[No public or meaningful name is available]

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:

February 22, 2010 to July 5, 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: guideline study on Gas-to-liquids (GTL) substance covering the carbon range from C18 to C50

Qualifier:

according to guideline

Guideline:

other: Test Method Relating to New Chemical Substances , which prescribes the procedure for testing new chemical substances as required by the Chemical Substance Control Law of Japan

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Details on sampling:

- Sampling intervals/frequency for test organisms:
On days 8, 12, 15, 22 and 28 of the exposure period, four fish were taken from each test chamber and analysed for test substance.

- Sampling intervals/frequency for test medium samples:
On days 0, 8, 12, 15, 21 and 28 of the exposure period, test water was analysed for test substance.

- Details on sampling and analysis of test organisms and test media samples (e.g. sample preparation, analytical methods):
Test substance in test water was analysed by GC/MS.
The test substance in test fish was analysed by GC/MS.
For details of pretreatment methods please see attached background material (Analytical methods).

Vehicle:

no

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

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:
Preparation of Feed Solutions:
High concentration level -
Test substance (10 g) dissolved and diluted to 500 mL with 50% (w/v) Tween-80/tetrahydrofuran solution (400 mL) and tetrahydrofuran.
Concentration of test substance: 20000 mg/L
Concentration of Tween-80: 40000 mg/L

Low concentration level -
Feed solution of high concentration level (50 mL) diluted to 500 mL with tetrahydrofuran.
Concentration of test substance: 2000 mg/L
Concentration of Tween-80: 40000 mg/L

Feed solutions were renewed within 21 days and the 44 days stability was confirmed beforehand.

- Controls:
50% (w/v) Tween-80/tetrahydrofuran solution (400 mL) diluted to 500 mL with tetrahydrofuran.
Concentration of Tween-80: 400000 mg/L

Please see attached background material (Preparation of Test Solutions) for full details.

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM FOR ACUTE TOXICITY TEST
- Common name: Medaka
- Strain: Oryzia latipes
- Source: Tsunashima Fishing Co., 5-18-1 Tsunashimanishi, Kouhoku-ku, Yokohama, Kanagawa, Japan
- Length at study initiation: 2 ± 1 cm
- Weight at study initiation: Approximately 0.2 g
- Health status: At the start of acclimation, visual observation was made and abnormal fish were removed.
- Description of housing/holding area: Fish were reared in an aquarium with flowing water.


ACCLIMATION
- Acclimation period: 18 days.
- Acclimation conditions (same as test or not): Water temperature 24 ± 1ºC
- Type and amount of food: Food was given in an amount of 2% of the fish weight everyday. Feeding was discontinued during the 24 hours before testing.
- Health during acclimation (any mortality observed): Mortality during the 1 week before testing : <5%

TEST ORGANISM
- Common name: Carp
- Strain: Cyprinus carpio
- Source: Niikura Fish Farm, 1217, Shimoya, Isehara-eity, Kanagawa, Japan
- Age at study initiation: Less than a year after hatching
- Length at study initiation: 8 ± 4 cm
- Weight at study initiation: Approximately 6 g
- Health status: At the start of aaclimation, visual observation was made and abnormal fish were removed.
- Description of housing/holding area: Fish were reared in an aquarium with flowing water.

ACCLIMATION
- Acclimation period: Janaury 13, 2010 to March 4, 2010
- Acclimation conditions (same as test or not): Water temperature 24 ± 2ºC
- Type and amount of food: The food (Babygold, Kyorin) was given in an amount of 1.5% if the fish weight everyday.
- Health during acclimation (any mortality observed): Mortality during the 1 week before testing: <5%

Route of exposure:

aqueous

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

29 d

Hardness:

49 mg/l as CaCO3

Test temperature:

24±2°C

pH:

6.0 - 8.5

Dissolved oxygen:

≥ 60% of the saturation (≥ 5 mg/l at 24°C)

TOC:

Dilution water quality results give TOC as <0.3 mg/l

Salinity:

Dilution water quality results give conductivity as 14 mS/m

Details on test conditions:

TEST SYSTEM
- Test vessel:
The bioconcentration test system was set up in duplicate for different concentrations and two groups of carp were kept in water containing the test substance. In addition, the system was also set up for control and a group of carp was kept in water without the test substance.

Each feed solution prepared was supplied to a mixing glass tube with a metering pump for the feed solution, diluted to the respective nominal concentration by mixing with dilution water delivered through a metering pump for dilution water, and then supplied to each test chamber. The test system is shown in Figure 1 (attached background material - Figure 1 bioconcentration test system).

- Aeration: : Continuous

- Type of flow-through (e.g. peristaltic or proportional diluter): Continuous flow-through dilution system
- Renewal rate of test solution (frequency/flow rate): Feed solutions were renewed within 21 days.

- No. of organisms per vessel:
Number of fish (at the start of the exposure period)L
High concentration level: 28 fish in 50 L test water (>1 L/day/g)
Low concentration level: 28 fish in 50 L test water (>1 L/day/g)
Control: 12 fish in 50 L test water (>1 L/day/g)


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
To prepare the dilution water, municipal tap water was treated with activated charcoal, and sodium thiosulfate solution was added to neutralize trace free chlorine.

OTHER TEST CONDITIONS
- Photoperiod: Approximately 16 hr/day
- Light intensity: Hf fluorescent lamp with wavelengths of 400-700 nm

RANGE-FINDING / PRELIMINARY STUDY
ACUTE TOXICITY TEST-

- Test concentrations:
0 (control) and 50 mg/l.

- Results used to determine the conditions for the definitive study:
In order to confirm that the concentration levels of bioconcentration test were less than a hundredth and a thousandth of the 96-hour LC50 to Medaka, the acute toxicity test was performed.

The test was conducted according to the OECD Guideline for Testing of Chemicals No. 203 "Fish, Acute Toxicity Test" (1992). The concentration of the test substance was not measured, and LC50 was evaluated based on the nominal concentration.

Exposure period: 96 hours
Concentration level: 0 (control), 50 mg/l
Number of fish: 10 /group
Volume of test water: 3.0 L
Water temperature: 24 ± 1°C
Test mode: Semi-static, renewal of test water every 24 hours.
Aeration: Continuous

Results:
The 96-hour LC50 of the test substance to Mefaka was > 50 mg/l. (No mortality in 50 mg/l exposure group ).

From the result of the preliminary acute toxicity test with Medaka, the 50% lethal concentration (LC50) for 96 hours of the test substance was >50 mg/L. Therefore the bioconcentration test was accordingly conducted at exposure concentration levels of 0.5 and 0.05 mg/L, less than a hundredth and a thousandth ofthe 96-hour LC50.

Determination of the Exposure Period:
The exposure period was set to be 29 days, it means that the last sampling for analysis was perfonned on day 28, because the bioaccumulation possibility of the test substance in fish was presumed low under the prescribed test condition.

Nominal and measured concentrations:

Nominal concentrations:
High concentration level: 0.5 mg/l
Low concentration level: 0.05 mg/l

Measured concentrations:
The mean concentration of the test substance during the exposure period was 0.432 mg/l at the high concentration level, and 0.0436 mg/l at the low concentration level.

Reference substance (positive control):

no

Details on estimation of bioconcentration:

The bioconcentration factor (BCF) is calculated by the following equation:
BCFn = Cf,n / Cw,s

The steady-state bioconcentration factor (BCFss) is calculated by the following equation:
BCFss = Cf,s / Cw,s

Where :
BCFn: The bioconcentration factor after n days
BCFss:The steady-state bioconcentration factor
Cf,n: Concentration ofthe test substance in test fish after n days (µg/g)
Cw,n: Mean concentration ofthe test substance in test water over n days (mg/L)
Cf,s,: Mean concentration in test fish at steady-state (µg/g)
Cw,s: Mean concentration in test water at steady-state (mg/L)

Steady state:
Steady-state is decided to be reached, when three successive BCF at intervals of 48 hours or longer are within 20% of each other. Cf,s, is calculated from last three successive data. Cw,s is the average of all data during steady-state period.

When the obtained BCF are all less than 100, it is also considered that steady-state is reached. In this case, three successive BCF are not
necessary within 20% of each other. The BCFss is not calculated by above equation and BCFss is just declared "< (maximum BCF value) "or
≤ (maximum BCF value).

Lipid content:

7.3 - 8.4 %

Time point:

start of exposure

Lipid content:

3.9 - 5 %

Time point:

end of exposure

Type:

BCF

Value:

<= 6 dimensionless

Basis:

whole body d.w.

Time of plateau:

28 d

Calculation basis:

steady state

Remarks on result:

other: Conc.in environment / dose:0.5 mg/l (hign concentration level)

Type:

BCF

Value:

<= 29 dimensionless

Basis:

whole body d.w.

Time of plateau:

28 d

Calculation basis:

steady state

Remarks on result:

other: Conc.in environment / dose:0.05 mg/l (low concentration level)

Results with reference substance (positive control):

No positive control.

Details on results:

Monitoring of Test Conditions:
During the test period, the dissolved oxygen in all water chambers was kept at > 60 % (7.2-8.4 mg/L) of the saturated oxygen concentration and the water temperature was kept at 24°C ± 2°C (23.9-24.4°C), both of which satisfied the test conditions. The pH value was kept at 7.3-7.6, which was within the proper range for fish rearing (6.0-8.5).

During the test period, there was no abnormality in shape of the body or in swimming and eating behaviour in the fish at all concentration levels throughout the test period. Therefore, it was confirmed that the test fish had been reared appropriately.

Lipid content:
7.9% (n=3, 7.3 - 8.4%) at the start of the exposure period
4.3% (n=3, 3.9-5.0%) at the end of the exposure period
The Standard Operation Procedure of the laboratory establishes the criteria of lipid content as below.
- The average of lipid content falls within 3.0% to 6.0%.
- The variation of lipid content at the end against at the start falls within ±25%.
The average oflipid content at the start was 7.9% and exceeded the criterion. With the high average of start lipid content, the variation of lipid content was 54% and also exceeded the criterion. The feeding through the acclimation and the exposure period was performed properly. Therefore, the reason for the high lipid content at the start was unexplained.

Concentration of Test Substance in Test Water:
The mean concentration of the test substance during the exposure period was 0.432 mg/L at the high concentration level, and 0.0436 mg/L at the low concentration level, both of which were > 80% of the nominal concentrations. The variations of the test substance concentrations in the test waters at both concentration levels were kept within ±20% of the mean measured values.

BCF of Test Substance in Test Fish:
The bioconcentration factor (BCF) during the exposure period was <2 to 6 at the high concentration level, and <11 to 29 at the low concentration level.

At both of the high concentration level and low concentration level, the variations of mean BCF at the three consecutive measurements were not confirmed to fall within ±20%. However, all of the BCF values during the exsposure period were less than 100, thus the BCF were considered to reach the steady-state. The bioconcentration factor at the steadystate (BCFss) was ≤6 at the high concentration level, and ≤29 at the low concentration level.

Please see attached background material for result tables.

Based on the results, the bioaccumulation potential of the test substance in fish tissues is judged to be low.

Validity criteria fulfilled:

yes

Remarks:

All results satisfied the validity criteria specified for this study.

Conclusions:

The bioconcentration factor at the steadystate (BCFss) was ≤6 at the high concentration level, and ≤29 at the low concentration level.

Based on the results, the bioaccumulation potential of the test substance in fish tissues is judged to be low.

Executive summary:

Methods:

The study was conducted in accordance with the Test Method Relating to New Chemical Substances <Bioconcentration test of chemical substances in fish and shellfish> (Yakushokuhatsu No.1121002, Heisei 15.11.13 Seikyoku No.2, Kanpokihatsu No.031121002, November 21, 2003; the latest revision, November 20, 2006), which prescribes the procedure for testing new chemical substances as required by the Chemical Substances Control Law of Japan.

Test substance in test water (nominal concentrations)

High concentration level: 0.5 mg/L

Low concentration level: 0.05 mg/L

Control: 0 mg/L

Vehicles in test water

High concentration level: Tween-80 10 mg/l, Tetrahydrofuran 15 ppm (v/v)

Low concentration level: Tween-80 1 mg/l, Tetrahydrofuran 24 ppm (v/v)

Control: Tween-80 10 mg/l, Tetrahydrofuran 15 ppm (v/v)

Supply of test water: Continuous flow-through dilution system (800 L/day)

Exposure period: 29 days.

The last sampling date was day 28. On day 29, the bioconcentration factors were confirmed to have reached the steady-state, which enable exposure to be completed.

Analytical Method:

After pretreatment, the test substacne was determined by gas chromatography mass spectrometer (GC/MS)

Lipid content in test fish:

7.9% (n=3, 7.3 -8.4%) at the start of the exposure period

4.3% (n=3, 3.9 -5.0%) at the end of the exposure period

96 -hour LC50 to Medaka: >50 mg/l

Results:

The bioconcentration factors (BCF) obtained in this study were as follows:

Exposure period		8	12	15	21	28
High concentration level
Mean conc. In test water (mg/l)		0.433	0.448	0.436	0.434	0.432
	Group 1	<2	1	1	<2	<2
	Group 2	6	2	<2	<2	<2
Low concentration level
Mean conc. In test water (mg/l)		0.0462	0.0452	0.0442	0.0431	0.0436
	Group 1	<11	11	<11	12	<11
	Group 2	<11	<11	16	13	29

 

BCFss: The steady-state bioconcentration factor

 

High concentration level BCFss: ≤6

Low concentration level BCFss: ≤29

At both of the high concentration level and low concentration level, the variations of mean BCF at the three consecutive measurements were not confirmed to fall within ±20%. However, all of the BCF values during the exsposure period were less than 100, thus the BCF were considered to reach the steady-state. The bioconcentration factor at the steadystate (BCFss) was ≤6 at the high concentration level, and ≤29 at the low concentration.

Based on the results, the bioaccumulation potential of the test substance in fish tissues is judged to be low.

Description of key information

bioconcentration factor for closely related substance 'Distillates (Fischer-Tropsch), heavy, C18-50 - branched, cyclic and linear' (covering the carbon range from C18 to C50) at the steady state (BCFss):  - ≤6 at the high concentration level (0.5 mg/L nominal) - ≤29 at the low concentration level (0.05 mg/L nominal).

Additional information

Bioaccumulation data (aquatic / sediment) are not available for 'Paraffin waxes (Fischer-Tropsch), full-range, C15-50 - branched and linear':

A bioaccumulation fish test was carried out on the closely related substance 'Distillates (Fischer-Tropsch), heavy, C18-50 - branched, cyclic and linear (covering the carbon range from C18 to C50) using a Test Method relating to New Chemical Substances <Bioconcentration test of chemical substances in fish and shellfish>, which prescribes the procedure for testing new chemical substances as required by the Chemical Substance Control Law of Japan.

The bioconcentration factor at the steady state (BCFss) was ≤6 at the high concentration level (0.5 mg/L nominal), and ≤29 at the low concentration level (0.05 mg/L nominal).

Based on the results, the bioaccumulation potential of the substance 'Distillates (Fischer-Tropsch), heavy, C18-50 - branched, cyclic and linear' in fish tissues is judged to be low.

Some further information is available regarding bioaccumulation, from a solid phase micro extraction (SPME) study. SPME is a biomimetic extraction system that tries to mimic the way organisms extract chemicals from water.

A biomimetic extraction technique using SPME in conjunction with gas chromatography (GC) was employed to assess the level of water soluble hydrocarbons and hence the potential toxicity of water accommodated fractions (WAFs) for a range of GTL products. In addition, the toxicity of WAFs was also monitored using an in-vitro MicrotoxTMassay, a biosensor-based measurement system based on Vibro Fischeri bacteria. In this study the majority of GTL samples analysed using SPME-GC indicated a low total peak area for soluble hydrocarbons (on average less than 12 000) and hence toxicity, with no detectable toxicity using the MicrotoxTM assay. The low total peak area for soluble hydrocarbons for these substances is indicative of a low potential to bioaccumulate.

It can be concluded that the test materials consisting of shorter chain hydrocarbons (i.e. C6-C16) sample were the only samples to cause a sufficient decrease in light emission for a detectable toxicity. All the other GTL samples - including the supporting substance GTL Gasoil covering the carbon range from C8 to C26 - did not cause a greater than 25% change in light emission indicating no detectable toxicity.

For the purposes of PBT assessment, a logistic function QSAR model has been used to predict BCF values for the individual constituents (See Endpoint Study Record PFA, 2014). Use of the logistic function QSAR model predicts a worst-case scenario for high log Kow constituents as it assumes a plateau at log BCF 3.3 (BCF 2000) for constituents with log Kow >4.3 (equivalent to carbon chain length ~9).

The logistic function QSAR model indicates that all constituents may reach the criteria for B. There are no vB constituents.

Finally, supporting information is available about the feasibility to undertake an OECD 305 study on whole substance. On the basis of the results of this study (which was conducted on the closely related substance GTL Gasoil covering the carbon range from C8 to C26), it is not feasible to undertake such a study.

In addition, based on OECD 307 (Aerobic and Anaerobic Transformation in Soil) studies (refer to Section 5.2.3), it is not possible to identify any constituents as P or vP, and thus warranting further investigation for their bioaccumulation potential either in a fish bioconcentration or dietary study.