Cobalt lithium dioxide

Acute Toxicity: other routes

Administrative data

Endpoint:

acute toxicity: other routes

Remarks:

oro-pharyngeal aspiration

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

other: not rated according to Klimisch et al.

Rationale for reliability incl. deficiencies:

other: Please refer to the field 'Remarks'

Remarks:

The investigated mechanistic parameters (i. e. bronchoalveolar lavage (BAL) fluid immune response marker analysis after oro-pharyngeal aspiration of the test material) have no direct value for fulfilling data requirements under the REACH regulation, the methodical setup is not adequately designed for risk assessment purposes of the test substance. The non-physiological route of administration via oro-pharyngeal aspiration is not guideline conform and not suitable to assess acute inhalation toxicity. Moreover, the publication shows reporting and methodological deficiencies. Test animal were obtained from two different sites. Since it is not stated how the mice are assigned to the test groups, it could be assumed that results are influenced by the origin of the mice, potentially leading to different response due to variation. Description of methodology lacks details. Information on housing of test animals are insufficient. Weights of the test animals are not specified for any point in time. Clinical signs and body weight development are not stated. No information on necropsy. Dosing is not justified. HCD is not specified. Results are not shown for the low dose group. Histological analysis was performed with already lavaged lungs. Histological analysis was performed with already lavaged lungs, which could have influenced tissues and particle deposition results.

Data source

Reference

Reference Type:

publication

Title:

Respiratory hazard of Li-ion battery components: elective toxicity of lithium cobalt oxide (LiCoO2) particles in a mouse bioassay

Author:

Sironval, V., Reylandt, L., Chaurand, P., Ibouraadaten, S., Palmai-Pallag, M., Yakoub, Y., Ucakar, B., Rose, J., Poleunis, C., Vanbever, R., Marbaix, E., Lison, D., van de Brule, S.

Year:

2018

Bibliographic source:

Archives of Toxicology (2018) 92:1673-1684

Materials and methods

Test guideline

Qualifier:

no guideline followed

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Cobalt lithium dioxide

EC Number:

235-362-0

EC Name:

Cobalt lithium dioxide

Cas Number:

12190-79-3

Molecular formula:

CoO2.Li

IUPAC Name:

cobalt(3+) lithium(1+) dioxidandiide

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited by the study): Lithium cobalt oxide (LiCoO2 or LCO)
- Supplier: MTI Corporation (Richmond, USA)
- State of aggregation: No aggregation
- Particle size distribution: 8.04 ± 0.17 µm (d50, wet); 6.73 ± 0.59 µm (d50, dry)
- Mass median aerodynamic diameter (MMAD): 9.93 ± 2.27 (experimental)
- Surface properties:
- Density: 2.37 g/cm³
- Contaminants: No contaminants detected
- Fine particle fraction (FPF): 7.21 ± 1.55 % of total mass

Test animals

Species:

mouse

Strain:

other: C57BL/6

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Janvier Labs (St Bertevin, France) and local breeding facility (Animalerie Centrale, Universite catholique de Louvain, Brussels, Belgium)
- Age at study initiation: 8 weeks
- Diet: Sterile rodent feed
- Water: Acidified water

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25°C
- Humidity (%): 50%
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

other: oro-pharyngeal aspiration

Vehicle:

other: sterile 0.9% saline solution

Details on exposure:

After anaesthesia with a mix of Nimatek, 1 mg/mouse (Eurovet, Bladel, Nederland) and Rompun, 0.2 mg/mouse (Bayer, Kiel, Germany) given intraperitoneally, 50 μl suspensions of LCO particles, silica, LiCl or NaCl (control groups) were once directly administered by oro-pharyngeal aspiration. Mice were sacrificed 18 h, 3 days and 2 months after administration with an intraperitoneal injection of 12 mg sodium pentobarbital (Certa, Braine-l’Alleud, Belgium).

Doses:

0.5 and 2 mg/mouse

No. of animals per sex per dose:

5 female mice per dose

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 2 months
- Time points of sacrifices: 18 hours, 3 days, and 2 months
- Other examinations performed: histopathology, and bronchoalveolar lavage fluid (BALF) analysis

BALF AND LUNG SAMPLING
Broncho-alveolar lavage (BAL) was performed by cannulating the trachea and infusing the lungs with 1 ml NaCl 0.9%. Whole lungs were then perfused with NaCl 0.9% and excised. Left lobes were placed in 3.65% paraformaldehyde (Sigma-Aldrich, St Louis, Missouri, USA) in phosphate buffered saline (PBS) for later histological analysis, and remaining lobes in liquid nitrogen or lysis buffer for homogenization. Lungs were homogenized on ice with an Ultra-Turrax T25 (Janke and Kunkel, Brussels, Belgium) and stored at − 80 °C. Particle biopersistence was assessed by inductively coupled plasma mass spectrometry (ICPMS), time-of-flight secondary ion mass spectrometry (ToFSIMS),scanning electron microscopy/energy dispersive X-ray spectrometry (SEM-EDX) and X-ray micro fluorescence (μ-XRF) (Supplementary methods). BAL was centrifuged 10 min at 4 °C (240 g). Cell-free supernatant (BALF) was used for biochemical measurements. After resuspension in PBS, total BAL cells were counted in Turch (crystal violet 1%, acetic acid 3%) and cytocentrifuged for differentiation by light microscopy after Diff-Quick staining (200 cells counted, Polysciences, Warrington, UK). Total proteins and lactate dehydrogenase (LDH) activity were assayed on BALF as described previously*.


QUANTIFICATION OF CYTOKINES, HIF‑1Α, HO‑1 AND LUNG COLLAGEN
IL-1β, IL-6, tumour necrosis factor (TNF)-α, IL-1α, transforming growth factor (TGF)-β1 and platelet-derived growth factor (PDGF)-bb were quantified by enzyme-linked immunosorbent assay (ELISA) (DuoSet ELISA, R&D Systems, Minneapolis, USA) in BALF following manufacturer’s instructions. Hypoxia-inducible factor (HIF)-1α (Duo-Set ELISA, R&D Systems) and heme oxygenase (HO)-1
(Immunoset, Enzo Life Sciences, Lausen, Switzerland) were assessed in supernatant (SN) of lung homogenates (centrifuged 10 min at 240g, 4 °C) following manufacturer’s instructions. Collagen deposition was assessed by measuring the OH-proline content in lung homogenates by highpressure liquid chromatography analysis on hydrolyzed lung homogenates as previously described*.


HISTOLOGY
Paraffin-embedded lung sections were stained with Masson’s trichrome blue (total collagen staining), Sirius Red (type I collagen staining) or Perl’s Prussian blue (Fe3+ staining). The stained sections were scanned (Leica SCN400, Brussels, Belgium) and examined with Tissue Image Analysis 2.0 (Leica Biosystems).

*References: Arras M, Huaux F, Vink A, Delos M, Coutelier JP, Many MC, Barbarin V, Renauld JC, Lison D (2001) Interleukin-9 reduces lung fibrosis and type 2 immune polarization induced by silica particles in a murine model. Am J Respir Cell Mol Biol 24:368–375.
Biondi PA, Chiesa LM, Storelli MR, Renon P (1997) A new procedure for the specific high-performance liquid chromatographic determination of hydroxyproline. J Chromatogr Sci 35:509–512

Statistics:

Differences between control and treated groups were evaluated using one-way analysis of variance (ANOVA) followed by a Dunnett’s multiple comparison or a Newman-Keuls multiple comparison test. Statistical significance was considered at P < 0.05.

Results and discussion

Effect levels

Sex:

female

Remarks on result:

other:

Remarks:

Please refer to the field 'Other findings'

Mortality:

No data

Clinical signs:

No data

Body weight:

No data

Gross pathology:

No data

Other findings:

BALF AND LUNG HOMOGENATES
Acute inflammatory response after 3 days (2 mg/mouse): significant induction of lactate dehydrogenase (cytotoxicity, transient), protein amount (total, transient), macrophages (total), neutrophils (total, transient), IL-1β (transient), IL6 (transient), IL-1α (transient), HO-1 (transient), and HIF-1 α

Notably, LiCl dose with a equimolar Li level to 2 mg/mouse LiCl did not result in an acute inflammatory response after 18 hours.

Sub-chronic inflammatory response after 2 months (2 mg/mouse): significantly increased levels of macrophages (total) and HIF-1 α

Sub-chronic fibrotic response after 2 months (2 mg/mouse): significant induction of OH-proline (accumulation of lung collagen)

HISTOLOGY
Sub-chronic fibrotic response after 2 months (2 mg/mouse): -Accumulation of collagen in lungs, -numerous lymphoid nodules around the bronchioles with focal collagen and macrophage accumulation

Particle accumulation: very few particles were visible in lungs

SEM-EDX AND µ-XRF ANALYSIS
Fe co-localised with Co in LCO-treated lungs 2 months after administration. No Fe was, however, present in the original particles alone or after 18 hours, thus suggesting a progressive deposition of endogenous Fe on LCO particles. These observations suggested the formation of ferruginous bodies (FB).

Applicant's summary and conclusion

Conclusions:

The authors considered lithium cobalt oxide to elicit acute to sub-chronic inflammatory response and fibrosis. Further, they stated that the toxicity is presumably dictated by the Co ions.
The investigated mechanistic parameters (i. e. bronchoalveolar lavage (BAL) fluid immune response marker analysis after oro-pharyngeal aspiration of the test material) have no direct value for fulfilling data requirements under the REACH regulation, the methodical setup is not adequately designed for risk assessment purposes of the test substance. The non-physiological route of administration via oro-pharyngeal aspiration is not guideline conform and not suitable to assess acute inhalation toxicity.
Moreover, the publication shows reporting and methodological deficiencies. Test animal were obtained from two different sites. Since it is not stated how the mice are assigned to the test groups, it could be assumed that results are influenced by the origin of the mice, potentially leading to different response due to variation. Description of methodology lacks details. Information on housing of test animals are insufficient. Weights of the test animals are not specified for any point in time. Clinical signs and body weight development are not stated. No information on necropsy. Dosing is not justified. HCD is not specified. Results are not shown for the low dose group. Histological analysis was performed with already lavaged lungs, which could have influenced tissues and particle deposition results. A dose of 2 mg/animal in BL/6 mice induces a massive overload in the lungs, leading to non-physiological response. It needs to be questioned that these results represent meaningful data for hazard and risk assessment purposes.