1-Octadecanaminium,N,N-Dimethyl-N-Octadecyl-,(Sp-4-2)-[29h,31h-Phthalocyanine2-Sulfonato(3-)-N 29,N 30,N 31,N 32]Cuprate(1-)

Bioaccumulation: aquatic / sediment

Administrative data

Endpoint:

bioaccumulation in aquatic species: fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1981

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to standard procedure ; well documented study report

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1981

Report date:

1981

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 305 (Bioconcentration: Flow-through Fish Test)

Deviations:

not applicable

Principles of method if other than guideline:

The test was composed of two parts: (1) Target lipid model (TLm) test in Japanese Rice Fish. This test was conducted as an independent study, and TLm (48 h) value found of . 1350 mg/L. (2) accumulation test. Accumulation test adopted flow through test apparatus similar to the prescribed in the protocol laid down by the Japanese Minister of International Trade and Industry of Health and Welfare. The test substance is an organic pigment containing complexed copper. This metal may be analyzed in fish by digestion of the whole dried fish in nitric acid and measurement of the copper content of the liquor so obtained by atomic absorption spectrophotometer. After adjustment for the background level of cupper in carp the level of any accumulated test substance may be calculated.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Details on test material:

- Physical state: Blue powder

Specific details on test material used for the study:

It should be noted that inconsistencies of high log Kow and low BCF are not uncommon and may be explained by limited solubility of polar moieties in n-octanol (and correspondingly, lipids) and molecular size constraints that limit cross membrane transport. Anliker and Moser (1987) and Anliker et. al. 1988.
Thus, the relatively large size of the substance (MW > 1200) combined with its amphiphobic nature may explain apparent BCF and Kow inconsistencies.

Radiolabelling:

no

Sampling and analysis

Details on sampling:

- Sampling intervals/frequency for test organisms: taken 2 fish from control and two treated groups and analyzed at weeks 2, 4, 6 of the test. At end of the test on week 8, all the reminding fish was sacrificed.
- Sampling intervals/frequency for test medium samples: the copper content of water from the control and two treated groups was determined on day 1, 2, 3, 6, 7, 8, 9, 10, 13, 14, 17, 21, 22, 24, 27, 30, 34, 38, 41, 44, 49, 52, 56.
- Sample storage conditions before analysis: the fish was weighed, and then individually deep frozen in “polythene” bas until the end of the test period.
- Details on sampling and analysis of test organisms and test media samples (e.g. sample preparation, analytical methods): at end of the test, the deep frozen fish were thawed, and all fish were analyzed following this procedure: fish samples with knowing weights were dried at 105 oC for 48 h, and re-weighted. Each fish was placed in a quartz/silica flask and 40 ml of “Aristar” nitric acid added. The top of the flask was covered with a watch glass and the mixture placed on a 140 oC hot plate overnight. The watch glass was then removed and the nitric acid allowed to evaporate, taking care not to char the residue. 40 ml of 2N nitric acid was now added, the watch glass replaced and the flask again stood on the hot plate overnight. After cooling the contents were then filtered through a Whatman GF-B glass fiber filter and made up to 50 ml with distilled water. These solutions were analyzed for copper using Perkin Elmer Model 2380 atomic absorption spectrophotometer.

Test solutions

Vehicle:

yes

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

(1) A dispersion of the test substance in water was sapcially prepared by Organic Division for use in the test. The formulation was as follows:
Test substance: 9.7%
Tween 40: 4.85%
Water: 85.45%.
This dispersion was allocated the Brixham Code “H250”.
(2) The stock dispersion were prepared by diluting 25.75 g and 257.5 g fo the dispersion “H250” in 5 L of distilled water.

Test organisms

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM
- Common name: Carp
- Strain: Cyprinus Carpio
- Source: The fish were hatched during early 1980 at a commercial fish hatchery and kept in stock tanks in the Brixham Laboratory form July 1980 to the start of the accumulation test in August, 1981.
- Age at study initiation (mean and range, SD): about 1.5 yr old
- Weight at study initiation (mean and range, SD): 23.6 g
- Weight at termination (mean and range, SD): 27.3 g
- Method of breeding: Hatched in a commercial fish hatchery.
- Health status: The fish in good health were selected for the test.
- Description of housing/holding area: Rectangular glass tanks of capacity approcimately 88 L were used.
ACCLIMATION
- Acclimation period: 8 days
- Acclimation conditions (same as test or not): same (25 C)
- Health during acclimation (any mortality observed): Healthy.

Study design

Route of exposure:

aqueous

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

8 wk

Test conditions

Hardness:

No data

Test temperature:

Test Tank No. 8 (1.0 mg/L nominal test concentration): Mean 24.8°C, range 24.5 to 25.1 °C.
Test Tank No. 9 (10.0 mg/L nominal test concentration): Mean 24.8°C, range 24.4 to 25.2 °C.

pH:

No data

Dissolved oxygen:

No data

TOC:

No data

Salinity:

No data

Details on test conditions:

TEST SYSTEM
- Test vessel: rectangular glass tanks of capacity of approximately 88 liter
- Type of flow-through: proportional diluter
- No. of organisms per vessel: 12
- 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: quartz distilled water
RANGE-FINDING / PRELIMINARY STUDY
- Test concentrations: 1350 mg/L
- Results used to determine the conditions for the definitive study: TLm (48 h) > 1350 mg/L

Nominal and measured concentrations:

Nominal concentrations: 1.0 and 10.0 mg/L with a conrol

Reference substance (positive control):

no

Details on estimation of bioconcentration:

Fish were exposed to nominal concentrations of 1 and 10 mg/L, well above the tested solubility of 0.02 mg/L.

Results and discussion

Lipid content

Remarks on result:

other: No data

Bioaccumulation factor

Bioaccumulation factor 1

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

4.4

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: used the mean L value of 2 fish at day 14 of the test

Remarks:

Conc.in environment / dose:1.09 mg/L

Bioaccumulation factor 2

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

2.1

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: used the mean L value of 2 fish at day 28 of the test

Remarks:

Conc.in environment / dose:1.13 mg/L

Bioaccumulation factor 3

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

8.4

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: At day 42 of the test (N=1)

Remarks:

Conc.in environment / dose:1.14 mg/L

Bioaccumulation factor 4

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

4.9

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Used mean L value of 6 fish at day 56 of test

Remarks:

Conc.in environment / dose:1.13 mg/L

Bioaccumulation factor 5

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

3.4

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Used mean L value of 2 fish at day 14 of the test

Remarks:

Conc.in environment / dose:10.3 mg/L

Bioaccumulation factor 6

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

1.3

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Used mean l value of 2 fish at day 28 of the test

Remarks:

Conc.in environment / dose:10.5 mg/L

Bioaccumulation factor 7

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

6.4

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Mean L value of 2 fish at day 42 of the test

Remarks:

Conc.in environment / dose:10.6 mg/L

Bioaccumulation factor 8

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

3.5

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Used mean L value of 3 fish at day 50 of the test

Remarks:

Conc.in environment / dose:10.5 mg/L

Bioaccumulation factor 9

Type:

other: Accum Factor L defined as: L = J/Kn where, J= ug/g test material and Kn is the mean conc. of test substance (mg/L) over the preceeding days of the test

Value:

5.5

Basis:

whole body d.w.

Calculation basis:

other: Used copper as an indicator to measure the test substance bioaccumulation poetential

Remarks on result:

other: Used mean L value of 3 fish at day 56 of the test.

Remarks:

The maximum value (9.9 whole body d.w) was experimentally obtained based on nominal concentration used in testing. In addition, a theoretical BCF (4,590 whole body d.w.) was calculated from the known solubility of 0.02 mg/L to provide a precautionary estimate of distribution to fish. Conc.in environment / dose:10.5 mg/L

Depuration

Parameter:

other: No data

Details on kinetic parameters:

No data

Metabolites:

No data

Results with reference substance (positive control):

Not applicable

Details on results:

No data

Reported statistics:

No data

Any other information on results incl. tables

Table 1. Fish Sample Analysis in Test Substance Treated Group

Day of Test	Test vessle	Body Weight (w.w)	Copper concn. in fish (ug/g)	Copper concn. Control (ug/g)^1	F1	F2	Test Substance Conc. in fish (ug/g)	Mean conc. of water (mg/L)	Accumulation Facotr (L)
14	8	24.46	0.8	0.1	100/4.2	100/98.8	2.6	11.09	2.2
14	8	25.25	1	0.3	100/4.2	100/98.8	7.2	1.09	6.6
28	8	34.28	0.8	0.1	100/4.2	100/98.8	2.4	1.13	2.1
28	8	25.62	0.8	0.1	100/4.2	100/98.8	2.4	1.13	2.1
42	8	22.15	0.4	-0.3	100/4.2	100/98.8	N.D	1.14	N.D.*
42	8	21.12	1.1	0.4	100/4.2	100/98.8	9.6	1.14	8.4
56	8	32.46	0.8	0.4	100/4.2	100/98.8	2.4	1.13	2.1
56	8	30.45	1.1	0.4	100/4.2	100/98.8	9.6	1.13	8.5
56	8	29.93	0.8	0.1	100/4.2	100/98.8	2.4	1.13	2.1
56	8	39.49	0.9	0.2	100/4.2	100/98.8	4.8	1.13	4.2
56	8	36.87	0.9	0.2	100/4.2	100/98.8	4.8	1.13	4.2
56	8	26.32	1.1	0.4	100/4.2	100/98.8	9.6	1.13	8.5
14	9	20.92	2.2	1.5	100/4.2	100/98.8	36.2	1.03	3.5
14	9	18.66	2.1	1.4	100/4.2	100/98.8	33.7	10.3	3.3
28	9	21.83	1.2	0.5	100/4.2	100/98.8	12.1	10.5	1.2
28	9	22.78	1.3	0.6	100/4.2	100/98.8	14.5	10.5	1.4
42	9	26.23	3.1	2.4	100/4.2	100/98.8	57.8	10.6	5.5
42	9	26.43	3.9	3.2	100/4.2	100/98.8	77.1	10.6	7.3
50^2	9	32.44	2.6	1.9	100/4.2	100/98.8	45.8	10.5	4.4
50^2	9	25.5	2.9	2.2	100/4.2	100/98.8	53	10.5	5
50^2	9	28.24	1.2	0.5	100/4.2	100/98.8	12.1	10.5	1.2
56	9	25.44	2.6	1.9	100/4.2	100/98.8	45.8	10.5	4.4
56	9	22.5	5	4.3	100/4.2	100/98.8	103.6	10.5	9.9
56	9	27.5	1.6	0.9	100/4.2	100/98.8	21.7	10.5	2.1

Table 2. Table 1. Fish Sample Analysis in Control Group

Day of Test	Test vessel	Body Weight (w.w)	Copper content (ug/g)
14	10	21.18	0.6
14	10	29.08	0.5
28	10	36.62	0.4
28	10	31.83	0.7
42	10	23.98	0.7
42	10	31.19	0.4
56	10	27.42	0.9
56	10	34.18	0.9
56	10	27.79	0.6
56	10	26.24	0.7
56	10	31.57	0.8
56	10	30.33	0.7

Note1: the copper concentration of the control fish in Vessel 10 did not show trend with time, the overall mean value of 0.7 ug/g was used for L calculation.

Note 2: 3 Fish died on this day.

*: No substance detected in fish.

Applicant's summary and conclusion

Validity criteria fulfilled:

not specified

Conclusions:

The accumulation factor of the test material was evaluated during a lower 8 week bioaccumulation test at concentrations of 1.0 and 10.0 mg/L in Carp. Accumulation factor ranges from 2.1 to 8.5, and 2.1 to 9.9 were obtained for low concentration or high concentration, respectively.

Executive summary:

In an 8 week bioaccumulation test, Carp were exposed to the test material at nominal concentrations of 1.0 and 10.0 mg/L, with a control, under flow-trough conditions according to procedures similar to that described in OECD 305. The test substance is an organic pigment containing complexed copper. This metal was analyzed in fish by digestion of the whole dried fish in nitric acid and measurement of the copper content of the liquor so obtained by atomic absorption spectrophotometer. After adjustment for the background level of copper in carp the level of any accumulated test substance may be calculated. And the following accumulation factor was obtained:

  

Test Vessel	2 Weeks	4 Weeks	6 Weeks	8 Weeks
8 8 8 8 8 8 Mean   9 9 9 9 9 9 Mean	2.2** 6.6** ─ ─ ─ ─ 4.4  3.5 3.3 ─ ─ ─ ─ 3.4	2.1** 2.1** ─ ─ ─ ─ 2.1  1.2 1.4 ─ ─ ─ ─ 1.3	N.D.* 8.4** ─ ─ ─ ─  8.4 (n =1) 5.5 7.3 ─ ─ ─ ─ 6.4	2.1** 8.5** 2.1** 4.2** 4.2** 8.5**  4.9 4.4 9.9 2.1 (4.4) (5.0) (1.2) 5.5 (N = 3, day 56)

*: N.D.: Not test substance detected in fish;

**: The values are only a guide since it is below detection limit.

( ): Results for the 3 fish which died on Day 50 of the test.