Copper, [29H,31H-phthalocyaninato(2-)-.kappa.N29,.kappa.N30,.kappa.N31,.kappa.N32]-, sulfonated, sodium salts

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 29 October 2020 to 17 November 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

adopted18 June 2019

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Refrigerated (2 – 8°C), protected from light
- Stability during storage: The stability of the test substance under storage conditions over the test period was guaranteed by the sponsor.
- Stability in light: photo sensitive

Analytical monitoring:

yes

Details on sampling:

At the start (0, 24, 48 and 72-hours) and end (24, 48, 72 and 96-hours) of each media renewal, 10 mL samples were taken from the control and test media tanks. The samples were taken using an air displacement pipette fitted with a plastic pipette tip. Samples were standardly sampled into 20 mL amber glass scintillation vials. In each case triplicate samples were taken, one for immediate chemical analysis and two as ‘back-up’ samples should further analysis be required.
The samples were transported and stored ambient, the maximum storage length was 6 days (storage stability was guaranteed for 7 d)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The fresh test solutions were prepared by combining test substance (1589.96 - 1590.07 mg) with 25 L of treated mains water. Following preparation, the test solutions were checked for the Tyndall effect using a Hach 2100 N turbidity meter.

Evidence of undissolved material:
On the initial day of preparation, it was found that the test substance solution gave a substantially higher turbidity measurement than the control (6.94/7.31 NTU (Nephelometric turbidity units) compared with the control 0.147/0.282 NTU). Before starting the test, the following work was completed to confirm if the high NTU values for the test substance were an artefact of the test substance colour or if there was undissolved material present.
As the results of the investigation work gave no evidence to suggest that the high turbidity was related to undissolved material following discussion with the study monitor, the test was started with no alterations to the test substance preparation procedures and a higher than normal turbidity was accepted for this study.

- Test concentration separation factor:
- Evidence of undissolved material (e.g. precipitate, surface film, etc.):
- Other relevant information:

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Source: Northern Trout, United Kingdom
All fish used were of the same age and originated from the same source and population
- Length mean (10 randomly Selected Fish from All Surviving Fish): 4.6 cm
- Weight mean (10 randomly Selected Fish from All Surviving Fish): 0.7926 g


ACCLIMATION
- Acclimation period: 18 days
- Acclimation conditions (same as test or not): yes
- Type and amount of food during acclimation: a proprietary fish food, which was added to the holding tank in quantities dictated by the size of the fish
- Health during acclimation: The mortality rate of the stock batch of fish was 0% in the 7 days prior to the test.


FEEDING DURING TEST (as applicable)
fish were not fed for at least 24 hours before the start of the test and were not fed throughout the test

MAINTENANCE
The fish were held in a temperature controlled room under artificial light, with a 16-hour light: 8-hour dark photo-period, in holding tanks at a density appropriate to their size, under continuous water renewal (flow-through) conditions:

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

see table 1

Test temperature:

see table 1

pH:

see table 1

Dissolved oxygen:

see table 1

Salinity:

see table 1

Conductivity:

see table 1

Nominal and measured concentrations:

Nominal: control and 16.8 mg (active ingredients)/L
See table 2 for measured concentrations
Measured:
non filtered: 99.4 % recovery
filtered: 92.8 % recovery

Details on test conditions:

TEST SYSTEM
- Test vessel: Single 30 L constructed glass aquaria, each fitted with clear Perspex lid, containing 25 L of media, were used for the control and test concentration.
- Type: closed
- Renewal rate of test solution: daily
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates):1
- Biomass loading rate: 0.22 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The water used for the holding of fish stocks and for the toxicity tests was treated mains water. The water was pumped to the laboratory through a particulate filter, an activated carbon filter and a UV steriliser
Water parameters: see table 1

- Culture medium different from test medium: no
- Intervals of water quality measurement: before every media renewal

OTHER TEST CONDITIONS
- Photoperiod: 16-hour light: 8-hour dark photo-period
Due to the photo sensitive nature of the test substance, the test substance was prepared under red light and the preparation vessels were protected from light until the fish were added.


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : see table 3
All fish in each vessel were observed at ca 3 and 5 hours after addition and then at 24-hour intervals (24, 48, 72 and 96 hours) throughout the duration of the test. Additional observations were also conducted at 24-hour + ca 4-hours intervals (i.e. ca 28, 52 and 76-hours). The number of dead fish and those fish exhibiting toxic symptoms or modified behaviour were recorded, if applicable.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: N.A. Limit test
- Justification for using less concentrations than requested by guideline: The limit test concentration was derived from the EC50 values for L. gibba and D. magna toxicity tests (Smithers ERS Study Numbers: 3202796 and 3202797, respectively) according to the threshold concept (OECD Series on Testing and Assessment No. 126 (2010)). As the L. gibba study was the most sensitive test, the limit test was conducted using the frond number yield EC50 value (16.8 mg active ingredient (AI)/L).

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 17.4 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 16.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other:

Details on results:

- Observations on body length and weight: see table 4
- Mortality of control: 1 fish had to be euthanized for welfare purposes, this fish death was not considered to be test substance related (see section overall remarks for details)
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: On the initial day of preparation, it was found that the test substance solution gave a substantially higher turbidity measurement than the control. As the results of the investigation work gave no evidence to suggest that the high turbidity was related to undissolved material following discussion with the study monitor, the test was started with no alterations to the test substance preparation procedures and a higher than normal turbidity was accepted for this study.

Reported statistics and error estimates:

No statistical analysis was performed on this test, as there was 14% mortality during the test which was considered to indicate no test substance effects, as one death would be acceptable if this vessel had been a control and there was no sub lethal effects noted in any of the surviving fish. The EC0, EC50 and EC100 concentrations were determined empirically by observing the data and were reported to be at or above the limit test concentration, as applicable.

Sublethal observations / clinical signs:

Table 4: Lengths and weights of a sub-sample of ten fish taken at random from all surviving fish after 96 hours

Fish Number	Total Length (cm)	Wet Weight (g)
1	4.9	0.8477
2	4.2	0.5943
3	5.4	1.2826
4	4.7	0.9389
5	4.1	0.5778
6	4.5	0.6874
7	4.3	0.6399
8	4.3	0.6820
9	4.7	0.7261
10	4.9	0.9494
Mean	4.6	0.7926
Minimum	4.1	0.5778
Maximum	5.4	1.2826

Validity criteria fulfilled:

yes

Conclusions:

No adverse effects of the test material to rainbow trout was observed during the 96 h acute limit test. Therefore, the LC50 was determined to be greater than 16.8 mg/L (nominal concentration, refers to the active ingredients (AI).

Executive summary:

The 96-hour acute toxicity of the test material to the freshwater fish species, Oncorhynchus mykiss, was determined under semi-static test conditions over a 96-hour period as limit test in accordance with the requirements of OECD Chemicals Testing Guideline No. 203 Fish, Acute Toxicity Test (adopted 18 June 2019).
Based on nominal active ingredient concentrations, the 96-hour LC50 value was considered to be ˃16.8 mg AI/L. The corresponding LC0 value was considered to be ≥16.8 mg AI/L.
Based on arithmetic mean measured active ingredient concentrations, the 96-hour LC50 value was considered to be ˃17.4 mg/L. The corresponding LC0 value was considered to be ≥17.4 mg/L.
All validity criteria were satisfied, therefore, the test was considered to be valid.

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- The concentrations determined by spectrophotometry

Vehicle:

no

Details on test solutions:

A stock solution of 3g/L (based on the active component) is prepared by disolving 16.077g of test substance in 500 mL demineralised water.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

- Source of supply: Hasselts aquarium
- The stock was adapted to the test conditions in LISEC lab for at least 12 days
- Toatl length: 3.2 +/-0.3 cm

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

20.0 - 20.9°C

pH:

7.75 - 8.26

Dissolved oxygen:

>= 7.2 mg/L

Nominal and measured concentrations:

Nominal test concentrations based on the active component: 0, 6.25, 12.5, 25, 50 and 100 mg/L

Details on test conditions:

- Renewal of the test solutions: every 24 hours
- Test containers: 3L borosilicate glass beakers
- Volume of test solution: 3 L per vessel
- Number of organisms: 10 fish for the control and for each test concentration
- Feeding: the fish are not fed during the test
- Aeration: moderate bubble-aeration for each test vessel
- Because of the photodegradation of the test substance under light, the test was carried out entirely in the dark

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Validity criteria fulfilled:

yes

Description of key information

With high probability acutely not harmful to fish.

LC50 (96h) >= 100 mg/L (nominal) for Brachydanio rerio (OECD 203)

LC50 (96h) >= 16.8 mg/L (nominal) for Oncorhynchus mykiss (OECD 203)

 

 

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

> 100 mg/L

Additional information

The acute toxicity of the test item to fish was assessed in two different key studies according to GLP regulations and OECD guideline 203.

Since the test item is an UVCB the concentrtaions based on the active ingredients (AI). In both tests, test materials with diffferent concentrations and and differernt test species were used. In both tests no adverse effects were observed up to highest measured concentrations.

 

 The test organism Oncorhynchus mykiss (rainbow trout) was exposed to a limit concentration of 16.8 mg AI/L (nominal) over 96 h. A test material with 26.4 % active ingredients was used (BASF, 2021). The LC 50 (96 h) was set to be > 16.8 mg AI/L.

Similar results were  obtained in a study with a test material of 9.33 % AI and the test organism Danio rerio  (Procter & Gamble 1994).

The fish were exposed to the substance for 96 hours in a semi-static test system. At test termination a LC50 >= 100 mg/L (nominal) was determined.