[No public or meaningful name is available]

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Study plan: 27/07/2018
Draft report: 31/01/2019
Final report: 18/02/2019

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Meets generally accepted scientific standards, well documented and acceptable for assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Objective of study:

bioaccessibility (or bioavailability)

Test guideline

Qualifier:

according to guideline

Guideline:

other: Standard Operating Procedure (SOP) for Bioelution Testing of Metals, Inorganic Metal Compounds, and MetalContaining Complex Materials: Simulated Gastric Fluid

Version / remarks:

Eurometaux, February 16, 2018

Deviations:

yes

Remarks:

The samples were preserved by adding 0.12 mL concentrated HNO3 per 12 mL instead of no preservation in the SOP.

Principles of method if other than guideline:

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”.
The objective of this study was to obtain knowledge about the bio-elution characteristics of G233A-HDE
in simulated gastric fluid. This study has been conducted according to the recommended Standard
Operating Procedure (SOP) for Bioelution Testing of Metals, Inorganic Metal Compounds, and MetalContaining Complex Materials: Simulated Gastric Fluid (Eurometaux, February 16, 2018) which is based
on ASTM D5517-07: Standard Test method for determining the extractability of metals from art materials
- ASTM, 2007 (American Society for Testing and Materials). The extent of dissolution of G233A-HDE as
received, was tested in a simulated gastric fluid at 37 °C and pH 1.5 for 2 hours (0.2 g/L and 2 g/L loading)
at an agitation speed of 100 revolutions per minute (rpm) for 1 hour followed by 1 hour of settling
(without shaking). The bio-elution endpoints were based on the dissolved boron (B), tellurium (Te),
tungsten (W) and zinc (Zn) concentrations obtained after 2 hours of extraction.
The following deviation from the SOP was made:
• The samples were preserved by adding 0.12 mL concentrated HNO3 per 12 mL instead of no
preservation in the SOP.
The study was performed at ECTX. Analysis of the concentrations of dissolved boron, tellurium, tungsten
and zinc has been performed at WLN Business B.V. (The Netherlands), an ISO 17025 accredited laboratory,
as delegated by ECTX.
The measured pH of the test medium at the start of the test was 1.50, i.e. within the specifications of
pH 1.5 ± 0.1. The pH measured in the additional test item vessel at the start of the test was 1.50 for both
loadings, 0.2 g/L and 2 g/L, i.e. within the specifications of pH 1.5 ± 0.1. At the 2 hours sampling point of
the test, the pH in the blank control vessel and the test item vessels was between 1.50 and 1.53.
The temperature of the sampled test solutions including the blank test vessel was between 36.9 °C and
37.0 °C and corresponded to the required test conditions of 37 °C ± 1 °C.
The blank control vessel showed no concentrations of boron, tellurium, tungsten and zinc above the limit
of quantification (i.e. accredited reporting limit) of 5 µg/L B, 0.5 µg/L Te, 0.5 µg/L W and 2 µg/L Zn

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Amorphous boron lithium zinc tellurite

Molecular formula:

Not applicable as UVCB

IUPAC Name:

Amorphous boron lithium zinc tellurite

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

The test item, G233A-HDE as received fulfilled the recommended <100 µm particle size. Therefore, no further preparation of the test item was needed. The Sponsor also provided the required document with the characteristics of the test item (Certificates of Analysis, MSDS).


Element(s) of interest: Boron, tellurium, tungsten and zinc
Chemical analysis regarding the elements of
interest:
1. 1.2 % B
2. 70.44 % TeO2 (i.e. 56.3 % Te)
3. 2.44 % WO3 (i.e. 1.93 % W)
4. 15.23 % ZnO (i.e. 12.2 % Zn)
A certificate of analysis of the tested batch is added
to this study report.
Physical form: Solid, (in the powder form)
Colour: White
Particle size distribution (volume): d10: 0.41 µm
d50: 1.04 µm
d90: 2.18 µm
A certificate of analysis of the particle size
distribution is added to this study report.
Specific surface area (N-BET): 5.30 m²/g
Homogeneous: Yes
Density: n.a.
Storage conditions
Room temperature, in the dark.

Radiolabelling:

no

Test animals

Species:

other: not applicable

Strain:

other: not applicable

Details on test animals or test system and environmental conditions:

Not applicable

Administration / exposure

Route of administration:

other: not applicable

Vehicle:

other: not applicable

Duration and frequency of treatment / exposure:

Not applicable

No. of animals per sex per dose / concentration:

Not applicable

Control animals:

other: not applicable

Positive control reference chemical:

Not applicable

Details on study design:

Test set-up
One PETG Erlenmeyer flask of 250 mL was used for the blank control vessel without test item. For each loading, 0.2 g/L and 2 g/L, 10 mg and 100 mg test item were weighed in triplicate into three separate 250 mL Erlenmeyer flasks. For each loading, a fourth replica with test item (named replica X) was prepared only for the measurement of the initial pH (at the start of the test). This additional test vessel was set up to avoid cross contaminations and did not have further utility in the test. 50 mL of extraction fluid (at 37 °C ± 1 °C) was added to the blank control vessel and each test item vessel resulting in a final loading of 0.2 g/L and 2 g/L, respectively. After covering the vessels with a screwcap and swirling the flasks to mix the test item and the medium, the flasks were placed into a thermostatic orbital shaker (37 °C ± 1 °C) at an agitation rate of 100 revolutions per minute for one hour. After that, the flasks were settled at 37 °C ± 1 °C for another hour.
Sampling during the test
The following sampling procedure was used at the sampling time to collect and preserve the samples for ICP-MS analyses: At the end of the incubation period (2 hours = 1 hour shaking at 100 rpm and 1 hour settling), each vessel was swirled to homogenise the solution prior to sampling, in order to avoid a concentration gradient after settling. Sampling was performed for the blank control and each test item vessel as indicated in the sampling scheme below. In each test vessel, a 12-mL sample was taken twice with a 12 mL syringe from the test vessel at a depth of two-thirds of the supernatant. The samples were filtrated through a 0.2 µm syringe filter and transferred to uniquely labelled 15-mL PP sample tubes. The samples were preserved by adding 0.12 mL concentrated HNO3 per 12 mL. The samples were covered (to avoid evaporation and concentration) and stored at room temperature in the dark until shipment to the Test Site (see 3.3 Test Site identification).
Observations and measurement of temperature and pH
At the start of the test (0h), pH was measured in the additional (fourth) replica with test item of each loading (named replica X). The addition of the test item to the test system had no significant influence on the initial pH of the test system. After sampling, temperature and pH were measured in the remaining blank control and test solutions after sampling.
Shipment of the samples to the Test Site
The samples and datasheet DSH026 Sample Transfer (external), which contains a chain of custody and a detailed sample list with storage conditions were transferred to the Test Site (see 3.3 Test Site identification) in an appropriate manner according to SOP AB-14 (see 5.2.1 SOPs of ECTX).
Analyses of the samples by the Test Site
The determination of dissolved boron, tellurium, tungsten and zinc concentrations in the blank control and test item vessels were carried out at the Test Site (see 3.3 Test Site identification), using an 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 ECTX (Ecotoxicology and Biodegradation) TB-2018-R-TB X02A-183 section IV – page 17 from 46 accredited reporting limit with an accuracy of 90 % and 110 % is 0.5 µg/L for tellurium and tungsten, 5 µg/L for boron, and 2 µg/L for zinc. 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.

Details on dosing and sampling:

Not applicable

Statistics:

Not applicable

Results and discussion

Preliminary studies:

Not applicable

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Not applicable

Details on distribution in tissues:

Not applicable

Details on excretion:

Not applicable

Metabolite characterisation studies

Metabolites identified:

not measured

Details on metabolites:

Not applicable

Any other information on results incl. tables

Bioaccessibility results

The temperature of the test solutions including the blank test vessel at the 2-hours sampling point was between 36.8 °C and 37.0 °C, which was in line with the test conditions of 37 °C ± 1 °C.

The measured pH of the test medium at the start of the test was 1.50, i.e. within the specifications of pH 1.5 ± 0.1. The pH measured in the additional test item vessel at the start of the test was 1.50 for both loadings, 0.2 g/L and 2 g/L i.e. within the specifications of pH 1.5 ± 0.1. At the 2 hours sampling point of the test, the pH in the blank control vessel and the test item vessels was between 1.50 and 1.53.

The blank control vessel showed no concentrations of boron, tellurium, tungsten and zinc above the limit of quantification (i.e. accredited reporting limit) of 5 µg/L B, 0.5 µg/L Te, 0.5 µg/L W and 2 µg/L Zn.

The following observations could be made in the test vessels with a loading of 0.2 g/L G233A-HDE:

Test Substance	Gastric Bioaccessibility- 2 hours (mg/l)	surface area (m2/g)	release per surface (mg/m2)	Gastric Bioaccessibility- 2 hours as % released of total content
Boron	2.26	5.30	2.1	94
Tellurium	39.9	5.30	38	35
Tungsten	0.736	5.30	0.69	19
Zinc	19.7	5.30	19	80

The following observations could be made in the test vessels with a loading of 2 g/L G233A-HDE:

Test Substance	Gastric Bioaccessibility- 2 hours (mg/l)	surface area (m2/g)	release per surface (mg/m2)	Gastric Bioaccessibility- 2 hours as % released of total content
Boron	17.4	5.30	1.6	72
Tellurium	165	5.30	16	15
Tungsten	0.580	5.30	0.055	1.5
Zinc	88.7	5.30	8.4	36

Applicant's summary and conclusion

Conclusions:

Acute Classification assessment – Inhalation route:
By solving the equation, an ATEmix of 14.54 (mg/l) is obtained. This value is higher than the threshold of 5 (mg/l) set by CLP Regulation, thus, according to the Acute INH classification criteria, this classification does not apply.

Reprotox assessment:
The effective concentation (Bioaccessible concentration-INH route/GCL) is far above (34.4%) than the Generic Concentration Limit of 0,3% set by the CLP Regulation triggering Reproductive toxicity Cat.1B. Therefore according to the CLP classification criteria, this classification shall apply.

Executive summary:

Acute Classification assessment – Inhalation route, linked to TeO2

HH Assessment applying the most conservative approach for deriving hazard classification: Application of the CLP mixture rules.

According to section 3.1.3.6.2, In order to ensure that classification of the mixture is accurate, and that the calculation need only be performed once for all systems, sectors, and categories, the acute toxicity estimate (ATE) of ingredients shall be considered as follows:

3. include ingredients with a known acute toxicity, which fall into any of the acute hazard categories shown in Table 3.1.1; (our case);


4. ignore ingredients that are presumed not acutely toxic (e.g., water, sugar);

ignore components if the data available are from a limit dose test (at the upper threshold for Category 4 for the appropriate route of exposure as provided in Table 3.1.1) and do not show acute toxicity.

By solving the equation, an ATE_mix of 14.54 (mg/l) is obtained. This value is higher than the threshold of 5 (mg/l) set by CLP Regulation, thus, according to the Acute INH classification criteria, this classification does not apply.

Reprotox assessment, linked to TeO2.

HH Assessment applying the most conservative approach for deriving hazard classification: Application of the CLP mixture rules.

Table 3.7.2 of CLP Regulation which establishes the Generic concentration limits of ingredients of a mixture classified as reproduction toxicants or for effects on or via lactation that trigger classification of the mixture.

The effective concentation (Bioaccessible concentration/GCL) is far above (34.4%) than the Generic Concentration Limit of 0,3% set by the CLP Regulation triggering Reproductive toxicity Cat.1B, being the Tellurium dioxide the main driver.

Therefore according to the CLP classification criteria, this classification shall apply.