Olivine, cobalt silicate blue

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2012-12-03 to 2012-12-14

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)

Version / remarks:

adopted 1992-07-17

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD Series on testing and assessment, Number 29. Guidance document on transformation/dissolution of metals and metal compounds in aqueous media. ENV/JM/MONO (2001)9, 23-July 2001.

Version / remarks:

According to OECD Series on Testing and Assessment # 29 (2001), “For hazard classification purposes the results of the dissolution/transformation protocol are compared with existing ecotoxicity data for metals and metal compounds. However, for purposes such as data validation, there might be cases where it may be appropriate to use the aqueous medium from a completed transformation test directly in an OECD 202 and 203 daphnia and fish ecotoxicity test. If the CaCl2.2H2O and MgSO4.7H2O concentrations of the transformation medium are reduced to one-fifth of the ISO 6341 medium, the completed transformation medium can also be used (upon the addition of micronutrients) in an OECD 201 algae ecotoxicity test.” Therefore, to assess the hazard potential of this pigment containing several different metals, an ecotoxicity validation test was performed applying the dissolved fraction collected after 7-d of transformation/dissolution of the pigment in the respective ecotoxicity medium.

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

signed 2011-02-07

Specific details on test material used for the study:

not applicable

Analytical monitoring:

yes

Details on sampling:

- Sampling method/Sample storage conditions before analysis:
Dissolved cobalt concentrations (the Co content of the test item is 41.4 %) were measured in all test solutions including the control at the start and end of the exposure period.

Duplicate samples of approximately 20 mL were taken. Solutions were examined for soapy bubbles or particles floating on the surface. The samples were filtered (0.22 µm polyether sulphone membrane syringe filter (DIAFIL PS, DIA-Nielsen GmbH & Co. KG, Düren, Germany) at room temperature (20-25°C), transferred into disposable polyethylene vials (Scintillation vials, Sarstedt, Nuembrecht, Germany ), acidified with HNO3 (final concentration 1- 3% HNO3) and were stored until further analysis. The concentrations of dissolved cobalt were determined by ICP-OES.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method:
As the test item is a multi-component substance, toxicity was determined using the dissolved fraction(s) after 7-d Transformation/dissolution (T/D fraction). The T/D fractions for the various test concentrations were prepared individually in accordance with the OECD guidance document on transformation/dissolution of metals and metal compounds in aqueous media No. 29 (2001)*.

Appropriate amounts of test item were transferred into glass flasks and filled up with dilution water to obtain T/D loadings of 10, 32, 100, 320, and 1000 mg/L, respectivley. One flask was filled with dilution water only for the control. Flasks were stirred at 100 rpm for 7 days at 21.5°C. For each solution the insoluble parts were then separated from the aqueous phase and sterilised by filtration through a 0.22 μm filter (Syringe Filters DIAFIL PS (Polyether sulfone, PES) DIA-Nielsen GmbH & Co. KG, Düren, Germany) under sterile conditions. The filtrated T/D fractions were directly used for the acute fish toxicity test. The test vessels were filled with 2.5 L of the filtered T/D fractions containing the test item. Dilution water was used as blank control.

- Controls: dilution water was used as negative control.

*Reference:
- OECD Series on testing and assessment, Number 29. Guidance document on transformation/dissolution of metals and metal compounds in aqueous media. ENV/JM/MONO (2001)9, 23-July 2001.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Species: Danio rerio (Teleostei, Cyprinidae)
- Source: test facility breed
- Length at study initiation (subsample of the fish used in the test was weighed (n = 7)): 1.9 to 2.5 cm; mean ± SD: 2.2 ± 0.3 cm
- Weight at study initiation: 0.08 to 0.12 g; mean ± SD: 0.104 g ± 0.02 g
- No feeding of the fish occurred during the test and 24 hours prior to test start.

ACCLIMATION
- Acclimation period: at least 12 days prior to the test
- Acclimation conditions: fish were held under equivalent water quality and illumination conditions to those proposed for use in the test.
- Type and amount of food: fish were fed throughout the holding period with live brine shrimp (Artemia spp.) nauplii and ground flake food TetraMinR (Tetra Werke, Melle, Germany).
- Feeding frequency: once daily; ad libitum
- Health during acclimation (any mortality observed): only healthy fish without diseases and abnormalities were used in the study. Mortality of the batch was less than 5 % in the week preceding the start of the study.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

total hardness (holding and dilution water): 1.1 mmol/L

Test temperature:

23.1 to 23.2°C

pH:

between 8.09 and 8.45

Dissolved oxygen:

oxygen saturation: 92 to 99%

Salinity:

not applicable.

Nominal and measured concentrations:

Nominal concentrations: 10, 32, 100, 320, and 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 3 L glass beakers filled with 2.5 L of test solution
- Aeration: test media in the vessels was slightly aerated via a glass capillary.
- No. of organisms per vessel: 7 fish
- The mean weight resulted in a loading of 0.3 g/L test medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: purified drinking water was used according to the OECD-guideline 203 (1992). The purification inlcudes filtration with activated charcoal, passage through a lime-stone column and aeration until oxygen saturation. To avoid copper contamination, plastic water pipes are used in the test facility.
Typical water chemistry data of the holding and dilution water are presented below.
- Conductivity: 212.5 µS/cm
- pH: 8.9
- dissolved oxygen content: 106%
- total residual chlorine: <0.02 mg/L
- nitrate: 1.1 mg/L
- nitrite: <0.005 mg/L
- ammonium: <0.01 mg/L
- phosphate: 1.57 mg/L
- total hardness: 1.1 mmol/L
- alkalinity: 2 mmol/L
- calcium: 0.8 mmol/L
- magnesium: 0.3 mmol/L
- NPOC: 0.537 mg/L
- copper: <2.02 µg/L*
- iron: < 2.20 µg/L*
- manganese: <2.99 µg/L*
- zinc: <0.471 µg/L*
<: value below the respective detection limit

OTHER TEST CONDITIONS
- The pH of solutions in test vessel was not affected by the test substance and remained constant over the course of study.
- Photoperiod: 12 light / 12 dark

EFFECT PARAMETERS MEASURED:
- mortality and clinical signs (sub-lethal and abnormal behaviour) were recorded at 3, 24, 48, 72, and 96 hours of exposure of the fish to the test item.

TEST CONCENTRATIONS
- Range finding study
A non-GLP range-finding test was performed with dissolved fractions of the loadings of 100, 10, and 1.0 mg/L.Mortality, clinical signs or any distress could not be observed throughout the test.

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

dissolved fraction at a loading of 1000 mg/L after 7 days (T/D fraction)

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

dissolved fraction at a loading of 1000 mg/L after 7 days (T/D fraction)

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: neither clinical signs nor mortality were observed throughout the study
- Mortality of control: none of the fishes in the control group died.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: measured Co concentrations after 96h ranged from 95% to 105% of initially measured levels, confirming continuous exposure.
Please also refer to the field "Any other information on results incl. tables" below.

Results with reference substance (positive control):

no data

Reported statistics and error estimates:

A statistical analysis to determine LC10 and LC50 values was not required. The LC50 is above the highest concentration tested and reported as such. The evaluation of the effects is based on loadings of the T/D fractions.

Sublethal observations / clinical signs:

Clinical signs during the test period of 96 h (number of fish per treatment n=7)

T/D concentration Olivine, cobalt silicate blue		Test duration
[mg/L]		3 h	24 h	48 h	72 h	96 h
Control		nd.	nd.	nd.	nd.	nd.
10		nd.	nd.	nd.	nd.	nd.
32		nd.	nd.	nd.	nd.	nd.
100		nd.	nd.	nd.	nd.	nd.
320		nd.	nd.	nd.	nd.	nd.
1000	[EBRC1]	nd.	nd.	nd.	nd.	nd.

nd.: no symptoms detected

 Cumulative mortality during the test period of 96 h (number of fish per treatment n=7).

T/D loading Olivine, cobalt silicate blue		Test duration
[mg/L]		3 h	24 h	48 h	72 h	96 h
Control		0	0	0	0	0
10		0	0	0	0	0
32		0	0	0	0	0
100		0	0	0	0	0
320		0	0	0	0	0
1000	[EBRC2]	0	0	0	0	0

Validity criteria fulfilled:

yes

Conclusions:

In an acute toxicity test according to OECD 203, neither clinical signs nor mortality were observed at any of the tested loadings up to 1000 mg/L. Thus, the LC50 is >1000 mg/L whereas the NOEC based on visual observation is ≥ 1000 mg/L (all: dissolved fraction at a loading of 1000 mg/L after 7 days (T/D fraction)).

Description of key information

Results of a supporting study with Reaction mass of cobalt olivine and crystalline silicon dioxide (Olivine, cobalt silicate blue pigment) indicate a low potential for bioavailability and acute toxicity to fish. The respective 96h-LC50 for the effect of Reaction mass of cobalt olivine and crystalline silicon dioxide on freshwater fish (Danio rerio) derived in a GLP-conform OECD 203 test is estimated with > 1000 mg/L (dissolved fraction at a loading of 1000 mg/L after 7 days of dissolution) (Teigeler, 2013). Nevertheless, read-across to the assessment entity soluble cobalt substances is applied since cobalt cations determine fate and toxicity in the environment.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Remarks:

dissolved fraction at a loading of 1000 mg/L after 7 days of dissolution

Effect concentration:

> 1 000 mg/L