Diquat dibromide

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

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

Deviations:

yes

Remarks:

exposure period 14 days, study pre-dates OECD guidance establishment

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Details on sampling:

WATER SAMPLING
- Total radioactivity: Three 10 mL aliquots of water were sampled daily at a depth of approximately 5 cm, and analysed by liquid scintillation counting. Samples were taken from the control to determine "background".
- Characterisation of radioactivity in the water: On days 0, 8 and 14, water was sampled from the treated tank by siphoning from approximately 5 cm below the surface. A sub-sample, 280 mL, was transferred to a round-bottomed flask. The flask had previously been silanized (using 10% dichlorodimethyl silane in toluene) and the sample was spiked with 10 mg of unlabelled test substance cation to minimise losses of 14C-test substance cation by adsorption onto glassware during rotary evaporation. Before rotary evaporation aliquots were samples for LSC.

The sample was reduced in volume by rotary evaporation under vacuum to < 50 mL and made up to 50 mL in a graduated flask with distilled water; 3 x 500 µL aliquots were sampled for LSC to determine recovery after rotary evaporation. A sub-sample (approximately 5 mL) was taken for analysis by HPLC. A 100 µL aliquot of the concentrated sample was analysed by TLC.

FISH SAMPLING
Fish (whole and muscle) were sampled on days 0, 1, 3, 7, 8, 10 and 14 of the exposure phase, and on days 1, 3, 7, 14 and 28 of the depuration phase. At each sampling date, 7 fish were removed from the treated tank and asphyxiated with carbon dioxide. They were then rinsed with tap water and their lengths and weights determined.
- Whole fish: Three fish were sample, cut into pieces and combusted separately to determine total 14C-content.
- Viscera: The remaining four fish were dissected and the viscera wet weighed and combusted separately. The viscera constituted approximately 7% of the weight of the whole fish.
- Muscle: After evisceration, the head (with gills) and fins were removed from the dissected fish. The remainder, referred to as muscle (lateral musculature, skin and bones), was wet weighed, cut into pieces if necessary and combusted separately. The muscle constituted approximately 60% of the weight of the whole fish.

Vehicle:

yes

Remarks:

ethanol:water (1:1)

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

See 'Details on test conditions'

Test organisms (species):

Lepomis macrochirus

Details on test organisms:

Bluegill sunfish (Lepomis macrochirus) were obtained from a commercial supplier and held in flowing, dechlorinated water for at least 5 weeks before introduction into the test system. The fish were acclimatised to the test temperature (20 °C) for 1 week prior to the start of the test. There were no fish dead from approximately 300 in the week before testing. Measurements of fish removed for analysis during the study gave a mean weight of 4.3 g (standard deviation 1.6) and a mean length of 53 mm.

Route of exposure:

aqueous

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

14 d

Total depuration duration:

28 d

Hardness:

244 mg CaCO3/L

Test temperature:

18 - 22 °C

pH:

7.8 - 8.3

Dissolved oxygen:

> 7.0 mg O2/L

TOC:

not reported

Salinity:

not applicable

Conductivity:

540 µS/cm

Details on test conditions:

EXPOSURE PHASE
Dechlorinated water was supplied to treated and control tanks at approximately 250 mL/min, using calibrated flow meters. Tanks were all glass aquaria, with a capacity of approximately 80 litres to the overflow. A reservoir (20 litres) of 14C-test substance cation was prepared every 2 days for dosing directly into the treated tank using a peristaltic pump. The flow rate was approximately 6 mL/min, giving a concentration of nominally 1 mg test substance cation/L and 0.001% methanol in the treated tank. The control tank was dosed with dechlorinated water containing 0.03% methanol, at approximately 6 mL/min to give a nominal methanol concentration of 0.001% in the tank .

DEPURATION PHASE
After exposure, the remaining fish from each tank were transferred to separate tanks (80 L, all glass aquaria), with a flow of approximately 250 mL/min of dechlorinated water. The water in each tank was aerated with a single air stone and heated with a glass aquarium heater.

MEASUREMENTS OF WATER QUALITY
- Dissolved oxygen and pH: Dissolved oxygen and pH were measured in each test tank at regular intervals (every 2-4 days) throughout the study.
- Temperature: The temperature of the water bath was monitored “continuously” using a maximum/minimum thermometer and recorded every 2-4 days.

Nominal and measured concentrations:

- Nominal concentration: 1 mg test substance cation/L
- Mean measured concentration: 1.03 mg test substance cation/L. See 'Any other information on materials and methods incl. tables'.

Reference substance (positive control):

no

Remarks on result:

not measured/tested

Key result

Conc. / dose:

1.03 mg/L

Temp.:

20 °C

pH:

8

Type:

BCF

Value:

1.3 L/kg

Basis:

whole body w.w.

Time of plateau:

7 d

Calculation basis:

steady state

Remarks on result:

other: Exposure phase

Conc. / dose:

1.03 mg/L

Temp.:

20 °C

pH:

8

Type:

BCF

Value:

0.7 L/kg

Basis:

edible fraction

Remarks:

Muscle

Time of plateau:

10 d

Calculation basis:

steady state

Remarks on result:

other: Exposure phase

Conc. / dose:

1.03 mg/L

Temp.:

20 °C

pH:

8

Type:

BCF

Value:

2.5 L/kg

Basis:

non-edible fraction

Time of plateau:

10 d

Calculation basis:

steady state

Remarks on result:

other: Exposure phase

Remarks on result:

not measured/tested

Remarks on result:

not measured/tested

Metabolites:

On neither HPLC nor TLC did any of the radiochemical residue chromatograph in the region of the photodegradation products M4 or M1. HPLC / quantification of the radioactivity in the eluates showed < 2% of the radioactivity in the water associated with fractions corresponding to the retention time of the photodegradation products.

Details on results:

An overview of the results was provided in Table 2 – Table 4 in ‘Any other information on results incl. tables’
- Concentration and Characterisation of Radioactivity in Water: During exposure the mean daily measured concentration of the test substance, based on total radioactivity, in the treated tank was 1.03 mg cation/L This value was used to calculate the bioconcentration factors. The approximate limit of detection for 14C-labelled test substance in water was 0.002 mg/L. The test substance was the only compound detected and accounted for > 90% of the radioactivity injected onto the HPLC column and > 76% of that applied to TLC plates at all sample dates.

14C-LABELLED RESIDUES IN FISH; AND BIOCONCENTRATION FACTORS
- Exposure Phase: In whole fish, muscle and viscera there was little change in the level of accumulated 14-labelled residues throughout exposure and it was apparent that a 'steady state' had been reached. Furthermore, the BCF was low, being < 3 in all tissues. Therefore, exposure was terminated on day 14.
a) Whole fish: The maximum level of residues measured in whole fish during exposure was 1.30 mg test substance equivalents/kg wet weight on day 7. equivalent to a BCF of 1.3.
b) Muscle: The maximum level of residues measured in muscle was 0.69 mg test substance/kg wet weight on day 10, equivalent to a BCF of 0.7.
c) Viscera: The maximum level of residues measured in viscera was 2.59 mg test substance/kg wet weight on day 10, equivalent to a BCF of 2.5.

- Depuration Phase: The level of accumulated residues in viscera fell by 50% within one day and in the whole fish and muscle within three days. After 28 days depuration almost 80% of the accumulated residues in whole fish had been eliminated.

Table 2. Characterisation of Radioactivity in Water from Treated Tank

Study and Exposure Phase Day	Recovery of Radioactivity in Water after Rotary Evaporation prior to HPLC and TLC (%)	HPLC Analysis	TLC Analysis in solvent system 1
		Radioactivity injected onto HPLC column co-chromatographing with the test substance^a(%)
			Radioactivity applied to TLC co-chromatographing with test substance standard
0	97	93	81
8	102	91	80
14	95	92	76

a. Mean of two injections

Table 3. Concentration of 14C-Residues in Fish During Exposure Phase; and Bioconcentration Factors

Exposure Phase Day	Mean^aconcentration in fish tissue (mg cation equivalents/kg wet weight)^b			Bioconcentration factor (BCF)
	muscle^c	viscera^c	whole fish^c	muscle	viscera	whole fish
1	0.32 (0.10)	1.05 (0.90)	0.60 (0.27)	0.3	1.0	0.6
3	0.46 (0.16)	0.59 (0.21)	0.69 (0.29)	0.4	0.6	0.7
7	0.48 (0.13)	1.70 (1.11)	1.30 (0.54)	0.5	1.7	1.3
10	o.69 (0.27)	2.59 (2.37)	0.94 (0.34)	0.7	2.5	0.9
14	0.36 (0. 19)	2.41 (1.44)	0.63 (0.12)	0.3	2.3	0.6

a mean of :

whole fish - 3 samples

muscle - 4 samples

viscera - 4 samples

b values in parentheses are standard deviations

c approximate limits of detection are 0.02, 0.1 and 0.02 mg cation equivalents/ kg wet weight for muscle, viscera and whole fish, respectively.

Table 4. Concentration of 14C-Residues in Fish During Depuration phase

Study Day	Depuration Phase Day	Meana concentration in fish tissue (mg cation equivalents/kg wet weight)b			Concentration in tissue as percentage of exposure day 14 values
		muscle^c	viscera^c	whole fish^c	muscle	viscera	whole fish
15	1	0.39 (0.22)	0.72 (0.53)	0.34(0.01)	108	30	54
17	3	0.18 (0.07)	0.26 (0.19)^d	0.32(0.17)	50	11	51
21	7	0.10 (0.02)	0.13 (0.05)	0.16(0.06)	28	5	25
28	14	0.10 (0.02)	0.24 (0.15)	0.19(0.03)	28	10	30
42	28	0.05 (0.01)	< 0.1^e	0.14(0.05)	14	1	22

a mean of :

whole fish - 3 samples 

muscle - 4 samples

viscera - 4 samples

b values in parentheses are standard deviations

c approximate limits of detection are 0.02, 0.1 and 0.02 mg cation equivalents/kg wet weight for muscle, viscera and whole fish respectively.

d based on 2 samples; 2 samples “lost” on combustion

e 3 of the 4 samples were at background radioactivity values

Validity criteria fulfilled:

yes

Conclusions:

Based on the findings, the maximum measured concentrations were 1.30, 0.69 and 2.59 mg test substance cation equivalents per kg wet weight for whole fish, muscle ("edible” tissue) and viscera, respectively; equivalent to bioconcentration factors of 1.3 / 0.7 and 2.5 L/kg.

Executive summary:

The bioaccumulation potential of the test substance was determined in a study performed similar to OECD TG 305 and in compliance with GLP criteria. Bluegill sunfish (Lepomis macrochirus) were exposed to 14C-ring-labelled test material in a flow-through system. A mean measured concentration of 1.03 mg cation equivalents per litre was maintained in the water during the 14 days exposure period. After exposure the fish were transferred to test material-free, flowing water for a 28 days depuration period. Water and fish samples were analysed for 14C-residues at regular intervals. Analysis of the water throughout the exposure period confirmed that 14C-labelled test substance cation was the only compound present. In the fish, the maximum measured concentrations were 1.30, 0.69 and 2.59 mg test substance cation equivalents per kg wet weight for whole fish, muscle ("edible” tissue) and viscera, respectively; equivalent to bioconcentration factors of 1.3 / 0.7 and 2.5 L/kg. There was no fish mortality or unusual behaviour in control or exposed fish. During the depuration period the 14C-residues fell by 50% in the viscera within one day, and in the whole fish and muscle within three days.

Description of key information

BCF = 1.3 L/kg in whole fish (bluegill sunfish), equivalent to OECD TG 305, Hamer 1987

Key value for chemical safety assessment

BCF (aquatic species):

1.3 L/kg ww

Additional information

The bioaccumulation potential of the test substance was determined in a study performed similar to OECD TG 305 and in compliance with GLP criteria. Bluegill sunfish (Lepomis macrochirus) were exposed to 14C-ring-labelled test material in a flow-through system. A mean measured concentration of 1.03 mg cation equivalents per litre was maintained in the water during the 14 days exposure period. After exposure the fish were transferred to test material-free, flowing water for a 28 days depuration period. Water and fish samples were analysed for 14C-residues at regular intervals. Analysis of the water throughout the exposure period confirmed that 14C-labelled test substance cation was the only compound present. In the fish, the maximum measured concentrations were 1.30, 0.69 and 2.59 mg test substance cation equivalents per kg wet weight for whole fish, muscle ("edible” tissue) and viscera, respectively; equivalent to bioconcentration factors of 1.3 / 0.7 and 2.5 L/kg. There was no fish mortality or unusual behaviour in control or exposed fish. During the depuration period the 14C-residues fell by 50% in the viscera within one day, and in the whole fish and muscle within three days.

The bioaccumulation was also tested in additional species including invertebrates and sediment dwelling organisms. The highest observed BCF was found to be 32 L/kg und thus indicating the low bioaccumulation potential of the substance.