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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:
April 30, 2015 through July 13, 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 305 (Bioaccumulation in Fish: Aqueous and Dietary Exposure) -I: Aqueous Exposure Bioconcentration Fish Test
Version / remarks:
Test method according to Japanese Test Guideline : "Method for Testing the Degree of Accumulation of Chemical Substances in Fish Body" stipulated in the "Testing Methods for New Chemical Substances" (March 31, 2011, No. 0331-7, Pharmaceutical and Food Safety Bureau, Ministry of Health, Labour and Welfare, March 29, 2011.
Principles of method if other than guideline:
No depuration-phase performed since BCFss established to be < or = 200 L/kg.
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
Nectaryl :
Lot Number : PE00106438
Purity : 97.6% (by GC area %)
Impurity : Unknown components 2.4% (by GC area %), all impurities less than 1% (by GC area %)
Expiry Date : 07 October 2015
Radiolabelling:
no
Details on sampling:
Water Samples :
Aqueous exposure samples were taken for Level 1 and Level 2 exposure solutions on Day -3 (prior to introduction of fish to the test system), 7, 14, 19, 22 and Day 28 after introduction of fish.

Fish Tissue Samples :
Samples of fish for analysis of tissue for Parent content were taken at Day -2 (prior to the start of exposure) and Day 29 for the Control Fish, and at Day 7, 14, 19, 22 and 28 in the Level 1 and Level 2 exposure set-ups.

Samples of fish from the Control set-up on Day -2 and Day 29 were also used for Lipid analysis.
Vehicle:
yes
Remarks:
N,N-dimethylformamide and HCO-40 (Ethoxylated Hydrogenated Castor Oil)
Details on preparation of test solutions, spiked fish food or sediment:
Two exposure levels and a Control system were run.

Level 1 (High Exposure Level) was 20 ppb
Level 2 (Low Exposure Level) was 2 ppb.

Exposure was performed via the aqueous phase.

Preparation of Level 1 (20 ppb) Stock Solution :
Nectaryl (500 mg) and HCO-40 (5.0 g) were dissolved in N,N-dimethylformamide to prepare a 1000 mg/L stock solution of Nectaryl (in 500 mL final volume).

Preparation of Level 2 (2 ppb) Stock Solution :
Nectaryl (100 mg) and HCO-40 (1.0 g) were dissolved in N,N-dimethylformamide to prepare a 1000 mg/L stock solution of Nectaryl (in 100 mL final volume). Then, 50 mL of this stock solution were diluted 10-times with N,N-dimethylformamide to prepare a 100 mg/L stock solution (final volume 500 mL).

Control :
HCO-40 (5.0 g) was dissolved in N,N-dimethylformamide to prepare a solution containing 10000 ppm of HCO-40 (final volume = 500 mL).


Test organisms (species):
Cyprinus carpio
Details on test organisms:
Species : Cyprinus carpio
Supplier : CERI, Kurume
Acclimatisation : External disinfection in aqueous solution containing OTC for fisheries (oxytetracycline hydrochloride) and Sodium Chloride. Fish were, thereafter, acclimatised in the following conditions : 18 days, Temperature 25 +/- 2°C. Mortality during acclimatisation was less than 5%.
Lot No. : TFC-150331-I
Length : 6.5 - 7.7 cm (at the beginning of the exposure 6.5 - 6.8 cm)
Age : > 6 months and < 12 months old
Route of exposure:
aqueous
Justification for method:
aqueous exposure method used for following reason: Test item suffiiciently water soluble and expected pathway of exposure.
Test type:
flow-through
Water / sediment media type:
natural water: freshwater
Total exposure / uptake duration:
28 d
Hardness:
Total Hardness (Ca, Mg) 15 mg/L
Test temperature:
24.2 to 24.8 °C during study.
pH:
pH 7.5 to 7.7 during study.
Dissolved oxygen:
Lowest dissolved oxygen concentration measured during study was 79% of the dissolved oxygen saturation concentration.
TOC:
Total Organic Carbon 0.6 mg/L
Conductivity:
Electric Conductivity 310 microS/cm
Details on test conditions:
In order to confirm the test levels to be applied in the aqueous exposure uptake phase, an acute toxicity study was performed with Ricefish (Oryzias latipes) which were exposed under semi-static conditions (renewal every 8 - 16 hours) for 96 hours. The 96 hour LC50 was determined to be 3.21 mg/L.

Test Tank : 100L glass containing 90L of exposure medium
Flow Rate of Test Water : 0.04 mL/minute for stock solution and 2000 mL/minute for dilution water, 2880L per day of test water equivalent to 32-volume exchanges per 24 hour period.
Stock Solution Bottle : 500 mL brown glass bottle (frequency of renewal 1 - 2 times per week)
Temperature of Test Water : Level 1 24.4 - 24.8 °C
Level 2 24.2 - 24.4 °C
Control 24.4 - 24.8 °C
Concen. of DO2 : Level 1 6.6 - 7.3 mg/L
Level 2 6.7 - 7.4 mg/L
Control 7.3 - 7.6 mg/L
pH of Test Water Level 1 7.5, 7.6
Level 2 7.5, 7.6
Control 7.7, 7.6
Aeration : No aeration was applied to the tanks due to the volatile nature of the test item. The lowest Dissolved Oxygen concentration measured was 79% of the Dissolved Oxygen saturation concentration which remains within the validity criteria (> 60%).
Nominal and measured concentrations:
Concentrations of the test item were maintained at > or = 91% of the nominal concentration and the variations were within +/- 20% of the average measured concentration.

The average exposure concentration of the three separated peaks of Nectaryl were, as follows :
Level 1 Peak 1 19.6 ppb
Peak 2 19.3 ppb
Peak 3 19.4 ppb
Level 2 Peak 1 2.05 ppb
Peak 2 2.00 ppb
Peak 3 2.02 ppb
Reference substance (positive control):
no
Details on estimation of bioconcentration:
BCF values have been calculated and are reported based on steady-state and also based on lipid normalisation to 5% content.

The lipid content determined in the fish :
Before initiation of experiment : 4.06%
After termination of experiment : 4.10%

BCF steady-state :
Level 1 (20 ppb) : Peak 1 150 L/kg (wet wt.)
Peak 2 92 L/kg (wet wt.)
Peak 3 200 L/kg (wet wt.)
Level 2 (2 ppb) : Peak 1 150 L/kg (wet wt.)
Peak 2 92 L/kg (wet wt.)
Peak 3 180 L/kg (wet wt.)

BCF Steady-State, Normalised to 5% Lipid Content :
Level 1 (20 ppb) Peak 1 180 L/kg (wet wt.)
Peak 2 110 L/kg (wet wt.)
Peak 3 240 L/kg (wet wt.)
Level 2 (2 ppb) Peak 1 180 L/kg (wet wt.)
Peak 2 110 L/kg (wet wt.)
Peak 3 230 L/kg (wet wt.)
Lipid content:
4.06 %
Time point:
start of exposure
Lipid content:
4.1 %
Time point:
end of exposure
Key result
Conc. / dose:
other: Maximum value of BCF Steady-State at both Levl 1 and Level 2 exposure, obtained for Peak 3
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
200 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: This value represents the maximum BCF steady state obtained for the three individual peaks of the test item.
Conc. / dose:
20.1 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
150 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 1
Conc. / dose:
19.2 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
92 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 2
Conc. / dose:
19.1 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
200 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 3
Conc. / dose:
2.01 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
150 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 1
Conc. / dose:
1.96 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
92 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 2
Conc. / dose:
1.96 µg/L
Temp.:
24 °C
pH:
7.6
Type:
BCF
Value:
180 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
steady state
Remarks on result:
other: Peak 3
Conc. / dose:
other: Maximum BCF steady state normalised to 5% lipid content
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
240 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Conc. / dose:
20.1 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
180 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 1
Conc. / dose:
19.2 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
110 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 2
Conc. / dose:
19.1 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
240 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 3
Conc. / dose:
2.01 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
180 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 1
Conc. / dose:
1.96 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
110 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 2
Conc. / dose:
1.96 µg/L
Temp.:
24 °C
pH:
7.6
Type:
other: BCF steady state normalised to 5% lipid content
Value:
230 L/kg
Basis:
whole body w.w.
Time of plateau:
7 d
Calculation basis:
other: BCF steady state normalised to 5% lipid content
Remarks on result:
other: Peak 3
Metabolites:
Two peaks derived from metabolites of the test item were detected on the mass chromatograms with a fragment of m/z 121 in test fish at exposure Levels 1 and 2. Chemical structures of these metabolites could not be elucidated. BCFs calculated using the sum of the test item and the metabolite on Day 28 were 190 and 200 L/kg (wet wt.) at Level 1 (20 ppb nominal exposure) and 170 and 210 L/kg (wet wt.) at exposure Level 2 (2 ppb nominal). The two metabolite peaks, as for Peaks 1, 2 and 3 of the Parent, had attained steady-state before the end of the 28-day exposure period.
Validity criteria fulfilled:
yes
Conclusions:
All Bioconcentration factors of Peaks 1, 2 and 3 were below 300 L/lg wet wt. throughout the entire uptake phase. A steady-state was reached by 28 days and steady state BCF values were 150 L/kg (wet wt.) for Peak 1, 92 L/kg (wet wt.) for Peak 2 and 180 - 200 L/kg (wet wt.) for Peak 3.

Two peaks derived from metabolites of the test item were detected on the mass chromatograms with a fragment of m/z 121 in test fish at exposure Levels 1 and 2. Chemical structures of these metabolites could not be elucidated. BCFs calculated using the sum of the test item and the metabolite on Day 28 were 190 and 200 L/kg (wet wt.) at Level 1 (20 ppb nominal exposure) and 170 and 210 L/kg (wet wt.) at exposure Level 2 (2 ppb nominal). The two metabolite peaks, as for Peaks 1, 2 and 3 of the Parent, had attained steady-state before the end of the 28-day exposure period.

Under the test conditions applied, Nectaryl (Peaks 1, 2 and 3) was observed to have a low potential to bioaccumulate in Common Carp. Although two metabolites with unknown chemical structures were detected on the chromatograms of test fish analysis, BCFs calculated using the sum of the Test Item and the metabolites were under 210 L/kg (wet wt.).

These results indicate that Nectaryl presents only low risk to bioaccumulate in fish.
Executive summary:

All Bioconcentration factors of Peaks 1, 2 and 3 were below 300 L/lg wet wt. throughout the entire uptake phase.  A steady-state was reached by 28 days and steady state BCF values were 150 L/kg (wet wt.) for Peak 1, 92 L/kg (wet wt.) for Peak 2 and 180 - 200 L/kg (wet wt.) for Peak 3.

Two peaks derived from metabolites of the test item were detected on the mass chromatograms with a fragment of m/z 121 in test fish at exposure Levels 1 and 2.  Chemical structures of these metabolites could not be elucidated.  BCFs calculated using the sum of the test item and the metabolite on Day 28 were 190 and 200 L/kg (wet wt.) at Level 1 (20 ppb nominal exposure) and 170 and 210 L/kg (wet wt.) at exposure Level 2 (2 ppb nominal).  The two metabolite peaks, as for Peaks 1, 2 and 3 of the Parent, had attained steady-state before the end of the 28-day exposure period.

Under the test conditions applied, Nectaryl (Peaks 1, 2 and 3) was observed to have a low potential to bioaccumulate in Common Carp.  Although two metabolites with unknown chemical structures were detected on the chromatograms of test fish analysis, BCFs calculated using the sum of the Test Item and the metabolites were under 210 L/kg (wet wt.).

These results indicate that Nectaryl presents only low risk to bioaccumulate in fish.

Description of key information

BCFsteady state Common Carp :

Peak 1 - 150 L/kg (wet wt)

Peak 2 - 92 L/kg (wet wt)

Peak 3 - 180 - 200 L/kg (wet wt)

Key value for chemical safety assessment

BCF (aquatic species):
200 L/kg ww

Additional information