved in N,N-dimethylformamide to prepare 2000 mg/L stock solution of HCO-40.
Test organisms (species):
Cyprinus carpio
Details on test organisms:
TEST ORGANISM
- Common name: Common carp
- Source: Kurume Laboratory, Chemicals Evaluation and Research Institute, Japan.
- Lot No.: TFC-031222-VI
- Age at study initiation: Yearling fish
- Length at study initiation: 8.3 - 10.1 cm
- Feeding during test
- Food type: Feed for fry of common carp (Proteins content ≥ 4 3. 0 %; Lipid content ≥ 3.0 %)
- Amount: Amount corresponding to about 2 % of total body weight was fed twice a day in halves (once a day in all at holiday). The fish were starved for 24 h before sampling.

ACCLIMATION
- Conditions for acclimatization: After rearing, the fish were transferred to an acclimatization tank and acclimatized there after the external disinfection. The fish showing any abnormalities during this period were removed and the remainder was reared for 33 days in a flow-through system at the temperature of 25 ± 2 °C. The fish were checked for health conditions and transferred to test tanks. Thereafter the fish were reared at the same temperature in the flow-through system for 22 days, following the external disinfection.
- External disinfection: The external disinfection in the acclimatization tank was carried out in an aqueous solution containing 20 mg/L ELBAZIU and 7 g/L sodium chloride for 24 h. The external disinfection in test tanks was carried out in an aqueous solution containing 20 mg/L ELBAZIU and 7 g/L sodium chloride for 24 h.
Route of exposure:
aqueous
Test type:
flow-through
Water / sediment media type:
natural water: freshwater
Total exposure / uptake duration:
28 d
Hardness:
12 mg/L
Test temperature:
Temperature of test water
- Level 1: 24.2-25.0 °C
- Level 2: 24.0-24.5 °C
- Control: 24.0-24.7 °C
pH:
pH of test water
- Level 1: 7.9 - 8.0
- Level 2: 7.9 - 8.0
- Control: 8.0
Dissolved oxygen:
Concentrations of dissolved oxygen in test water
- Level 1: 7.2 - 7.7 mg/L
- Level 2: 7.2 - 7.6 mg/L
- Control: 7.7 - 8.3 mg/L
TOC:
0.1 mg/L
Salinity:
No data
Details on test conditions:
TEST SYSTEM
- Test vessel: Flow-through system assembled at this laboratory was used; Level 1 and Level 2: 70-L glass tank for volatile substance; Control: 70 L glass tank
- Flow rate of test water: 0.05 mL/min for stock solution and 2000 mL/min for dilution water, 2880 L/day of test water, were supplied.
- Renewal rate of test solution (frequency/flow rate): 1 - 2 times/week
- Number of fish (at the beginning of exposure): Level 1 and 2: 29 fish; Control: 12 fish

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Groundwater from the premises of Kurume Laboratory
- Residual Chlorine: < 0.01 mg/L
- Alkalinity: 100 mg/L
- Conductance: 260 µs/cm

OTHER TEST CONDITIONS
- Photoperiod: 14 h light/10 h dark (artificial light of white fluorescent lamp)

OBSERVATION, MEASUREMENT AND CLEANING OF TEST TANK
- Observation of test fish: Condition of test fish was observed visually twice a day (once a day at holiday).
- Flow rate of test water: Flow rate of stock solution and dilution water were measured with graduated cylinder and recorded once a day.
- Temperature of test water: Temperature of test water was measured with alcohol thermometer and recorded once a day.
- Concentration of dissolved oxygen in test water: Concentration of dissolved oxygen in test water was measured with dissolved oxygen probe and recorded twice a week.
- pH of test water: pH of test water was measured with pH meter once a week or more.
- Cleaning of test tank: In experimental period, excreta of common carp, dirt on test tank, etc. were removed about once a day.

RANGE-FINDING / PRELIMINARY STUDY
- Based on preliminary test results for the 96 h LC50 value and analytical detection limits, test concentrations of the test item were decided as follows. The control was set as a blank test.
Level 1: 5 μg/L
Level 2: 0.5 μg/L
Nominal and measured concentrations:
High exposure level (Level 1): 5 μg/L
Low exposure level (Level 2): 0.5 μg/L
Reference substance (positive control):
no
Details on estimation of bioconcentration:
Calculation of bioconcentration factor:
BCF = Cf/Cw

BCF: Bioconcentration factor
Cf : Concentration of the test item in test fish (subtracting FB) (ng/g)
Cw : The average concentration of the test item in test water for calculating BCF (μg/L)
FB : Average concentration of the test item or blank in the control test fish analyzed before and after the experiment (ng/g)

The average bioconcentration factor in m-th analysis
BCFm = (BCFa + BCFb) / n

BCFm : The average bioconcentration factor in m-th analysis (number of individual or group 2 (a,b))
BCFa,b : Each bioconcentration factor in m-th analysis of test fish
n: Number of individual or group in m-th analysis of test fish
The date of the analysis for test fish containing value when were not more than calculable BCF, BCFm is not calculated.
Lipid content:
2.69 %
Time point:
start of exposure
Remarks on result:
other: The measured lipid contents in the test fish before initiation of experiment
Lipid content:
2.97 %
Time point:
end of exposure
Remarks on result:
other: The measured lipid contents in the test fish after termination of experiment
Key result
Conc. / dose:
5 µg/L
Type:
BCF
Value:
> 8.4 - < 20 dimensionless
Basis:
not specified
Time of plateau:
28 d
Calculation basis:
steady state
Key result
Conc. / dose:
0.5 µg/L
Type:
BCF
Value:
> 8.4 - < 20 dimensionless
Basis:
not specified
Time of plateau:
28 d
Calculation basis:
steady state
Details on kinetic parameters:
None
Metabolites:
No data
Results with reference substance (positive control):
No data
Details on results:
Bioconcentration factors: BCFs of the test item were 8.4 - 20 at Level 1 and 8.4 - 20 at Level 2.

Results of test fish observation: No abnormality in behavior or appearance was noted.
Reported statistics:
No data

Concentration of the test item in test water: Concentrations of the test item were maintained at more than 78% of nominal concentrations. The variation of the concentrations of the test item was within± 20% of the average of the measured concentrations.

 

Table 5.3.1/1: Measured concentrations of test item in test water (µg/L) = Cw

 

Level

After 3 days

After 6 days

After 12 days

After 19 days

After 24 days

After 28 days

Average (Standard deviation)

1

4.63

4.44

4.55

4.66

4.85

4.95

4.68

(0.189)

2

0.448

0.417

0.424

0.403

0.437

0.388

0.420

(0.0217)

 

Table 5.3.1/2: Measured concentrations of test item in test fish (ng/g) = Cf

Level

After 3 days

After 6 days

After 12 days

After 19 days

After 24 days

After 28 days

1

Not measured

55.4

42.9

74.0

78.5

92.0

52.1

59.0

56.3

40.6

52.7

2

Not measured

3.65

8.74

5.79

6.46

6.90

6.61

7.68

7.15

6.00

3.99

Table 5.3.1/3: BCFs = Cf / Cw

 

Level

After 6 days

After 12 days

After 19 days

After 24 days

After 28 days

1

12

9.5

(11)

16

17

(17)

20

11

(16)

13

12

(12)

8.4

11

(9.7)

2

8.4

20

(14)

13

15

(14)

17

16

(16)

18

17

(18)

15

9.7

(12)

 

Table 5.3.1/4: Variation of BCFs (the values are indicated for five figures)

 

Level

BCF

After 19 days

After 24 days

After 28 days

Average of three results

1

The average BCF

15.836

12.306

9.6833

12.610

Variation rate from the average of three results (%)

25.581

2.4093

23.172

-

2

The average BCF

16.272

17.591

12.198

15.354

Variation rate from the average of three results (%)

5.9837

14.569

20.553

-

 

Because the variation of BCFs (average value) for the last three successive analyses was over ± 20%, the BCFss could not be calculated. However, it was evaluated that a steady-state was reached after 28 days because all BCFs were less than 100.

Validity criteria fulfilled:
yes
Conclusions:
According to the results of the bioconcentration test, it was confirmed that the steady state in both concentration levels was achieved on the 28th day of exposure. The bioconcentration factors (BCFs) were 8.4 - 20 at Level 1 and 8.4 - 20 at Level 2.
Executive summary:

The bioaccumulation study was performed according to the OECD Guideline No. 305. The study was carried out to determine the extent of accumulation of test item by the carp, Cyprinus carpio during the exposure period of 28 days.  

 

Nominal exposure levels: Level 1: 5 µg/L; Level 2: 0.5 µg/L.

Test was performed at 24.0-25.0 °C and dissolved oxygen was 7.2 - 8.3 mg/L. Analyses of test water and fish were conducted on Days 3, 6, 12, 19, 24 and 28 after exposure using gas chromatography-mass spectrometry (GC-MS). Bioconcentration factor (BCF) was calculated.

 

Concentrations of test substance in test water:

Mean concentration during the test period were as follows

Concentration Level 1:4.68± 0.189 µg/L

Concentration Level 2: 0.420±0.0217 µg/L

 

Bioconcentration factors: BCFs of the test item were 8.4 - 20 at Level 1 and 8.4 - 20 at Level 2. No abnormality in behavior or appearance was noted.

 

According to the results of the bioconcentration test, it was confirmed that the steady state in both concentration levels was achieved on the 28th day of exposure. The bioconcentration factors (BCFs) were 8.4 - 20 at Level 1 and 8.4 - 20 at Level 2.

Description of key information

OECD Guideline 305, GLP, key study, validity 2:

BCF = 8.4 to 20

Not bioaccumulable

Key value for chemical safety assessment

BCF (aquatic species):
20 dimensionless

Additional information

To assess the bioaccumulation potential of the registered substance, one key study is available. This study was performed according to the OECD Guideline 305 with GLP statement in oder to determine the extent of accumulation of registered substance by the carp, Cyprinus carpio, during the exposure period of 28 days. Two nominal exposure levels of the test substance were used (5 µg/L for the Level 1 and 0.5 µg/L for the Level 2, corresponding to mean measured concentrations of 4.68 +/- 0.189 µg/L and 0.420 +/- 0.0217 µg/L, respectively).

Analyses of test water and fish were conducted on Days 3, 6, 12, 19, 24 and 28 after exposure using gas chromatography-mass spectrometry (GC-MS) and the Bioconcentration factor (BCF) were calculate. According to the results of this study, it was confirmed that the steady state in both concentration levels was achieved on the 28th day of exposure and the BCFs of the test substance were comprised between 8.4 and 20 for both concentration levels (same result). No abnormality in behavior or appearance was noted.