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Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics, other
Remarks:
in vitro bioaccessibility
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018-06-21 to 2018-07-04 and 2018-08-19 to 2018-08-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2019
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Objective of study:
bioaccessibility
Test guidelineopen allclose all
Qualifier:
equivalent or similar to
Guideline:
other: ASTM D5517-07: Standard Test method for determining the extractability of metals from art materials
Version / remarks:
ASTM, 2007 (American Society for Testing and Materials)
Qualifier:
according to
Guideline:
other: Standard Operating Procedure (SOP) for Bioelution Testing of Metals, Inorganic Metal Compounds, and Metal-Containing Complex Materials: Simulated Gastric Fluid
Version / remarks:
(Eurometaux, February 16, 2018)
GLP compliance:
yes
Remarks:
The study was performed and conducted acc. to GLP (ECTX), the delegated analytical phase was not performed in compliance with the OECD principles of GLP but acc. to the criteria of the ISO 17025 standard by the accredited test side (WLN Business B.V.).

Test material

Reference
Name:
Unnamed
Type:
Constituent
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Constituent
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Constituent
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Constituent
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Constituent
Test material form:
other: - solid (crystalline): Co(CH3COO)2; - solid, moist granular paste; Co(OH)O; - viscous paste: CoBNeo; - powder: CoCl2, CoS, CoLiO2, Co(NO3)2, Co(OH)2, Co3O4
Details on test material:
1) Cobalt diacetate tetrahydrate (Co(CH3COO)2)
- Physical form: solid (crystalline)
- Colour: reddish-violet
- Particle size distribution: d10: 26.81 μm, d50: 238.16 μm, d90: 445.35 μm
- Relative Density: 1.76

2) Cobalt hydroxide oxide (Co(OH)O):
- Physical form: solid, moist granular paste
- Colour: back - brown

3) Cobalt dichloride (CoCl2):
- Physical form: solid (in the powder form)
- Colour: rose red
- Relative Density: 1.924 g/cm³

4) Cobalt sulphide (CoS):
- Batch: HM4414
- Expiry date: August 2020
- 65.1 % cobalt
- Physical form: solid (in the powder form)
- Colour: grey
- Density: 5.2 g/cm³

5) Cobalt borate neodecanoate (CoBNeo):
- Physical form: viscous paste
- Colour: dark blue
- Relative Density: 1.2-1.4

6) Cobalt lithium dioxide (CoLiO2):
- Physical form: solid (in the powder form)
- Colour: black
- Particle size distribution: D10: 5.3 μm, D50: 10.2 μm, D90: 18.8 μm
- Specific surface area (N-BET): 0.33 m²/g
- Density: 5.1 g/cm³

7) Cobalt dinitrate (Co(NO3)2):
- Physical form: solid (in the powder form)
- Colour: red
- Relative Density: 1.87 g/cm³ (at 20 °C)

8) Cobalt dihydroxide (Co(OH)2):
- Physical form: solid (in the powder form)
- Colour: pink
- Particle size distribution (laserdiffraction: hexane): D10: 8.8 μm, D50: 34.1 μm, D90: 77.3 μm
- Particle size distribution (laserdiffraction: acetone): D10: 1.6 μm, D50: 31.4 μm, D90: 67.1 μm
- Specific surface area (N-BET): 19.607 m²/g
- Relative Density: 3.6

9) Tricobalt tetraoxide (Co3O4):
- Physical form: solid (in the powder form)
- Colour: grey/black
- Particle size distribution (Volume based): D10: 1.6 μm, D50: 5.4 μm, D90: 7.0 μm
- Specific surface area (N-BET): 3.3 m²/g
- Relative Density: 6.11 g/cm³ at 20 °C
Specific details on test material used for the study:
1) Cobalt diacetate tetrahydrate:
- Storage conditions: room temperature, in the dark

2) Cobalt hydroxide oxide:
- Storage conditions: room temperature, in the dark

3) Cobalt dichloride:
- Storage conditions: room temperature, in the dark

4) Cobalt sulphide:
- Storage conditions: room temperature, in the dark

5) Cobalt borate neodecanoate:
- Storage conditions: room temperature, in the dark.

6) Cobalt lithium dioxide:
- Storage conditions: room temperature, in the dark

7) Cobalt dinitrate:
- Storage conditions: room temperature, in the dark

8) Cobalt dihydroxide:
- Storage conditions: room temperature, in the dark

9) Tricobalt tetraoxide:
- Storage conditions: room temperature, in the dark

Test animals

Species:
other: in vitro (simulated gastric fluid)
Details on test animals and environmental conditions:
Test principle in brief for each of the nine test item
- the test item was tested in a simulated gastric fluid (0.07 N HCl, pH 1.5)
- loading of 0.2 and 2 g/L
- samples were taken after 2 hours: 1 hour agitation at 100 rpm and 1 hour of settling (without shaking) at 37 +/- 1 °C in the dark,
- one additional replicate per loading was prepared to avoid cross contamination and sampled prior to the test to control the pH
- one method blank and 3 replicates per loading were sampled, filtered and preserved by adding HNO3, stored at room temperature in the dark until shipment to the test side for the analytical measurement.
- The delegated analytical phase was carried out by the accredited Dutch laboratory WLN Business B.V. ( Rijkstraatweg 85, NL-9756 AD Glimmen, The Netherlands)
- measurement of dissolved cobalt was conducted by IPC-MS

The objective of this studies was to obtain knowledge about the bio-elution characteristics of nine cobalt substances in simulated gastric fluid.
Bio-elution refers to the in vitro extraction methods used to measure the degree to which a substance (e.g., metal or mineral ion) is dissolved in artificial biological fluids. Simulated biological fluids represent relevant exposure routes. The resulting value is the “bio-accessibility”, and is defined as the “fraction of a substance that is soluble under physiological conditions and therefore potentially available for absorption into systemic circulation”.

Administration / exposure

Duration and frequency of treatment / exposure:
Samples were taken after 2 h.
Doses / concentrationsopen allclose all
Dose / conc.:
0.2 other: g test item /L simulated gastric fluid
Dose / conc.:
2 other: g test item/L simulated gastric fluid
Details on study design:
The determination of dissolved cobalt concentrations in the blank control and test item vessels were carried out at the test site (WLN Business B.V.), using an ICP-MS (Thermo Fischer Scientific iCAP Q ICP-MS; SOP WLN-CM.W.11.1 Elemental determination by ICP-MS technique, SFS-EN ISO 17294-2). To describe the dissolution behaviours, even below the accredited reporting limits of WLN and to allow more accurate toxicity calculations, raw data were used to perform calculations. Rounded values were used to at least one more decimal place than the corresponding accredited reporting limit of WLN, if relevant. The accredited reporting limit with an accuracy of 90 % and 110 % for cobalt is 0.5 μg/L. For values between the detection limit and the accredited reporting limit, an accuracy of 90 % - 110 % cannot be guaranteed.
The analyses were not performed in compliance with the GLP principles, but according to the ISO 17025 standard.
Each sample, possibly after proper dilution, was measured once.
To prepare blanks, calibrations and control solutions/dilutions de-ionised water (d.i. H2O) at a purity level of 18 MΩ was used.
The ICP-MS was calibrated at the start of each run.
Recalibration was performed every 25 samples and at the end of the run and also used for instrument drift corrections.
The linearity (2x calibration level) and the factor of interference were determined and checked after the initial calibration.
The calibration standard solution used was a custom made multielement standard (iV inorganic ventures).
Matrix corrections were performed by the online addition of an internal standard.
Each run also contained at least one blank procedure.
All measurements were checked for "carry over" as also the RSD of the three-integrations of each measurement.
For each measurement the `drift´ of the internal standard was checked.
The calibrations were checked by means of an independent control standard (another supplier: either SCP Science or J.T. Baker), which was prepared using single element standards.
All measurement values of the control standards were evaluated against the control charts. These values were expected to be within 2 sigma levels of the control.
The last control sample value was transferred to the WLN laboratory information management system (LIMS)

Thermo Fischer Scientific iCAP Q ICP-MS
- Intsrument: Thermo Fischer Scientific iCAP Q ICP-MS (serial no.: 730R, WLN no.: 730000/ serial no. 731R, WLN no.: 731000)
- Forward power: 1550 W
- Cool gas flow rate: 14.00 L/min
- Auxiliary gas flow rate: 0.80 L/min
- Number of replicates analyses: 1 (average of 3 integrations)
- Isotopes monitored : 59Co

Performed characteristics - Measured parameters accredited for drinking water, groundwater, surface water
- Reporting limit: 0.5 µg/L for Co
- Repeatability: <= 5 %
- Reproducibility: <= 10 %
- Accuracy: 90-100 %
Details on dosing and sampling:
Further details are given in "Any other information on materials and methods incl. tables".

Results and discussion

Toxicokinetic / pharmacokinetic studies

Toxicokinetic parametersopen allclose all
Key result
Test no.:
#1
Toxicokinetic parameters:
other: Bioaccessibility / dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2 g test item/L: 46.1 ± 0.8 mg/L Co (230 ± 4 mg/g test item), Co release: 97 %; loading of 2 g test item /L: 472 ± 1 mg/L Co (236 ± 1 mg/g test item), Co release:100 %
Remarks:
Test item: Cobalt diacetate tetrahydrate
Key result
Test no.:
#2
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2 g test item/L: 1.81 mg/L (8.98 ± 0.29 mg/g test item), Co release: 3.1 %; loading of 2 g test item/L: 8.45 mg/L (4.22 ± 0.13 mg/g test item), Co release: 1.5 %
Remarks:
Test item: Cobalt hydroxide oxide
Key result
Test no.:
#3
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2 g test item/L: 49.1 ± 1.4 mg/L (245 ± 7 mg/g test item), Co release: 99 %; loading of 2 g test item/L: 486 ± 4 mg/L (243 ± 2 mg/g test item), Co release: 98 %
Remarks:
Test item: Cobalt dichloride
Key result
Test no.:
#4
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading 0.2g test item/L: 0.786 ± 0.008mg/L (3.93 ± 0.04mg/g test item), Co release: 0.60%; loading of 2g test item/L: 7.65 ± 0.09mg/L (3.82 ± 0.05mg/g test item), Co release: 0.59%
Remarks:
Test item: Cobalt sulphide
Key result
Test no.:
#5
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2g test item/L: 8.99 ± 0.13 mg/L (44.9 ± 0.7 mg/g test item), Co release: 7.5%; loading of 2g test item/L: 70.3 ± 1.0 mg/L (35.2 ± 0.5 mg/g test item), Co release: 5.9%
Remarks:
Test item: Cobalt lithium dioxide
Key result
Test no.:
#6
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid: loading of 0.2 g test item/L: 36.7 ± 1.0 mg/L (183 ± 5 mg/g test item), Co release: 82 %; loading of 2 g test item/L: 371 ± 6 mg/L (185 ± 3 mg/g test item), Co release: 83 %
Remarks:
Test item: Cobalt borate neodecanoate
Key result
Test no.:
#7
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2 g test item/L: 34.9 ± 0.3 mg/L (175 ± 1 mg/g test item), Co release: 88 %; loading of 2 g test item/L: 375 ± 4 mg/L (187 ± 2 mg/g test item), Co release: 94 %
Remarks:
Test item: Cobalt dinitrate
Key result
Test no.:
#8
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading of 0.2 g test item/L: 114 ± 2 mg/L (569 ± 13 mg/g test item), Co release: 92 %; loading of 2 g test item/L: 692 ± 3 mg/L (346 ± 1 mg/g test item), Co release: 56 %
Remarks:
Test item: Cobalt dihydroxide
Key result
Test no.:
#9
Toxicokinetic parameters:
other: Bioaccessibility/ Dissolution of Co in simulated gastric fluid, after 2h: loading 0.2 g test item/L: 0.233 ± 0.003mg/L (1.16 ± 0.02mg/g test item), Co release: 0.16%; loading 2 g test item/L: 2.16 ± 0.10mg/L (1.08 ± 0.05mg/g test item), Co release: 0.15%
Remarks:
Test item: Tricobalt tetraoxide

Bioaccessibility

Bioaccessibility testing results:
1) Cobalt diacetate tetrahydrate
- loading of 0.2 g/L: 46.1 mg/L Co; Cobalt release: 97 %; CV(between-vessel) = 2 %; N = 3
- loading of 2 g/L: 472 mg/L Co; Cobalt release: 100 %; CV(between-vessel) = <0.5 %; N = 3
The results can be assumed reliable since the test conditions of the stayed constant during the experiment. However, the pH of the test item vessels with a loading of 2 g/L rose to 1.89 but that was due to inherent properties of the test item itself.

2) Cobalt hydroxide oxide
- loading of 0.2 g/L: 1.81 mg/L Co; Cobalt release: 3.1 %; CV(between-vessel) = 3 %; N = 3
- loading of 2 g/L: 8.45 mg/L Co; Cobalt release: 1.5 %; CV(between-vessel) = 3 %; N = 3
The results can be assumed reliable since the test conditions stayed constant during the experiment.

3) Cobalt dichloride
- loading of 0.2 g/L: 49.1 mg/L Co; Cobalt release: 99 %; CV(between-vessel) = 3 %; N = 3
- loading of 2 g/L: 486 mg/L Co; Cobalt release: 98 %; CV(between-vessel) = 1 %; N = 3
The results can be assumed reliable since the test conditions stayed constant during the experiment.

4) Cobalt sulphide
- loading of 0.2 g/L: 0.786 mg/L Co; Cobalt release: 0.60 %; CV(between-vessel) = 1 %; N = 3
- loading of 2 g/L: 7.65 mg/L Co; Cobalt release: 0.59 %; CV(between-vessel) = 1 %; N = 3
The results can be assumed reliable since the test conditions stayed constant during the experiment

5) Cobalt lithium dioxide
- loading of 0.2 g/L: 8.99 mg/L Co; Cobalt release: 7.5 % (or 136 mg/m²); CV(between-vessel) = 1 %; N = 3
- loading of 2 g/L: 70.3 mg/L Co; Cobalt release: 5.9 % (or 107 mg/m²); CV(between-vessel) = 1 %; N = 3
The results can be assumed reliable since the test conditions stayed rather constant during the experiment.

6) Cobalt borate neodecanoate
- loading of 0.2 g/L: 36.7 mg/L Co; Cobalt release: 82 %; CV(between-vessel) = 3 %; N = 3
- loading of 2 g/L: 371 mg/L Co; Cobalt release: 83 %; CV(between-vessel) = 2 %; N = 3
The results can be assumed reliable since the test conditions of the stayed constant during the experiment. However, the pH of the test item vessels with a loading of 2 g/L rose to 1.80 but that was due to inherent properties of the test item itself.

7) Cobalt dinitrate
- loading of 0.2 g/L: 34.9 mg/L Co; Cobalt release: 88 %; CV(between-vessel) = 1 %; N = 3
- loading of 2 g/L: 375 mg/L Co; Cobalt release: 94 %; CV(between-vessel) = 1 %; N = 3
The results can be assumed reliable since the test conditions stayed constant during the experiment.

8) Cobalt dihydroxide
- loading of 0.2 g/L: 114 mg/L Co; Cobalt release: 92 % (or 29 mg/m²); CV(between-vessel) = 2 %; N = 3
- loading of 2 g/L: 692 mg/L Co; Cobalt release: 56 % (or 18 mg/m²); CV(between-vessel) = <0.5 %; N = 3
The results can be assumed reliable since the test conditions of the stayed constant during the experiment. However, the pH of the test medium with a loading of 2 g/L rose to 6.92 after addition of the test item but that was due to inherent properties of the test item itself.

9) Tricobalt tetraoxide
- loading of 0.2 g/L: 0.233 mg/L Co; Cobalt release: 0.16 % (or 0.35 mg/m²); CV(between-vessel) = 1 %; N = 3
- loading of 2 g/L: 2.16 mg/L Co; Cobalt release: 0.15 % (or 0.33 mg/m²); CV(between-vessel) = 5 %; N = 3
The results can be assumed reliable since the test conditions stayed rather constant during the experiment

Any other information on results incl. tables

1) Cobalt diacetate tetrahydrate

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank, 0.2 g/L loading: 1.52, 2 g/L loading: 1.84

- pH after 2 h: blank: 1.50, 0.2 g/L loading: 1.58 - 1.59, 2 g/L loading: 1.88 - 1.89

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 46.1 ± 0.8 mg/L Co (or 230 ± 4 mg/g test item), with a between-vessel variation of 2 %.

- Based on the cobalt content of the test item (i.e. 23.67 % Co) and the average dissolved cobalt concentration, a cobalt release of 97 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 472 ± 1 mg/L Co (or 236 ± 1 mg/g test item), with a between-vessel variation of 0 %.

- Based on the cobalt content of the test item (i.e. 23.67 % Co) and the average dissolved cobalt concentration, a cobalt release of 100 % was calculated.

2) Cobalt hydroxide oxide

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank, 0.2 and 2 g/L loading: 1.5

- pH after 2 h: blank: 1.50; 0.2 g/L loading: 1.49 - 1.50; 2 g/L loading: 1.48 - 1.49

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (black-brown) could be observed on the filter. The Erlenmeyer flasks contained a dark suspension showing still pieces of test item.

- Average dissolved cobalt concentration of 1.81 ± 0.06 mg/L Co (or 8.98 ± 0.29 mg/g test item), with a between-vessel variation of 3 %.

- Based on the cobalt content of the test item (i.e. 28.96 % Co) and the average dissolved cobalt concentration, a cobalt release of 3.1 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (black-brown) could be observed on the filter. The Erlenmeyer flasks contained a dark suspension showing still pieces of test item.

- Average dissolved cobalt concentration of 8.45 ± 0.26 mg/L Co (or 4.22 ± 0.13 mg/g test item), with a between-vessel variation of 3 %.EBRC

- Based on the cobalt content of the test item (i.e. 28.96 % Co) and the average dissolved cobalt concentration, a cobalt release of 1.5 % was calculated.

3) Cobalt dichloride

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 g/L loading: 1.50, 2 g/L loading: 1.48

- pH after 2 h: blank, 0.2 and 2 g/L loading: 1.47 - 1.51

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 49.1 ± 1.4 mg/L Co (or 245 ± 7 mg/g test item), with a between-vessel variation of 3 %

- Based on the cobalt content of the test item (i.e. 24.74 % Co) and the average dissolved cobalt concentration, a cobalt release of 99 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks

- Average dissolved cobalt concentration of 486 ± 4 mg/L Co (or 243 ± 2 mg/g test item), with a between-vessel variation of 1 %.

- Based on the cobalt content of the test item (i.e. 24.74 % Co) and the average dissolved cobalt concentration, a cobalt release of 98 % was calculated.

4) Cobalt sulphide

- Temperature after 2 h: 36.8 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 and 2 g/L loading: 1.49

- pH after 2 h: blank, 0.2 g/L loading: 1.49 - 1.50, 2 g/L loading: 1.49 - 1.50

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (grey particles) could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 0.786 ± 0.008 mg/L Co (or 3.93 ± 0.04 mg/g test item), with a between-vessel variation of 1 %.

- Based on the cobalt content of the test item (i.e. 65.1 % Co) and the average dissolved cobalt concentration, a cobalt release of 0.60 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (grey particles) could be observed on the filter and in the Erlenmeyer flasks. Also a silver-coloured film was visible on the liquid surface.

- Average dissolved cobalt concentration of 7.65 ± 0.09 mg/L Co (or 3.82 ± 0.05 mg/g test item), with a between-vessel variation of 1 %.

- Based on the cobalt content of the test item (i.e. 65.1 % Co) and the average dissolved cobalt concentration, a cobalt release of 0.59 % was calculated.

5) Cobalt lithium dioxide

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 and 2 g/L loading: 1.49

- pH after 2 h: blank: 1.50; 0.2 g/L loading: 1.52 - 1.53; 2 g/L loading: 1.62 - 1.63

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (dark particles) could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 8.99 ± 0.13 mg/L Co (or 44.9 ± 0.7 mg/g test item), with a between-vessel variation of 1 %.

- Based on the specific surface area of Cobalt lithium dioxide (0.33 m²/g test item) a cobalt release per surface of 136 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 60 % Co) and the average dissolved cobalt concentration, a cobalt release of 7.5 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (dark particles) could be observed on the filter and in the Erlenmeyer flasks. A silver-grey film was visible on the liquid surface.

- Average dissolved cobalt concentration of 70.3 ± 1.0 mg/L Co (or 35.2 ± 0.5 mg/g test item), with a between-vessel variation of 1 %.

- Based on the specific surface area of Cobalt lithium dioxide (0.33 m²/g test item) a cobalt release per surface of 107 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 60 % Co) and the average dissolved cobalt concentration, a cobalt release of 5.9 % was calculated.

6) Cobalt borate neodecanoate

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 g/L loading: 1.50, 2 g/L loading: 1.51

- pH after 2 h: blank, 0.2 g/L loading: 1.56 - 1.57, 2 g/L loading: 1.78 - 1.80

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item could be observed at the bottom of the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 36.7 ± 1.0 mg/L Co (or 183 ± 5 mg/g test item), with a between-vessel variation of 3 %.

- Based on the cobalt content of the test item (i.e. 22.2 % Co) and the average dissolved cobalt concentration, a cobalt release of 82 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item could be observed at the bottom of the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 371 ± 6 mg/L Co (or 185 ± 3 mg/g test item), with a between-vessel variation of 2 %.

- Based on the cobalt content of the test item (i.e. 22.2 % Co) and the average dissolved cobalt concentration, a cobalt release of 83 % was calculated.

7) Cobalt dinitrate

- Temperature after 2 h: 36.9 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 g/L loading: 1.52, 2 g/L loading: 1.50

- pH after 2 h: blank, 0.2 g/L loading: 1.56 -1.58, 2 g/L loading: 1.54 - 1.55

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 34.9 ± 0.3 mg/L Co (or 175 ± 1 mg/g test item), with a between-vessel variation of 1 %

- Based on the cobalt content of the test item (i.e. 19.91 % Co) and the average dissolved cobalt concentration, a cobalt release of 88 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles of test item could be observed on the filter and in the Erlenmeyer flasks

- Average dissolved cobalt concentration of 375 ± 4 mg/L Co (or 187 ± 2 mg/g test item), with a between-vessel variation of 1 %.

- Based on the cobalt content of the test item (i.e. 19.91 % Co) and the average dissolved cobalt concentration, a cobalt release of 94 % was calculated.

8) Cobalt dihydroxide

- Temperature after 2 h: 36.8 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 g/L loading: 1.58, 2 g/L loading: 6.92

- pH after 2 h: blank: 1.50; 0.2 g/L loading: 1.59 - 1.60; 2 g/L loading: 6.99 - 7.05

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- No undissolved particles on the filter and some flakes were visible in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 114 ± 2 mg/L Co (or 569 ± 13 mg/g test item), with a between-vessel variation of 2 %.

- Based on the specific surface area of Cobalt dihydroxide (19.607 m²/g test item) a cobalt release per surface of 29 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 62.1 % Co) and the average dissolved cobalt concentration, a cobalt release of 92 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Pink precipitate or remaining undissolved test item could be observed on the filter and in the Erlenmeyer flasks.

- Average dissolved cobalt concentration of 692 ± 3 mg/L Co (or 346 ± 1 mg/g test item), with a between-vessel variation of <0.5 %

- Based on the specific surface area of Cobalt dihydroxide (19.607 m²/g test item) a cobalt release per surface of 18 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 62.1 % Co) and the average dissolved cobalt concentration, a cobalt release of 56 % was calculated.

9) Tricobalt tetraoxide

- Temperature after 2 h: 36.8 - 37.0 °C

- pH at the start 0 h: blank: 1.50; 0.2 and 2 g/L loading: 1.52

- pH after 2 h: blank: 1.50; 0.2 g/L loading: 1.66 - 1.67; 2 g/L loading: 1.65 - 1.66

- Concentration of cobalt in blank control vessel: <LOQ (i.e.accredited reporting limit) of 0.5 μg/L Co

Loading of 0.2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (dark grey particles) could be observed on the filter and at bottom of the Erlenmeyer flasks. Floating particles were visible at the liquid surface.

- Average dissolved cobalt concentration of 0.233 ± 0.003 mg/L Co (or 1.16 ± 0.02 mg/g test item), with a between-vessel variation of 1 %.

- Based on the specific surface area of Tricobalt tetraoxide (3.3 m²/g test item) a cobalt release per surface of 0.35 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 72.4 % Co) and the average dissolved cobalt concentration, a cobalt release of 0.16 % was calculated.

Loading of 2 g/L, dissolution in simulated gastric fluid (pH 1.5) for 2h:

- Remaining test item (dark grey particles) could be observed on the filter and at bottom of the Erlenmeyer flasks. A floating dark film was visible at the liquid surface.

- Average dissolved cobalt concentration of 2.16 ± 0.10 mg/L Co (or 1.08 ± 0.05 mg/g test item), with a between-vessel variation of 5 %.

- Based on the specific surface area of Tricobalt tetraoxide (3.3 m²/g test item) a cobalt release per surface of 0.33 mg/m² was calculated.

- Based on the cobalt content of the test item (i.e. 72.4 % Co) and the average dissolved cobalt concentration, a cobalt release of 0.15 % was calculated.

Applicant's summary and conclusion

Conclusions:
On the basis of ASTM D5517-07 as well as according to recommended Standard Operating Procedure (SOP) for Bioelution Testing of Metals, Inorganic Metal Compounds, and Metal-Containing Complex Materials: Simulated Gastric Fluid (Eurometaux, February 16, 2018), the dissolution of nine Cobalt substances in simulated gastric fluid with a loading of 0.2 and 2 g test item/L, agitation (100 rpm) at 37 °C ± 1 °C and sampling after 2 h in the dark, were determined. The measurement of dissolved cobalt concentrations after filtration were performed by ICP-MS. The study was performed in triplicate with one additional method blank per loading for each substance.

1) Cobalt diacetate tetrahydrate
- loading of 0.2 g/L: 46.1 mg/L Co; Cobalt release: 97 %
- loading of 2 g/L: 472 mg/L Co; Cobalt release: 100 %

2) Cobalt hydroxide oxide
- loading of 0.2 g/L: 1.81 mg/L Co; Cobalt release: 3.1 %
- loading of 2 g/L: 8.45 mg/L Co; Cobalt release: 1.5 %

3) Cobalt dichloride
- loading of 0.2 g/L: 49.1 mg/L Co; Cobalt release: 99 %
- loading of 2 g/L: 486 mg/L Co; Cobalt release: 98 %

4) Cobalt sulphide
- loading of 0.2 g/L: 0.786 mg/L Co; Cobalt release: 0.60 %
- loading of 2 g/L: 7.65 mg/L Co; Cobalt release: 0.59 %

5) Cobalt lithium dioxide
- loading of 0.2 g/L: 8.99 mg/L Co; Cobalt release: 7.5 % (or 136 mg/m²)
- loading of 2 g/L: 70.3 mg/L Co; Cobalt release: 5.9 % (or 107 mg/m²)

6) Cobalt borate neodecanoate
- loading of 0.2 g/L: 36.7 mg/L Co; Cobalt release: 82 %
- loading of 2 g/L: 371 mg/L Co; Cobalt release: 83 %

7) Cobalt dinitrate
- loading of 0.2 g/L: 34.9 mg/L Co; Cobalt release: 88 %
- loading of 2 g/L: 375 mg/L Co; Cobalt release: 94 %

8) Cobalt dihydroxide
- loading of 0.2 g/L: 114 mg/L Co; Cobalt release: 92 % (or 29 mg/m²)
- loading of 2 g/L: 692 mg/L Co; Cobalt release: 56 % (or 18 mg/m²)

9) Tricobalt tetraoxide
- loading of 0.2 g/L: 0.233 mg/L Co; Cobalt release: 0.16 % (or 0.35 mg/m²)
- loading of 2 g/L: 2.16 mg/L Co; Cobalt release: 0.15 % (or 0.33 mg/m²)